DiSA_ttivo © DiSA - Università degli studi di Foggia - Agraria ...

Sinigaglia Milena 

Altieri Clelia 

Beneduce Luciano 

Corbo Maria Rosaria 

Massa Salvatore 

Spano Giuseppe 

Vernile Anna 

La Notte Ennio 

Del Nobile Matteo Alessandro 

Di Luccia Aldo 

Baiano Antonietta 

Lamacchia Carmela 

Conte Amalia 

Pati Sandra 

la Gatta Barbara 

Lecce Lucia 

Sepielli Grazia 

Laverse Janine 

Frisullo Pierangelo 

DiSA_ttivo


sommario 6 Disa: le persone 

13 Disa: rassegna delle ricerche (2007-2010) 

14 Studio degli effetti metabiotici tra muffe 

e patogeni in alimenti 

17 Effetti delle alte pressioni di omogeneizzazione 

su microrganismi di interesse alimentare 

in relazione a fattori intrinseci ed estrinseci 

20 Implicazioni di biofilm microbici 

nell’industria alimentare 

22 Caratterizzazione della microflora 

di formaggi tradizionali pugliesi 

24 Modellazione dello sviluppo microbico 

negli alimenti e nei processi fermentativi 

29 Aspetti microbiologici connessi ai prodotti 

di quarta gamma 

33 Alicyclobacillus acidoterrestris: 

un alterante emergente 

38 Antimicrobici naturali: studio delle proprietà 

antimicrobiche e delle potenzialità 

applicative in alimenti 

44 Bioattività di acidi grassi nei confronti 

di microrganismi alteranti e patogeni 

47 Studio della microflora protecnologica 

e alterante nelle olive da mensa 

49 Tolleranza agli stress abiotici 

in microrganismi starter e probiotici 

55 Identificazione e tracciabilità 

di microrganismi patogeni e tolleranza 

agli stress in alimenti minimamente 

processati 

58 Identificazione e caratterizzazione 

di starter enologici e analisi dei meccanismi 

coinvolti nella produzione di ammine 

biogene in vino 

63 Innovazione nella produzione di pasta 

alimentare (fresca e secca) mediante l’utilizzo 

di microrganismi (batteri) selezionati 

66 Ricerca e caratterizzazione di batteri 

patogeni emergenti negli alimenti 

e nelle acque 

69 Caratterizzazione della microflora 

in formaggi tipici 


sommario 73 Caratterizzazione degli alimenti ed effetti 

dei processi tecnologici sulla materia 

prima mediante studi proteomici, lipidomici 

e metabolomici 

85 Studio della componente polifenolica 

e volatile nelle uve e nei vini 

89 Studio della componente fenolica delle matrici 

alimentari e misura dell’attività antiossidante 

97 Studio delle produzioni alimentari tipiche 

attraverso l’applicazione di tecniche innovative 

100 Applicazione degli standard di certificazione 

101 Studio degli effetti dei processi tecnologici 

sulle proprietà strutturali e funzionali 

delle proteine degli alimenti 

104 Studio delle relazioni esistenti tra 

le proteine e le proprietà tecnologiche 

di sfarinati di frumento e sfarinati 

gluten free 

107 Film edibili 

111 Packaging attivo 

116 Confezionamento di Prodotti 

di IV Gamma 

124 Tecnologie innovative per prolungare 

la shelf life di alimenti freschi 

134 Alimenti Funzionali 

138 Caratterizzazione chimico-fisica 

di matrici alimentari 


Prefazione 

Il Dipartimento di Scienze degli Alimenti (DiSA) 

è stato costituito con D.R. del 5/11/2002 nel con- 

testo della Facoltà di Agraria per promuovere e 

coordinare ricerca scientifica e impegno didattico 

nei settori delle tecnologie e della microbiologia 

degli alimenti. 

La ricerca viene finanziata da enti pubblici nazionali 

e internazionali; inoltre la presenza di docenti con 

qualificata e comprovata esperienza nel settore 

alimentare agevola il non sempre facile rapporto 

con le industrie alimentari del territorio favorendo 

l’attivazione di numerosi contratti di convenzione 

con le imprese private. 

Il Dipartimento dispone di laboratori di base e spe- 

cializzati per analisi chimiche, fisiche e microbiolo- 

giche. Possiede impianti pilota per la simulazione di 

operazioni e processi. Possiede inoltre un laboratorio 

di analisi sensoriale, dove vengono organizzati ed 

addestrati panel e un laboratorio attrezzato per 

biologia molecolare. 

L’attività di ricerca riguarda prevalentemente i 

seguenti temi: messa a punto di modelli matema- 

tici per la previsione della shelf-life degli alimenti; 

analisi chimica degli alimenti, con particolare riferi- 

mento alle filiere del vino e dell’olio e al settore lat- 

tiero-caseario; sviluppo e messa a punto di materiali 

polimerici per l’imballaggio alimentare; processi 

di trasformazione di conserve e semiconserve di 

origine animale e vegetale e loro effetti sulla qua- 

lità dei prodotti; microbiologia e biotecnologie ali- 

mentari e agro-ambientali; biologia molecolare dei 

microrganismi. I risultati conseguiti sono dimostrati 

attraverso le numerose pubblicazioni internazionali 

e nazionali ed i brevetti ottenuti. 

Lo staff del dipartimento conta 15 docenti e ricerca- 

tori, 6 tecnici, 2 amministrativi e numerosi dotto- 

randi, borsisti e laureati frequentatori. 

Si tratta quindi di un Dipartimento con un numero di 

docenti tale che, nell’ottica di una razionalizzazione 

dei Centri di spesa, è destinato alla “disa…attiva- 

zione”. Lungi da me la volontà di criticare una logica 

di risparmio, mi piacerebbe tuttavia che tale logica 

fosse applicata non meramente al numero di docenti 

ma, anche e soprattutto, alla loro operosità. 

Da qui è nata l’idea di una raccolta che mostrasse le 

attività del nostro “DiSA..ttivando” dipartimento, una 

sorta di canto del cigno del nostro DiSA fatto di per- 

sone attive …. fatto di noi. 

Il direttore pro-tempore 

Prof.ssa Milena Sinigaglia 

5 


Disa: 

le Persone 

Cognome: Sinigaglia 

Nome: Milena 

Ruolo: Professore Ordinario 

Cell: 320 43 94 679 

Tel.: 0881 589 233 

m.sinigaglia@unifg.it 

6 


Cognome: Altieri 

Nome: Clelia 

Ruolo: Ricercatore Confermato 

Cell: 320 43 94 810 

Tel.: 0881 589 235 

c.altieri@unifg.it 

Cognome: Beneduce 

Nome: Luciano 


Cell: 338 37 26 208 

Tel.: 0881 589 234 

l.beneduce@unifg.it 

Cognome: Corbo 

Nome: Maria Rosaria 


Cell: 320 43 94 680 

Tel.: 0881 589 232 

m.corbo@unifg.it 

7 


Cognome: Massa 

Nome: Salvatore 


Cell: 349 62 95 987 

Tel.: 0881 589 361 

s.massa@unifg.it 

Cognome: Spano 

Nome: Giuseppe 

Ruolo: Professore Associato 

Cell: 338 32 14 987 

Tel.: 0881 589 303 

g.spano@unifg.it 

Cognome: Vernile 

Nome: Anna 

Ruolo: Tecnico di Laboratorio 

Cell: 328 82 32 565 

Tel.: 0881 589 234/352 

a.vernile@unifg.it 

8 


Cognome: La Notte 

Nome: Ennio 


Cell: 320 43 94 517 

Tel.: 0881 589 217 

e.lanotte@unifg.it 

Cognome: Del Nobile 

Nome: Matteo Alessandro 


Cell: 320 43 94 676 

Tel.: 0881 589 242 

ma.delnobile@unifg.it 

Cognome: Di Luccia 

Nome: Aldo 

Ruolo: Professore Associato 

Cell: 347 12 63 965 

Tel.: 0881 589 243 

a.diluccia@unifg.it 

9 


Cognome: Baiano 

Nome: Antonietta 


Tel.: 0881 589 117 

a.baiano@unifg.it 

Cognome: Lamacchia 

Nome: Carmela 


Cell.: 349 61 80 642 

Tel.: 0881 589 243 

c.lamacchia@unifg.it 

Cognome: Conte 

Nome: Amalia 

Ruolo: Ricercatore non Confermato 

Cell.: 349 05 33 439 

Tel.: 0881 589 240 

a.conte@unifg.it 

10 


Cognome: Pati 

Nome: Sandra 

Ruolo: Ricercatore non Confermato 

Tel.: 0881 589 240 

s.pati@unifg.it 

Cognome: la Gatta 

Nome: Barbara 

Ruolo: Tecnico di laboratorio 

Tel.: 0881 589 350 

b.lagatta@unifg.it 

Cognome: Lecce 

Nome: Lucia 


Tel.: 0881 589 246 

l.lecce@unifg.it 

11 


Cognome: Sepielli 

Nome: Grazia 


Cell.: 347 68 45 359 – 328 55 56 389 

Tel.: 0881 338 575 

g.sepielli@unifg.it 

Cognome: Laverse 

Nome: Janine 


Cell: 339 82 49 511 

Tel.: 0881 338 575 

j.laverse@unifg.it 

Cognome: Frisullo 

Nome: Pierangelo 


Cell: 329 63 11 546 

Tel.: 0881 338 575 

p.frisullo@unifg.it 

12 


Disa: 

rassegna 

Delle ricerche 

(2007-2010) 

13 


stuDio Degli effetti metabiotici 

tra muffe e Patogeni in alimenti 

comPosizione Del gruPPo Di ricerca 

Milena Sinigaglia, Professore Ordinario 

Claudio Ciccarone, Professore Associato 

Clelia Altieri, Ricercatore Confermato 

Antonio Bevilacqua, Borsista post-doc 

Persona Da contattare 


Dipartimento di Scienze degli Alimenti (DISA), 

Facoltà di Agraria, Via Napoli 25, 71100 Foggia; 

tel. 0881 589 233; m.sinigaglia@unifg.it 

Parole chiave 

ATTIVITà PROTEOLITICA 

SALMOnELLA 

ESChERiChiA COLi O157:H7 

FuSARiuM SPP. 

breve Descrizione 

La metabiosi è definita come l’interazione fra due microrganismi, in cui uno si 

avvantaggia del metabolismo dell’altro, per la produzione di metaboliti utili e/o 

la rimozione di composti tossici. Il gruppo di ricerca si è occupato dell’interazione 

di specie afferenti ai generi Aspergillus, Fusarium e Penicillium, studiandone 

l’interazione con alcuni patogeni alimentari, quali Salmonella sp., Escherichia coli 

O157:H7 e Listeria monocytogenes. 

In una prima fase è stata studiata l’attività proteolitica di 18 ceppi fungini, 

appartenenti ai generi citati, sia in un sistema modello che in succo di pomodoro. 

Il genere Fusarium, con qualche eccezione, mostrava l’attività proteolitica più 

spiccata, seguito dal genere Aspergillus, caratterizzato da un proteolisi blanda e 

specie-dipendente; tutti i penicilli, al contrario, non erano in grado di idrolizzare le 

proteine del pomodoro. L’attività proteolitica, causando un innalzamento del pH del 

mezzo, esercitava un effetto protettivo su Salmonella sp. coinoculata nel mezzo. 

La metabiosi, tuttavia, non dipende solo dall’innalzamento del pH del mezzo, 

ma anche da altri fattori non ancora del tutto chiariti (produzione di metaboliti 

non alcalini, idrolisi delle pectine), come dimostrato nel caso dell’interazione E. 

coli O157:H7/F. oxysporum-F. proliferatum-F. avenaceum su pomodori di quarta 

gamma. Le muffe, infatti, rallentavano la cinetica di morte del patogeno, sebbene 

il pH del mezzo non cambiasse significativamente. 

Dati Pubblicati inerenti il tema Di ricerca 

1 

Proteolytic activity of moulds and their metabiotic association with 

Salmonella sp. in a model system. Cibelli, F., Ciccarone, C., Altieri, C., 

Bevilacqua, A., Sinigaglia, M. Journal of Food Protection, 71, 2129-2132 

(2008). Impact Factor: 1, 886. 

The aim of this study was to investigate the proteolytic ability of some strains of 

aspergilli, fusaria and penicilli and the metabiotic effect of Fusarium oxysporum 

14 




and Penicillium expansum on Salmonella sp. The proteolytic activity of the target 

molds was determined on Tomato Juice Agar (TJA) and Tomato juice, whereas the 

metabiotic effect of F. oxysporum and P. expansum on Salmonella sp. was assessed 

in a model system consisting of Tryptone Soya Broth (TSB) added with different 

amounts of Tomato Juice. Fusaria, some aspergilli and one strain amongst penicilli 

showed a proteolytic activity on TJA. In addition, Salmonella survival was enhanced 

in TSB+20 or 50% of tomato juice, when the model system had been inoculated 

with F. oxysporum. 

2 Metabiotic 

effects of Fusarium spp. on Escherichia coli O157.H7 and Listeria 

monocytogenes on raw portioned tomatoes. Bevilacqua, A., Cibelli, F., 

Cardillo, D., Altieri, C., Sinigaglia, M. Journal of Food Protection, 71, 1366- 

1371 (2008). Impact Factor: 1,886. 

The metabiotic effects of Fusarium proliferatum, F. avenaceum and F. oxysporum 

on Escherichia coli O157:H7 and Listeria monocytogenes in fresh tomatoes were 

investigated. Tomatoes were pre-inoculated with the molds and incubated at 15°C 

for 7 days; then, they were inoculated separately with the pathogens, packaged 

in air and modified atmosphere (MA, 5% O 2 ; 30% CO 2 ; 65% N 2 ) and stored at 4, 

8 and 12°C for 9 days. The cell loads of pathogens and lactic acid bacteria (LAB) 

and the pH were evaluated periodically. The data were modeled through some 

different mathematical models, to assess the shoulder length (S.L., time before 

the beginning of the exponential death phase), the 1-log reduction time (δ) and 

the pathogen death time (δ stand ). The pre-inoculation of tomatoes with the molds 

enhanced the survival of E. coli O157:H7, by prolonging S.L. and δ parameters; 

this effect, however, was not observed for L. monocytogenes. pH values did not 

undergo significant changes within the storage time and the LAB increased from 5 

to 7 log CFU/g, without significant differences amongst the storage temperatures 

or the packaging atmospheres. The results of this research showed that the use of 

fresh tomatoes colonized by fusaria (even if the contamination is not visible) could 

15 




increase significantly the risk of outbreaks due to some pathogens, that could be 

on the surface of fruits and vegetables as a results of a cross-contamination at 

home or for not-correct post-harvest operations. 

3 Modelling 

the survival of Escherichia coli O157:H7 on raw portioned tomatoes, 

inoculated with Aspergillus fumigatus and Emericella nidulans. Cardillo, D., 

Bevilacqua, A., Cibelli, F., Altieri C., Sinigaglia M. Journal of Biomedicine and 

Biotechnology (2009), article ID 184130. Impact Factor: 2,563. 

The metabiotic interactions occurring among two fungi (Aspergillus fumigatus 

and Emericella nidulans) and Escherichia coli O157:H7 on raw portioned tomatoes 

were studied. Tomatoes, pre-inoculated with the moulds and inoculated with the 

pathogen, were packaged in air and stored at 4, 8 and 12°C for 9 days; pathogen 

cell number and pH were monitored throughout the storage and the data 

were modeled using three different equations (Geeraerd, Weibull and modified 

Weibull), to assess the shoulder length, the 1-log reduction time and the death 

time. Both A. fumigatus and E. nidulans increased the survival of E. coli O157:H7 

through the prolongation of the shoulder length; in contrast, the death time was 

significantly increased. 

The results of this paper suggested that the metabiotic interactions aspergilli/E. 

coli O 157:H7 could be of public concern, as the consumption of tomatoes (or 

other fruits and vegetables) contaminated both by the moulds and the pathogen 

is a possible scenario. 

16 


effetti Delle alte Pressioni 

Di omogeneizzazione su 

microrganismi Di interesse 

alimentare in relazione a fattori 

intrinseci eD estrinseci 



Maria Rosaria Corbo, Ricercatore Confermato 



Milena Sinigaglia, Professore Ordinario, 




Parole chiave 

OMOGENEIzzAzIONE 

MICRORGANISMI ALTERANTI 

BEVANDE ACIDE 

TRATTAMENTI MULTI-STEP 

SCALE DI SUSCETTIBILITà 


Proposta dal 1900, l’omogeneizzazione (High Pressure Homogenization, HPH) 

è una tecnologia emergente, utilizzata dall’industria alimentare, cosmetica e 

farmaceutica, per la preparazione di emulsioni e l’inattivazione di microrganismi 

patogeni e alteranti gli alimenti. L’efficacia di un trattamento HPH dipende da tre 

diversi tipi di fattori: parametri di processo (pressione, temperatura, numero di 

passaggi nell’omogeneizzatore e tipo di valvola), stato fisiologico delle cellule e 

reologia del fluido. I trattamenti HPH sono stati utilizzati come metodo alternativo 

per l’inattivazione dei microrganismi alteranti le bevande acide, quali Alicycloacillus 

acidoterrestris, Lactobacillus plantarum, L. brevis, Bacillus coagulans, Saccharomyces 

bayanus, Rhodotorula bacarum e Pichia membranaefaciens, Fusarium oxysporum, 

Penicillium expansum e Emericella nidulans. In particolare, nel caso di A. acidoterrestris 

i dati sperimentali hanno mostrato come trattamenti HPH a 150 MPa siano in grado 

di garantire una riduzione significativa delle cellule; al contrario la germinazione delle 

spore può essere evitata combinando il trattamento HPH con benzoato di sodio. 

I risultati ottenuti per gli altri microrganismi alteranti dei succhi hanno mostrato 

come in generale i lieviti siano estremamente sensibili. I batteri e le muffe, al 

contrario, appaiono più resistenti; nel caso dei batteri lattici è necessario utilizzare 

trattamenti HPH multi-step, applicando in ciascuna fase una pressione di 150 MPa, 

per ottenere una riduzione della popolazione microbica di circa 1 log ufc/ml. 


1 

Effects of high pressure homogenization on the survival of 

Alicyclobacillus acidoterrestris in a laboratory medium. Bevilacqua, A., 

Cibelli, F., Corbo, M.R., Sinigaglia, M. Letters in Applied Microbiology, 45, 

382-386 (2007). Impact Factor: 1,623. 

Aims - This study was aimed to investigate the effectiveness of high pressure 

homogenization (HPH) against Alicyclobacillus acidoterrestris. 

17 







Methods and Results - The susceptibility of three different strains of A. 

acidoterrestris (DSMz 2498, c4 and c8) to HPH (500-1700 bar) was studied. The 

experiments were performed in a laboratory medium (Malt Extract broth) on cells 

and spores. HPH caused a significant reduction of the initial cell number (1-2 Log 

CFU/ml at the highest pressures) in γ4 and DSMz 2498 strains, whereas the effect 

on the spores was less significant. 

Conclusions - This study showed that the susceptibility of A. acidoterrestris to HPH 

was strain-dependent: DSMz 2498 seemed the most susceptible strain, whereas 

c8 was the most resistant one. 

Significance and impact of the study - The results of our study will provide useful 

information on the sensitivity of an emerging spoilage microorganism, as A. 

acidoterrestris, to HPH. 

2 Effects 

of the high pressure of homogenization on some spoiling 

microorganisms, representative of fruit juices microflora, inoculated in 

saline solution. Bevilacqua, A., Costa, C., Corbo, M.R., Sinigaglia, M. Letters 

in Applied Microbiology, 48, 261-267 (2009). Impact Factor: 1,623. 

Aims - This study was aimed to investigate the effects of a high-pressure 

homogenization (HPH) treatment on some microorganisms, involved in the 

spoilage of fruit juices. 

Methods and results - Lactobacillus plantarum, Lact. brevis, Bacillus coagulans 

cells, Saccharomyces bayanus, Pichia membranifaciens and Rhodotorula bacarum 

were separately inoculated in a saline solution (0.9% NaCl); the initial inoculum 

was ca. 5 log CFU mL -1 . Then, the samples were processed through a homogenizer 

at 10-150 MPa for 1, 2 or 3 times. Yeasts were completely inactivated at 50-110 

MPa with a single pass treatment, while lactic acid bacteria counts were reduced 

of approximately 1 log CFU mL -1 after a 3-steps HPH processing. 

Conclusions - Yeasts were the most sensitive microorganisms, followed by B. 

coagulans. On the other hand, lactic acid bacteria appeared resistant to HPH. 

18 







Significance and impact of the study - The results of this study provided some 

useful information on the susceptibility of microflora of juices to homogenization; 

moreover, they suggested that HPH could be used successfully to inactivate yeasts. 

3 Use 

of high pressure homogenization as a mean to control the growth 

of foodborne moulds in tomato juice. Corbo, M.R., Bevilacqua, A., 

Campaniello, D., Ciccarone, C., Sinigaglia, M. Food Control, DOI: 10.1016/ 

j.foodcont.2010.04.023 

The effect of High Pressure Homogenization (HPH) on Fusarium oxysporum, 

Emericella nidulans and Penicillium italicum was studied. Moulds were inoculated 

in Tween 80, tomato juice and diluted tomato juice (ratio juice/water: 1:3) and 

homogenized with different pressure levels for 1, 2 or 3 times (between 30 and 

150 MPa): the number of conidia decreased with the progressive increase of the 

pressure applied. This effect was more evident when a multi-step process was 

applied; furthermore the decrease of the fluid viscosity in tomato juice reduced 

the effectiveness of homogenization. Finally, a discolouration of inoculated P. 

italicum was observed probably due to the stress caused by high pressure levels. 

19 


imPlicazioni Di biofilm microbici 

nell’inDustria alimentare 





Barbara Speranza, Assegnista di ricerca 






Parole chiave 

CELLULE PLANCTONICHE 

CELLULE SESSILI 

BIOFILM UTILI 

NSLAB 

ACETOBATTERI 


Il termine biofilm viene utilizzato per definire un’associazione di microrganismi 

adesi a una superficie e intrappolati all’interno di sostanze polimeriche extracellulari 

(EPS) da essi stessi prodotte. Nell’industria alimentare l’adesione di microrganismi a 

prodotti e/o a superfici a contatto con i prodotti durante il processo può comportare 

seri problemi igienico-sanitari nonché perdite economiche. Tuttavia, non sempre i 

biofilm sono nocivi; ad esempio, in molti ambienti naturali la qualità dell’acqua è 

garantita proprio dal metabolismo di biofilm microbici in grado di biodegradare 

molti composti tossici e numerose sostanze inquinanti. Inoltre, i biofilm microbici 

costituiscono un esempio naturale di immobilizzazione cellulare, utile per migliorare 

l’efficienza dei processi di fermentazione. Il gruppo di ricerca ha focalizzato 

l’attenzione proprio sui biofilm utili, prodotti da batteri acetici e da batteri lattici 

non starter. In particolare, nel caso dei batteri acetici, come target microbico è stato 

utilizzato un ceppo di Acetobacter cerevisiae isolato da aceto balsamico tradizionale; 

i dati hanno mostrato come il ceppo fosse in grado di formare biofilm solo in assenza 

di acido acetico. 

I biofilm di NSLAB (Lactobacillus plantarum, L. casei, L. curvatus, L. paracasei), 

invece, si sono dimostrati un utile mezzo per ritardare lo sviluppo di Listeria 

monocytogenes nei formaggi freschi. 


1 

Survival of planktonic and sessile Acetobacter cerevisiae. Altieri, C., Speranza, 

B., Cardillo, D., Sinigaglia, M. International Journal of Food Science and 

Technology, 43, 484-489 (2008). Impact Factor: 1,065. 

This study was designed to investigate the individual or combined effects 

of ethanol (4–12%), glucose (10–30%) and acetic acid (0–6%) on survival 

of planktonic and sessile cells of one strain belonging to the new species 

Acetobacter cerevisiae isolated from traditional balsamic vinegar. The results 

20 


imPlicazioni Di biofilm microbici 

nell’inDustria alimentare 

confirm that the planktonic cells’ survival was negatively affected by ethanol 

and by acetic acid; moreover, there was a positive correlation between survival 

and glucose concentration. On the contrary, biofilm formation was observed 

only in the absence of acetic acid, which was the most affecting variable on 

the natural ability of cells to attach and grow on surfaces. The obtained results 

supply new information about the physiology of a novel Acetobacter species 

and about its natural capability to develop biofilms so that improved control 

over balsamic vinegar fermentation could be implemented in the future. 

2 Non 

starter lactic acid bacteria biofilms: a means to control the growth of 

Listeria monocytogenes in soft cheese. Speranza, B., Sinigaglia, M., Corbo, 

M.R. Food Control , 20, 1063-1067 (2009). Impact Factor: 1,823. 

The possibility to consider non starter lactic acid bacteria (NSLAB) biofilms as 

a means to control the growth of Listeria monocytogenes in soft cheeses was 

evaluated. In particular, the aptitude to form biofilm of four NSLAB strains 

(Lactobacillus plantarum DSM1055, Lactobacillus casei DSM20011, Lactobacillus 

curvatus DSM20019 and Lactobacillus paracasei DSM20207) was investigated. 

All tested strains were able to form biofilm on stainless steel and the highest 

quantities were produced when NSLAB were present simultaneously (about 6 Log 

CFU cm -2 ). Then, experimental cheeses were made in presence of chips containing 

7-days NSLAB biofilms and inoculated with L. monocytogenes (about 2 Log CFU 

g -1 ). Results demonstrated that NSLAB biofilms can be considered a useful means 

to delay the growth of L. monocytogenes in soft cheeses: the maximum cell load 

attained at the stationary phase was about 6 Log CFU g -1 in experimental cheeses 

against about 7 Log CFU g -1 observed in control samples. 

21 


caratterizzazione Della 

microflora Di formaggi 

traDizionali Pugliesi 




Daniela D’Amato, Borsista post-doc 






Parole chiave 

CANESTRATO PUGLIESE 

TRATTAMENTI SUPERFICIALI 

CANESTRELLO PUGLIESE 

CARATTERIzzAzIONE 

MICROBIOLOGICA 


Le caratteristiche chimico-fisiche e microbiologiche di formaggi tradizionali 

pugliesi, quali il Canestrato Pugliese DOP e il Canestrello, rivestono grande 

importanza nella qualità di tali prodotti. Nel caso del Canestrato Pugliese, dopo 

una fase preliminare di studio della microflora autoctona, l’attenzione è stata 

focalizzata sugli effetti di trattamenti superficiali con pimaricina, olio di oliva 

e “fumo liquido” sulla microflora fungina, sulle caratteristiche chimico-fisiche 

del formaggio e sull’attività proteolitica e lipolitica nel corso della maturazione. 

Mentre come l’uso di olio di oliva favoriva limitatamente lo sviluppo delle muffe, al 

contrario la pimaricina e il “fumo liquido” ne rallentavano la crescita. Relativamente 

alla caratterizzazione della microflora del Canestrello Pugliese, oltre allo studio 

delle cinetiche di sviluppo della microflora autoctona durante la maturazione 

(batteri lattici mesofili e termofili, Enterobacteriaceae, micrococchi e stafilococchi), 

è stata isolata, identificata e caratterizzata la microflora lattica. In particolare è stata 

studiata la capacità di produrre ammine biogene (tiramina, cadaverina, putrescina) 

sia in vivo che durante la maturazione del formaggio. 


1 

Effects of different surface treatments on ripening of Canestrato Pugliese 

cheese. Quinto, M., Spadaccino, G., Rotunno, T., Sinigaglia, M., Ciccarone, C., Fox, 

P.F. International Dairy Journal, 17, 1240-1247 (2007). Impact Factor: 2,198. 

The effect of different surface treatments (with olive oil, liquid smoke or pimaricin) 

on a number of physico-chemical characteristics (namely, moisture, NaCl content, 

pH, total and soluble nitrogen fractions, lipolytic processes and mould production) 

of the core and the rind zones of Canestrato Pugliese cheese were studied during 

the whole ripening period (100 days). Treated cheese data were statistically 

compared with data obtained for untreated cheese. While for the core zone 

proteolytic and lipolytic processes, together with all the other physico-chemical 

22 



microflora Di formaggi 

traDizionali Pugliesi 

properties studied, remained unaffected by the surface treatment, in the layer 

close to the rind these were influenced significantly by the surface treatment in a 

manner correlated with the fungal growth. 

2 Microbiological 

and physicochemical characterisation of Canestrello 

Pugliese cheese, a traditional Apulian cheese made from cows’ milk. 

D’Amato, D., Sinigaglia, M., Corbo, M.R. Australian Journal of Dairy 


A microbiological and physicochemical characterization of Canestrello Pugliese 

cheese, obtained from cows’ milk by heating the curd in hot whey, according 

to a traditional protocol, is reported. The numbers of total mesophilic bacteria, 

and mesophilic and thermophilic lactic acid bacteria were high during ripening, 

while at the end of ripening, the cheese contained less than 1.0 log cfu/g of total 

and fecal coliforms and Enterobacteriaceae. The evolution of the major physico- 

chemical parameters investigated during ripening contributed to the decrease 

in Enterobacteriaceae, total and fecal coliforms, and micrococci but they did 

not affect the growth of the microflora adapted to the hostile conditions of the 

ripened cheese environment. 

Non-Starter Lactic Acid Bacteria (NSLAB) were essentially comprised of 

Lactobacillus rhamnosus, Lactobacillus plantarum, Lactobacillus delbrueckii ssp. 

lactis and Lactococcus lactis ssp. lactis. 

Only two strains of 84 lactic acid bacteria tested exhibited tyrosine decarboxylase 

activity and five strains were able to decarboxylate arginine. Low concentrations 

of tyramine, cadaverine and putrescine were found in cheese samples at the end 

of ripening. 

23 


moDellazione Dello sviluPPo 

microbico negli alimenti 

e nei Processi fermentativi 



Matteo Alessandro Del Nobile, Professore Ordinario 











Parole chiave 

MODELLI MATEMATICI 

PROCESSI FERMENTATIVI 

CICLO DI SVILUPPO MICROBICO 

BIOMASSA 


La microbiologia predittiva si propone di determinare le relazioni matematiche 

che correlino lo sviluppo di microrganismi durante la conservazione di un 

alimento alle variabili fisiche, chimiche e biologiche che lo caratterizzano come 

ecosistema. L’impiego di piani sperimentali a più variabili (CCD o Centroide) ha 

consentito la valutazione dell’effetto dei nutrienti sulla produzione di biomassa 

e la capacità acidificante di un ceppo probiotico di Lactobacillus plantarum. 

Inoltre, sono stati messi a punto modelli meccanicistici per la previsione del ciclo 

di sviluppo di Saccharomyces cerevisiae, i cui risultati hanno evidenziato che la 

concentrazione iniziale di azoto influenza sia la fermentazione alcolica che l’intero 

ciclo di sviluppo dei lieviti con interessanti ricadute sulla resistenza all’etanolo e 

sulla “senescenza” . 

Per la determinazione della “shelf-life”, un nuovo approccio, basato sulla 

“riparametrizzazione” dell’equazione di Gompertz, ha permesso una valutazione 

corretta della significatività delle differenze riscontrabili in risposta ai diversi 

trattamenti effettuati. 

L’uso di cinetiche di morte (equazione di Weibull e di Geerarerd) e un approccio 

innovativo basato su un’equazione polinomiale ha consentito di modellare le 

interazioni fra i bifidobatteri e i microrganismi starter dello yogurt. 


1 

A novel approach for calculating shelf life of minimally processed vegetables. 

Corbo, M.R., Del Nobile, M.A., Sinigaglia M. International Journal of Food 

Microbiology, 106, 69-73 (2006). Impact Factor: 2,608. 

Shelf life of minimally processed vegetables is often calculated by using the kinetic 

parameters of Gompertz equation as modified by zwietering et al. [zwietering, 

M.H., Jongenburger, F.M., Roumbouts, M., van’t Riet, K., 1990. Modelling of the 

bacterial growth curve. Applied and Environmental Microbiology 56, 1875–1881.] 

24 





taking 5*10 7 CFU/g as the maximum acceptable contamination value consistent 

with acceptable quality of these products. As this method does not allow 

estimation of the standard errors of the shelf life, in this paper the modified 

Gompertz equation was re-parameterized to directly include the shelf life as a 

fitting parameter among the Gompertz parameters. Being the shelf life a fitting 

parameter is possible to determine its confidence interval by fitting the proposed 

equation to the experimental data. The goodness-of fit of this new equation was 

tested by using mesophilic bacteria cell loads from different minimally processed 

vegetables (packaged fresh-cut lettuce, fennel and shredded carrots) that differed 

for some process operations or for package atmosphere. The new equation was 

able to describe the data well and to estimate the shelf life. The results obtained 

emphasize the importance of using the standard errors for the shelf life value to 

show significant differences among the samples. 

2 Combined 

effects of temperature, ammonium, and glucose concentrations 

on yeast growth-cycle in a model wine system. D’Amato, D, Corbo, M.R., 

Del Nobile, M.A., Sinigaglia, M. International Journal of Food Science and 


In enology, alcoholic fermentation is a complex process involving several 

mechanisms. Slow and incomplete alcoholic fermentation is a chronic problem 

for the wine industry and factors leading to sluggish and stuck fermentations have 

been extensively studied and reviewed. The most studied cause of sluggish and 

stuck fermentation is the nitrogen content limitation. Nevertheless, other factors, 

such as temperature of fermentation and sugar concentration can affect the 

growth of yeasts. In this study we modelled the yeast growth-cycle in wine model 

system as a function of temperature, sugar and ammonium concentrations; the 

individual effects and the interaction of these factors were analysed by means of 

a quadratic response surface methodology. Cell concentrations and weight loss 

were monitored in the whole wine fermentation process. The results of central 

25 





composite design show that the lower is the availability of nitrogen, the higher 

is the cell growth rate; moreover, initial nitrogen concentration also influences 

survival time of Saccharomyces cerevisiae. 

3 Combined 

effects of pH, yeast extract, carbohydrates and di-ammonium 

hydrogen citrate on the biomass production and acidifying ability of a 

probiotic Lactobacillus plantarum strain, isolated from table olives, in a 

batch system. Bevilacqua, A., Corbo, M.R., Mastromatteo, M., Sinigaglia, M. 

World Journal of Microbiology and Biotechnology, 24, 1721-1729 (2008). 

Impact Factor: 0,745. 

A four variables-five levels Central Composite Design was developed to model 

the individual and interactive effects of carbohydrates (lactose or maltose), yeast 

extract, di-ammonium hydrogen citrate and pH on the biomass production 

(Abs 600nm ), viable and cultivable cell number and acidifying ability of a probiotic 

strain of Lactobacillus plantarum, isolated from table olives “Bella di Cerignola”. 

pH values were modeled through a negative Gompertz equation, in order to obtain the 

parameter α (metabolic adaptation time). This value and the biomass were submitted to 

a stepwise procedure and second order polynomial equations were derived. 

The parameter α was affected by the initial pH and lactose; the effect of the 

maltose, however, was not significant. The biomass production increased 

with increasing of yeast extract, di-ammonium hydrogen citrate and maltose 

concentrations and was maximum at pH 6.0 and 20 g L -1 of lactose. 

4 Modelling 

the survival of starter lactic acid bacteria and Bifidobacterium 

bifidum in single and simultaneous cultures. Altieri, C., Bevilacqua, A., 

D’Amato, D., Del Nobile, M.A., Sinigaglia, M. Food Microbiology, 25, 729-734 

(2008). Impact Factor: 2,039. 

The inactivation kinetics at 4°C of Bifidobacterium bifidum, Lactobacillus delbrueckii 

subsp. bulgaricus and Streptococcus thermophilus, cultured alone or consociated 

26 





in a laboratory medium (modified MRS broth), were modelled through the 

Weibull model and a second order polynomial equation. The initial cell number 

of S. thermophilus and L. delbrueckii subsp. bulgaricus was approximately 6-7 log 

(CFU/mL); the viability loss after 30 days of storage was 2.87 and 1.99log (CFU/L) 

for L. delbruckii subsp. bulgaricus and S. thermophilus, cultured alone, respectively; 

whereas the consociation of lactobacilli and streptococci with bifidobacteria 

reduced viability loss during storage (0.28 and 0.54 log CFU/L for lactobacilli and 

streptococci, respectively). 

Finally, the consociation of lactobacilli and streptococci with B. bifidum improved 

their oxygen uptake. 

5 Modelling 

the interactions between Pseudomonas putida and Escherichia 

coli O157:H7 in fish-burgers: use of the lag-exponential model and of a 

combined interaction index. Speranza, B., Bevilacqua A., Mastromatteo M., 

Sinigaglia M., Corbo, M.R., Journal of Applied Microbiology, DOI: 10.1111/ 

J.1365-2672.2010.04692.x (2010). Impact Factor: 2,028. 

Aims - The objective of the current study was to examine the interactions 

between Pseudomonas putida and Escherichia coli O157:H7 in co-culture studies 

on fish-burgers packed in air and under different modified atmospheres (30:40:30 

O 2 :CO 2 :N 2 , 5:95 O 2 :CO 2 and 50:50 O 2 :CO 2 ), throughout the storage at 8°C. 

Methods and Results - The lag-exponential model was applied to describe 

the microbial growth. In order to give a quantitative measure of the occurring 

microbial interactions, two simple parameters were developed: the combined 

interaction index (CII) and the partial interaction index (PII). Under air, the 

interaction was significant (P




Conclusions - The interactions occurring into the system affected both E. coli 

O157:H7 and pseudomonads sub-populations. The packaging atmosphere 

resulted a key-element. 

Significance and impact of study - The paper provides some useful information 

on the interactions occurring between E. coli O157:H7 and Ps. putida on fish- 

burgers. The proposed index describes successfully the competitive growth of both 

microorganisms, giving also a quantitative measure of a qualitative phenomenon. 

28 


asPetti microbiologici connessi 

ai ProDotti Di quarta gamma 





Daniela Campaniello, Contrattista 


Persona da contattare 

Maria Rosaria Corbo, Ricercatore Confermato, 



tel. 0881 589 232; m.corbo@unifg.it 

Parole chiave 

VEGETALI DI IV GAMMA 

FRUTTA MINIMAMENTE 

PROCESSATA 

COATING EDIBILI 

STABILITà MICROBIOLOGICA 


La richiesta di prodotti con un elevato contenuto di servizio, pronti per il consumo 

e freschi, ha portato alla diffusione dei cosiddetti “minimally processed foods”, 

entro cui si inseriscono i prodotti della IV gamma. Si tratta di “prodotti vegetali 

crudi pronti per l’uso alimentare umano, mondati, tagliati e confezionati”, la cui 

conservabilità è di 7-15 giorni. 

Per quanto riguarda gli ortaggi, è stata valutata l’influenza delle operazioni 

tecnologiche che caratterizzano la produzione industriale della IV gamma nonché 

la possibilità dell’uso di un dipping in soluzione di timolo per il prolungamento 

della stabilità microbiologica di insalata caprese. 

La preparazione di prodotti di IV gamma a base di frutta richiede una strategia di 

interventi che includono l’uso di film idonei, di atmosfere modificate, l’aggiunta 

di antimicrobici e/o antiossidanti, la conservazione a basse temperature. In 

particolare, numerosi studi sono stati condotti per valutare qualità e sicurezza 

di mele, noce di cocco, ficodindia, kiwi, fragole, uva e macedonia. I risultati 

ottenuti hanno dimostrato che, partendo da materia prima di buona qualità, la 

combinazione di bassa temperatura, atmosfera modificata e in alcuni casi “coating 

edibili” (chitosano) o antimicrobici naturali (oli essenziali o molecole volatili) 

può garantire sicurezza e prolungare la stabilità microbiologica per un tempo 

compatibile con la commercializzazione. 


1 

Combined effects of modified atmosphere and thymol for prolonging the 

shelf-life of caprese salad. Bevilacqua, A., Corbo, M.R., Sinigaglia, M. Journal 

of Food Protection, 70, 722-728 (2007). Impact Factor: 1,886. 

The aim of this study was to investigate the influence of packaging atmosphere 

and a thymol dipping solution on the shelf-life of caprese salad. Caprese salad 

was prepared with sliced tomatoes and mozzarella cheese. The samples were pre- 

29 




treated by dipping in a 400 ppm thymol solution and then packaged in air and 

modified atmosphere (MA, 65% N 2 ; 30% CO 2 ; 5% O 2 ). Changes in natural microflora 

of mozzarella and tomato as well as O 2 and CO 2 in the head space, pH and sensory 

characteristics were monitored during storage. The combination of a thymol dip 

and MA decreased the coliform populations in caprese salad from 5.65 to 4.23 log 

CFU.*g -1 and extended the shelf-life from 3.77 to 12 days. A decrease in numbers 

of Pseudomonadaceae from 7.03 to 5.09 log CFU*g -1 was observed, along with 

prolongation of the lag phase to approximately 3 days. The combination of MA and 

thymol did not affect the growth kinetics of lactic acid bacteria and enterococci, 

thus preserving the functionality of mozzarella cheese in the salad. 

2 Chitosan: 

antimicrobial activity and potential applications for preserving 

minimally processed strawberries. Campaniello, D., Bevilacqua, A., 

Sinigaglia, M., Corbo, M.R Food Microbiology, 25, 992-1000 (2008). Impact 

Factor: 2,039. 

In this research the possible use of chitosan coating on fresh-cut strawberries 

was investigated. Manually sliced strawberries were treated with a solution 

of 1% chitosan, packaged in modified atmosphere with high (80%) and low 

(5%) percentage of oxygen and then stored at 4, 8, 12 and 15°C. Changes in 

microbiological quality were measured and the shelf life of the samples, as 

stability time, was kinetically modelled in order to check the effects of storage 

temperature on the most relevant microbial indices for product quality. 

A chitosan coating inhibited the growth of microorganisms and affected 

significantly and positively the stability time of the products, above all when the 

samples were packaged in modified atmosphere (with low and high percentage 

of oxygen). Besides, the presence of high percentage of oxygen, combined with 

chitosan coating, seemed to affect positively the colour. 

The data revealed that applying a chitosan coating prolonged effectively the 

quality and extended the shelf life of fresh-cut strawberries. 

30 




3 Growth 

of Listeria monocytogenes in fresh-cut coconut as affected by 

storage conditions and inoculum size. Sinigaglia, M., Bevilacqua, A., 

Campaniello, D., D’Amato, D., Corbo, M.R. Journal of Food Protection 69, 

820-825 (2006). Impact Factor: 1,921. 

In this work the effects of packaging atmosphere (air and modified atmosphere, 

65% N 2 -30% CO 2 -5% O 2 ), temperature (2°, 4°, 8° and 12°C) and inoculum size (low, 

102 CFU/g, and high inoculum, 5*10 5 CFU/g) on the growth of L. monocytogenes 

in minimally processed coconut were investigated. Growth data were fitted to the 

Gompertz equation and the sanitary risk, as time necessary to have an increase of 

two logarithmic units, was also calculated. At higher inoculum the health risk was 

more marked; moreover, as showed by the parameter A of the Gompertz equation 

(maximum increase of cell load data), the risk time seemed to be influenced 

neither by temperature nor by type of atmosphere. At low inoculum level and 

in air-stored product, the risk time was strongly influenced by temperature and 

the use of modified atmosphere caused a significant increase of the maximum 

cell load reached in the stationary phase. The results show that L. monocytogenes 

not only survives but is able to proliferate on fresh-cut coconut stored both in 

air and in modified atmosphere also at very low temperature (2°C); moreover, 

the presence of high cell load can lead to a health risk since this pathogen can 

increase in number to a risk level during shelf-life of product. 

4 Use 

of chitosan, honey and pineapple juice as filling liquids for increasing 

the microbiological shelf life of a fruit-based salad. D’Amato, D., Sinigaglia, 

M., Corbo, M.R. International Journal of Food Science and Technology 45, 

1033-1041 (2010). Impact Factor: 1,065 

The activity of some natural compounds for extending the storage life of fresh- 

cut fruits was examined. In particular, the effect of chitosan, honey and pineapple 

juice used as filling solution on the growth of mesophilic bacteria, psychrotrophic 

bacteria, lactic acid bacteria and yeasts, in fruit-based salads stored at 4°C, 8°C 

31 




and 12°C was investigated. Honey showed the greatest antibacterial effect on 

mesophilic and psychrotrophic bacteria, with non significant effects on lactic acid 

bacteria and yeasts. The antimicrobial activity of chitosan affected the growth 

of all microbial groups considered (mesophilic and psychrotrophic bacteria, 

lactic acid bacteria and yeast), particularly, in conditions of refrigerated storage. 

In contrast, no difference in the growth of monitored microorganisms between 

samples containing pineapple juice and control was detected. 

32 


AlicyclobAcillus Acidoterrestris: 











Parole chiave 

ALiCyCLOBACiLLuS 

SPORE 

ACidOtERREStRiS 

GERMINAzIONE 

BEVANDE ACIDE 


Alicyclobacillus acidoterrestris è un microrganismo alterante, sporigeno e acidofilo, 

in grado di alterare le bevande acide. Le spore sono estremamente termoresistenti 

e sopravvivono ai tradizionali trattamenti termici utilizzati nell’industria 

alimentare. Il gruppo di ricerca ha focalizzato l’attenzione su metodologie non 

convenzionali per il controllo della germinazione delle spore, come l’impiego di 

antimicrobici naturali, microonde, alte pressioni di omogeneizzazione. Una prima 

fase di screening ha consentito di selezionare l’aldeide cinnamica e il benzoato di 

sodio come gli antimicrobici più efficaci nell’inibire la germinazione delle spore 

per 8-13 giorni; successivamente, l’effetto dell’aldeide cinnamica è stato studiato 

in combinazione con il pH del mezzo colturale e con un blando trattamento 

termico. I risultati hanno mostrato come la riduzione del pH del mezzo potenzi 

notevolmente l’efficacia dell’antimicrobico; tuttavia, la combinazione dell’aldeide 

cinnamica con un trattamento termico non determina un effetto sinergico delle due 

variabili, poiché il trattamento termico riduce la concentrazione di spore iniziale, 

mentre l’antimicrobico inibisce la germinazione nel corso delle conservazione. 

Per limitarne gli effetti organolettici indesiderati, l’aldeide cinnamica è stata 

combinata con l’eugenolo e un blando trattamento termico, evidenziando come 

la stabilizzazione di un succo si possa ottenere addizionando 40 ppm di aldeide 

cinanmica e 80 ppm di eugenolo. 


1 

Inhibition of Alicyclobacillus acidoterrestris spores by natural compounds. 

Bevilacqua, A., Corbo, M.R., Sinigaglia, M. International Journal of Food 

Science and Technology, 43, 1271-1275 (2008). Impact Factor: 1,065. 

The aim of this study was to evaluate the antimicrobial effectiveness of some 

natural compounds (cinnamaldehyde, eugenol, limonene) and sodium benzoate 

against two strains of Alicyclobacillus acidoterrestris (c8 and c4). The antimicrobial 

33 




compounds (10–500 ppm) were solved in malt extract broth, inoculated separately 

(with 10 3 spores ml) of each strain; the samples were incubated at 44°C and the 

outgrowth of spores was evaluated every day by measuring the absorbance of 

the medium at 420 nm; inoculated samples without active compounds were used 

as controls. The results pointed out that limonene was not effective in inhibiting 

the outgrowth of A. acidoterrestris spores; 100 ppm of cinnamaldehyde or sodium 

benzoate slowed the spore germination, whereas 500 ppm of eugenol inhibited 

the growth of microbial targets for 13 days. Strain c8 was more resistant than 

isolate c4 and cinnamaldehyde was the most effective compound in inhibiting 

the germination of A. acidoterrestris spores. 

2 Alicyclobacillus 

acidoterrestris: new methods for inhibiting spore 

germination. Bevilacqua, A., Sinigaglia, M., Corbo, M.R. International Journal 

of Food Microbiology, 125, 103-110 (2008). Impact Factor: 2,581. 

For a long period the thermal processing has been considered as the only 

way to reduce the initial spore number of Alicyclobacillus acidoterrestris 

and prevent the spoilage of acidic beverage. New methods, however, were 

proposed by the literature to control spore germination both in laboratory 

media and in real systems. After a brief introduction on the impact of 

A. acidoterrestris in food microbiology and a description of enumeration 

methods and heat processing applied by the juices manufactures, a review 

of innovative approaches to inhibit and/or control spore germination is 

proposed. In particular, this paper focuses on two different topics; the 1 st is the 

use of some natural compounds (monolaurin, lysozyme, nisin and essential 

oils) or some chemicals, conventional (like sodium-benzoate, organic acids, 

surfactants and chlorine dioxide) or not conventional (chlorine dioxide as 

gas). The 2 nd topic is a description of some innovative methods to reduce 

the initial spore number (high hydrostatic and homogenisation pressures, 

radiation and microwaves). 

34 




3 Combined 

effects of pH and cinnamaldehyde in inhibiting Alicyclobacillus 

acidoterrestris spores. Bevilacqua, A., Corbo, M.R., Sinigaglia, M. Journal of 

Food Processing and Preservation, 32, 839-852 (2008). Impact Factor: 0,389. 

This study has focused on the inhibition of Alicyclobacillus acidoterrestris spores 

through the use of cinnamaldehyde at different pH values. The assays were 

performed in vitro in Malt Extract broth and two strains of A. acidoterrestris 

(c8 and c24) were used as target microorganisms. The amount of the active 

compound (0-200 ppm) and the pH (3.5-5.5) of the medium were modulated 

according to a two variables-five levels central composite design. Experimental 

data were modelled through the modified Gompertz equation; then, the derived 

parameters were submitted to a stepwise regression. The results pointed out 

that a low pH enhanced the antimicrobial activity of cinnamaldehyde; moreover, 

its effectiveness was dose dependent, as low doses caused a reversible inhibition 

of germination (i.e the prolongation of the lag phase), whereas high amounts 

(150-200 ppm) determined a loss of the germinating ability. An intra-species 

sensitivity of A. acidoterrestris to cinnamaldehyde was recorded and the strain 

c24 was more sensitive to the active compound. 

4 Spore 

viability of Alicyclobacillus acidoterrestris in a laboratory medium: 

effects of pH, cinnamaldehyde and heat-shock temperature. Bevilacqua, 

A., Sinigaglia, M. Corbo, M.R. International Journal of Food Science and 


A three variables-five levels Central Composite Design (CCD) was used to study 

the effects of pH (3.5, 4.0, 4.5, 5.0 and 5.5), cinnamaldehyde (0, 40, 80, 120 and 

160 ppm) and heating temperature (80, 84, 88, 92 and 96°C) on the spores of 

Alicyclobacillus acidoterrestris in malt extract broth (MEB). The heat-shock resulted 

in a slight decrease of spore number from 3.2 to 2.5 log CFU/mL and occurred at 

80-85°C depending on the pH of the medium. Otherwise, cinnamaldehyde acted 

as an additional hurdles within the storage time; the critical amounts to be used 

35 




were included in the range 41.35-44.42 ppm of cinnmaldehyde and were related 

positively to the pH, i.e. the critical amount of active compound decreased with 

the decreasing of the pH. 

5 Combining 

eugenol and cinnamaldehyde to control the growth of 

Alicyclobacillus acidoterrestris. Bevilacqua, A., Corbo, M.R., Sinigaglia, M. 

Food Control, 21, 172-177 (2010). Impact Factor: 1,823. 

This paper proposes the use of a combination of cinnamaldehyde and 

eugenol to control spore germination of Alicyclobacillus acidoterrestris. In 

the 1 st phase a three variables-five levels Central Composite Design was 

used to study the individual and additive effects of cinnamaldehyde (0-80 

ppm), eugenol (0-160 ppm) and pH (3.5-5.5). The assays were carried out in 

a laboratory medium (Malt Extract Broth). Cinnamaldehyde appeared as the 

most effective antimicrobial; otherwise, eugenol acted as a strengthening 

element and allowed a reduction of cinnamaldehyde amount in the system. 

Then, the experiments were performed in a commercial apple juice, thus 

highlighting that spore germination could be inhibited through the use of 80 

ppm of eugenol and 40 ppm of cinnamaldehyde or alternatively through the 

combination of 40 ppm of eugenol with 20 ppm of cinnamaldehyde. 

6 Use 

of microwave processing to reduce the initial contamination by 

Alicyclobacillus acidoterrestris in a cream of asparagus and effect of the 

treatment on the lipid fraction. Giuliani, R., Bevilacqua, A., Corbo, M.R., 

Severini, C. Innovative Food Science and Emerging Technologies, 11, 328- 

334 (2010). Impact Factor: 1,474. 

The effects of a short-time microwave (MW) treatments on the survival of 

Alicyclobacillus acidoterrestris spores (10 5 spores/g) inoculated in a cream of 

asparagus, along with the oxidation of the fat component (olive oil), was 

investigated. 

36 




The samples were MW-treated at 60-100% of power (2450 MHz) for 3-7 min; 

power and processing time changed according to a Central Composite Design 

at two variables/five levels. The combinations resulting in a 2 –fold reduction 

in the number of alicyclobacilli spores (100% of power-5 min; 80%-6 min; 

80%-7 min) were stored at different temperatures and times (25°C/27 days, 

37°C/18 days and 50°C/9 days), to determine the decay of the qualitative 

characteristics of the lipid fraction. Peroxides value, K 232 , K 270 and Rancimat 

test (induction time) were chosen as indexes of primary and secondary 

oxidation. MW-effect on the spores depended upon both the power and 

the treatment time; moreover the interaction [power] x [time] was the most 

significant variable. With regard to the effect of MW on the lipid fraction, the 

results suggested that MW affected slightly induction time, peroxide values 

and spectrophotometric determinations, especially at low powers. 

37 


antimicrobici naturali: stuDio Delle 

ProPrietà antimicrobiche e Delle 

Potenzialità aPPlicative in alimenti 



Matteo Alessandro Del Nobile, Professore Ordinario 



Daniela Campaniello, Contrattista 








Parole chiave 

ANTIMICROBICI “FRIENDLY” 

MIC 

NIC 

MICRO-DILUIzIONE 

PROVE “IN VIVO” 


Negli ultimi anni nei Paesi industrializzati si è diffusa una nuova tendenza, nota 

come consumismo verde, caratterizzata dalla richiesta di alimenti contenenti 

basse concentrazioni di additivi e composti chimici. In riposta a questa tendenza, 

è cresciuto l’interesse per i cosiddetti antimicrobici friendly (naturali e a basso 

impatto sull’ambiente e sulla salute del consumatore); gli oli essenziali, il lisozima, 

le batteriocine, gli acidi grassi e il chitosano sono i composti naturali, su cui si è 

concentrata l’attenzione del gruppo di ricerca. In particolare, risultati interessanti 

sono stati ottenuti con il timolo e il lemon extract nei confronti di microrganismi 

patogeni (Listeria monocytogenes, Staphylococcus aureus, Bacillus cereus, Escherichia 

coli O157:H7, yersinia enterocolitica) alteranti (coliformi, Lactobacillus plantarum, 

L. curvatus, B. licheniformis, B. subtilis) e di batteri malolattici (Lactobacillus plantarum 

e Oenococcus oeni) inoculati in mezzo di laboratorio; utilizzando l’approccio della 

micro-diluizione e la modellazione dei dati con l’equazione di Gompertz e la formula 

di Lambert, è stato possibile calcolare per ciascuna specie la Minima Concentrazione 

Inibente (MIC) e la Massima Concentrazione che non sortiva nessun effetto sul 

target microbico (NIC). Timolo e carvacrolo sono stati, invece, con successo utilizzati 

per preservare la qualità di hamburger a base di carne di struzzo. 


1 

Prolonging microbial shelf life of foods through the use of natural compounds 

and non-thermal approaches - a review. Corbo, M.R., Bevilacqua, A., 

Campaniello, D., D’Amato, D., Speranza, B., Sinigaglia, M. International Journal 

of Food Science and Technology, 44, 223-241 (2009). Impact Factor: 1,065. 

This paper proposes a review of some alternative approaches for food stabilization 

and shelf life prolonging (based on the use of natural compounds and/or non- 

thermal techniques). After a brief description of food structure implication on the 

way of using the alternative approaches, two paragraphs summarize the topics 

38 





of natural molecules (essential oils, lysozyme, lactoferrin and lactoperoxidase 

system, fatty acids, chitosan) and non thermal approaches (high hydrostatic and 

homogenization pressures, pulsed electric fields, high power ultrasound and 

irradiation). Finally, the last sections deal with the use of combined hurdles (along 

with the proposal of three possible modes of action of a multi-target preservation), 

the mathematical approaches for shelf life evaluating and some critical issues to be 

addressed in the future for a real scaling up of the proposed techniques. 

2 Evaluating 

in vitro antimicrobial activity of thymol toward hygiene- 

indicating and pathogenic bacteria. Falcone, P.M., Mastromatteo, M., 

Del Nobile, M.A., Corbo, M.R., Sinigaglia, M. Journal of Food Protection, 70, 

425-431 (2007). Impact Factor: 1,886. 

Results of a study of the kinetics of bacterial inhibition by thymol in order to 

develop appropriate applications for the compound in food systems are presented. 

A modeling-based approach was used to provide a quantitative description of the 

antimicrobial activity of thymol toward some foodborne pathogens and hygiene- 

indicating bacteria, which could be postprocessing contaminants of ready-to-eat 

meat products. The effect of the active compound on the bacterial growth was 

assessed from growth kinetics curves and dose-response profiles in a wide range 

of thymol concentrations, i.e., from 50 to 1,000 ppm. Inhibitory data were produced 

using a macrodilution methodology based on a turbidimetric technique. Microbial 

response was discussed in terms of Gompertz’s parameters as well as in terms of the 

active concentration of thymol affecting the growth status of microbial suspension 

(noninhibitory concentration and MIC). Results suggested that thymol can be 

successfully used as an alternative antimicrobial to increase the lag time as well as 

to decrease the maximum value of the growth index as reached in the stationary 

phase of the growth cycle for all investigated bacteria. Due to their high sensitivity 

to the antimicrobial stress as observed at sub-MIC, it is arguably a potential use 

of thymol for assurance of food safety and hygiene in combination with other 

39 





preservative technologies. A quantitative evaluation of the antimicrobial properties 

of the active compound was performed using a macrodilution methodology based 

on a turbidimetric technique to produce inhibitory data. Both the growth kinetics 

and inhibition profile in a wide range of thymol concentrations were obtained 

for each test bacterium, mathematically modeled, and analyzed. Noninhibitory 

concentration and MIC were determined to investigate both the microbial sensibility 

and resistance toward thymol, and Gompertz’s parameters were evaluated to assess 

the microbial response at each phase of growth cycle. The in vitro-obtained results 

suggested that thymol may be successfully used as a alternative preservative to 

increase the lag time as well as to decrease the maximum cell load reached in the 

stationary phase of growth cycle for all investigated bacteria. 

3 Effect 

of lemon extract on foodborne microorganisms. Conte, A., Speranza, 

B., Sinigaglia, M., Del Nobile, M.A. Journal of Food Protection, 70, 1896-1900 


A quantitative investigation was conducted on the antimicrobial effect of lemon 

extract against some food spoilage microorganisms: yeasts, Bacillus species, and 

lactic acid bacteria. Growth kinetics and dose-response profiles were determined 

from experimental data obtained with a suitable macrodilution methodology 

based on a turbidimetric technique. Growth and no-growth status of microbial 

suspensions were expressed in terms of non inhibitory concentration (NIC) and MIC. 

Lemon extract was effective in inhibiting the growth of the investigated vegetative 

cells and spores of microorganisms; effects were similar for bacteria and yeasts. The 

NICs for all microorganisms were very small, at around 10 ppm. Based on MICs, 

among the Bacillus species, the more resistant was Bacillus licheniformis. For yeasts, 

Saccharomyces cerevisiae was the least resistant, and similar results were obtained 

for Pichia subpelliculosa. Candida lusitaniae had an MIC of more than 100 ppm. 

Both Lactobacillus species were more resistant to lemon extract; concentrations 

necessary to provoke complete inhibition were approximately 150 ppm. 

40 





4 Use 

of lemon extract to inhibit the growth of malolactic bacteria. Conte, A., 

Sinigaglia, M., Del Nobile, M.A. Journal of Food Protection, 70, 114-118 


The potential use of lemon extract as a natural preservative to inhibit the growth 

of Oenococcus oeni and Lactobacillus plantarum, microorganisms involved in the 

malotactic fermentation of wine, was studied. Growth tests were run at 30°C using 

a laboratory media. Carbon dioxide concentration in the vial headspace was used 

as metabolic activity index of the investigated microorganisms. The MIC and 

the noninhibiting concentration (NIC) were calculated for each microorganism. 

Results suggest that lemon extract was active on each phase of the growth cycle 

for the tested microorganisms. It was also shown that lemon extract exhibits a 

nonlinear dose-related inhibitory effect on microbial growth. In particular, the 

active compound could be added at concentrations slightly higher than the NIC 

levels in order to appreciably slow down the microbial growth rate as well as to 

reduce the maximum microbial growth level. 

5 Combined 

effects of thymol, carvacrol and temperature on the quality of 

non conventional poultry patties. Mastromatteo M., Lucera A., Sinigaglia 

M., Corbo M. R. Meat Science, 83, 246-254 (2009). Impact Factor: 2,183. 

The combined effect of thymol (0–300 ppm), carvacrol (0–300 ppm), and 

temperature (0–18°C) on the quality of non conventional poultry patties 

packaged in air and modified atmosphere (MAP: 40% CO 2 ; 30%O 2 ; 30% N 2 ) 

was investigated using a simplex centroid mixture design. The patties were 

monitored for microbiological (total viable count, Enterobacteriaceae, lactic 

acid bacteria, Pseudomonas spp.) physico-chemical (pH, colour) and sensory 

attributes. For the patties mixed with the antimicrobials and stored at low 

temperature (0–3°C) a reduction of the cell load of about 1–1.5 log cfu/g was 

observed. The log reduction was lower at the end of storage time and decreased 

with the increase of the temperature. For the poultry patties packaged in MAP 

41 





the higher log reduction for Pseudomonas spp. during all the storage time was 

observed. In both packaging atmospheres the combination of the essential oils 

and low temperature determined no modification for off-odour during the first 

4 days of storage. 

6 Combined 

effects of thymol, carvacrol and grapefruit seed extract on lipid 

oxidation and colour stability of poultry meat preparations. Lucera, A. L., 

Mastromatteo, M., Sinigaglia M., Corbo, M. R. International Journal of Food 

Science and Technology, 44 (11), 2256-2267 (2009). Impact Factor: 1,065. 

The combined effects of thymol, carvacrol and grapefruit seed extract (GFSE) 

on lipid oxidation and colour stability of poultry meat preparations packaged in 

air or modified atmospheres (MAP: 5% O 2 ; 30% CO 2 ; 65% N 2 ) were investigated 

using a simplex centroid mixture design. Lipid oxidation was evaluated through 

measurement of secondary oxidation products (malonaldehyde, MDA) and 

general appearance with visual assessment and instrumental measurement of 

colour. Thymol and carvacrol, as individual antioxidants, retarded the oxidation 

process by maintaining MDA values below 2 mg kg -1 meat. The effect of GFSE 

was less than thymol and carvacrol. Redness (a*) decreased in all treatments 

during storage but this reduction was more evident in the control and in samples 

containing GFSE than in thymol and carvacrol. Although colour acceptability 

decreased with time, all meat preparations packaged in air maintained desirable 

appearance better than samples in MAP. Also, off-odours developed faster in the 

samples packaged in MAP than in aerobically packaged samples. 

7 Bioactivity 

of grapefruit extract against Pseudomonas spp. Bevilacqua, A., 

Ficelo, S., Corbo, M.R., Sinigaglia M. Journal of Food Processing and Preservation, 

DOI: 10.1111/j.1745-4549.2008.00354.x (2010). Impact Factor: 0,313. 

The effectiveness of Grapefruit Seed Extract (GFSE) against 3 strains of 

Pseudomonas spp. (PSE 5 and PSE 8, two wild strains isolated from mozzarella 

42 





cheese and identified as P. fluorescens and P. putida, respectively, and P. fluorescens 

DSMz 50090) was studied. Biomass production and viable cell count were 

evaluated. Data of biomass production were modeled through the modified 

Gompertz equation; the lag phase was assumed as the “no spoiling time” (NST), 

whereas the parameter A (maximum amount of biomass produced) was used to 

quantify the inhibition exerted by the essential oil. MIC value (minimum inhibitory 

concentration) was evaluated, too. 

The addition of GFSE (50-100 ppm) resulted in a prolongation of the no spoiling 

time and MIC values were 150 ppm for the wild strains and 200 ppm for 

P. fluorescens DSMz 50090, respectively. 

8 In 

vitro evaluation of the antimicrobial activity of eugenol, limonene and 

citrus extract against bacteria and yeasts, representative of the spoiling 

microflora of fruit juices. Bevilacqua, A., Corbo, M.R., Sinigaglia, M. Journal 

of Food Protection, DOI: food-73-03-15.3d (2010). Impact Factor: 1,763. 

This paper investigated the bioactivity of eugenol, limonene, and citrus extract 

against three bacteria (Lactobacillus plantarum, Lactobacillus brevis, and Bacillus 

coagulans) and three yeasts (Saccharomyces bayanus, Pichia membranifaciens, 

and Rhodotorula bacarum), representing the spoilage microflora of fruit juices. 

The experiments were performed with laboratory media by using a microdilution 

method. Data were fitted using the Gompertz equation, and the kinetic parameters 

were used to evaluate the MIC and the dose-dependent effect (at suboptimal 

doses for each essential oil). Citrus extract was the most effective essential 

oil, and the results suggested the following susceptibility hierarchy, from the 

most sensitive microorganism to the most resistant one (values in parentheses 

represent MICs): S. bayanus (2 ppm)> R. bacarum (3 ppm)>P. membranifaciens 

(5 ppm)>B. coagulans (cells, 20 ppm)>L. brevis (40 ppm)>L. plantarum (>40 ppm). 

43 


ioattività Di aciDi grassi 

nei confronti Di microrganismi 

alteranti e Patogeni 






Clelia Altieri, Ricercatore Confermato, 



tel. 0881 589 235; c.altieri@unifg.it 

Parole chiave 

MUFFE 

BATTERI GRAM NEGATIVI 

DOSE SUB-LETALE 

MDT 

INDICE DI INIBIzIONE 


Gli acidi grassi e i loro monogliceridi sono composti bioattivi dalle interessanti 

potenzialità applicative, proposti come antimicrobici e antifungini. Utilizzati 

inizialmente come principi attivi per inibire lo sviluppo di patogeni (batteri e muffe) 

di interesse medico (ad esempio Chlamydia trachomatis e Candida albicans), il loro 

uso è stato proposto successivamente come additivi utili a ritardare e/o inibire lo 

sviluppo di microrganismi alteranti e/o patogeni di matrici alimentari. 

In particolare, l’attenzione del gruppo di ricerca è stata orientata allo studio della 

bioattività degli acidi laurico, miristico e palmitico e dei rispettivi monogliceridi 

(monolaurina, acido monomiristico e palmitina) nei confronti di muffe (Aspergillus 

spp., Fusarium spp. e Penicillium spp.) e batteri patogeni Gram negativi (Escherichia 

coli O157:H7, Salmonella sp., yersinia enterocolitica), inoculati in mezzo di laboratorio. 

Nei confronti delle muffe è stato utilizzato l’approccio del Minimum Detection Time 

(tempo necessario affinché la colonia fungina raggiunga il diametro di 1 cm), che ha 

consentito di individuare delle scale di sensibilità ben precise inter e intra-genere. 

Nel caso dei batteri Gram negativi, invece, è stato utilizzato l’approccio della 

microdiluizione e degli indici di inibizione che hanno permesso di evidenziare una 

bioattività dei composti anche a dosi sub-letali e nelle prime ore di incubazione. 


1 

Inhibition of Aspergillus spp. and Penicillium spp. by fatty acids and their 

monoglycerides. Altieri, C., Cardillo, D., Bevilacqua, A., Sinigaglia, M. Journal 

of Food Protection, 70, 1206-1212 (2007). Impact Factor: 1,886. 

The antifungal activity of three fatty acids (lauric, myristic and palmitic acids) 

and their monoglycerides (monolaurin, monomyristic acid and palmitin) against 

Aspergillus and Penicillium spp. in a model system was investigated. 

Data were modelled through a re-parameterized Gompertz equation and the 

maximum colony diameter attained within the experimental time (30 days) (A), 

44 





the maximal radial growth rate (μ max ), the lag value (i.e. the time-days- after which 

the radial fungal growth started) (λ) and the Minimum Detection Time (i.e. the 

time-days- to attain 1 cm colony diameter) (MDT) were evaluated. 

Fatty acids and their monoglycerides inhibited significantly molds growth, by 

increasing MDT and lag values. Moreover the effectiveness of the active compounds 

seemed to be strain/genus dependent: in fact, palmitic acid was the most effective 

chemical against aspergilli, whereas penicilli were strongly inhibited by myristic acid. 

Besides, aspergilli were more susceptible to fatty acids than penicilli, as showed 

by the higher increase of MDT value. 

2 Antifungal 

activity of fatty acids and their monoglycerides against 

Fusarium spp. in a laboratory medium. Altieri, C., Bevilacqua, A., Cardillo, 

D., Sinigaglia, M. International Journal of Food Science and Technology, 44, 

242-245 (2009). Impact Factor: 0,941. 

The antifungal activity of lauric, myristic and palmitic acids and their monoglycerides 

in laboratory media against fusaria was investigated. Data were modelled through 

a re-parameterized Gompertz equation and the Minimum Detection Time (i.e. 

the time-days-to attain 1 cm colony diameter) (MDT) was evaluated. Lauric acid 

exerted a strong bioactivity against moulds; otherwise, palmitic and myristic acids 

and their monoglycerides showed a moderate effectiveness and in a reversible 

manner. The results of this work could be considered quite promising; however, 

further investigations are proposed to validate these data in foods. 

3 Effectiveness 

of fatty acids and their monoglycerides against Gram- 

negative pathogens. Altieri, C., Bevilacqua, A., Cardillo, A., Sinigaglia, M. 

International Journal of Food Science and Technology, 44, 359-366 (2009). 


The effectiveness of lauric, myristic and palmitic acids and their monoglycerides 

against Escherichia coli O157:H7, yersinia enterocolitica and Salmonella sp. 

45 





20 ppm of myristic and palmitic acids and their monoglycerides showed a 

promising bioactivity (60-80%) against E. coli O157:H7 within 10-24 h; 50 ppm 

of monolaurin inhibited y. enterocolitica and E. coli O157:H7 by > 90% of control 

for 96 h; otherwise, 40 ppm of monolaurin and 30-50 ppm of lauric acid reduced 

y. enterocolitica growth by >65% of control. The effect of lauric acid and 

its monoglyceride against Salmonella spp. was moderate (inhibition of 

approximately 30%). 

The results of this paper suggested some interesting ideas: the same compound 

could exert an inhibition or a stimulation of microbial growth; moreover, the 

studied compounds seemed to act in a reversible manner, as the inhibition of 

microorganism was quite strong within the first 10-24 h and decreased for a 

prolonged incubation. 

46 


stuDio Della microflora 

Protecnologica e alterante 

nelle olive Da mensa 










Parole chiave 

OLIVE DA TAVOLA 

LIEVITI 

COMPOSTI FENOLICI 

ACIDI ORGANICI 


Le olive da mensa rappresentano uno dei prodotti tipici a più forte vocazione 

territoriale della Daunia; il sistema produttivo, tuttavia, si basa ancora oggi su un 

complesso di piccole imprese a conduzione familiare, che non riescono a imprimere 

una svolta in termini di innovazione e valorizzazione del prodotto. Tra le cultivar più 

conosciute, vi è senza dubbio la Bella di Cerignola, che ha ricevuto il marchio DOP. 

Dopo una prima fase di caratterizzazione della microflora lattica e di selezione 

di uno starter idoneo, è stata studiata e caratterizzata la microflora levurica 

autoctona, in relazione alle caratteristiche tecnologiche e al potenziale alterante. 

In particolare, i dati ottenuti hanno mostrato come le specie maggiormente 

rappresentate fossero Candida famata e C. guilliermondii, che risentivano in 

maniera preponderante dell’effetto della temperatura e della concentrazione di 

NaCl nelle salamoie, poiché i ceppi di C. guilliermondii apparivano resistenti al 

sale, mentre gli stipiti di C. famata sviluppavano a 15°C. Il gruppo di ricerca si 

è occupato, inoltre, delle caratteristiche tecnologiche di ceppi di Enterobacter 

cloacae, isolati da olive da mensa Bella di Cerignola. E. cloacae può essere 

classificato come un patogeno opportunista emergente, isolato frequentemente 

da prodotti vegetali. I ceppi isolati dalle olive da mensa apparivano resistenti a 

concentrazioni medio-alte di sale (7-8%) e di composti fenolici secondari, oltre 

alla capacità di sviluppo a 15°C e alla resistenza a pH 5.0, suggerendo così che 

questa specie potrebbe rappresentare un grave problema se la fermentazione 

non è accuratamente controllata. 


1 

Technological and spoiling characteristics of the yeast microflora isolated 

from Bella di Cerignola table olives. Bevilacqua, A., Perricone, M., Cannarsi, 

M., Corbo M.R., Sinigaglia M. International Journal of Food Science and 


47 


stuDio Della microflora 

Protecnologica e alterante 

nelle olive Da mensa 

This study focused to investigate the technological significance and the spoiling 

impact of diverse yeast strains, isolated from “Bella di Cerignola” Italian table olives. 

64 isolates (belonging mainly to the species Candida famata and C. guilliermondii) 

were studied to assess their growth at different temperatures (15, 25 and 37°C), 

pHs (4.0, 5.0 and 9.5) and NaCl concentrations (0-10.0%) in lab medium. Their 

pectolytic, xylanolytic, lipolytic and catalase activities were also evaluated. Most 

of the yeasts showed a moderate pectolytic activity and were able to grow at 

pH 9.5 after 120 h of incubation. Salt and temperature were the most important 

environmental variables affecting yeast growth, which could exert a strong 

selective pressure on yeast population. C. guilliermondii appeared more resistant 

to high salt concentrations, whereas C. famata was able to grow at 15°C. 

2 Characterization 

and implications of Enterobacter cloacae strains, isolated 

from Italian table olives “Bella di Cerignola”. Bevilacqua, A., Cannarsi, M., 

Gallo, M., Sinigaglia, M., Corbo, M.R. Journal of Food Science, 75, M53-M60 


Enterobacter cloacae can be recovered in the spontaneous fermentations of 

Italian table olives. In this research the effects of salt (20-100 g/L), temperature 

(10-37°C), pH (4.0-5.0 and 8.0-10.0), p-coumaric and vanillic acids (0.5-2.0 g/L) 

and the acidification of the medium through lactic, citric and ascorbic acids were 

investigated on 15 strains of E. cloacae, isolated from Italian table olives “Bella di 

Cerignola”. Finally, a confirmatory experiment in a synthetic brine was run. 

The strains were inhibited only by an NaCl amount of 70-80 g/L and by p-coumaric acid; 

on the other hand, they showed the ability to grow also at low temperatures (10-15°C). The 

confirmatory experiment highlighted their ability to survive both at 15°C and at pH 5.0 

E. cloacae could be a real problem for the fermentation of table olives in 

Southern Italy; some hurdles could be used (salt or brine acidification), but some 

environmental conditions (for example the temperature) should be controlled 

carefully to maintain olive safety at acceptable levels. 

48 


tolleranza agli stress abiotici in 

microrganismi starter e Probiotici 


Giuseppe Spano, Professore Associato 

Annalisa Vernile, tecnico di Laboratorio 

Vittorio Capozzi, Contrattista 

Pasquale Russo, Contrattista 

Maria Carmela Garofano, Contrattista 


Giuseppe Spano, Professore Associato, 

Dipartimento di Scienze degli alimenti (DISA), 

Facoltà di Agraria, Università degli Studi 

di Foggia, via Napoli 25, 71100 Foggia; 

tel. 0881 589 303; g.spano@unifg.it 

Parole chiave 

STRESS ABIOTICI 

PROBIOTICI 

CRIOCONSERVAzIONE 


La risposta agli stress abiotici è un comportamento comune da parte di 

organismi vegetali o animali che reagiscono con rapidi cambiamenti nel 

loro genoma e/o proteoma a stimoli esterni quali ad esempio alte e basse 

temperature, variazioni di pH o etanolo, presenza di metalli pesanti etc. La 

tolleranza agli stress è particolarmente importante nei microrganismi utilizzati 

come starter o probiotici. Infatti, oltre ai comuni stress incontrati durante 

i processi di fermentazione (starter) o durante il transito nel tratto gastro- 

intestinale (probiotici), tali microrganismi devono essere in grado di tollerare, 

rimanendo vitali, sia trattamenti di produzione (essiccazione con calore -spray 

drying- o liofilizzazione) che il periodo di conservazione (ad esempio crio- 

conservazione). In questa ricerca vengono analizzati, mediante metodiche 

quali overespressione di geni, knock out genico, analisi di proteine e produzione 

di proteine ricombinanti, utilizzo di osmoliti neutri (zuccheri e amminoacidi), 

i meccanismi adottati da alcuni microrganismi per tollerare stress abiotici 

comunemente presenti negli alimenti (sale, temperatura, pH ed etanolo) e nel 

tratto gastrointestinale (bile, enzimi pancreatici etc.). Si cerca inoltre di migliorare 

la tolleranza ai processi di conservazione (in particolare crioconservazione). 

Risultati principali: di base 1) identificazione di alcuni meccanismi coinvolti 

nella tolleranza agli stress in lattobacilli; 2) presenza di meccanismi di “cross 

protection”; 3) fluidità di membrana strettamente connessa al tipo di stress 

imposto; 4) identificazione di proteine da stress con funzione di lipochaperonine. 

Applicativi: 1) aumentata tolleranza agli stress e ai processi di conserva- 

zione in alcuni lattobacilli mediante induzione mirata di alcuni “marcatori” 

da stress; 2) formulazione di nuovi sistemi di conservazione per la tolleranza 

alle basse temperature; 3) è in corso lo sviluppo di un sistema che mima 

gli stress gastro-intestinali per analizzare, in vitro, la tolleranza agli stress in 

microrganismi probiotici. 

49 





1 

Environmental stress response in wine Lactic Acid Bacteria: beyond Bacillus 

subtilis, Spano G., Massa S. Critical Review in Microbiology, 32, 77-86, (2006). 

Impact Factor 3,368. 

Lactic acid bacteria (LAB) constitute a heterogeneous group of bacteria that 

are traditionally used to produce fermented foods. The industrialization of food 

transformations increased the economical importance of LAB, as they play a 

crucial role in the development of the organoleptique and hygienic quality 

of fermented products. However, the strains selected for industrial pourpose, 

should tolerate adverse conditions encountered in industrial processes, either 

during starter handling and storage (freeze-drying, freezing or spray-drying) or 

are food processing in which abiotic stresses such as heat, cold, acidity and high 

concentration of NaCl or ethanol are common. Wine LAB have to deal with several 

stresses including an acidic pH, a high alcoholic content, non optimal growth 

temperatures, and growth-inhibitory compounds such as fatty acids and tannins, 

originated from yeast and bacteria metabolism. Also in the stressful environment 

generated, wine LAB are able to undergoing biochemical process such as 

malolactic fermentation and have, therefore, developed several mechanisms 

to escape or to tolerate wine conditions. In addition to the regulation of the 

expression of specific genes, bacteria have evolved adaptive networks to face the 

challenges of a changing environment and to survive under conditions of stress. 

This so called Global Regulatory Systems, control the simultaneous expression 

of a large number of genes in response to a variety of environmental stress. 

CIRCE sequences able to bind the HrcA repressor, σB dependent promoters 

and CtsR regulatory elements were observed in several genes identified from 

wine LAB. The knowledge of regulators and a better understanding of LAB stress 

responses could constitute a basis of comparison with the well known model 

microorganisms, Escherichia coli and Bacillus subtilis. Moreover, it can provide an 

important insight into improving current industrial starter strains. 

50 




2 Validation 

of an internal control gene to apply reverse transcription 

quantitative PCR to study heat, cold and ethanol stresses in Lactobacillus 

plantarum. Fiocco D., Crisetti E., Capozzi V., Spano G. World Journal of 

Microbiology and Biotechnology 24, 899-902 (2007). Impact Factor 0,745. 

The dissection of the stress tolerance in lactic acid bacteria (LAB) may improve 

our knowledge on the adaptive physiology of LAB and may facilitate the 

selection of species suitable for industrial application. In this work, we report 

the validation of the ldhd gene as an internal control for reverse transcription 

quantitative polymerase chain reaction (RT-qPCR) analysis in a model of LAB such 

as Lactobacillus plantarum. The expression of the ldhd gene was analysed in lag 

phase, early, mid, and late exponential phase and early and late stationary phase. 

Moreover, different stress conditions such as heat shock (40°C), cold shock (10°C) and 

ethanolic shock (10% v/v) were analysed. Validation of this method was performed 

by quantification of transcript levels of a small heat shock gene (hsp 18.55) after 

heat, cold and ethanol shock. RT-qPCR appeared as a powerful tool to study 

L. plantarum response in stress conditions, also the internal control that was used 

in this study may be extended to other specie of LAB. 

3 Improved 

Adaptation to Heat, Cold and Solvent Tolerance in Lactobacillus 

plantarum. Fiocco D., Capozzi V., Goffin P., Hols P., Spano G. Applied 

Microbiology and Biotechnology, 77, 909-915 (2007). Impact Factor 2,475. 

The effect of overproducing each of the three small heat shock proteins (Hsp) (Hsp 

18.5, Hsp 18.55 and Hsp 19.3) was investigated in Lactobacillus plantarum strain WCFS1. 

Overproduction of the three genes, hsp 18.5, hsp 18.55 and hsp 19.3 translationally 

fused to the start codon of the ldhL gene yielded a protein of approximately 19 kDa, as 

estimated from Tricine sodium dodecyl sulfate - polyacrylamide gel electrophoresis 

(SDS-PAGE) in agreement with the predicted molecular weight of sHsp. Small Hsp 

overproduction alleviated the reduction in growth rate triggered by exposing 

exponentially growing cells to heat shock (37°C or 45°C) and cold shock (12°C). 

51 




Moreover, overproduction of Hsp 18.55 and Hsp 19.3 led to an enhanced survival in 

presence of butanol (1% v/v) or ethanol (12% v/v) treatment suggesting a potential 

role of L. plantarum small Hsps in solvent tolerance. 

4 The 

Lactobacillus plantarum ftsh gene is a novel member of the CtsR regulon. 

Fiocco D., Msadek T., Collins M., Sacco M., Hols P., Kleerebezem M., Spano G. 

Journal of Bacteriology, 191, 1688-1694 (2009). Impact Factor: 4,12. 

FtsH proteins have dual chaperone-protease activities and are involved in protein 

quality control under stress conditions. Although the functional role of FtsH proteins 

has been clearly established, the regulatory mechanisms controlling ftsh expression 

in gram-positive bacteria remain largely unknown. Here we show that ftsh of 

Lactobacillus plantarum WCFS1 is transiently induced at the transcriptional level upon 

a temperature upshift. In addition, disruption of ftsh negatively affected the growth 

of L. plantarum at high temperatures. Sequence analysis and mapping of the ftsh 

transcriptional start site revealed a potential operator sequence for the CtsR repressor, 

partially overlapping the _35 sequence of the ftsh promoter. In order to verify whether 

CtsR is able to recognize and bind the ftsh promoter, CtsR proteins of Bacillus subtilis 

and L. plantarum were overproduced, purified, and used in DNA binding assays. CtsR 

from both species bound specifically to the ftsh promoter, generating a single protein- 

DNA complex, suggesting that CtsR may control the expression of L. plantarum ftsh. In 

order to confirm this hypothesis, a _ctsR mutant strain of L. plantarum was generated. 

Expression of ftsh in the _ctsR mutant strain was strongly upregulated, indicating that 

ftsh of L. plantarum is negatively controlled by CtsR. This is the first example of an ftsh 

gene controlled by the CtsR repressor, and the first of the low-G_C gram-positive 

bacteria where the regulatory mechanism has been identified. 

5 The 

hsp 16 gene of the probiotic Lactobacillus acidophilus is differently 

regulated by single and combined stress as revealed by reverse transcription 

quantitative PCR (qRT-PCR). 

52 




V. Capozzi, D. Fiocco, S. Weidmann, G. Felis, and G. Spano Annals of 


Lactobacillus acidophilus is used extensively for the production of dairy foods 

and is increasingly applied in the area of health improvernent, as probiotics, 

in the form of yogurts and dietary supplements. It is also being considered as 

a potential vaccine-delivery vehicle to the gastrointestinal tract (Sanders and 

Klaenhammer, 2001). The probiotic effects of this organism include the treatment 

of various types of diarrhoea, alleviation of Crohn’s disease, and balancing of 

intestinal microflora through the growth or modulation of bacteria present in 

the gastrointestinal tract (Pfeiler et al., 2007). Because of the importance of this 

organism as probiotics, studies into different mechanisms of stress response may 

be useful in order to select or to improve L. acidophilus strains able to growth 

under harsh stress condition. In this study, a reverse transcription quantitative 

PCR (RT-qPCR) was developed and used in order to quantify the transcript level of 

a small heat shock gene (shs) in L. acidophilus ATCC 4356 under stress conditions 

such as heat (45 and 53 T), bile (0,3% v/v), NaCI (1 M and 2.5 M), and low pH value 

(pH 4). The shs gene of L. acidophilus ATCC 4356 was induced either by salt or by 

high temperature stress and repressed by bile stress. Interestingly, the shs gene 

was repressed when combined salt and high temperature stresses were applied. 

The effect of overproducing the small heat shock protein (sHsp) was investigated 

in L. acidophilus ATCC 4356. Overproduction of the shs gene yielded a protein 

of approximately 16 kDa, as estimated from Tricine sodium dodecyl sulphate - 

polyacrylarnide gel electrophoresis (SDS-PAGE) in agreement with the predicted 

molecular weight of sHsp. Small Hsp overproduction alleviated the reduction in 

growth rate triggered by exposing exponentially growing cells to heat shock 

(45°C) and salt stress (1 M and 2.5 M). Therefore, the observations reported in 

our study could be of general interest in developing Lactobacillus strains with 

an improved resistance to multiple stresses for applications where growth 

under harsh stress condition would be an attribute. 

53 





of the CtsR stress response regulon in Lactobacillus plantarum. 

D. Fiocco, V. Capozzi, A. Gallone, P. Hols, J. Guzzo, S. Weidmann, A. Rieu, T. 

Msadek and G. Spano. Journal of Bacteriology, (2010) 196, 896-900. Impact 

Factor: 4,12. 

Lactobacillus plantarum ctsR was characterized. ctsR was found to be cotranscribed 

with clpC and induced in response to various abiotic stresses. ctsR deletion 

conferred a heat-sensitive phenotype with peculiar cell morphological features. 

The transcriptional pattern of putative CtsR regulon genes was examined in the ctsR 

mutant. Direct CtsR-dependent regulation was demonstrated by DNA-binding assays 

using recombinant CtsR and the promoters of the ctsR-clpC operon and hsp1. 

7 Cold 

stress response in Lactic Acid Bacteria. 

V. Capozzi, D. Fiocco and G. Spano 

In “Stress response of Lactic Acid Bacteria”, eds K. Papadimitriou and E. 

Tsakalidou, Springer Science-Business Media, LLC, New York. (2010), 1-36. 

In response to temperature downshift, a number of changes occur in cellular 

physiology such as decrease in membrane fluidity, inefficient folding of some 

proteins, and hampered ribosome function. Cold stress leads to distinct 

responses depending on the actual temperature and in particular whether it 

is greater than or less than 0°C. At subzero temperatures, which are associated 

with water freezing, the response of most prokaryotes is passive, leading slowly 

to death of the cells. This is associated with damage to the cell membrane and to 

DNA. Exposure to low temperatures above 0°C usually triggers active response 

by bacteria typically based on the synthesis of specific proteins, generally called 

cold shock proteins, to counteract the harmful effects of temperature downshift, 

leading to a transient metabolic adaptation. Usually, the specific response is 

species or strains specific. Cold stress, which takes place during the cooling and 

freezing steps and throughout the frozen storage of industrial strains, is also the 

main cause of loss of bacterial activity. 

54 


iDentificazione e tracciabilità 

Di microrganismi Patogeni e 

tolleranza agli stress in alimenti 

minimamente Processati 



Annalisa Vernile, tecnico di laboratorio 




Facoltà di Agraria, Università degli 

Studi di Foggia, via Napoli 25, 71100 Foggia; 


Parole chiave 

MICRORGANISMI PATOGENI 

StAPhyLOCOCCuS AuREuS 

LiStERiA MOnOCytOgEnES 

FRESH CUT 


L’identificazione e la tracciabilità di microrganismi patogeni quali Escherichia 

coli O157:H7 e Staphylococcus aureus in prodotti derivati dal latte, diverse 

specie di Listeria inclusa Listeria monocytogenes in prodotti “ready to use” 

(RTE), patogeni veicolati da prodotti ittici come Vibrio vulnificus, permette non 

solo di migliorare la sicurezza di un prodotto alimentare e del consumatore, 

ma anche di identificare eventuali punti critici della filiera attraverso i quali 

un microrganismo patogeno può contaminare l’alimento. Gli studi si sono 

inizialmente concentrati su patogeni emergenti e non contribuendo allo 

sviluppo di marcatori molecolari adatti ad analisi quantitative (qPCR) e a una 

rapida identificazione di E. coli O 157:h7 ed helicobacter pilory in alimenti solidi 

e liquidi. Attualmente, oltre al lavoro di identificazione e caratterizzazione, il 

laboratorio sta cercando di elucidare i meccanismi che permettono a Listeria 

monocytogenes di adattarsi ai processi di lavorazione e conservazione di 

prodotti vegetali. Inoltre, si stanno valutando gli effetti inibenti di miscele di 

batteri lattici sullo sviluppo e crescita di Listeria monocytogenes in prodotti 

ready to use (RTU). Il gruppo di ricerca si occupa inoltre della trasmissione 

di geni codificanti per antibiotico resistenza e geni di patogenicità tra specie 

geneticamente affini. 

Risultati principali: Applicativi 1) Sviluppo di metodiche basate sulla Real Time 

PCR per l’identificazione e la quantificazione di alcuni microrganismi patogeni 

direttamente negli alimenti. 


1 

Bacterial stressors in minimally processed food. Vittorio Capozzi, Daniela 

Fiocco, Maria Luisa Amodio, Anna Gallone and Giuseppe Spano Special 

issues “Biotic and Abiotic Stress” International Journal of Molecular Sciences, 

(2009),10, 3076-3105. Impact Factor: 1. 

55 






Stress responses are of particular importance to microorganisms, because their 

habitats are subjected to continual changes in temperature, osmotic pressure, and 

nutrients availability. Stressors (and stress factors), may be of chemical, physical, 

or biological nature. While stress to microorganisms is frequently caused by the 

surrounding environment, the growth of microbial cells on its own may also 

result in induction of some kinds of stress such as starvation and acidity. During 

production of fresh-cut produce, cumulative mild processing steps are employed, 

to control the growth of microorganisms. Pathogens on plant surfaces are already 

stressed and stress may be increased during the multiple mild processing steps, 

potentially leading to very hardy bacteria geared towards enhanced survival. 

Cross-protection can occur because the overlapping stress responses enable 

bacteria exposed to one stress to become resistant to another stress. A number 

of stresses have been shown to induce cross protection, including heat, cold, acid 

and osmotic stress. Among other factors, adaptation to heat stress appears to 

provide bacterial cells with more pronounced cross protection against several 

other stresses. Understanding how pathogens sense and respond to mild stresses 

is essential in order to design safe and effective minimal processing regimes. 

2 Diversity 

and enterotoxigenicity of Staphylococcus spp. associated with 

Domiati cheese. Walid M. El-Sharoud, Spano G. Journal of Food Protection, 

71, 2567–2571 (2008). Impact Factor: 1,89. 

A total of 87 samples of fresh and stored Domiati cheese (an Egyptian soft cheese) 

were examined for the presence of Staphylococcus spp. Fifteen Staphylococcus 

isolates identified as S. aureus (2 isolates), S. xylosus (4), S. caprae (4), and S. 

chromogenes (5) were recovered from 15 cheese samples. The S. aureus isolates 

were resistant to penicillin G and ampicillin, and one isolate was also resistant 

to tetracycline. S. aureus isolates harbored classical staphylococcal enterotoxin 

(SE) genes (sea and seb) and recently characterized SE-like genes (selg, seli, selm, 

and selo). One S. aureus isolate contained a single SE gene (sea), whereas another 

56 






isolate contained five SE genes (seb, selg, seli, selm, and selo). These results suggest 

that Domiati cheese is a source for various Staphylococcus species, including S. 

aureus strains that could be enterotoxigenic. 

3 Horizontal 

Gene Transfer in gut: is it a risk? 

V. Capozzi and G. Spano Food Research International, 42, 1501-1502 (2009). 

Impact Factor: 2, 073. 

The human gastro-intestinal microbiota involves the highest concentration of 

microorganisms in the human body and contains both probiotic and pathogenic 

bacteria. The co-presence of these different bacteria may lead to the occurrence of 

the so-called horizontal gene transfer (HGT) between these microorganisms.This 

phenomenon has been shown to allow the transfer of genes encoding virulence 

traits and antimicrobial drug resistance from pathogenic bacteria to other non- 

pathogenic strains, which highlight edits significance to human health. It could 

be envisaged that HGT may occur between pathogenic and probiotic bacteria 

in the gut, where pathogens could receive genetic elements from probiotics 

that enhance the pathogen’s colonization in the gut. HGT may also allow for the 

acquisition of virulence genes by probiotics from neighboring gut pathogens. 

Pertinent literature on HGT events within gut microbiota is controversial and 

the present paper aims to critically address this aspect. In light of an increased 

production of probiotics-containing foods, addressing HGT would be a vital act 

to ensure food safety. 

57 


iDentificazione e caratterizzazione 

Di starter enologici e analisi 

Dei meccanismi coinvolti nella 

ProDuzione Di ammine biogene in vino 



Luciano Beneduce, Ricercatore 


Pasquale Russo, Assegnista di ricerca 

Cosimo Vetrana, Assegnista di ricerca 







Parole chiave 

STARTER ENOLOGICI 

AMMINE BIOGENE 

PCR-DGGE 

ARGININA 


L’ utilizzo di “starter” microbici quali Saccharomyces cerevisae (nella fermentazione 

alcolica) ed Oenococcus oeni (nella fermentazione malolattica) è diventato 

uso comune nella pratica vinicola per il miglioramento delle caratteristiche 

organolettiche del vino. L’isolamento e la caratterizzazione delle principali specie 

microbiche (lieviti e batteri) viene effettuato sia mediante metodi colturali e saggi 

biochimici ampiamente descritti e riportati in letteratura che mediante tecniche 

molecolari, PCR specie-specifica (amplificazione e sequenziamento del gene 16S 

rDNA) e Real time PCR. I ceppi microbici isolati e caratterizzati, rappresentativi 

delle specie S. cereviseae ed O. oeni vengono successivamente analizzati per la 

loro capacità di tollerare stress abiotici comuni in vino e per le loro proprietà 

fisiologiche e biochimiche. I ceppi caratterizzati vengono poi utilizzati in prove di 

microvinificazione ed eventuali prodotti per essere posti in commercio. 

Alcuni microrganismi inoltre, pur partecipando marginalmente alle fermentazioni 

alcolica e malolattica, risultano importanti per potenziali effetti “spoilage” che 

potrebbero avere sul prodotto finale portando alla formazione di ammine 

biogene (BA). Poiché la produzione di ammine biogene dipende da molteplici 

fattori quali ad esempio pratiche agronomiche ed enologiche (utilizzo o 

meno di starter microbici) e dalla presenza/frequenza di batteri lattici in 

grado di decarbossilare amminoacidi quali histidina e tiramina, e produrre le 

corrispondenti ammine, risulta importante il monitoraggio e la caratterizzazione 

di batteri lattici in grado di produrre ammine biogene, per un eventuale controllo 

e riduzione delle stesse. 

Risultati principali: di base 1) Sviluppo di metodiche molecolari per 

l’identificazione simultanea di batteri lattici del vino in grado di produrre ammine 

biogene quali tiramina, istamina e putrescina; 2) identificazione di induttori e/o 

inibitori della produzione di ammine biogene in alimenti fermentati. 

Applicativi 1) Utilizzo di marcatori da stress per l’identificazione di starter enologici; 

2) costituzione di una ceppoteca di Oenococcus oeni; 3) formulazione di coinoculi 

58 






(lieviti-batteri) vinari; 4) utilizzo di batteri lattici per la riduzione del contenuto di 

ammine biogene in alimenti fermentati. 


1 

Characterization of wine Lactobacillus plantarum by PCR-DGGE and RAPD- 

PCR analysis and identification of Lactobacillus plantarum strains able 

to degrade arginine. Spano, G., Vernile, A., Beneduce, L., Tarantino, D., De 

Palma, L., Massa, S. World Journal of Microbiology and Biotechnology, 22, 

769-773 (2006). Impact Factor: 0,745. 

Screening of strains isolated from red wine undergoing malolactic fermentation 

allowed the identification of lactic acid bacteria able to degrade arginine. A 

denaturing gradient gel electrophoresis approach, using the rpoB gene as the 

molecular target, was developed in order to characterize the isolated strains. 

Several strains were identified as Lactobacillus plantarum and were typed by 

RAPD-PCR with several randomly designed primers. Almost all of the L. plantarum 

strains identified were able to produce citrulline and ammonia, suggesting that 

the ability of L. plantarum to degrade arginine is a common feature in wine. 

During the characterization of the newly identified L. plantarum strains, the 

presence of genes coding for the arginine deiminase pathway was observed in 

the strains able to produce citrulline, while the lack of this genes was observed in 

strain unable to produce citrulline. These results suggest that the degradation of 

arginine in L. plantarum is probably strain-dependent. 

2 in 

vivo PCR-DGGE analysis of Lactobacillus plantarum and Oenococcus oeni 

populations in red wine. Spano G., Lonvaud-Funel A., Classe O., Massa S. 

Current Microbiology, 54, 9-13 (2007). Impact Factor: 1,2. 

In order to monitor Lactobacillus plantarum and Oenococcus oeni in red 

wine produced with Italian grape (variety “Primitivo di Puglia”), a PCR-DGGE 

59 






approach using the rpoB as gene target was established. Wine was treated 

or not with potassium metabisulphite and supplemented with a commercial 

bacterial starter of Oenococcus oeni to encourage malolactic fermentation. 

Samples were taken from the vinification tanks at 4, 10, 16, 22 and 28 days after 

the start of alcoholic fermentation. Genomic DNA was directly isolated from 

wine and identification of lactic acid bacteria was performed using primers 

rpoB1, rpoB1O and rpoB2 able to amplify a region of 336 bp corresponding to 

the rpoB gene. Amplified fragments were separated in a 30-60% DGGE gradient 

and the ability of the PCR-DGGE analysis to distinguish Lactobacillus plantarum 

and Oenococcus oeni was assessed. The results reported suggest that PCR-DGGE 

method, based on the rpoB gene as molecular marker, is a reproducible and 

suitable tool and may be of great value for wine makers in order to monitor 

spoilage microrganisms during wine fermentation. 

3 Arginine 

metabolism in wine Lactobacillus plantarum: in vitro activities 

of the enzymes arginine deiminase (ADI) and ornithine transcarbamilase 

(OTCase). Spano G., Massa S., Arena M. E., Manca de Nadra M. C. Annals of 


This work was carried out to determine the activity of enzymes involved 

in arginine metabolism in Lactobacillus plantarum isolated from wine and 

previously characterised at molecular level. The activity of the enzymes arginine 

deiminase and ornithine transcarbamylase was determined and citrulline and 

ornithine formed were analysed by HPLC analysis. Although the enzymatic 

activity was detected in all the strains analysed, a strong variability was 

observed between strains. Lactobacillus plantarum strain Lp60 is the strain with 

more possibilities to accumulate citrulline, precursor of the carcinogenic ethyl- 

carbamate, as showed by its high arginine deiminase activity and low ornithine 

transcarbamylase activity. 

60 







of a Lactobacillus plantarum strain able to produce 

tyramine and partially cloning of a putative tyrosine decarboxylase gene. 

Arena M. E., Fiocco D., Manca de Nadra M. C., Pardo I., Spano G. Current 


The aim of this article was to analyze the ability of wine Lactobacillus plantarum 

strains to form tyramine. Preliminary identification of L. plantarum strains was 

performed by amplification of the recA gene. Primers pREV and PlanF, ParaF and 

PentF were used respectively as reverse and forward primers in the polymerase 

chain reaction tests as previously reported. Furthermore, the gene encoding for 

the tyrosine decarboxylase (TDC) was partially cloned from one strain identified 

as L. plantarum. The strain was further analyzed by 16S rDNA sequence and 

confirmed as belonging to L. plantarum species. The tyrosine decarboxylase 

activity was investigated and tyramine was determined by the high-performance 

liquid chromatography method. Moreover, a negative effect of sugars such as 

glucose and fructose and L-malic acid on tyrosine decarboxylase activity was 

observed. The results suggest that, occasionally, L. plantarum is able to produce 

tyramine in wine and this ability is apparently confined only to L. plantarum 

strains harboring the tdc gene. 

5 Technological 

properties of Oenococcus oeni strains isolated from tipical 

southern italian wines. Capozzi V., Russo, P., Beneduce L., Weidmann S., 

Grieco F., Guzzo J., Spano G. Letters in Applied Microbiology, 50, 327-334 

(2010). Impact Factor 1.68 

Aims - To isolate indigenous Oenococcus oeni strains suitable as starters for 

malolactic fermentation (MLF), using a reliable polyphasic approach. 

Methods and Results - Oenococcus oeni strains were isolated from Nero di 

Troia wines undergoing spontaneous MLF. Samples were taken at the end 

of alcoholic fermentation and during MLF. Wine samples were diluted in 

a sterile physiological solution and plated on MRS and on modified FT80. 

61 






Identification of O. oeni strains was performed by a polymerase chain reaction 

(PCR) experiment using strain-specific primers. Strains were further grouped 

using a multiplex RAPD-PCR analysis. Then, six strains were inoculated in two 

wine-like media with two different ethanol concentrations (11 and 13% vol/ 

vol) with a view to evaluate their capacity to grow and to perform MLF. In 

addition, a quantitative PCR (qRT-PCR) approach was adapted to monitor the 

physiological state of the strains selected. 

Conclusion - A positive correlation between the malolactic activity performance 

and the ability to develop and tolerate stress conditions was observed for two 

selected O. oeni strains. 

Significance and Impact of the Study - The results reported are useful for the 

selection of indigenous MLF starter cultures with desired oenological traits from 

typical regional wines. It should be the base for the improvement in organoleptic 

quality of typical red wine. 

62 


innovazione nella ProDuzione 

Di Pasta alimentare (fresca 

e secca) meDiante l’utilizzo Di 

microrganismi (batteri) selezionati 



Vittorio Capozzi, Contrattista 







Parole chiave 

PASTA 

CEREALI 

BATTERI LATTICI 

ALIMENTI FUNzIONALI 


L’obiettivo di questa linea di ricerca è quello di mettere a disposizione del 

settore nazionale di produzione della pasta un processo in grado di permettere 

la creazione del segmento “pasta funzionale” e “pasta probiotica” mediante 

l’utilizzo di coadiuvanti (inoculi microbici) naturalmente presenti nelle semole. 

Nello specifico il programma si pone tre obiettivi: 1) la caratterizzazione della 

microflora presente durante la produzione delle paste artigianali con periodici 

prelievi di campioni delle materie prime della pasta durante le fasi di lavorazione 

e in diversi periodi dell’anno; 2) l’ utilizzo (inoculo) di microrganismi in grado di 

arricchire l’alimento in composti aventi attività biologica; 3) l’ottenimento del 

segmento pasta funzionale e/o probiotica. 

Tutto il processo è caratterizzato sia mediante approccio metabolico (HPLC, Gas 

cromatografia etc.) per l’analisi dei composti volatili e delle molecole ad attività 

biologica apportate dagli inoculi batterici, che nutraceutico e probiotico per la 

valutazione della funzionalità dell’alimento e della capacità dei microrganismi 

probiotici utilizzati nel raggiungere il tratto gastro-intestinale, svilupparsi ed 

aderire allo stesso epitelio intestinale. 

Risultati attesi - Produzione di paste probiotiche e paste FUNzIONALI mediante 

utilizzo di batteri lattici. 


1 

Molecular analysis of a durum wheat ‘stay green’ mutant: expression pattern 

of photosynthesis-related genes. Rampino P., Spano G., Pataleo S., Mita G., 

Napier J. A., Di Fonzo N., Shewry P. R., Perrotta C. Journal of Cereal Science, 

43, 160–168 (2006). Impact Factor: 3.026. 

Elucidation of molecular mechanisms underlying the ‘stay green’ trait is not 

only relevant to understanding the senescence phenomenon itself, but also has 

potential significance for yield improvement. A mutant of durum wheat (triticum 

63 






durum Desf.) cultivar Trinakria (designated 504) was characterised by delayed 

leaf senescence, and analysis of photosynthetic parameters showed that it was 

functionally ‘stay green’. An in vitro system was established to mimic senescence 

by incubating wheat leaves in the dark, allowing the expression of genes known 

to be differentially regulated during senescence to be determined. Moreover, 

cDNAs found to be differentially expressed in the mutant were cloned and 

sequenced, showing homologies with photosynthesis-related genes. Reverse 

Transcription-Polymerase Chain Reaction (RT-PCR) analyses were performed 

using RNA samples from 504 mutant and parental Trinakria plants, during both 

natural and artificially induced senescence, confirming the altered expression 

profiles of these genes in the ‘stay green’ mutant 504. The exploitation of such 

‘stay green’ phenotype in breeding of durum and bread wheats has the potential 

to increase yields by extending the period of active photosynthesis during grain 

filling, especially under unfavourable environmental conditions. 

2 Metabolic 

profiling and analysis of volatile composition of durum wheat 

semolina and pasta. Beleggia R., Platani C., Spano G., Monteleone M., Cattivelli 

L. Journal of Cereal Science, 49, 301–309 (2008). Impact Factor: 3,026. 

Although pasta is generally not considered for its aromatic properties, some 

evidence proves that cereal flours release volatile compounds and they might 

have an effect on the aroma of the transformed products. This work reports 

on the characterization of the volatile components of semolina and pasta 

obtained from four durum wheat cultivars (Triticum durum Desf., cvs. PR22D89, 

Creso, Cappelli, Trinakria). Semolina samples were characterized through polar 

metabolite profiling and fatty acid analysis to identify potential precursors of the 

volatile components. The results show significant differences among the samples 

tested with cv. Trinakria characterized by the highest content of sugars and fatty 

acids. Volatile composition was investigated both in semolina and in cooked 

pasta using headspace solidphase micro-extraction (HS-SPME) and identified by 

64 






GC–MS. Thirty-five volatile compounds including aldehydes, ketones, alcohols, 

terpenes, esters, hydrocarbons and a furan were identified. Significant differences 

were observed between semolina and pasta samples in terms of composition 

and amount of the volatile compounds. During cooking an increase in aldehyde 

content, the appearance of ketones and a decrease in alcohol content were 

observed. Correlations between metabolites and volatiles demonstrate that the 

flavour of cooked pasta may differ significantly depending on the durum wheat 

cultivar employed. 

3 A 

polyphasich approach in order to identify dominant lactic acid bacteria 

and their evolution during pasta manufacturing. 

P. Russo, S. Ferrè, I. Pardo, R. Beleggia and G. Spano. LTW – Food Science 


Using a polyphasic approach, we have isolated and identified, lactic acid bacteria 

(LAB) in samples directly collected from an artisanal pasta-making manufactory 

located in Puglia (South Italy). Samples were collected during several steps of 

pasta processing and LAB were isolated on MRS and M17 plates. Furthermore, 

strains were grouped in a total of eight species by means of randomly amplified 

polymorphic DNA (RAPD)-polymerase chain reaction (PCR) typing and 16S rDNA 

sequencing. The majority of strains were identified as belonging to Pediococcus 

pentosaceus and Enterococcus faecium species. The remaining strains were 

characterized and assigned to Weissella confusa, Pediococcus acidilactici, 

Leuconostoc mesenteroides, Leuconostoc citreum, Lactobacillus fermentum and 

Lactobacillus plantarum species. Strains were identified from all the analysed 

samples, and growth of LAB was monitored from extrusion to pre-heating steps. 

65 


icerca e caratterizzazione 

Di batteri Patogeni emergenti 

negli alimenti e nelle acque 


Salvatore Massa, Professore Ordinario 

Luciano Beneduce, Ricercatore Confermato 



Luciano Beneduce, Ricercatore Confermato, 




tel. 0881 589 234; l.beneduce@unifg.it 

Parole chiave 

BATTERI PATOGENI EMERGENTI 

DELLE ACQUE E DEGLI 

ALIMENTI 

CARATTERIzzAzIONE 

GENOTIPICA 

REAL TIME PCR 


La linea di ricerca ha operato nel triennio 2006-2009 verso lo sviluppo di 

metodiche di identificazione e caratterizzazione molecolare di batteri patogeni 

emergenti negli alimenti e nelle acque. Sebbene la maggior parte delle 

tossinfezioni alimentari nei paesi sviluppati siano causate da Salmonella spp. e 

Campylobacter spp. si fanno strada sempre più i cosiddetti “patogeni emergenti” 

tra cui E. coli eneteroemorragica (EHEC), helicobacter pylori, Vibrio vulnificus e 

Vibrio parahaemolyticus. Il nostro gruppo di ricerca si è interessato allo sviluppo 

e l’impiego di sonde molecolari (PCR e real-time PCR) per l’identificazione di 

alcuni di questi microrganismi, insieme a tecniche di caratterizzazione, sempre 

mediante metodi di biologia molecolare (RAPD, PFGE). 

Le pubblicazioni sono il frutto di collaborazione con il CRA di Fiorenzuola d’Arda 

(1) e con l’ Institut Agronomique et Vétérinaire Hassan II, Rabat, Morocco (2). 

Risultati di base - Studio sulla presenza di sierotipi di E. coli enteroemorragica nei 

prodotti a base di carne cruda commercializzati nella regione di Rabat. 

Valutazione della presenza di h. pylori nelle acque reflue di tre impianti di 

depurazione della provincia di Foggia. 

Applicativi - Messa a punto una metodica real-time PCR per l’identificazione di h. 

pilori nelle acque, potenzialmente adatta a monitorare la presenza del patogeno 

sia nell’ambiente che negli alimenti. 

Ottimizzazione di metodiche per ricerca e caratterizzazione di E. coli O157:H7 ed 

altri sierotipi coinvolti in episodi epidemici in Europa e nord-Africa. 


1 

Assessment of a quantitative PCR method for the detection of helicobacter 

pylori in environmental samples. Beneduce L., Spano G., Nabi A., Terzi V., 

Lasalandra G., Massa S. Fresenius Environmental Bulletin 16(7), 749-755 


66 





Urban wastewater samples were collected from three urban wastewater 

treatment plant located in Puglia (south of Italy). helicobacter pylori strains 

DSM 4867 and 26695 were used as positive control. Wastewater samples were 

inoculated with tenfold dilution from 10 1 to 10 7 cell ml -1 of h. pylori strain DSM 

4867, and real-time PCR experiments were performed using the Sybr Green PCR 

mix and primer related to vacA, cagA and ureC genes. The sensitivity limit of the 

assay was 10 -3 pg µl -1 for ureC gene. The efficiency of amplification of total DNA 

isolated from wastewater ranged between 90-96%, and the squared regression 

coefficient (correlation) for ureC gene was 0.99. No positive sample was found 

among wastewaters collected from the three depuration basins. The suitability 

and utility of a rapid semi-automated real-time PCR assay for detection of h. pylori 

in waste-water has been evaluated. Although we were unable to detect any DNA 

corresponding to h. pylori in unspiked collected samples, the method developed 

may be useful to assess the presence of this pathogenic microorganism in water 

and wastewater. ureC gene is the most suitable candidate that may be used in 

R-PCR experiments in order to detect h. pylori in environmental samples. VacA 

and CagA genes can be targeted in order to evaluate the presence of strains 

harbouring pathogenicity factors among positive samples. 

2 Occurrence 

and characterization of Escherichia coli O157 and other 

serotypes in raw meat products in Morocco. Beneduce L., Spano G., 

Nabi A.Q., Lamacchia F., Massa S., Aouni R., Hamama A. Journal of Food 

Protection, 71(10), 2082-2086 (2008). Impact Factor: 1,886. 

In this study, 100 raw meat samples were collected from 15 local Moroccan 

butcheries in five different areas of the city of Rabat during a period of 4 

months. Overall, 7 of 15 butcheries from three areas of the city yielded 

strains of Escherichia coli O157. Single isolates from 9 (9%) of 100 raw meat 

samples were biochemically and serologically confirmed as E. coli O157. Using 

molecular techniques, two strains were positive for the Shiga toxin, with two 

67 





additional strains containing an attaching-effacing gene. All potentially virulent 

serotypes isolated from these meat samples showed distinct pulsed-field gel 

electrophoresis profiles. Based on antibiotic susceptibility testing, more than 

70% of the isolates were resistant to ampicillin and clavulanic acid–amoxicillin. 

Moreover, one strain was resistant to more than three antibiotics. Our study 

represents the first survey of E. coli O157 and related serotypes in raw meat 

products in Morocco. 

3 Occurrence 

of giardia and Cryptosporidium in Italian water supplies. 

Vernile A., Nabi A. Q., Bonadonna L., Briancesco R., Massa S. Environmental 

Monitoring Assessment, 152: 203-207 (2009). Impact Factor: 0,793. 

A total of 21 samples: raw water (RW) samples; water samples after 

coagulation with aluminium sulfate (clarified water: CW); and water after 

chlorination (treated water: TW) from a water purification plant that 

treats river surface water from the neighbourhood of Foggia (Italy), were 

analysed for the presence of giardia cysts and Cryptosporidium oocysts. 

Bacteriological indicator of faecal contamination (total and faecal coliforms, 

faecal streptococci), total bacterial count at 22 and 36°C and physicochemical 

parameters (turbidity, temperature, pH) were evaluated. Cryptosporidium 

oocysts were not found in any samples examined, while giardia cysts were 

found only in RW samples, with the maximal concentration of 8 cysts/100 l. 

A positive correlation was found between the giardia densities and quality 

parameters such as TC, FC and TBC at 22°C. giardia levels in raw water samples 

correlated (p


microflora in formaggi tiPici 


Salvatore Massa, Professore Ordinario 

Luciano Beneduce, Ricercatore Confermato 



Salvatore Massa, Professore Ordinario, 




tel. 0881 589 234; s.massa@unifg.it 

Parole chiave 

LACTIC ACID BACTERIA 

CHEESE RIPENING 

NSLAB 


La linea di ricerca ha operato nel triennio 2006-2009 verso l’isolamento, l’identificazione e 

la caratterizzazione della microflora autoctona di formaggi tipici, ottenuti mediante latte 

crudo senza l’aggiunta di colture starter, in particolar modo del pecorino siciliano, e sullo 

studio delle caratteristiche chimiche di questo formaggio. Sono state impiegate tecniche 

di biologia molecolare, quali reazione a catena della polimerasi (PCR) ed elettroforesi in 

campo pulsato (PFGE), come strumenti affidabili e riproducibili per la caratterizzazione 

genotipica di isolati di batteri lattici e di stafilococchi del pecorino siciliano. 

La reazione a catena della polimerasi e l’elettroforesi in campo pulsato hanno 

rivelato la presenza di numerosi ceppi di batteri lattici e di stafilococchi presenti 

nel pecorino siciliano. Questa microflora autoctona è la sola responsabile delle 

caratteristiche chimico-fisiche dei formaggi tipici. Questi risultati, pertanto, 

suggeriscono che la microflora autoctona del pecorino siciliano dovrebbe essere 

preservata, al fine di valorizzare i formaggi tipici e per selezionare nuove colture 

starter, che possono rivelarsi importanti per l’industria lattiero-casearia. 


1 

Microbial study of Pecorino Siciliano cheese throughout ripening. Vernile 

A., Spano G., Beresford, T. P., Fox, P. F., Beneduce, L., Massa, S. Milk Science 

International 61, 169-172 (2006). Impact Factor: 0,36. 

The microbiological characteristics of Pecorino Siciliano raw ewe’s milk cheese 

were studied. Six batches of cheeses, three made in winter and the others in 

spring, were examined during ripening. Analyses of the main microbial groups 

were conducted on the internal part of cheese, whereas micrococci/staphylococci, 

halophilic bacteria, yeasts and moulds were detected on the rind cheese. In 

general, the microbial counts in the interior of the cheeses during winter at 1, 

30 and 90 d of ripening were slightly higher than during the spring, except for 

mesophilic and thermophilic lactobacilli whose counts were higher in spring than 

69 




in winter. During the ripening, high numbers of lactic acid bacteria were found: 

the numbers of lactococci, lactobacilli and enterococci were the most abundant 

organisms in Pecorino Siciliano cheese. The number of micrococci/staphylococci 

and halophilic bacteria decreased slowly during ripening. Coliforms disappeared 

in two batches of spring cheeses, whereas in the others these decreased during 

ripening to values between 2.3 and 4.1 log units. Values of some physicochemical 

indices, such as pH, % moisture content and % NaCI, were also determined. 

2 Chemical 

Studies of Pecorino Siciliano Cheese throughout Ripening. Vernile 

A., Beresford T. P., Spano G., Massa S., Fox P. F. Milk Science International, 62, 

280-284 (2007). Impact Factor: 0,36. 

Changes in the composition and level of proteolysis in the internal portions 

of Pecorino Siciliano cheese were studied. At the end of ripening, the average 

moisture, fat and protein concentrations were 33.36, 30.9 and 27.73%, respectively, 

for winter-made cheese and 35.63, 30.2 and 27.56%, respectively, for spring-made 

cheese. The average level of salt-in-moisture (g 100 g-1) was 11.55 and 6.16 in 

winter-and spring-made cheese, respectively. The average pH of both winter- and 

spring-made cheese was 5.36. Urea-polyacrylamide gel electrophoresis of the 

cheese samples indicated that α s1 –casein was degraded in all samples leading 

to the formation of products with higher electrophoretic mobility. β−Casein was 

also degraded to equivalent levels in all cheeses at the end of ripening. Peptide 

profiles obtained by reversed phase high performance liquid chromatography 

indicated significant changes during ripening. The principal free amino acids 

(FAAs) found in all samples were Glu, Leu, Phe, Lys and Val. 

3 Genotypic 

characterization of lactic acid bacteria isolated from traditional 

Pecorino Siciliano cheese. Vernile A., Giammanco G., Spano G., Beresford 

T.P., Fox P.F., Massa S. Dairy Science & Technology, 88, 619-622 (2006). 


70 




A total of 468 lactic acid bacteria (LAB) isolates from the interior of 6 

traditional Pecorino Siciliano cheeses during ripening (1, 30 and 90 days) were 

characterized genotypically in order to assess the biodiversity within this wild 

microbial population. Two DNA-based technique, PCR and PFGE were used 

for genetic typing of isolates. Of the 468 isolates, species-specific PCR analysis 

showed that 79, 58, 2, 9 and 4 isolates reacted with primers for Lactobacillus 

paracasei, Lb. plantarum, Lb. pentosus, Lb. rhamnosus and Lb. curvatus, 

respectively and no isolates reacted with the Lb. casei primers. Genus-specific 

PCR analysis showed that 59 isolates reacted positively with the lactococcal 

primers, 221 with the enterococcal primers and 34 with the Leuconostoc 

primers, 1 with the pediococcal primers and 4 with the streptococcal primers. 

Enterococci were characterized at species level and twelve of the 221 

enterococci isolates showed positive reaction with the E. faecalis species- 

specific primers, and the remainder 209 isolates positively with the E. faecium 

species-specific primers. PFGE analysis allowed to identify different strains 

of the same species of Lb. plantarum and Lb. paracasei. The strains which 

reacted positively with Lb. curvatus, Lb. pentosus or Lb. rhamnosus primers 

gave a unique PFGE pattern. PFGE indicated 52 different band patterns for 

enterococci, 9 for lactococci, 5 for leuconostocs and 1 for streptococci and 

pediococci. The results suggest that wild bacterial populations should be 

preserved in order to protect the traditional raw milk cheeses, and to select 

new specific strains for the dairy industry. 

4 Biodiversity 

of Staphylococcus species in Pecorino Siciliano Cheese. Vernile 

A., Spano G., Beresford T.P., Fox P.F., Massa S. Milk Science International, 64, 

67-70, (2009). Impact Factor: 0,36. 

Some species of Staphylococcus found in dairy products can be important in 

the cheese ripening and particularly for the Protected Designation of Origin 

(PDO) cheeses. Bacteriological analysis was carried out and the bacteria were 

71 




enumerated by surface plating on PCA and on Baird Parker. Fifty-four Gram- 

positive, catalase positive isolates from the core of Pecorino Siciliano cheese, a raw 

ewe’s milk cheese, at different stages of ripening were identified phenotypically as 

members of the genus Staphylococcus. All the isolates were coagulase-negative, 

facultative anaerobes and were, therefore, Staphylococcus species. No effort was 

made to identify them to species level. Pulsed Field Gel Electrophoresis (PFGE) 

analysis was used as a tool for examining overall genome similarity or divergence 

between isolates. Twenty-five different profiles were obtained with by PFGE, 

revealing the presence of a wide biodiversity among the strains. The results of the 

first study gave an overview of the population of staphylococci associated with 

Pecorino Siciliano cheese. Further studies will be required to study interaction 

among the predominant biotypes in relation to the physicochemical properties 

of cheese, in order to protect the traditional raw milk cheeses. 

72 


caratterizzazione Degli alimenti 

eD effetti Dei Processi tecnologici 

sulla materia Prima meDiante stuDi 

Proteomici, liPiDomici e metabolomici 


Ennio La Notte, Professore Ordinario 

Carmela Lamacchia, Ricercatore confermato 

Antonietta Baiano, Ricercatore confermato 

Sandra Pati, Ricercatore non confermato 

Barbara La Gatta, tecnico laureato 


Aldo Di Luccia, Professore Associato, 



di Foggia, Via Napoli 25, 71122 Foggia; 

tel. 0881 589 243, a.diluccia@unifg.it 

Parole chiave 

MARCATORI MOLECOLARI 

SPETTROMETRIA DI MASSA 

PROTEOLISI E LIPOLISI 

METABOLOMICA 

ALIMENTI E PROCESSI 

TECNOLOGICI 


Le proprietà organolettiche e nutrizionali degli alimenti derivano essenzialmente 

da due categorie: 

• intrinseche del prodotto, frutto dei metabolismi cellulari come proteine, lipidi 

e polisaccaridi e piccoli metaboliti di natura organica; 

• l’altra è rappresentata da molecole che possono essere assorbite dall’ambiente 

esterno e veicolate dai fluidi biologici o essere di neo formazione dovuta 

ad aggregazioni molecolari o a cambiamenti molecolari per effetto delle 

tecnologie di trasformazione e di conservazione. 

Le scienze omiche in campo alimentare hanno permesso di studiare la struttura 

delle molecole di base che costituiscono un alimento e le trasformazione che esse 

subiscono nel corso della manipolazione ancorché l’uomo consumi l’alimento 

fresco, trasformato e/o conservato. 

L’applicazione di queste scienze ha portato alla definizione dei processi 

proteolitici e lipolitici negli alimenti di origine animale fermentati e non 

fermentati, all’individuazione e alla caratterizzazione molecolare di componenti 

marcatori di prodotto e processo nonché all’attitudine della materia prima alla 

trasformazione. Attualmente sono in corso ricerche sulla identificazione delle 

cagliate fresche e conservate, profilo proteico e peptidico delle uve e dei vini, in 

particolare degli spumanti, al fine di definire l’importanza tecnologica dei queste 

frazioni. Inoltre si stanno proseguendo le ricerche sulla capacità di pastificazione 

delle diverse semole e di farine provenienti da altre fonti. 


1 

Proteomic study of muscle sarcoplasmic proteins using aut-page/sds-page as 

two-dimensional gel electrophoresis. Picariello G., De Martino A., Mamone G, 

Ferranti P., Addeo F., Faccia M., Spagna Musso S. and Di Luccia A. J. Chromatogr. B 

Analyt. technol. Biomed. Life Sci. 833: 101-108. (2006). Impact Factor: 2,500 

73 






In the present study, an alternative procedure for two-dimensional (2D) 

electrophoretic analysis in proteomic investigation of the most represented basic 

muscle water-soluble proteins is suggested. Our method consists of Acetic acid- 

Urea-Triton polyacrylamide gel (AUT-PAGE) analysis in the first dimension and 

standard sodium dodecyl sulphate polyacrylamide gel (SDS-PAGE) in the second 

dimension. Although standard two dimensional Immobilized pH Gradient- 

Sodium Dodecyl-Sulphate (2D IPG-SDS) gel electrophoresis has been successfully 

used to study these proteins, most of the water-soluble proteins are spread on the 

alkaline part of the 2D map and are poorly focused. Furthermore, the similarity 

in their molecular weights impairs resolution of the classical approach. The 

addition of Triton X-100, a non-ionic detergent, into the gel induces a differential 

electrophoretic mobility of proteins as a result of the formation of mixed micelles 

between the detergent and the hydrophobic moieties of polypeptides, separating 

basic proteins with a criterion similar to reversed phase chromatography based 

on their hydrophobicity. The acid pH induces positive net charges, increasing 

with the isoelectric point of proteins, thus allowing enhanced resolution in the 

separation. By using 2D AUT-PAGE/SDS electrophoresis approach to separate 

water-soluble proteins from fresh pork and from dry-cured products, we could 

spread proteins over a greater area, achieving a greater resolution than that 

obtained by IPG in the pH range 3–10 and 6–11. Sarcoplasmic proteins undergoing 

proteolysis during the ripening of products were identified by Matrix Assisted 

Laser Desorption/Ionization–Time of Flight (MALDIToF) mass spectrometry 

peptide mass fingerprinting in a easier and more effective way. Two-dimensional 

AUT-PAGE/SDS electrophoresis has allowed to simplify separation of sarcoplasmic 

protein mixtures making this technique suitable in the defining of quality of dry- 

cured pork products by immediate comparison of 2D maps to define the events 

occurring during their ripening and individuate candidate molecular markers 

of the characteristic proteolytic processes. Considering that, essentially, muscle 

endogenous enzymic activity, calpains and cathepsins, occur in the ripening 

74 






process of dry-cured ham, whereas a combined action between endogenous 

and microbial enzymes takes place in the case of sausage ripening, these results 

provide deeper insight into the respective role of endogenous and microbial 

enzymes in performing proteolysis. Finally, image analysis and creation of data 

bank could be achieved to quickly identify and protect typical products. 

2 Evaluation 

of gamma rays influence on some biochemical and microbiological 

aspects in black truffles. Nazzaro F., Fratianni F., Picariello G., Coppola R., Reale 

A. and Di Luccia A. Food Chemistry, 103: 344-354. (2007). Impact Factor: 2.696. 

The effects of two gamma-ray doses (1.5 kGy and 2.0 kGy) on some biochemical 

aspects and on the microbiological profile of black truffles was monitored, 

immediately after treatment and after 30 days of storage at 4°C. Electrophoretic 

and chromatographic analyses of proteins and peptides, just like monitoring of 

polyphenol content, peroxides formation and microbial profile, allowed for the 

first time a better understanding of the mechanisms responsible for biochemical 

alterations and bacterial pattern in black truffles during their storage. 

Treatment at 1.5 kGy appeared to better preserve the characteristics of the fresh 

product. In 2.0 kGy-samples, the protein profile was characterised by a 20 kDa- 

polypeptide, which could be considered as an useful marker of the irradiation 

treatment and of the storage time of the product. MALDI-TOF mass spectrometry 

analysis did not permit a correct identification from tryptic peptides in databases, 

although the nano-ES/MS/MS analyses performed on the 10 kDa tryptic digest 

peptides showed an amino acidic sequence entirely contained in a protein of 

filamentous fungus neurospora crassa. 

3 Mutagenic 

and antimutagenic properties of aqueous and ethanolic extracts 

from fresh and irradiated tuber aestivum black truffle: A preliminary study. 

F. Fratianni, A. Di Luccia, R. Coppola and F. Nazzaro Food Chemistry, 102: 

471-474. (2007). Impact Factor: 2.696. 

75 






A preliminary study was performed on fresh and irradiated Tuber aestivum black 

truffles to investigate the presence of mutagenic and antimutagenic activities in 

the fresh product and to examine the possible effects of treatment with gamma 

rays. The study was performed on aqueous and ethanolic extracts from truffles, 

untreated or irradiated with a final dose of 1.5 kGy. Two Salmonella typhimurium 

His – strains, TA 98 and TA100, were used. The preliminary results indicate that 

black truffles may contain compounds having an inhibitory effect against direct 

and indirect acting mutagenic compounds. The irradiation did not lead to the 

formation of mutagenic compounds, but the level of antimutagenic activity was 

slightly decreased after the treatment. 

4 Influence 

of type of milk and ripening time on proteolysis and lipolysis in a 

cheese made from overheated milk. M. Faccia, , G. Gambacorta, F., Caponio, 

S., Pati and A. Di Luccia international Journal of Food Science and technology, 

42: 427–433. (2007). Impact factor: 1,065. 

A study was carried out in order to assess the influence of the type of milk 

and of the ripening time on the biochemical changes in Cacioricotta, a craft 

cheese manufactured from overheated milk. The features of the cheese were 

strongly influenced by the peculiar cheese-making process, which is also 

characterized by absence of added starters and ‘forced drying’ in ventilated 

room. The electrophoretic patterns revealed heterogeneity of as-casein, marked 

degradation of β-casein and slight hydrolysis of β-lactoglobulin in goat’s cheese, 

whereas presence of a-lactalbumin and strong hydrolysis of as-casein were found 

in the ovine cheese. In spite of a significant primary proteolysis, the ripening 

indices of matured cheeses were low, because of poor secondary proteolysis. 

Lipolysis proved to be the most significant biochemical event of ripening, and 

was strongly influenced by the type of milk used. The possible reasons of this 

particular ripening profile are discussed. 

76 






5 Changes 

in pasta proteins induced by drying cycles and their relationship 

to cooking behaviour. Lamacchia C., Di Luccia A., Baiano A., Gambacorta 

G., la Gatta B., Pati S., La Notte E. Journal of Cereal Science 46: 58–63. (2007). 


Spaghetti produced in a pilot plant was dried using three different drying 

conditions: 60°C for 7.5 h; 75°C for 5.5 h; 90°C for 5 h. Proteins of spaghetti were 

characterized by size-exclusion chromatography (SEC) and size-exclusion–high- 

performance liquid chromatography (SE-HPLC). As the temperature of the drying 

cycles increased, a progressive decrease of the small and large monomeric 

proteins and an increase in molecular size of the large polymeric proteins (LPPs) 

were observed. Drying temperature at 60°C already induces a significant increase 

in the molecular size of the LPPs and a significant decrease of the other protein 

fractions. At 70 and 90°C, large and small monomeric proteins as well as LPPs 

polymerized as seen by a progressive shift towards higher molecular weights in 

the SEC profile as temperature increased. The changes in the chromatographic 

profiles were accompanied by increasing amount of total unextractable polymeric 

protein (UPP). A decrease in stickiness and an increase in firmness corresponded 

to the formation of large and insoluble protein aggregates. 

6 Application 

of capillary electrophoresis to determine the technological 

properties of wheat flours by a glutenin index. Di Luccia A., Lamacchia C., 

Mamone G., Picariello G., Trani A., Masi P., Addeo F. Journal of Food Science, 

74: C307-311. (2009). Impact Factor: 1,496 

Capillary electrophoresis was used to characterize glutenin proteins from ancient 

varieties of Southern Italy common wheat and to determine the technological 

properties of wheat flours based on a glutenin index. Three zones were identified 

in the electropherograms, indicated as A, B, and C according to electroelution 

order. The three zones corresponded to the low molecular weight glutenin 

subunits and to the y- and x-type high molecular weight subunits, respectively. 

77 






The ratio B/C was correlated to the alveographic parameter P/L. These results 

indicated that flours resulting in a B/C ratio lower than 2 produced elastic doughs 

whereas flours resulting in a B/C ratio higher than 2 produced doughs more 

resistant to extension. This study showed that capillary electrophoresis is useful 

for determining the types and quantities of gluten proteins in the evaluation of 

wheat-flour technological properties of a limited number of non commercial 

varieties as evidenced by the x-type content which seems to strongly influence 

the flour technological parameters. 

7 Occurrence 

of β-Casein Fragments in Cold Stored and Curdled River 

Buffalo (Bubalus Bubalis, L.) Milk. Di luccia A., Picariello G., Trani A., Alviti G., 

Loizzo P., Faccia M., Addeo F. Journal of dairy Science, 92: 1319-1329. (2009). 


The safeguard of river buffalo Mozzarella cheese, a Protected Designation of 

Origin dairy product, has prompted an analytical study to trace the milk and 

curd used as raw material in cheesemaking. This is to prevent the illegal use of 

milk or curd from different geographical areas outside of those indicated in the 

official production protocol. For this purpose, we studied primary proteolysis 

occurring in fresh and frozen milk and curd to identify a molecular marker that 

could indicate the raw material used. Whole casein from frozen river buffalo milk 

was separated using cation-exchange chromatography and sodium dodecyl 

sulfate-PAGE, and a protein component with an estimated molecular weight of 

15.3 kDa was detected. This protein component was revealed in fresh river buffalo 

milk as a faint electrophoresis band, which drastically increased in intensity in 

refrigerated and frozen milk as well as in curd and was found to be associated 

with beta-CN through hydrophobic interaction. By using matrix-assisted laser 

desorption/ionization-time of flight peptide mass mapping, this component was 

identified as the C-terminal fragment f(69-209) of beta-CN (expected molecular 

weight of 15,748.8 Da). β-Casein f(69-209), originating from the early hydrolysis 

78 






of Lys(68)-Ser(69) by plasmin, has no counterpart in bovine milk. The increased 

rate of hydrolysis by plasmin toward the cleavage site Lys(68)-Ser(69) has to 

be ascribed to the elevated proline content of the peptide 61-73. The favored 

production of beta-CN f(69-209) has also drawn attention to the complementary 

proteose peptone beta-CN f(1-68) that is presumed to play a physiological role 

in inducing milk secretion similar to that of beta-CN f(1-29). The higher in vivo 

and in vitro production rate, compared with gamma(1)-CN formation, indicates 

that beta-CN f(69-209) and its complementary fragment are candidate molecular 

markers to evaluate milk and curd freshness. We suggested [corrected] indirect 

ELISA analysis based on the determination of remaining nonhydrolyzed beta-CN 

to perform a quantitative evaluation of proteolysis. 

8 Relationships 

between milk quality and acidification in the production of table 

Mozzarella without starters. Faccia M., Trani A., Di Luccia A. Journal of dairy 

Science, 92: 4211-4217. (2009). Impact Factor: 2,486. 

The effect of some quality parameters of the milk (refrigeration time, pH, 

protein, and fat/protein ratio) on the extent of acidification in the production 

technology of table Mozzarella without starters was investigated. A screening 

phase carried out at the laboratory level demonstrated that variations of the 

milk characteristics require different levels of acidification to keep constant 

the quality of the cheese. The elaboration of the data collected throughout 

the successive experimentation on industrial scale allowed us to find a 

mathematical model to describe the relationships between the pH of the 

curd at stretching time and the milk characteristics, of which the protein 

concentration and the refrigeration time play the main roles. 


and Genetic Study of the Ovine aS2-Casein (CSn1S2) Allele B. 

Picariello G., Rignanese D., Chessa S., Ceriotti G., Trani A., Caroli A., and Di Luccia 

A. the Protein Journal, 28: 333-340. (2009). Impact Factor: 0,940 

79 






An interesting and quite complex protein pattern has been described at ovine milk 

proteins but the genetic control of the variation observed was assessed only in few 

cases. The aim of this work was to characterize the ovine as2-casein (CSN1S2) B variant, 

first observed in the Italian Gentile di Puglia, a fine-wooled ovine breed, and to 

investigate its occurrence in two further breeds, the Sarda and Camosciata, which are 

the most widespread dairy breeds in Italy. The B variant differs from the most common 

form A with two amino acid exchanges: Asp75 → Tyr75 and Ile105 → Val105. The 

first substitution, resulting in a loss of a negative charge, is responsible for the 

higher isoelectric point of the B protein variant, which allows its detection by 

isoelectric focusing electrophoresis (IEF). The occurrence of CSN1S2*B in Sarda 

and Comisana was demonstrated. Since the Asp75 → Tyr75 substitution modifies 

the protein electric charge, milk properties may result affected to some extent. 

10 

Proteomic-based analytical approach for the characterization of glutenin 

subunits in durum wheat. Mamone G., De Caro S., Di Luccia A., Addeo F. 

and Ferranti P. Journal of Mass Spectrometry, 44: 1709-1723. (2009). Impact 

Factor: 2.94 

One of the main objectives of wheat glutenin subunit (GS) analysis is the 

identification of protein components linked to wheat quality. 

The proteomic characterization of glutenin has to consider the relatively low 

levels of arginine and lysine residues and the close sequence similarity among 

the different groups of these subunits, which hinders or even prevents the 

identification of the GS. 

In this study, a proteomic approach has been applied to resolve the 

heterogeneity of wheat glutenin components. Proteins extracted from triticum 

durum flour were first analyzed by two-dimensional gel electrophoresis, which 

greatly reduced glutenin complexity. 

The identity of each spot was confirmed by nano liquid chromatography 

tandem mass spectrometry analysis of tryptic peptides. 

80 






In parallel, measurements of the high mass range by matrix-assisted laser 

desorption/ionization time-of-flight analysis allowed detection of the large 

tryptic peptides. Gathering all data from search engine interrogation, very 

high sequence coverage was obtained for high molecular weight GS, including 

Bx7 and By8, in agreementwith the known genetic profile of durum wheat. 

In addition, a truncated form of By8, never detected before, was also found. 

Low molecular weight GS (LMW-GS) B-type was identified with reasonable 

sequence coverage, while a clear identification of LWM-GS C- and D-type 

was hindered by the incompleteness of the wheat DNA databases. This study 

represents the first comprehensive analysis of the glutenin proteome and 

provides a reliable method for classifying wheat varieties according to their 

glutenin profile. 

11 

Gene duplication at the alpha-lactalbumin locus: finding the evidence 

in water buffalo (Bubalus bubalus, L.). Rullo R., Di Luccia A., Chianese L., 

Pieragostini E. Journal of dairy Science, 93: 2161-2167. (2010). Impact 

Factor: 2,486 

Studies on milk proteins revealed that a qualitative and quantitative polymorphism 

may be often found regarding alpha-lactalbumin (α-LA). In mammals, this 

phenomenon was widely documented in the alpha globin system as the result 

of a gene duplication. The presence of several, differently expressed alpha- 

lactalbumin gene (LALBA) products, suggests that the phenomenon may be 

the result of non allelic genes. To check this hypothesis, an experiment was 

set up to investigate LALBA gene arrangement of a water buffalo, exhibiting 

an α-LA phenotype characterized by a double-band pattern; particularly, the 

relative amount of protein, inferred from the different intensity of the bands, 

was remarkably similar to the amount observable in the presence of a gene 

duplication. Thus, leukocyte DNA was extracted from a blood sample of this 

subject and amplified with four primers (IIRV- IIIFW for PCR and IIIFW- IRV for 

81 






nested PCR). The intergenic segments of the assumed duplicated gene were 

then analyzed with two different PCR protocols. First, the segment limited by 

the 3 rd exon in the upstream gene and the 2 nd exon in the downstream gene 

was amplified by simple-PCR that gave aspecific results. The nested PCR of 

the segment limited by the 4 th exon in the upstream gene and the 1 st exon 

in the downstream gene yielded in the presence of an amplified nucleotide 

fragment of about 6500 bp; this fragment was then purified, cloned in 

pGEM-Teasy vector system and sequenced by an ABI Sequencer. The sequences 

of the extremities of the large DNA segment, containing the intergenic portion, 

were aligned with the LALBA gene (accession number AF194373 NCBI data 

bank). They were found to coincide, with the portion containing the exon 

4 and the untranslated region (UTR) at the 3’end of the upstream gene and with 

the portion containing exon 1 and UTR at the 3’end of the downstream gene. 

These results confirm the hypothesis that a tandemly repeated copy of the 

LALBA gene is present in water buffalo. 

12 

Study on the interactions between soy and semolina proteins during pasta- 

making. Lamacchia C., Baiano A., Lamparelli S., Paladino L., La notte E., Di 

luccia A. Food Research international, in press. (2010). Impact Factor: 2,073. 

Spaghetti produced in a pilot plant were made from semolina and semolina 

blended with 10%, 15%, 25% or 50% of defatted soy flour (DSF) or toasted soy flour 

(TSF). Proteins of spaghetti were characterized by size exclusion-high-performance 

liquid chromatography (SE-HPLC). Results showed that soy globulins interact 

with semolina proteins during pasta making, forming polymers of high molecular 

weight. Of these, the sodium dodecyl sulphate-unextractable components were 

significantly higher (p





different classes of proteins involves interaction by sulphydryl residues in blends 

with above 15% of DSF and that soy proteins tend to disrupt own gluten S–S 

interchange system. In the TSF–semolina spaghetti the increase of S–S bonds was 

higher with respect to that of –SH free, suggesting that the heat treatment, to 

which the TSF proteins were subjected, allowed them to cross link to semolina 

proteins by disulphide bonds. 

13 

Peptides surviving the simulated gastrointestinal digestion of milk 

proteins: biological and toxicological implications. Picariello G., Ferranti 

P., Fierro O., Mamone G., Caira S., Di Luccia A., Monica S., Addeo F. Journal 

of Chromatography B Analyt technol Biomed Life Sci. 878(3-4): 295-308. 


Resistance to proteases throughout the gastrointestinal (GI) tract is a prerequisite 

for milk-derived peptides to exert biological activities. In this work an in vitro 

multi-step static model to simulate complete digestion of the bovine milk proteins 

has been developed. The experimental set-up involved the sequential use of: 

(i) pepsin, (ii) pancreatic proteases, and (iii) extracts of human intestinal brush 

border membranes, in simulated gastric, duodenal and jejuneal environments, 

respectively. Enzymatic concentrations and reaction times were selected in order 

to closely reproduce the in vivo conditions. The aim was to identify the peptide 

candidates able to exhibit significant bioactive effects. Casein and whey protein 

peptides which survived the in vitro GI digestion have been identified by the 

combined application of HPLC and mass spectrometry techniques. While the 

permanence of the main potentially bioactive peptides from both casein and 

whey proteins was found of limited physiological relevance, the high resistance to 

proteolysis of specific regions of beta-lactoglobulin (beta-Lg), and especially that 

of the peptide beta-Lg f125-135, could have implications for the immunogenic 

action of beta-Lg in the insurgence of cow’s milk allergy. 

83 






14 

Changes in durum wheat kernel and pasta proteins induced by toasting 

and drying processes. Lamacchia C., Baiano A., Lamparelli S., La Notte E., 

Di Luccia A. Food Chemistry, 118: 191–198. (2010). 

Durum wheat kernels were subjected to a toasting process and the proteins 

characterised by size exclusion-high performance liquid chromatography (SE- 

HPLC) and sodium dodecyl sulphate–polyacrylamide gel electrophoresis. With 

this physical process, albumins and globulins, as well as glutenins and gliadins, 

polymerised as seen by a shift of the SE-HPLC profile to lower elution times. 

The polymerisation seemed to happen mainly through disulphide bonds, 

even though the participation of x-gliadins to the aggregation suggested the 

involvement of other kinds of interactions. It led to the revelation of a new peak 

originated by thermal aggregation of small polymeric proteins. The changes 

in the chromatographic profile were accompanied by increasing amounts of 

total unextractable polymeric proteins. The replacement of semolina with 

toasted durum wheat flour (5%, 10%, 15%, 20% and 30%) for the production of 

pasta in the shape of spaghetti significantly (p0.05) change the unextractable polymeric proteins (UPP) when 

compared with spaghetti made with 100% durum semolina. On the other hand, 

the replacements of semolina with 15–30% TDWF showed significant (p

stuDio Della comPonente 

Polifenolica e volatile 

nelle uve e nei vini 



Aldo Di Luccia, Professore Associato 



Barbara La Gatta, tecnico laureato 


Ennio La Notte, Professore Ordinario, 


Facoltà di Agraria, Università di Foggia, 

Via Napoli 25, 71122 Foggia; 

tel. 0881-589217; e.lanotte@unifg.it 

Parole chiave 

POLIFENOLI 

UVA 

VINO 

COMPOSTI VOLATILI 

SPETTROMETRIA DI MASSA 


Tra i differenti composti chimici presenti nelle uve, i polifenoli quali antociani, 

tannini e proantocianidine giocano un ruolo importante sulle caratteristiche 

sensoriali e salutistiche dei relativi vini. Da qui l’importanza della caratterizzazione 

della frazione fenolica di uve derivanti da vitigni autoctoni e/o introdotti nella 

regione Puglia. Inoltre, durante la conservazione e invecchiamento del vino la 

concentrazione delle varie classi fenoliche cambia perché reagiscono dando 

origine a composti più stabili, responsabili dei mutamenti del colore e della 

perdita di astringenza del vino. La conoscenza delle strutture molecolari coinvolte 

risulta fondamentale per la comprensione della complessa evoluzione del vino e 

la messa a punto di strategie idonee al miglioramento della sua qualità. 


1 

Simultaneous separation and identification of oligomeric procyanidins and 

anthocyanin-derived pigments in raw red wine by HPLC-UV-ESI-MSn. Pati 

S., Losito I., Gambacorta G., La Notte E., Palmisano F., zambonin P.G., Journal 

of Mass Spectrometry, 41, 861-871. (2006). Impact Factor: 2.945. 

Samples of raw red wine (Primitivo di Manduria, Apulia, Southern Italy) were analysed 

without any pre-treatment (except 1:2 dilution with water) using HPLC with detection 

based on UV absorbance and Electrospray Ionisation Sequential Mass Spectrometry 

(ESI-MS n , with n = 1-3) in a series configuration. In particular, absorbance at 520 nm 

was monitored for UV detection in order to identify pigments responsible for wine 

colour. On the other hand, two subsequent stages of MS detection based on positive 

ions were adopted. The first consisted of an explorative MS acquisition, aimed at the 

individuation of the m/z ratios for positively charged compounds; the second was 

based on fragmentation of the detected ions within an ion trap analyser, followed by 

MS/MS and, if required, MS 3 acquisitions. The synergy between UV detection and MS n 

analysis led to the identification of 41 pigments, which can be classified into five groups: 

85 





grape anthocyanins, pyranoanthocyanins, vinyl-linked anthocyanin-flavanol pigments, 

ethyl-bridged anthocyanin-flavanol pigments and flavanol-anthocyanin compounds. 

Many isomeric and oligomeric structures were found within each group. A further class 

of compounds, not absorbing in the visible spectrum, could be also characterised by 

ESI-MS n and corresponded to B-type procyanidins, i.e. proanthocyanidins arising from 

C4-→C8/C4-→-C6 couplings between catechin or epicatechin units. In particular, 

oligomeric structures (from dimers to pentamers), often present with several isomers, 

were identified and their fragmentation patterns clarified. 

2 Phenolic 

composition of red grapes grown in Southern Italy. Lovino R., 

Baiano A., Pati S., Faccia M., Gambacorta G. italian Journal of Food Science, 

18, 177-186 (2006). Impact Factor: 0.506. 

The phenolic composition of red grapes native to Southern Italy (Aglianico, 

Carignano, Frappato, Gaglioppo, Negro Amaro, Nero d’Avola, Primitivo, Tintilia, 

and Uva di Troia) and an “international” grape (Cabernet Saouvignon) introduced 

into Apulia region were investigated. Results showed that these cultivars could be 

divided into two groups on the basis of both their anthocyanin content and the 

presence of ortho-hydroxylated groups. Further differences regarded the ratio 

between flavans reacting with vanillin and proanthocyanidins.The anthocyanin 

profile of the skin of Negroamaro, Primitivo and Uva di Troia grapes was found to be 

a specific characteristics of the grape variety which was affected only slightly by the 

place of growing. The different phenolic composition of the cultivars determines a 

different aptitude to wine production. The Cabernet Sauvignon grapes, due to their 

high concentration in polyphenolic substances, could be added to the native grape 

varieties in order to produce wines with a more complex aroma. 

3 Rapid 

screening for anthocyanins and anthocyanin dimers in crude grape 

extracts by high performance liquid chromatography coupled with diode 

array detection and tandem mass spectrometry. Pati S., Liberatore M.T., 

86 





Gambacorta G., Antonacci D., La Notte E.. J. Chromatogr. A, 1216, 3864– 

3868 (2009). Impact Factor: 3.554. 

A rapid and efficient method using High Performance Liquid Chromatography 

coupled with Diode Array detection and Tandem Mass Spectrometry (HPLC-DAD- 

MS/MS) for fast screening large numbers of anthocyanins and anthocyanin dimers 

in different grape skin extracts, without further sample clean-up procedures, was 

developed. A good separation of most detected anthocyanins was achieved 

in a run time of 15 min. Identification of anthocyanins pigments required a 

combination of several information: UV-vis spectra, MS and MS/MS spectra, and 

elution pattern. Many compounds have been here detected for the first time and 

their structures tentatively elucidated. 

4 Chemical 

changes in Apulia table wines as affected by plastic packages. 

Mentana A., Pati S., La Notte E., del Nobile M. A. LWt - Food Science and 

technology, 42, 1360–1366 (2009). Impact Factor: 1.887 

The influence of polymeric packages on the quality decay of red and white Apulia 

table wine is addressed in this work. In particular, polyethylene terephthalate with 

oxygen scavenger package compared with polyethylene terephthalate, with a 

different permeability to oxygen, was used in this investigation. Wine packaged in 

glass and stored under the same environmental conditions served as the ‘‘control’’ 

sample. Classical oenological parameters including the anthocyanin fraction and 

the volatile fraction, were monitored over 7 months assessing the chemical changes 

related to oxidation and flavor scalping. Polyethylene terephthalate with oxygen 

scavenger showed a behavior in preserving wine much closer to the glass one with 

respect to polyethylene terephthalate as evidenced from all the selected descriptors. 

5 Influence 

of ageing on lees on polysaccharide glycosyl-residue composition 

of Chardonnay wine. Pati S., Liberatore M.T., Lamacchia C., Gambacorta G., 

La Notte E.. Carbohydrate Polymers, 80, 333-337 (2010). Impact Factor: 2.644. 

87 





The influence of different ageing conditions on lees, and in particular of 

lees obtained with and without a rough raking, on polysaccharide glycosyl- 

residues of Chardonnay wine is addressed in this work. Methanolysis and 

derivatisation procedures of ethanol precipitated polysaccharides, followed by 

gas-chromatography coupled to mass spectrometry, allowed the determination 

of all the main sugars known to constitute wine polysaccharides. The aging on 

lees led to an increase of the contents of all the glycosyl residues, except for both 

galacturonic acid, whose concentration slightly decreased, and glucose and myo- 

inositol, which were not affected by the ageing process. Results suggest that the 

use of aging on lees without a rough raking enriched wine of mannoproteins 

meanwhile contributing to the solubilisation of grape polysaccharides more than 

aging on lees with a rough raking. 

6 Aroma 

quality improvement of chardonnay white wine by fermentation and 

aging in barrique on lees. Influence of ageing on lees on polysaccharide glycosyl- 

residue composition of Chardonnay wine. Liberatore M.T., Pati S., del Nobile M. 

A., La Notte E. Food Research international, in press. Impact Factor: 2.073. 

The aroma quality of Chardonnay white wines as affected by different processing 

conditions is assessed in this work. Three technologies were tested: fermentation 

and ageing in stainless steel, fermentation and ageing in barrique on lees, 

fermentation and ageing in barrique on lees after partial removal of lees after 

fermentation. Volatile profile by means of gas-chromatography coupled to mass 

spectrometry and sensory analysis by means of descriptive and triangle tests 

were evaluated in order to discriminate between the tested wines. The ageing 

on lees, most without raking, led to an enhancement of flavor active compounds, 

such as lactones, terpenoids, ketones, aldehydes and esters, and to a likely related 

improvement of taste. Change in odor with respect to traditionally made wine 

was not easily established, because the presence of lees attenuated the impact of 

wood, which is generally strong, on the aroma balance of wine. 

88 



fenolica Delle matrici 

alimentari e misura 

Dell’attività antiossiDante 



Aldo Di Luccia, Professore associato 


Carmela Terracone, dottore di Ricerca 






tel. 0881 589 217; e.lanotte@unifg.it 

Parole chiave 

ATTIVITà ANTIOSSIDANTE 

COMPOSTI FENOLICI 

PRODOTTI OLIVICOLO-OLEARI 

PRODOTTI VITI-VINICOLI 

SOIA 


I composti fenolici sono metaboliti secondari prodotti dalle piante e dotati di 

attività antiossidante. I composti fenolici sono definiti sostanze “nutraceutiche” in 

quanto esplicano effetti benefici sull’organismo e gli alimenti che li contengono 

sono detti “funzionali” in quanto, alla normale funzione nutrizionale, uniscono 

connotazioni nutraceutiche. Le tendenze della moderna tecnologia alimentare 

vanno in molte direzioni: aggiunta agli alimenti di composti fenolici esogeni; 

arricchimento con fenoli endogeni (a titolo di esempio si può citare l’aggiunta 

delle frazioni di perlatura dei cereali, agli impasti destinati alla panificazione o 

alla pastificazione); applicazioni di tecnologi capaci di preservare il contenuto 

in sostanze fenoliche. Poiché l’attività antiossidante si esplica attraverso diversi 

meccanismi (radical scavenging, protezione di substrati lipidici), appare 

evidente la necessità di applicare un ampio ventaglio di metodi per la misura 

dell’attività antiossidante. La determinazione quali-quantitativa dei composti 

fenolici di prodotti vitivinicoli, olivicolo-oleari, cerealicoli e a base di soia, gli 

effetti delle tecnologie di produzione e della conservazione sul contenuto 

fenolico e la messa a punto di metodiche analitiche per la misura dell’attività 

antiossidante sono state e sono tuttora oggetto di sperimentazioni, di progetti 

di ricerca finanziati, di diverse tesi di laurea e di dottorato, e sono illustrati in 

pubblicazioni e presentazioni a convegni. 


1 

Phenolic Content and Antioxidant Activity of Primitivo Wine: Comparison 

among Wine-Making Technologies. Baiano A., Terracone C., Gambacorta G., 

La Notte E. Journal of Food Science, 74, 3, 258-267 (2009). Impact Factor 1.255. 

The aim of this work was to assess the influence of nine wine-making technologies 

(traditional, delestage, saignée, delayed punching-down, addition of grape 

seed tannins, addition of ellagic-skin-seed tannins, heating of must-wine, cryo- 

89 






maceration, and prolonged maceration) on the phenolic content and antioxidant 

activity of Primitivo musts and wines. Three methods for the determination of the 

antioxidant activity were compared: DPPH, β-carotene bleaching assay and ABTS. 

Oenological parameters and composition of the phenolic fraction of 1-year wines 

was also determined. The addition of tannins allowed the increase of the phenolic 

content of musts and wines in a greater amount than the other technologies. The 

results concerning the antioxidant activity depended on the method applied. 

Concerning musts, the DPPH assay did not highlight great differences among 

technologies, whereas the addition of tannins allowed the obtainment of the 

highest antioxidant activity according to β-carotene and ABTS assays. The wine 

aging determined an increase of the antioxidant activity, independently on 

the method applied. Wine obtained through traditional technology, saignée 

and addition of tannins showed the highest antioxidant activities according 

to DPPH and β-carotene. The highest correlation coefficients (0.961 and 0.932) 

were calculated between phenolic content and ABTS values of musts whereas 

the lowest values (0.413 and 0.517) were calculated between phenolic content 

and ABTS values of wines. Wines produced through traditional technology were 

the richest in anthocyanins. The addition of tannins allowed to obtain high 

content in monomeric anthocyanins, flavonoids, flavans reactive to vanillin, and 

coumaroylated malvidin and a low content in acetylated malvidin. 

2 Changes 

in phenolic content and antioxidant activity of Italian extra-virgin 

olive oils during storage. Baiano A., Gambacorta G., Terracone C., Previtali 

M.A., Lamacchia C., La Notte E. Journal of Food Science, 74(2), 177-183, (2009). 

Impact Factor 1.255. 

Phenolic composition and antioxidant activity of 24 extra-virgin olive oils extracted 

from several Italian varieties were studied at production and during storage. The 

antioxidant activity was measured according to the following tests: in the aqueous 

phase, by radical scavenging of the 2,2’-azino-bis(3-ethylbenzthiazoline-6-sulfonic 

90 






acid (ABTS) radical cation; and in the lipid phase, using the β-carotene bleaching 

method. The phenolic content was not correlated to the oxidation indices (peroxide 

value and spectrophotometric constants). The phenolic contents and profiles of the 

various cultivars showed remarkable differences. The phenolic content was strongly 

correlated with the antioxidant activity measured according to the β–carotene test 

and weakly correlated with the radical scavenging ability. 

3 Aromatization 

of olive oil. Baiano A., Gambacorta G., La Notte E.. Monografia 

dr. S. g. Pandalai Editor of Research Signpost, (2009). 

A “flavoured olive oil” can be defined as an olive oil (generally an extra-virgin 

olive oil) that has been processed with vegetables, herbs, spices or other fruits 

in order to improve the nutritional value, enrich the sensory characteristics and 

increase shelf-life. 

Lots of different types of flavoured oil are available on the market. The 

assortment is very wide since it is possible to choose among oils flavoured with: 

vegetables, herbs, spices, mushrooms, fruits, nuts, aromas. Several methods of oil 

aromatization are available and the choice is very important since the extraction 

method affects both acceptability and stability oxidation of the flavoured oil. 

Infusion is a traditional method of oil aromatization. Another method uses a 

concentrated extract obtained by infusion of vegetables in extra-virgin olive oil. 

Then, the flavoured olive oils were prepared by adding different aliquots of the 

filtered extract to the pure extra-virgin olive oil. The safest method to obtain a 

flavoured oil is represented by the cold pressing of olives together with the herb, 

spice or vegetable. The flavoured olive oils available on sale often have a long 

shelf life guaranteed by the way of production. Unacquainted with the potential 

hazard, some consumers try to infuse flavoured oils by application of home- 

made procedure and store them also for long time at room temperature. Lots of 

techniques and methods for the evaluation of acceptability, shelf life, and quality 

of flavoured olive oils are available. 

91 






4 Evaluation 

of isoflavone content and antioxidant properties of soy-wheat 

pasta. Baiano A., Terracone C., Gambacorta G., La Notte E. international 

Journal of Food Science and technology, in press. Impact Factor 0.941. 

The purposes of this work were to determine isoflavones profile and antioxidant 

activity of two soy flours and spaghetti produced from semolina blended with 

increasing amount of these flours. Defatted and toasted soy flour (DSF and TSF, 

respectively) showed different isoflavone profiles though glucosides were the 

most abundant isoflavone forms in both the meals. The pasta-making procedure 

determined an increase of the aglycone forms that was at least 37 and 21% in 

TSF and DSF pasta, respectively. Cooking operations induced severe decreases 

of the isoflavone content (the losses ranged from 60.44 to 104.91 mg/kg for 

TSF pasta and from 85.41 to 469.33 mg/kg for DSF pasta). The results of the 

antioxidant activity were quite different as a function of the methods applied. 

A significant decrease of the antioxidant activity was caused by cooking for 

almost all samples. This result appears well correlated with the decrease of 

the total isoflavone content and the presence of antioxidant activity in the 

cooking water. 

5 Characteristics 

of drupes, phenolic content and antioxidant capacity of 

Italian olive fruits. Baiano A., Gambacorta G., Terracone C., Previtali M.A, La 

Notte E. Journal of Food Lipids, 16, 209-226, (2009). Impact Factor: 0.606. 

Phenolics, widely spread in the plant kingdom, are antioxidant compounds. With 

this study two purposes have been reached: the screening of phenolic content of 

five cereal species and the evaluation of the influence of genotype, environment 

and their interaction on the phenolic content. Drupe characteristics, phenolic 

content and antioxidant capacity of olive fruits were studied for 12 Italian oil 

cultivars grown into two areas (Cerignola and Torremaggiore) of Daunia district. 

The effects of plantation place were higher than those exerted by cultivars on drupe 

characteristics such as weight, pulp/stone ratio, oil content, and MI. Olive fruits 

92 






from Cerignola, which is located at a lower altitude than Torremaggiore, showed 

the highest phenolic content. The highest phenolic content (14 g gallic acid/kg 

dry olive) was detected on Peranzana and Cellina di Nardò grown in Cerignola 

groves. The lowest values were registered for FS17 and Cima di Melfi (about 7 

g/kg). 1-acetoxypinoresinol, 3,4-DHPEA-EDA (3,4-dihydroxyphenylethyl 

4-formyl-3-formylmethyl-4-hexenoate, dialdehydic form of elenolic acid linked 

to hydroxytyrosol), hydroxytyrosol, and luteolin were the phenolic compounds 

detected in a greater amount and in most of the cultivars. Independently on 

the assay applied, the highest antioxidant activity was detected for Cellina di 

Nardò (Cerignola). Instead, the lowest antioxidant activity values were different 

depending on the method applied. According to the ABTS (2,2-azinobis-(3- 

ethylbenzothiazoline)-6-sulfonic acid) assay, the lowest value was detected for 

Moraiolo from Torremaggiore whereas Nociara showed the lowest contribution 

to the inhibition of lipid oxidation (β-carotene beaching assay). A strong 

positive linear correlation (R=0.82) existed between total phenolic content 

and antioxidant activity measured according to the ABTS method whereas a 

low linear correlation coefficient (R=0.36) was obtained when the β-carotene 

bleaching assay was used. 

6 I 

composti fenolici degli alimenti. Baiano A., Terracone C. Aracne Editrice, 

in press. 

I composti fenolici sono un insieme estremamente disomogeneo di 

sostanze per le quali studi recenti hanno dimostrato un’azione protettiva 

sulla salute umana se assunti a livelli significativi. Essi influiscono inoltre 

su caratteristiche quali la qualità, l’accettabilità e la conservazione dei 

prodotti alimentari, agendo come aromatizzanti, coloranti e antiossidanti. Il 

contenuto e la disponibilità dei composti fenolici nei tessuti vegetali varia in 

funzione di specie, varietà, organo considerato, stadio fisiologico, condizioni 

pedoclimatiche, modalità di conservazione e trasformazione. Le autrici 

93 






hanno deciso di dedicare i 5 capitoli del libro alla classificazione dei composti 

fenolici, alla loro distribuzione negli alimenti e alle metodiche di estrazione, 

quantificazione e determinazione dell’attività antiossidante, proponendo 

un’opera che combina il rigore scientifico alla facilità di consultazione da parte 

di un pubblico eterogeneo. 

7 Analisi 

sensoriale. Baiano A., Perniola R. Capitolo del libro “uva da 

tavola”, 2010. 

L’analisi sensoriale è una disciplina scientifica che consente di quantificare, 

identificare e descrivere quelle caratteristiche di un prodotto alimentare capaci 

di dare origine a stimoli di natura chimica, chimico-fisica e fisica ai quali gli organi 

di senso (olfatto, gusto e vista, in particolare) reagiscono. La qualità dell’uva da 

tavola è correlata a caratteristiche quali: l’assenza di semi; il colore, la forma e 

l’uniformità degli acini; la consistenza e la croccantezza; la dolcezza, l’acidità e 

l’aroma. La valutazione sensoriale dell’uva da tavola è generalmente effettuata 

applicando la cosiddetta “Analisi Quantitativa Descrittiva” (QDA), che consiste 

nel proporre una serie di descrittori visivi, olfattivi e gustativo-tattili, da valutare 

generalmente su una scala strutturata. 

8 Succhi 

d’uva. Baiano A., La Notte E., Lovino R. Capitolo del libro “uva da 


Secondo la Direttiva 2001/112/CE, con il termine succo di frutta si intende “il 

prodotto fermentescibile ma non fermentato, ottenuto da frutta sana e matura, 

fresca o conservata al freddo, appartenente a una o più specie e avente il colore, 

l’aroma e il gusto caratteristici dei succhi di frutta da cui proviene. L’aroma, la 

polpa e le cellule del succo che sono separati durante la lavorazione possono 

essere restituiti allo stesso succo”. Oggi è possibile trovare sul mercato succhi 

d’uva propriamente detti e prodotti assimilabili ai succhi d’uva ma la cui 

composizione e le cui modalità produttive non ne consentono l’inquadramento 

94 






nell’ambito delle denominazioni previste dalla direttiva sopra citata. Questa 

diversificazione delle referenze è dovuta al fatto che i consumatori, quelli Italia- 

ni in particolare, trovano il succo d’uva tal quale eccessivamente dolce, poco 

acido e di aroma poco gradevole e quindi la ricerca alimentare si è orientata 

verso lo sviluppo di succhi a base d’uva ma con aggiunta di succhi di altri frutti 

e/o verdure che, oltre ad aumentarne l’accettabilità sensoriale, determinano 

un incremento della concentrazione delle sostanze nutraceutiche (vitamine, 

polifenoli ed altre molecole). 

9 Confetture. 

Baiano A., Milella R.A., Crupi P. Capitolo del libro “uva da 


Si definisce confettura (D. Lgs. N. 50 del 20 febbraio 2004) un prodotto a base di 

zucchero e frutta (ad eccezione degli agrumi) o, in taluni casi, di verdure (cipolle, 

melanzane, pomodori verdi, zucca) in cui la quantità di frutta impiegata deve 

essere pari ad almeno il 35% per la confettura e al 45% per la confettura extra. A 

causa della sua esclusione dal settore vitivinicolo, l’uva da tavola eccedente o non 

idonea al consumo fresco è avviata alla produzione di succhi d’uva (il cui mercato 

è però ancora poco sviluppato) o alla distillazione per la produzione di alcool. In 

considerazione della sua concentrazione in sostanze nutraceutiche (antiossidanti 

quali i polifenoli) e all’elevato tenore in solidi solubili, l’utilizzo di uva da tavola per 

l’ottenimento di confetture può rappresentare una valida alternativa al consumo 

del prodotto fresco. 

10 

Effects of processing on isoflavone content and profile in foodstuffs: a 

review. Baiano A. Capitolo del libro “isoflavones: Biosynthesis, Occurrence 

and health Effects”, accettato per la pubblicazione. 

Isoflavones, together with lignans and coumestans represent the three classes 

of phytoestrogens, compounds that have an estrogen-like and antioxidant 

activity and are structurally similar to estradiol. Isoflavones are present under 

95 






several chemical forms: the β-glucosides genistin, glycitin, daidzin, their 6”- 

O-malonyl-glucosides and a 6”-O-acetyl-glucosides and the aglycone forms 

genistein, daidzein and glycitein. Processing greatly affect the content of total 

isoflavones and the concentrations of each of their chemical forms. Among 

soy foods, the highest concentration of isoflavones is detectable in the whole 

soybeans whereas soy milk, tofu, tempeh, miso, fermented bean curd, bean 

paste and the so called “second-generation soyfoods” (soy hot dog, tempeh 

burger, soy noodles) show isoflavone contents ranging from 2 to 46% with 

respect to the whole soybeans. These reductions are mainly due to dilution 

of soy protein with other ingredients and heat treatments that also result in 

changes of the isoflavone profiles. In fact, the predominant isoflavones in 

soybeans and non-fermented soy products (soy protein, soymilk derivates, etc.) 

are malonyl glucosides, whereas fermented soy foods (miso, tempeh) contain 

mainly aglycones. These changes affect the metabolic activity of isoflavone- 

containing foods since the glycoside forms are biologically inactive and need to 

be cleaved by the β-glucosidases from intestine in the glucosyl residue and the 

biologically active aglycones. 

96 


stuDio Delle ProDuzioni 

alimentari tiPiche 

attraverso l’aPPlicazione 

Di tecniche innovative 






Persona da contattare 






Parole chiave 

CEREALI 

COMPORTAMENTO 

PASTA 

ALLA COTTURA 

PRODOTTI TIPICI 

TECNICHE ANALITICHE 

INNOVATIVE 


Sono definiti “tipici” quei prodotti differenziabili da altri presenti perché provvisti di 

particolari caratteri organolettici dovuti a fattori geografici, alla qualità della materia 

prima impiegata o, in altre parole, alle tecniche di lavorazione che distinguono la 

produzione di una località da quella conseguita altrove. In un’epoca caratterizzata 

dall’economia di mercato, dalla rivoluzione delle telecomunicazioni e da un rapido 

diffondersi dell’informazione, la globalizzazione crea certamente uno spazio 

commercialmente e finanziariamente integrato, ricco di scambi e dominato da 

aziende impegnate in strategie di fusione e acquisizione tali da assicurare loro 

posizioni dominanti sul mercato mondiale. Discorso diverso per quanto riguarda 

l’Italia, in cui il settore agro-alimentare è caratterizzato, per lo più, da aziende di piccole 

dimensioni, difficilmente in grado di competere all’interno di un siffatto mercato 

globale. In tal caso, la strategia vincente è rappresentata dalla diversificazione e dalla 

valorizzazione dei prodotti, puntando sul binomio qualità-informazione capillare 

del consumatore. L’obiettivo di questo lavoro di ricerca è stato quello di pervenire 

alla completa caratterizzazione dei prodotti tradizionali e tipici presenti sul mercato, 

alla valutazione degli effetti di modifiche composizionali sulle loro caratteristiche 

chimico-fisiche e alla tracciabilità attraverso la determinazione di indici chimico- 

fisici misurati mediante tecniche analitiche innovative. 


1 

Use of a toasted durum whole meal in the production of a traditional Italian 

pasta: chemical, mechanical, sensory and image analyses. Baiano A., Fares C., 

Peri G., Romaniello R., Taurino A.M., Siciliano P., Gambacorta G., Lamacchia 

C., Pati S., La Notte E. international Journal of Food Science & technology, 43, 

1610-1618, (2008). Impact Factor 0.941. 

The characterisation of traditional Italian pasta obtained by mixing amounts of 

toasted whole meal with remilled semolina and other ingredients was obtained 

97 






by means of physico-chemical, rheological, mechanical, sensory and image 

analyses. The toasted meal showed higher ash, fibre and protein contents than re- 

milled semolina. The replacement of percentages of re-milled semolina with the 

toasted meal and soft flour increased tenacity and decreased extensibility and 

strength, making the dough less suitable for pasta-making. The P⁄L values were 

indices of high starch damage. The replacement of part of re-milled semolina and 

water with toasted whole wheat meal, soft flour and eggs increased the optimal 

cooking time and the amount of water absorbed during cooking but made the 

other cooking parameters worse. The image analysis provided evidence of the 

changes induced by the use of toasted wholemeal, soft flour and eggs in the 

microscopic structure of pasta protein and starch. 

2 Electronic 

nose and GC-MS as powerful tools to investigate the volatile 

component of an italian traditional pasta made from toasted durum wheat. 

Baiano A., Siciliano P., Taurino A.M., Presicce D.S., Gambacorta G., Lamacchia 

C., Pati S., La Notte E. Cereal Foods World, May-June, 138-143, (2007). Impact 

Factor 0.545. 

The present work investigated the possibility of combining headspace sampling– 

solid phase microextraction with gas chromatography–mass spectroscopy (GC-MS) 

analysis and an array of semiconductor gas sensors to study the volatile fraction of 

traditional Italian cereal foods. In particular, it studied a pasta recognized as typical 

by the Italian laws and made with a semolina obtained from a toasted durum 

wheat. The GC-MS analysis allowed the identification of several volatile compounds, 

almost always in small concentrations, in raw materials and in fresh and dried pasta. 

The data deriving from the application of the GC-MS and electronic nose analyses 

were submitted to principal component analysis to evaluate the suitability of these 

techniques for emphasizing differences due to composition and/or processing. 

These preliminary results showed that both the analytical techniques allowed good 

discrimination among the meals but not among the pasta samples. 

98 






3 Physical 

and mechanical properties of bread loaves produced by 

incorporation of two types of toasted durum wheat flour. Baiano A., 

Romaniello R., Lamacchia C., La Notte E. Journal of Food Engineering. In 

press. Impact Factor 1.848. 

The aim was to study the effects of toasting of durum wheat kernels on the 

physical and mechanical properties of bread containing increasing amounts 

of toasted wholemeal. Two toasting procedures were considered: heating of 

the kernels arranged on tin cans (TDW1) and the stubble burning (TDW2). The 

addition of toasted flour determined a decrease in volume (greater when TDW2 

was used) and detrimental effects on mechanical properties caused by the 

thermal denaturation of the gluten-forming proteins and the interference of bran. 

Volume was negatively correlated with compressibility, gumminess, masticability 

and positively correlated with elasticity (0.824 < R < 0.971). The bread structural 

parameters were quantified by digital image analysis and correlated to the 

variations of the mechanical properties. The addition of increasing percentages 

of toasted wheat flour (especially, TDW1) determined significant increases of 

the cell density, and decreases of cell mean area, and cell equivalent diameter. 

Statistically significant correlations were obtained between cell density or 

equivalent diameter and the mechanical properties of all the bread types 

according to a second order polynomial regression but only the former allowed 

to predict approximately 60% of the variation of crumb compressibility and 87% 

of the variation of elasticity and gumminess both in bread produced with TDW1 

and in bread produced with TDW2. 

99 


aPPlicazione Degli stanDarD 

Di certificazione 









Parole chiave 

AzIONI CORRETTIVE 

AzIONI PREVENTIVE 

CERTIFICAzIONE 

CHECK-LIST 

QUALITà 


Gli standard di qualità, se applicati, consentono alle aziende di conseguire risultati 

sia in termini di immagine che in termini di migliore gestione interna. L’adozione 

degli standard di qualità è un’azione meramente volontaria e, dunque, solo chi 

desidera investire tempo, risorse economiche e umane in tale attività, ha accesso a 

un mondo di eccellenza. Le università sono oramai assimilabili a strutture aziendali 

in cui la conduzione necessita di un approccio manageriale alla prevenzione e alla 

risoluzione dei problemi. 


1 

Valutazione della didattica universitari. Baiano A., Carlucci G. Aracne 

Editrice (2009). 

La riforma degli ordinamenti didattici ha determinato l’esigenza di promuovere 

la qualità negli atenei, attraverso la razionalizzazione dei sistemi di gestione e 

il miglioramento della didattica erogata dai corsi di laurea. Il raggiungimento 

di tali obiettivi può indubbiamente essere agevolato dall’adozione di sistemi di 

gestione conformi alla norma ISO 9001:2008 e pertanto certificabili da parte di 

un ente terzo. Questo manuale si propone come guida e strumento operativo 

per l’implementazione dei requisiti di tale norma nei corsi di laurea triennali/ 

magistrali. Esso descrive dettagliatamente gli elementi di cui si compone un 

sistema di gestione per la qualità e offre una guida alla redazione del manuale e di 

tutti gli altri documenti della qualità. Particolare attenzione è riservata al sistema 

di monitoraggio degli stakeholders, da attuarsi mediante somministrazione di 

questionari, al fine di accrescere la soddisfazione del cliente e realizzare il principio 

del miglioramento continuo. 

100 


stuDio Degli effetti Dei Processi 

tecnologici sulle ProPrietà 

strutturali e funzionali 

Delle Proteine Degli alimenti 






Barbara La Gatta, tecnico Laureato 

Sara Lamparelli, Post doc 


Ennio La Notte, Professore Ordinario, Dipartimento 

di Scienze degli Alimenti (DISA), Facoltà di Agraria, 

Università di Foggia, Via Napoli 25, 71122 Foggia; 


Parole chiave 

PASTA PROTEINS; 

DRYING CYCLES; 

TOASTING PROCESS; 

UNEXTRACTABLE POLYMERIC 

PROTEINS 

TECHNOLOGICAL PROCESSES 


Le proprietà organolettiche e nutrizionali degli alimenti derivano essenzialmente 

da due categorie: 

• intrinseche del prodotto, frutto dei metabolismi cellulari come proteine, lipidi 

e polisaccaridi e piccoli metaboliti di natura organica; 

• l’altra è rappresentata da molecole che possono essere assorbite dall’ambiente 

esterno e veicolate dai fluidi biologici o essere di neo formazione dovuta ad 

aggregazioni molecolari a cambiamenti molecolari per effetto delle tecnologie 

di trasformazione e di conservazione. 

Le scienze ohmiche in campo alimentare hanno permesso di studiare la struttura 

delle molecole di base che costituiscono un alimento e le trasformazione che esse 

subiscono nel corso della manipolazione ancorché l’uomo consumi l’alimento 

fresco, trasformato e/o conservato. 

L’applicazione di queste scienze ha portato alla definizione dei processi 

proteolitici negli alimenti e all’individuazione e alla caratterizzazione molecolare 

di componenti marcatori di prodotto e processo nonché all’attitudine della 

materia prima alla trasformazione. Attualmente sono in corso ricerche sui 

cambiamenti strutturali indotti dalle temperature di essiccazione e/o di 

tostatura delle proteine del glutine e le relazioni con il comportamento delle 

paste prodotte alla cottura. 


1 

Changes in pasta proteins induced by drying cycles and their relationship 

to cooking behaviour. C. Lamacchia, A. Di Luccia, A. Baiano, G. Gambacorta, 

B. la Gatta, S. Pati, E. La Notte. Journal of Cereal Science, 46, 58-63. (2007). 

Impact Factor: 1.905. 

Spaghetti produced in a pilot plant were dried using three different drying 

conditions: 60°C for 7.5h; 75°C for 5.5h; 90°C for 5h. Proteins of spaghetti were 

101 






characterized by size exclusion chromatography (SEC) and size exclusion– 

high performance liquid chromatography (SE-HPLC). As the temperature of 

the drying cycles increased, a progressive decrease of the small and large 

monomeric proteins and an increase in molecular size of the large polymeric 

proteins were observed. Drying temperature at 60 o C already induces a 

significant increase in the molecular size of the large polymeric proteins and 

a significant decrease of the other protein fractions. At 70 and 90 o C, large and 

small monomeric proteins as well as large polymeric proteins polymerized as 

seen by a progressive shift towards higher molecular weights in the SEC profile 

as temperature increased. The changes in the chromatographic profiles were 

accompanied by increasing amount of total UPP. A decrease in stickiness and 

an increase in firmness corresponded to the formation of large and insoluble 

protein aggregates. 

2 Changes 

in durum wheat kernel and pasta proteins induced by toasting 

and drying processes. C. Lamacchia, A. Baiano, S. Lamparelli, E. La Notte, A. 

Di Luccia. Food Chemistry, 118, 191-198. (2010). Impact Factor: 3.054. 

Durum wheat kernels were subjected to a toasting process and the proteins 

characterized by size exclusion-high performance liquid chromatography (SE- 

HPLC) and sodium dodecyl sulphate-polyacrylamide gel electrophoresis. With 

this physical process, albumins and globulins, as well as glutenins and gliadins, 

polymerized as seen by a shift of the SE-HPLC profile to lower elution times. 

The polymerization seemed to happen mainly through disulphide bonds, 

even though the participation of ω-gliadins to the aggregation suggested 

the involvement of other kinds of interactions. It led to the revelation of a 

new peak originated by thermal aggregation of small polymeric proteins. 

The changes in the chromatographic profile were accompanied by increasing 

amounts of total unextractable polymeric proteins. The replacement of 

semolina with toasted durum wheat flour (5, 10, 15, 20 and 30%) for the 

102 






production of pasta in the shape of spaghetti significantly (p0.05) change the unextractable polymeric proteins 

(UPP) when compared with spaghetti made with 100% durum semolina. On 

the other hand, the replacements of semolina with 15-30% TDWF showed 

significant (p

stuDio Delle relazioni esistenti 

tra le Proteine e le ProPrietà 

tecnologiche Di sfarinati Di 

frumento e sfarinati gluten free 






Barbara La Gatta, tecnico Laureato 

Sara Lamparelli, Post doc 


Ennio La Notte, Professore Ordinario, Dipartimento 

di Scienze degli Alimenti (DISA), Facoltà di Agraria, 

Università di Foggia, Via Napoli 25, 71122 Foggia; 


Parole chiave 

WHEAT FLOUR 

NON CONVENTIONAL FLOURS 

GLUTENIN INDEX 

TECHNOLOGICAL PROPERTIES 


Le proteine del frumento rappresentano solo il 10-15% circa della granella 

matura. Tuttavia, esse determinano la qualità del prodotto finito in termini di 

caratteristiche nutrizionali e performance tecnologiche. Queste ultime sono 

influenzate da un gruppo di proteine di riserva denominate prolammine che 

rappresentano il 60% circa dell’azoto totale presente nella cariosside di frumento. 

Quando la farina è miscelata con acqua, le prolammine formano una massa visco- 

elastica, il glutine, che può essere frazionata, utilizzando soluzioni idroalcoliche, 

nelle gliadine solubili e monomeriche, e nelle insolubili e aggregate glutenine. 

Entrambe le frazioni sono coesive ma il loro contributo alle proprietà reologiche 

degli impasti è differente. L’attività di ricerca si prefigge: di descrivere le relazioni 

esistenti tra queste proteine e le proprietà tecnologiche delle farine di frumento in 

quanto risulta ancora difficile predire le performance tecnologiche degli impasti 

solo sulla base della composizione allelica delle varietà di frumento rispetto alla 

quantità di subunità gluteniniche presenti. Inoltre l’attività si prefigge di descrivere 

le relazioni esistenti tra le proteine di sfarinati non convenzionali gluten free e le 

loro proprietà tecnologiche al fine di predirne le performance in pastificazione. 


1 

Application of capillary electrophoresis to determine the technological 

properties of wheat flours by a glutenin index. A. Di Luccia, C. Lamacchia, G. 

Mamone, G. Picariello, A. Trani, P. Masi, F. Addeo. Journal of Food Science,74, 

4, 307-311. (2009). Impact Factor: 1.255. 

Capillary electrophoresis was used to characterise glutenin proteins from ancient 

varieties of Southern Italy common wheat and to determine the technological 

properties of wheat flours based on a glutenin index. Three zones were identified 

in the electropherograms, indicated as A, B and C according to electroelution 

order. The three zones corresponded to the low molecular weight glutenin 

104 






subunits and to the y- and x- type high molecular weight subunits, respectively. 

The ratio B/C, was correlated to the alveographic parameter P/L . These results 

indicated that flours resulting in a B/C ratio lower than 2 produced elastic doughs 

whereas flours resulting in a B/C ratio higher than 2 produced doughs more 

resistant to extension. This study showed that capillary electrophoresis is useful 

for determining the types and quantities of gluten proteins in the evaluation of 

wheat-flour technological properties of a limited number of non-commercial 

varieties as evidenced by the x-type content which seems to strongly influenced 

the flour technological parameters. 

2 Amaranth, 

quinoa and oat doughs: mechanical and rheological behaviour, 

polymeric size distribution and extractability. C. Lamacchia, S. Chillo, 

S. Lamparelli, N. Suriano, M.A. Del Nobile. Journal of Food Engineering, 

96, 97-106. (2010). Impact Factor: 1.848. 

The rheological characteristics, static and dynamic-mechanical properties of 

amaranth, quinoa and oat doughs and the relative size distribution of their 

polymeric proteins were evaluated. For the sake of comparison, semolina dough 

rheological and mechanical properties and the relative size distribution of 

proteins were also determined. 

From rheological results it was inferred that the tenacity of amaranth, oat and 

quinoa dough samples was lower than that of semolina dough. The elastic 

modulus (Ec) of amaranth, oat and quinoa doughs was higher than that of 

semolina dough. Amaranth and quinoa G' was found to be similar and significantly 

higher (p





lower (p

film eDibili 


Amalia Conte, Ricercatore non confermato 

Marcella Mastromatteo, Assegnista di ricerca 


Matteo Alessandro Del Nobile, Professore Ordinario, 




tel. 0881-589242; ma.delnobile@unifg.it 

Parole chiave 

FILM EDIBILI 

CHITOSANO 

AMIDO 

GLUTINE 

CRUSCA 


Questo filone di ricerca è finalizzato allo sviluppo e alla caratterizzazione 

di film edibili a base proteica e polisaccaridica e relativi compositi. I sistemi 

compositi sono ottenuti rinforzando le matrici di base con polveri edibili. Il 

lavoro di ricerca mira a determinare l’influenza della composizione chimica sulle 

proprietà viscoelastiche delle soluzioni polimeriche da cui si ottengono i film, 

sulla permeabilità al vapor d’acqua e sulle proprietà meccaniche delle matrici 

polimeriche e sulla velocità di rilascio di sostanze a basso peso molecolare 

inglobate nel polimero. 


1 

Rheological characterization of deacylated/acylated-gellan films carrying 

L-(+)-ascorbic acid. Leon P., Chillo S., Conte A., Gerschenson L., Del Nobile 

M.A., Rojas A.M. Food hydrocolloids. 23, 1660-1669. (2009) Impact 

Factor: 2.511. 

Films developed for supporting L-(+)-ascorbic acid (AA), natural antioxidant for 

the protection of foods, were rheologically evaluated. A film was formulated by 

mixing gellan gum with its acylated form to attain a higher AA stability and lower 

non-enzymatic browning. The polymer mixture allowed obtaining a less rigid 

film, permitting the usage of a lower proportion of glycerol. Mechanical spectra of 

films evidenced solid-like materials provided the time scale of the observation was 

shorter than the lifetime of the physical cross-links involved. There was evidence 

for a shorter time of relaxation in the films stored at 33.3–57.7% of relative 

humidity, which influenced small deformation (linear range) measurements. Out 

of linear viscoelasticity a greater hardening effect was observed for A- and B-films 

and related to enhanced polymer interactions in gellan based films. Its increase 

with the moisture content may be a result of macromolecular association. Higher 

rigidity characterized gellan based films (A- and B-networks) a intermediate 

107 


film eDibili 

deformations, which may also lead to higher softening degree till rupture. Smaller 

hardening and softening degrees were observed for C- and D-systems. This can 

be ascribed to spatial hindering in the film network by acylated gellan chains; this 

fact worsened polymer interactions. Atomic force microscopy images reinforced 

rheological conclusions. 

2 Influence 

of spelt bran on the physical properties of WPI composite films. 

Mastromatteo M., Chillo S., Buonocore G.G., Massaro A., Conte A., Bevilacqua 

A., Del Nobile M.A. Journal of Food Engineering. 92, 467-473. (2009). Impact 


The film properties of whey protein isolate (WPI) with different levels of glycerol 

and spelt bran were investigated using response surface methodology. Tests 

were carried out to determine to what extent the water vapour permeability 

(WVP) of whey protein isolate films can be improved by adding the bran. 

Moreover, mechanical and dynamical properties, colour of the films as well as 

the rheological properties of the film forming solutions were also investigated. 

Results highlight that the WVP was significantly affected by the spelt bran 

concentration. Increasing the spelt bran concentration led to decrease in 

water vapour permeability, whereas the glycerol concentration incremented 

the WVP values of the films. The elastic modulus (Ec) and complex modulus 

(E*) of the composite films increased with the decrease of the glycerol content 

and the increase of the bran concentration. Glycerol addition acting as a 

plasticizer led to more flexible films. Rheological characteristics of the film 

forming solutions were also studied. The WPI solutions with the lower bran 

concentration behaved as Newtonian fluid; as the bran concentration increased 

the solutions demonstrated a non-Newtonian behaviour. Colour results 

highlight that Yellow Index (YI), a and b parameter of Hunter scale increased 

with the bran concentration. On the contrary, L parameter decreased with the 

bran concentration increase. 

108 


film eDibili 

3 Influence 

of Glycerol and Chitosan on Tapioca Starch-based Edible Film Properties. 

Chillo S., Flores S., Mastromatteo M., Conte A., Gerschenson L., Del Nobile 

M.A. Journal of Food Engineering. 88, 159-168. (2008). Impact Factor: 1.848. 

The individual and interactive effects of glycerol and chitosan on the tapioca 

starch-based edible film properties were investigated using response surface 

methodology. Tests were run on the polymeric matrixes to determine film forming 

solution apparent viscosity, mechanical and dynamic-mechanical properties, 

water vapour permeability (WVP) and color. All film forming solutions presented 

pseudoplastic behaviour. The Elastic modulus (Ec) and the tensile strength were 

positively influenced by chitosan and negatively by chitosan-glycerol interaction. 

The glycerol concentration had no effect on the E’ (Storage modulus) values and 

had a positive influence on the E’’ (Loss modulus) values. Moreover, the chitosan 

percentage affected positively both E’ and E’’ values. Tanδ values increased with 

increasing glycerol concentration and a slight decrease was observed as the 

chitosan concentration decreased. Relating to WVP data, the chitosan addition 

had a negative effect, whereas the glycerol one had a positive influence. The color 

results showed that Yellow Index and b parameter of Hunter scale increased with 

chitosan concentration, whereas the L values decreased. The glycerol increasing 

concentration produced an increase in Yellow Index and b value. 

4 Effects 

of Spelt and Wheat Bran on the Performances of Wheat Gluten Films. 

Mastromatteo M., Chillo S., Buonocore G.G., Massaro A., Conte A., Del Nobile 

M.A. Journal of Food Engineering. 88, 202-212. (2008). Impact Factor: 1.848. 

The individual and interactive effects of the spelt and wheat bran as well as 

glycerol, on the properties of wheat gluten edible film were investigated using 

response surface methodology. Tests were run to determine water vapour 

permeability, mechanical and dynamical properties and colour of the film as well 

as the rheological properties of the film forming solution. Results highlight that the 

glycerol presence had a negative effect on water vapor permeability values of the 

109 


film eDibili 

films (increase of permeability), whereas the bran presence had a positive influence 

(decrease of permeability). The Elastic modulus of the composite film increased 

with the increase of bran concentration and with the decrease of glycerol. The 

tenacity increased with the increase of glycerol and spelt bran up to a threshold 

value after which a decrease was observed. The complex modulus of the composite 

film increased with the decrease of the glycerol concentration and with the increase 

of the wheat bran. All film forming solutions presented pseudoplastic behaviour; 

moreover, the apparent viscosity increased with the increase of glycerol and bran 

up to a threshold value. Both complex shear modulus and complex viscosity were 

affected by a positive interaction between spelt bran and wheat bran. Results also 

showed that Yellow Index (YI) and b parameter of Hunter scale increased with 

the bran concentration, whereas the L values decreased. The glycerol increase 

determined a decrease in the YI and b value and an increase in L value. 

5 Mass 

Transport Properties of tapioca-Based Active Edible Films. Flores 

S., Conte A., Campos, C., Gerschenson L., Del Nobile, M.A. Journal of Food 

Engineering. 81, 580-586 (2007). Impact Factor: 1.848. 

The release of potassium sorbate from tapioca starch-based edible films with 

glycerol as plasticizer was studied and modeled. Tapioca importance as a source 

of starch is growing rapidly, especially because its price in the world market is low 

when compared to starches from other sources. A semi-empirical approach was 

proposed to describe the diffusion of water molecules into the polymeric film, 

the counter diffusion of the antimicrobial compound and the polymeric matrix 

swelling kinetic. Despite the simplicity of the proposed model, it successfully 

fitted the experimental data. From the obtained results it was observed that 

casting technique, producing films with high amorphous degree, determined 

a greater contribution of matrix relaxation to sorbate release. The pH of the 

receiving media contributed slightly to antimicrobial release and its effects seem 

to be related to the influence of this variable on antimicrobial solubility. 

110 


Packaging attivo 



Incoronato Anna Lucia, dottoranda 


Sinigaglia Milena, Professore Ordinario 






tel. 0881 589 242; ma.delnobile@unifg.it 

Parole chiave 

FILM ATTIVI 

COMPOSTI NATURALI 

TRATTAMENTI VIA PLASMA 

AGENTI RETICOLANTI 

IMMOBILIzzAzIONE/RILASCIO 


Il lavoro di ricerca relativo a tale filone riguarda principalmente lo studio di due 

tipologie di imballaggi attivi: quelli basati sul rilascio di sostanze attive con azione 

antimicrobica, e quelli che prevedono l’immobilizzazione della sostanza attiva. 

Nell’ambito di quest’ultima tipologia di film attivi sono state messe a punto 

diverse tecniche di immobilizzazione basate sull’uso di agenti reticolanti o su 

tecniche di immobilizzazione via plasma. L’attività antimicrobica dei film attivi 

sviluppati è stata testata sia su sistemi simulanti l’alimento che in reali matrici 

alimentare a base liquida. 


1 

Active packaging by extrusion processing of recyclable and biodegradable 

polymers. Del Nobile M.A., Conte A., Buonocore G.G., Incoronato A.L., 

Massaro A., Panza O. Journal of Food Engineering. 93, 1-6. (2009). Impact 


In this work a preliminary study on the suitability of environmentally friendly active 

films to be produced by melt extrusion process is presented. In particular, low density 

polyethylene (LDPE), polylactic acid (PLA) and polycaprolactone (PCL) were used as 

environmentally friendly polymeric matrices, with the former being recyclable and 

the latter two biodegradable. Lemon extract, thymol, and lysozyme at three different 

concentrations were incorporated in each polymeric matrix, respectively, to have a 

total of nine active films. Results suggest that processing temperatures play a major 

role in determining the antimicrobial efficiency of the investigated active films. In 

particular, antimicrobials incorporated into PLA and LDPE showed retained slight 

antimicrobial activity, thus having lost some activity due to higher processing 

temperature. On the other hand, PCL was processed at a lower temperature, which 

allowed less degradation of antimicrobial activity. Among the natural compounds 

under study lysozyme showed the higher thermal stability. 

111 



2 Controlled 

release of thymol from zein based film. Mastromatteo M., 

Barbuzzi G., Conte A., Del Nobile M. A. innovative Food Science and Emergine 

technologies, 10, 222-227. (2009). Impact Factor: 1.713. 

Active packaging materials, able to release antimicrobial compounds into 

foodstuffs, can be used in order to inhibit or slow down bacterial growth during 

storage. zein-based mono and multilayer films were loaded with spelt bran 

and thymol (35%w/w) to obtain edible composite polymeric materials. Various 

composite systems were developed to control the release of thymol. In particular, 

the thickness of the layers and the biodegradable fibers amount were varied. 

Results highlight that thymol release rate decreased with the increase of the 

film thickness for both mono and multilayer films, without spelt bran addition. 

Conversely, a significant increase of thymol release rate with the increase of the 

bran concentration is recorded for both mono and multilayer films. 

3 Antimicrobial 

efficacy and release kinetics of thymol from zein Films. Del 

Nobile M.A., Conte A., Incoronato A.L., Panza O. Journal of Food Engineering. 

89, 57-63. (2008). Impact Factor: 1.848. 

A comprehensive study on thymol loaded zein films is presented in this work. 

In particular, the antimicrobial efficacy of developed active zein films was tested 

against three different spoilage microorganisms (cells and spores of Bacillus cereus, 

Candida lusitaniae and Pseudomonas spp) and one GRAS (Generally recognized as 

safe) microbial strain (Streptococcus thermophilus). Results show that the films 

are effective against the selected spoilage food borne microorganisms, without 

compromising the growth of S. thermophilus. Release tests were also run at 25°C 

to investigate the thymol release mechanism. Results show that the Fick’s Second 

Law satisfactorily describes the release kinetic. Moreover, it was also found that 

the thymol diffusion coefficient does not depend to a great extent on the thymol 

concentration. On the other hand, thymol sorption isotherm in zein showed an 

upward concavity as in the case of low molecular weight compound sorption 

112 



in polymeric matrix. Finally, antimicrobial effectiveness of developed active 

film as compared to thymol directly added to the inoculated growing medium 

was assessed on Pseudomonas spp by comparing results obtained from direct 

addiction of thymol to that obtained from released thymol. Results show that 

differences are negligible. 


activity of immobilized lysozyme on plasma treated 

polyethylene films. Conte A., Buonocore G.G., Sinigaglia M., Lopez L.C., 

Favia P., d’Agostino R., Del Nobile M.A. Journal of Food Protection. 71, 119- 

125. (2008). Impact Factor: 2.154. 

In this paper we tested the antimicrobial activity of polyethylene films modified 

by means of plasma processes followed by the chemical immobilization of 

lysozyme, an antimicrobial enzyme. In order to chemically immobilize the 

enzyme in its active form at the surface of polyethylene, substrates plasma- 

treated in different experimental conditions were soaked in lysozyme solutions 

at different concentrations. The immobilization of the enzyme was checked and 

the antimicrobial activity of the films was investigated by observing the death 

rate of Micrococcus lysodeikticus cells suspended in phosphate buffer in contact 

with the films. The results clearly indicate that plasma-treated films loaded with 

lysozyme are active against the selected microorganism. A modified version of 

the Gompertz equation was used to quantitatively valuate the dependence of 

the antimicrobial activity of the films, on both plasma treatment conditions and 

lysozyme concentrations. 

5 Development 

of immobilized lysozyme based active film. Conte A., 

Buonocore G.G., Sinigaglia M., Del Nobile M.A. Journal of Food Engineering. 

78, 741-745. (2007). Impact Factor: 1.848. 

The aim of this work is to develop an active packaging material in which the 

active compound, lysozyme, is completely immobilized onto the polymeric 

113 



material and acts directly from the film without being released into the packed 

foodstuff. Cross-linked films of polyvinyl alcohol, differing in the amount of 

both antimicrobial and binding agents, were prepared. In order to determine 

the amount of active substance completely bonded to the polymer backbone, 

lysozyme release tests were run. The antimicrobial activity of the developed active 

films was investigated in order to verify their effectiveness in controlling microbial 

spoilage. A modified version of Gompertz equation was used to quantitatively 

determine the antimicrobial activity of the films. Results indicate the developed 

active films are effective in inhibiting the growth of selected microorganism, and 

that the antimicrobial activity of the investigated films increases as the amount of 

enzyme incorporated increases. 


effectiveness of lysozyme immobilized on polyvinylalcohol- 

based film against Alicyclobacillus acidoterrestris. Conte A., Sinigaglia M., 

Del Nobile M.A. Journal of Food Protection. 69(4), 861-866. (2006). Impact 


In this work the effectiveness of active polyvinylalcohol (PVOH) based film against 

Alicyclobacillus acidoterrestris was assessed. The developed active film was 

obtained by immobilizing an active compound on the surface of the polymeric 

matrix and by putting the obtained film in contact with a medium inoculated 

with microbial cells. Microbiological tests were conducted with success both on a 

single strain and on culture cocktail of A. acidoterrestris at 44°C. By monitoring the 

viable cell concentration at three different packaging conditions it was possible 

to demonstrate that the active film, with a comparable efficacy for both single 

strain and culture cocktail, maintained its efficacy at different medium volumes. 

The same microbial tests were conducted also on viable spores of the investigated 

microorganism, inoculated both in laboratory medium and in apple juice. The 

results indicate that the developed active film successfully inhibits, more easily 

than cells, the viable spores in both investigated media. 

114 



7 Immobilization 

of lysozyme on polyvinylalcohol Ffilms for active packaging 

applications. Conte A., Buonocore G.G., Bevilacqua A., Sinigaglia M., Del 

Nobile M.A. Journal of Food Protection. 69(4), 866-870. (2006). Impact 


A new technique for the immobilization of lysozyme onto the surface of 

polyvinylalcohol films is presented. The active compound was sprayed along with 

a suitable bonding agent onto the surface of the cross-linked polymeric matrix. 

Active compound release tests determined the amount of lysozyme immobilized 

on the film surface. By using Micrococcus lysodeikticus the antimicrobial activity 

of the films was determined and the results correlated with the amount of 

immobilized lysozyme. This new technique was effective for immobilizing the 

enzyme and the developed films were active against the test microorganism. 

Results were compared with those obtained with a different immobilizing 

technique, in which the active compound was bound into the bulk of the 

polymeric film. As expected, the surface immobilized lysozyme films have a 

higher antimicrobial activity than bulk bound films. 

115 


confezionamento Di 

ProDotti Di iv gamma 



Annalisa Lucera, dottoranda 

Cristina Costa, dottoranda 

Janine Laverse, tecnico laureato 

Lucia Lecce, tecnico diplomato 

Sinigaglia Milena, Professore Ordinario 







Parole chiave 

PRODOTTI READY-TO-EAT/ 

READY-TO-COOK 

DIPPING 

COATING 

SHELF LIFE 


Questo filone di ricerca è finalizzato all’estensione della shelf life dei prodotti di IV 

Gamma. In particolare, l’attività di ricerca si prefigge di descrivere l’attività respiratoria 

di tali prodotti, determinare l’influenza della confezione sulla conservabilità di questi 

alimenti e individuare le condizioni ottimali di confezionamento per consentire un 

prolungamento della loro shelf life. Attraverso lo studio delle velocità di respirazione 

e lo sviluppo di modelli matematici è possibile compiere una scelta oculata del film 

e delle condizioni di imballaggio, in modo da raggiungere concentrazioni di gas e 

permeabilità delle matrici polimeriche ottimali per una conservazione che garantisca 

il mantenimento della qualità per tempi più lunghi. Lo sviluppo di modelli matematici 

che simulano il comportamento del prodotto confezionato nelle reali condizioni di 

conservazione serve a ridurre considerevolmente i tempi di ottimizzazione del sistema 

di confezionamento e rende possibile inoltre, progettare il sistema di imballaggio, 

specifico per le esigenze del singolo alimento in questione. 


1 

Ready-to-eat sweet cherries: study on different packaging systems. Conte 

A., Scrocco C., Lecce L., Mastromatteo M., Del Nobile M. A. innovative Food 

Science & Emerging technologies. 10, 564-571. (2009). Impact factor: 1.713. 

In this study the influence of different packaging systems on quality loss of ready- 

to-use cherries was assessed. In particular, the fruits were packaged in oriented 

polypropylene-based bag and in a bio-based polymeric matrix under ordinary 

and modified atmosphere conditions (MAP). Cherries quality during storage was 

determined by monitoring headspace gas concentration, weight loss, titrable 

acidity, total soluble solids, maturity index, antocyanins level, pH, viable cell 

load of various microbial groups and sensory quality. Results suggest that under 

ordinary atmosphere conditions, OPP shows the best performances. Otherwise, 

under MAP, both investigated films exert similar effects on the portioned fruit. 

116 




2 Different 

packaging strategies for fresh-cut “barattiere” melon cultivar 

(Cucumis melo L.). Conte A., Scrocco C., Brescia I., Del Nobile M. A. 

international Journal of Food Science and technology. 44, 1422-1428. (2009). 


Optimizing packaging for minimally processed “Barattiere” melon cultivar 

(Cucumis melo L.) is addressed in this work. Melon slices were dipped in water 

solution containing ascorbic acid (1%) and calcium chloride (0.2%). Dipped and 

untreated fresh-cut produce was packaged in 4 different polymeric materials (two 

biodegradable films, a commercially available multilayer film and polypropylene), 

before storing at 5°C. During storage, headspace gas composition, microbial 

quality, pH and mass loss were monitored for a period of 2 weeks. Results suggest 

that film characteristics strongly influenced gas concentrations in the headspace, 

microbial growth and weight. Among the tested packaging materials, the 

multilayer film seemed to delay significantly the detrimental phenomena involved 

in quality acceptance, even if clearly anaerobic atmospheres were recorded. The 

biodegradable films, combined with a dipping pre-treatment, could represent a 

good compromise between film performance and environmental impact. 

3 New 

strategies for minimally processed cactus pear packaging. Del Nobile 

M. A., Conte A., Scrocco C., Brescia I. innovative Food Science and Emerging 

technologies. 10, 356-362. (2009). Impact Factor: 1.713. 

This work presents a preliminary study on the packaging procedures to prolong 

the shelf life of minimally processed cactus pears. In particular, different packaging 

strategies were tested by combining coating and hydro-gels to different polymeric 

materials. Monitoring headspace gas concentrations, viable cell load of main 

spoilage microorganisms, sensory characteristics and weight loss, the quality decay 

in each packaging system was assessed. Results showed that the immersion of 

fresh-cut fruit into both hydro-gels strongly reduced the shelf life, most probably 

due to water migration from the surrounding hydro-gel to the crop. On the 

117 




contrary, the coating prolonged the shelf life of the minimally processed fruit to 

about 13 days, corresponding to an increase of about 40%, compared to the control 

sample. Results also suggested that the barrier properties of the selected films did 

not affect greatly the quality of the coated fresh-cut produce. 

4 New 

packaging strategies to preserve fresh-cut artichoke quality during 

refrigerated storage. Del Nobile M.A., Conte A., Scrocco C., Laverse J., Brescia 

I., Conversa G., Elia A. innovative Food Science and Emerging technologies. 

10, 128-133. (2009). Impact Factor: 1.713. 

The influence of both post-harvest treatments and film permeability on 

the quality loss kinetic of minimally processed artichokes is assessed in this 

study. In particular, fresh-cut artichoke heads were subjected to dipping in 

citric acid/calcium chloride water solution, and coating with citric acid loaded 

sodium alginate, respectively. Three different packaging materials were used: a 

polyester-based biodegradable film, an aluminum-based multi-layer film, and 

a commercially available oriented polypropylene film. Artichokes quality loss 

kinetic during storage was determined by monitoring produce appearance, 

weight loss, pH, and viable cell load of the main spoilage microorganisms. Results 

suggest that among the selected treatments, coating shows the best performance 

in terms of artichokes shelf life. As far as the packaging material is concerned, the 

biodegradable film tested in this work seems to be the most suitable packaging 

to preserve the quality of the coated fresh-cut produce. 

5 A 

study on the quality loss of minimally processed grapes as affected by 

film packaging. Del Nobile M. A., Conte A., Scrocco C., Brescia I., Speranza B., 

Sinigaglia M., Perniola R., Antonacci D. Postharvest Biology and technology. 

51, 21-26. (2009). Impact Factor: 1.590. 

The influence of film barrier properties on the quality loss of minimally processed 

grapes stored at 5°C is addressed in this study. Two table grapes (Vitis vinifera 

118 




cv. Italia), differing in quantity and frequency of irrigation, were tested with five 

different packaging films. Two commercially available films were used: a multi- 

layer film obtained by laminating nylon and a polyolefin layer (NP), and an 

oriented polypropilene film (OPP); along with three biodegradable polyester- 

based films (NVT-100, NVT-50 and NVT-35). The packed grape quality during 

storage was determined by monitoring the headspace oxygen and carbon dioxide 

concentration, the grape sensorial quality, and the viable cell concentration of the 

following spoilage microorganisms: total viable bacterial count, lactic acid bacteria, 

yeasts and molds. Results suggest that all the investigated films successfully 

preserve the quality of packed produce for the entire observation period (35 d). 

However, the best results were obtained using high barrier films such as NP and 

NVT-100. Slight differences were recorded between the two investigated table 

grapes in terms of respiratory activity and sensorial quality. 

6 Packaging 

strategies to prolong the shelf life of minimally processed 

lampascioni (Muscari comosum). Conte A., Scrocco C., Brescia I., Del Nobile 

M. A. Journal of Food Engineering. 90, 199-206. (2009). Impact Factor: 1.848. 

In this work different packaging strategies aimed to prolong the shelf life of minimally 

processed lampascioni are presented. In particular, two different treatments prior to 

packaging were tested: dipping in a solution containing citric acid (1%) and calcium 

chloride (8%), and coating with sodium alginate (5%), in combination with citric acid 

(1%) and calcium chloride solution (8%). The treated samples were packaged using 

two types of polymeric films: a commercially available oriented polypropylene film 

(OPP), and a polyester-based biodegradable film (NVT2). The investigated produce 

was stored at 5°C for approximately 20 days. Microbial populations, pH, weight loss 

and visual quality were monitored for the entire observation period. In order to 

determine the respiration activity, O 2 and CO 2 concentration were monitored not 

only in OPP and NVT2 packages but also in the headspace of an aluminum-based 

package. Results show that, among the packaging strategies investigated in this 

119 




work, the coated lampascioni packaged in NVT2 film were best preserved over the 

entire storage period. In fact, the alginate coating, combined with the gas barrier 

properties of the NVT2 film, can delay the lampascioni respiratory activity and the 

browning process, as well as the microbial growth, allows prolonging the shelf life 

of the investigated fresh-cut produce. 

7 Influence 

of growing periods on the quality of baby spinach leaves at harvest 

and during storage as minimally processed produce. Conte A., Conversa G., 

Scrocco C., Brescia I., Laverse J., Elia A., Del Nobile M. A. Postharvest Biology 

and technology. 50, 190-196. (2008). Impact Factor: 1.590. 

In this work the influence of different growing periods on quality and decay of 

fresh-cut baby spinach leaves during chilling storage was assessed. Three different 

spinach crops were sown on October, December and January, respectively. On 

January, February and March the vegetable leaves were harvested, analyzed 

for nitrate, oxalate, vitamin C and chlorophyll content and, after the minimal 

processing operations, the leaves were packaged and stored a 5°C. During 

storage, respiration rate, microbiological quality, dry matter, chlorophyll content 

and color parameters were monitored. Results show that in all harvests the initial 

quality of leaves is acceptable, although slight changes may be due to climatic 

conditions variability. 

8 Influence 

of postharvest treatments and film permeability on quality 

decay kinetics of minimally processed grapes. Del Nobile M. A., Sinigaglia, 

M., Conte A., Speranza, B., Scrocco C., Brescia, I., Bevilacqua, A., Laverse J., La 

Notte, E., Antonacci, D. Postharvest Biology and Technology. 47, 389-396. 

(2008). Impact Factor: 1.590. 

The influence of both postharvest treatments and film permeability on the quality 

decay kinetic of fresh processed grape fruit is addressed in this work. Ethanol, 

chlorinated water and hot water were tested to reduce microbiological spoilage. 

120 




The treated grapes were packaged in two polyester-based biodegradable films 

(NVT1, NVT2) and the composition of the atmosphere within the packages was 

determined. A multilayer film, made by laminating an aluminium foil with a 

polyethylene film (Aluminium) was used to evaluate separately the respiration 

activity of the packed grape, and the oxygen and carbon dioxide permeability 

coefficient of the investigated films in the real working conditions. The quality 

decay kinetic of fresh processed table grape packed at 5°C was assessed by 

monitoring the cell load of the main spoilage microorganisms (total mesophilic 

viable count, acid lactic bacteria, yeasts and moulds) and the product appearance 

for a period of about 30 days. Results suggest that ethanol was the most efficient 

treatment, as it successfully reduced the cell load of the spoilage microorganisms 

without affecting the respiration activity of the packed food and its appearance. 

Differences between the investigated biodegradable films were measured; 

however, they were not statistically significant. 

9 Use 

of biodegradable films for prolonging the shelf life of minimally 

processed lettuce. Del Nobile M.A., Conte A., Cannarsi M., Sinigaglia M. 

Journal of Food Engineering. 85, 317-325. (2008). Impact Factor: 1.848. 

The ability of biodegradable films to prolong the shelf life of minimally process 

lettuce stored at 4°C was addressed. Four different films were tested: two polyester- 

based biodegradable films (NVT1, NVT2), a multilayer film made by laminating an 

aluminum foil with a polyethylene film (All-PE), and an oriented polypropylene 

film (OPP). Package headspace, microbial load and color of the packed lettuce 

were monitored for a period of 9 days. A simple mathematical model was used to 

calculate the senescence level, whereas the re-parameterized Gompertz equation 

was used to calculate microbial shelf life of the fresh product. The fastest quality 

decay kinetic was observed for the lettuce packed in OPP, whereas the slowest 

one was detected for that in All-PE. Results suggest that the gas permeability of 

the investigated films plays a major role in determining the quality of the packed 

121 




produce. Moreover, it was observed that biodegradable films guarantee a shelf 

life longer than that of OPP. 

10 

Design of plastic packages for minimally processed fruits. Del Nobile 

M.A., Licciardello F., Muratore G., zappa M. Journal of Food Engineering. 

79, 217-224. (2007). Impact Factor: 1.848. 

In this work the possibility of using a simple mathematical model for designing 

plastic film for fresh processed fruits packaging is addressed. The study was 

conducted by packaging three different fresh processed fruits (prickly pear, 

banana and kiwifruit) with two different commercially available films: a laminated 

PE/aluminum/PET film, and a co-extruded polyolefinic film, and storing the 

packages at 5°C. The package headspace composition, in terms of oxygen and 

carbon dioxide concentration, was monitored for a period ranging between 4 

and 12 days, depending on film type. A simple mathematical model was used 

for describing and predicting the respiration behavior of packed fresh processed 

fruits during storage. Results showed that despite its simplicity the proposed 

model satisfactorily describes and predicts the respiration behavior of investigated 

fresh processed fruits. It was also found that the predictive ability of the proposed 

model depends on both the permeability of the package and the fresh processed 

fruit packed. Packaging optimization was performed taking the film thickness 

as variable. Results suggest that the optimal film thickness strongly depends on 

packed fruits. 

11 

Use of a simple mathematical model to evaluate dipping and MAP 

effects on aerobic respiration of minimally processed apples. Rocculi 

P., Del Nobile M.A., Romani S., Baiano A., Dalla Rosa M. Journal of Food 

Engineering. 76, 334-340. (2006). Impact Factor: 1.848. 

A new general respiratory model based on the Michaelis–Menten type enzyme 

kinetics was tested to describe changes in aerobic respiration of minimally processed 

122 




apple. Golden delicious apple slices, washed in distilled water or dipped in an 

antioxidant solution (1% ascorbic acid, 1% citric acid), were packed in multilayer 

pouches in passive or active modified atmosphere (respectively air and 90% N 2 O, 

5% CO 2 , 5% O 2 ). During four days of storage at 4°C, O 2 and CO 2 concentrations in the 

package headspace were monitored. The proposed model successfully fitted the 

experimental data, adequately describing the aerobic respiration of apple slices. 

Results suggest that both dipping treatment and active modified atmosphere 

affect the respiratory activity of the packed product. In particular, dipping reduced 

both the initial oxygen respiration rate (for about 10 h of storage) and the inhibitory 

effect that CO 2 had on O 2 consumption of packed product. The active modified 

atmosphere decreased the rate of oxygen consumption compared with passive 

MA, in particular at the beginning of storage. 

12 

Respiration rate of minimally processed lettuce as affected by packaging. 

Del Nobile M.A., Baiano A., Benedetto A., Massignan L. Journal of Food 

Engineering. 74, 60-69 (2006). Impact Factor: 1.848. 

A comparative study on the influence of films with different water and gas 

permeability on the quality decay of fresh processed lettuce is presented. The 

study was conducted by packaging two lettuce cultivars, Iceberg and Romaine, 

with three different plastic films, two commercially available polyolefines (PF1 

and PF2) and a biodegradable film (BF), and storing the packages at 5 package 

headspace composition, in terms of oxygen and carbon dioxide concentration, as 

well as the lettuce colour were monitored for 10 days. The film barrier properties 

(oxygen, carbon dioxide and water permeability coefficients) were measured. 

Results showed that over the period of observation the lowest respiration activity 

was found for lettuces packed with PF1, whereas lettuces packed using PF2 and 

BF films showed a similar respiration activity. On the other hand, no substantial 

differences in the colour variation kinetics due to the packaging with the three 

investigated films were detected. 

123 


tecnologie innovative 

Per Prolungare la shelf life 

Di alimenti freschi 



Daniela Gammariello, Post-doc 








Parole chiave 

SOSTANzE ANTIMICROBICHE 

SOSTANzE ANTIOSSIDANTI 

CONFEzIONAMENTO 

SISTEMA INTEGRATO PROCESSO/ 

CONFEzIONAMENTO 


Questo filone di ricerca è finalizzato allo studio di varie tecniche innovative di 

processo e/o confezionamento finalizzate a prolungare la shelf life di matrici 

alimentari altamente deperibili appartenenti al settore caseario, ittico e 

carneo. Gli studi effettuati sono stati focalizzati sull’utilizzo di sostanze attive 

naturali ad azione antimicrobica incorporate direttamente nella matrice 

alimentare o nel liquido di governo in cui viene solitamente conservato il 

prodotto, in combinazione, ove possibile, all’uso di atmosfere modificate in 

fase di confezionamento. 


1 

A combination of chitosan, coating and modified atmosphere packaging 

for prolonging Fior di Latte cheese shelf life. Del Nobile M.A. Gammariello 

D., Conte A., Attanasio M. Carbohydrate Polymers. 78, 151-156. (2009). 


In this work the addition of chitosan into cheese making, combined with 

either coating or active coating (lysozyme and ethylenediamine tetraacetic 

acid, disodium salt) and MAP (modified atmosphere packaging) was used 

to prolong the shelf life of “Fior di latte” cheese. On the packaged cheese 

stored at 4°C microbiological, pH, gas composition and sensory changes 

were monitored over an eight-day period. Results highlighted that the 

combination of chitosan, active coating and MAP improved “Fior di latte” 

cheese preservation by increasing the shelf life in comparison with the 

traditional packaging. In fact, the latter showed a very short shelf life limited 

to more or less 1 day, whereas the integrated approach developed in this 

study allowed us to obtain a significant shelf life prolongation to 5 days, most 

probably due to the synergic effect between the active compounds and the 

atmospheric conditions in the package headspace. 

124 





2 Combined 

Effects of Chitosan and MAP to Improve the Microbial Quality 

of Amaranth Home-Made Fresh Pasta. Del Nobile M. A., Di Benedetto N., 

Suriano N., Conte A., Corbo M. R., Sinigaglia M. Food Microbiology. 26, 587- 

591. (2009). Impact Factor: 2.847. 

In this work a study on the combined effects of chitosan and modified 

atmosphere packaging (MAP) to improve the microbiological quality of 

amaranth-based homemade fresh pasta is presented. In particular, two different 

chitosan concentrations were combined to three different MAP conditions and 

tested against the following spoilage microorganisms: mesophilic bacteria, 

Staphylococcus spp., yeasts, moulds and total coliforms. Their viable cell 

concentrations were monitored for about 2 months at 4°C. Results suggest that 

there is a combined effect between MAP and chitosan in delaying the microbial 

quality loss of pasta during storage. Moreover, it was also found that among the 

tested MAP conditions, the combination of 30:70 N 2 :CO 2 is the most efficient, 

promoting an extension of the microbial acceptability limit beyond two months. 

3 Thymol 

and modified atmosphere packaging to control microbiological 

spoilage in packed fresh cod hamburgers. Corbo M.R., Di Giulio S., Conte 

A., Speranza B., Sinigaglia M., Del Nobile M.A. international Journal of Food 

Science and technology. 44, 1553–1560. (2009). Impact Factor: 1.065. 

A study on the use of mild technologies to produce packaged fish hamburgers 

was presented. In particular, the antimicrobial effect of some natural compounds 

(carvacrol, eugenol, thymol, green tea extract, rosemary extract, grapefruit seed 

extract and lemon extract), at various concentrations (500-10.000 ppm), was 

screened in vitro against the main fish spoilage microorganisms (Shewanella 

putrefaciens and Photobacterium phosphoreum). Lemon extract and thymol, 

in combination with modified atmosphere packaging, showed the greatest 

inhibition activity, therefore, thymol was subsequently used as ingredient for 

producing fish hamburgers. Results pointed out that this combination is effective 

125 





in controlling the growth of microbial species mainly involved in fresh fish 

spoilage; in particular, it significantly (P




(MAP) on quality retention of blue fish burgers were studied and discussed. 

In particular, samples were packaged in air and in three different gas mix 

compositions: 30:40:30 O 2 :CO 2 :N 2 , 50:50 O 2 :CO 2 and 5:95 O 2 :CO 2 . During a 28- 

days storage period at 4°C, the nutritional, microbiological and sensorial quality 

of the packed products were assessed. The potential development of biogenic 

amines was also evaluated. The obtained results highlight the possibility to 

improve the microbial quality of blue fish burgers by using very small amount of 

thymol (110 ppm), GFSE (100 ppm) and lemon extract (120 ppm) in combination 

with MAP. Based primarily on microbiological results, the combined use of the 

tested natural preservatives and a packaging system characterized by a high 

CO 2 -concentration, was able to guarantee the microbial acceptability of fish 

burgers until the 28th day of storage at 4°C. On the other hand, results from 

sensory analyses showed that sensorial quality was the sub-index that limited 

the burgers shelf life (to about 22-23 days), even if the proposed strategy was 

also effective in minimizing the sensory quality loss of the product having no 

effect on its nutritional quality. 

6 Strategies 

for prolonging the shelf life of minced beef patties. Del Nobile 

M.A., Conte A., Cannarsi M., Sinigaglia M. Journal of Food Safety. 29, 14-25. 


The study of different strategies for prolonging the shelf life of fresh minced 

beef patties was addressed in this work. Fresh minced beef was supplemented 

with thymol (THY) at levels of 250, 500 and 750 mg per Kg of ground 

beef. Treated samples were packed using a high barrier film and stored in 

refrigerated conditions (4°C) under ordinary (OAP) and modified atmosphere 

(MAP) conditions for 16 days. The microbiological quality decay kinetic was 

determined by monitoring the following microorganisms: Enterobacteriaceae, 

Pseudomonas spp., lactic acid bacteria, Brochothrix thermosphacta, coliforms 

and total psychrotrophic viable count. Results show that thymol, working 

127 





alone, was effective on coliforms and Enterobacteriaceae; whereas, it does 

not seem to inhibit to a great extent the growth of the other microbial 

populations. On the other hand, an increased amount of thymol, under MAP 

conditions, had better effects on the product quality, with a consequent 

prolongation of the shelf life. 

7 Use 

of natural compounds to improve the microbial stability of amaranth 

based home-made fresh pasta. Del Nobile M.A., Di Benedetto N., Suriano 

N., Conte A., Lamacchia C., Corbo M.R., Sinigaglia M. Food Microbiology. 26, 

151-156. (2009). Impact Factor: 2.039. 

A study on the use of natural antimicrobial compounds to improve the 

microbiological stability of refrigerated amaranth-based homemade fresh pasta 

is presented in this work. In particular, the antimicrobial activity of thymol, lemon 

extract, chitosan and grapefruit seed extract (GFSE) has been tested against 

mesophilic and psychrotrophic bacteria, total coliforms, Staphyloccocus spp., 

yeasts and moulds. A sensory analysis on both fresh and cooked pasta was also 

run. Results suggest that chitosan and GFSE strongly increase the microbial 

acceptability limit of the investigated spoilage microorganisms, being the 

former the most effective. Thymol efficiently reduces the growth of mesophilic 

bacteria, psychrotrophic bacteria and Staphyloccocus spp., whereas it does not 

affect, substantially, the growth cycle of total coliforms. Lemon extract is the 

less effective in preventing microbial growth. In fact, it is able to delay only total 

mesophilic and psychrotrophic bacterial evolution. From a sensorial point of view 

no significant differences were recorded between the control samples and all the 

types of loaded amaranth-based pasta. 

8 Shelf 

life of Straciatella cheese under modified atmosphere packaging. 

Gammariello D., Conte A., Di Giulio S., Attanasio M., Del Nobile M.A. Journal 

of dairy Science. 92, 483-490. (2009). Impact Factor: 2.361. 

128 





The aim of this work is to evaluate the shelf-life of Stracciatella cheese packaged 

in a protective atmosphere, using four different CO 2 /N 2 /O 2 gas mixtures (50/50/0 

(M1), 95/5/0 (M2), 75/25/0 (M3) and 30/65/5 (M4) v/v) and stored at 8°C. Cheese 

in traditional tubs (Cnt) and under vacuum (VP) were used as the control. 

Results showed that the modified atmosphere packaging, in particular M1 and 

M2, delayed microbial growth of spoilage bacteria, without affecting the dairy 

microflora, and prolong the sensorial acceptability limit. 

9 Active 

coating and modified atmosphere packaging to extend the shelf 

life of Fior di latte cheese. Conte A., Gammariello D., Di Giulio S., Attanasio 

M., Del Nobile M.A. Journal of dairy Science. 92, 887-894. (2009). Impact 


In this work the combination of active coating and modified atmosphere packaging 

(MAP) was used to prolong the shelf life of “Fior di latte” cheese. The active coating 

was based on sodium alginate (8% w/v) containing lysozyme (0.25 mg*mL -1 ) and 

ethylenediamine tetraacetic acid, disodium salt (Na 2 -EDTA, 50mM). The MAP was 

made up of 30% CO 2 , 5% O 2 and 65% N 2 . The quality loss kinetic of the “Fior di latte” 

cheese, stored at 10°C, was assessed by monitoring pH and weight loss, as well as 

microbiological and sensorial evolution. Results highlighted that the combination 

of active coating and MAP improved “Fior di latte” cheese preservation, increasing 

the shelf life to more than 3 days. In addition, the substitution of brine with coating 

could allow us to gain a double advantage: on the one hand, it could preserve the 

product quality and on the other hand, it could represent a strategic solution to 

reduce the cost of its distribution, due to the lower weight of the package. 

10 

Study on the synergic effect of natural compounds on the microbial quality 

decay of packed fish hamburger. Corbo M.R., Speranza B., Filippone A., 

Granatiero S., Conte A., Sinigaglia M, Del Nobile M.A. international Journal 

of Food Microbiology. 127, 261-267. (2008). Impact Factor: 2.581. 

129 





The effectiveness of natural compounds in slowing down the microbial quality 

decay of refrigerated fish hamburger is addressed in this study. In particular, the 

control of the microbiological spoilage by combined use of three antimicrobials, 

and the determination of their optimal composition to extend the fish hamburger 

Microbiological Stability Limit (MAL) are the main objectives of this work. Thymol, 

grapefruit seed extract (GFSE) and lemon extract were tested for monitoring the 

cell growth of the main fish spoilage microorganisms (Pseudomonas fluorescens, 

Photobacterium phosphoreum and Shewanella putrefaciens), inoculated in fish 

hamburgers, and the growth of mesophilic and psychrotrophic bacteria. A 

Central Composite Design (CCD) was developed to highlight a possible synergic 

effect of the above natural compounds. Results showed an increase in the MAL 

value for hamburgers mixed with the antimicrobial compounds, compared to 

the control sample. The optimal antimicrobial compound composition, which 

corresponds to the maximal MAL value determined in this study, is: 110 mgL -1 of 

thymol, 100 mgL -1 of GFSE and 120 mgL -1 of lemon extract. The presence of the 

natural compounds delay the sensorial quality decay without compromising the 

flavor of the fish hamburgers. 

11 

Use of active compounds for prolonging the shelf life of mozzarella 

cheese. Sinigaglia M., Bevilacqua A., Corbo M.R., Pati S., Del Nobile M.A. 

international dairy Journal. 18, 624-630. (2008). Impact Factor: 2.198. 

The effectiveness of lysozyme and ethylenediaminetetraacetic disodium salt 

(Na2-EDTA) against the spoilage microorganisms of mozzarella cheese was 

studied. Mozzarella cheeses were packaged in a conditioning solution (diluted 

brine), which contained lysozyme (0.25 mg mL1) and different amounts of 

Na2-EDTA (10, 20 and 50 mmol L1), and stored at 4 1C for 8 days. The population 

of spoilage microorganisms (total coliforms and Pseudomonadaceae), along 

with the functional microbiota of mozzarella cheese (lactic acid bacteria) was 

enumerated. Lysozyme and Na2-EDTA significantly inhibited the growth of 

130 





coliforms and Pseudomonadaceae during the first 7 days of storage, whereas the 

functional microbiota (or lactic acid bacteria) were not affected. The results of 

this study showed that it is possible to extend the shelf life of mozzarella cheese 

through the use of lysozyme and Na2-EDTA in the conditioning brine. 

12 

Effects of natural compounds of microbial safety and sensory quality of 

Fior di latte cheese, a typical Italian cheese. Gammariello D., Di Giulio S., 

Conte A., Del Nobile M.A. Journal of dairy Science. 91, 4138-4146. (2008). 


This work presents a preliminary study to assess the efficiency of plant essential 

oils as natural food preservatives in “Fior di Latte” cheese. In particular, selected 

compounds were directly dissolved into “Fior di Latte” brine. Packaged “Fior 

di Latte” samples were stored at 10°C for about 6 days. The cell loads of both 

spoilage and useful microorganisms were monitored in order to calculate the 

microbial acceptability limit (MAL). Results show that some tested compounds 

were not acceptable by the panel from a sensorial point of view. Most of them 

do not affect, to a great extent, the MAL value, and only few of them, such as 

lemon, sage and thyme, markedly prolong the microbial acceptability limit 

of the investigated fresh cheese. Moreover, the above active agents exert an 

inhibitory effect on the microorganisms responsible for spoilage, without 

affecting the dairy microflora. 

13 

Use of nisin, lysozyme and EDTA for inhibiting microbial growth in chilled 

buffalo meat. Cannarsi M., Baiano A., Sinigaglia M., Ferrara L., Baculo R., 

Del Nobile M.A. international Journal of Food Science and technology. 43, 

573-578. (2008). Impact Factor: 0.941. 

The effect of nisin, lysozyme and lysozyme combined with disodium 

ethylenediaminetetraacetate salt (EDTA) for inhibiting the growth of spoilage 

micro-organisms in buffalo meat was investigated. Prior to packaging, the samples 

131 





were alternatively dipped in solutions containing different concentrations of nisin, 

lysozyme and combinations of a pre-fixed amount of lysozyme and different 

concentrations of EDTA. The packed meat samples were stored at 4°C and the 

microbial changes were monitored for a period of 8 days. The best results, in terms of 

microbial shelf-life extension, were obtained by dipping the meat steaks in a solution 

containing a combination of 0.5% of lysozyme and 2% of EDTA. This treatment was 

able to reduce the growth of all the investigated bacteria and showed a bactericidal 

effect on Brochotrix thermosphacta during the entire period of storage. 

14 

Innovative active packaging systems to prolong the shelf life of mozzarella 

cheese. Conte A., Scrocco C., Sinigaglia M., Del Nobile M. A. Journal of 

dairy Science. 90, 2126-2131. (2007). Impact Factor: 2.361. 

In this work the effectiveness of different antimicrobial packaging systems on 

the microbial quality decay kinetic during storage of mozzarella cheese was 

evaluated. Lemon extract, at three different concentrations, was used as active 

agent, in combination with brine and with a gel solution, made of sodium 

alginate. Shelf life tests were run at 15°C to simulate thermal abuse. The cell load 

of both spoilage and dairy functional microorganisms were monitored at regular 

time intervals during storage. By fitting the experimental data through a modified 

version of Gompertz equation, the shelf life of dairy product packaged in the 

different systems was calculated. Results show an increase in the shelf life of all 

active packaged mozzarella cheeses, confirming that the investigated substance 

may exert inhibitory effect on the microorganisms responsible for spoilage 

phenomena, without affecting the functional microbiota of the product. 

15 

The influence of using different packaging on the quality decay kinetics 

of Cúccìa Muratore G., Lanza C.M., Baiano A., Tamagnone P., Asmundo 

C.N., Del Nobile M. A. Journal of Food Engineering. 73, 239-245. (2006). 


132 





The renewed interest of consumer towards traditional and typical dishes leaded to 

an increase in the demand for Cúccía, which is a traditional Sicilian food made by 

boiling in water durum wheat kernels ‘‘complete’’ of its aleuronic and embryonic 

layer. A comparative study on the influence of using different packaging (PET, 

active PET packaging and glass) on the quality decay kinetics of Cúccıa is 

presented. In particular, the performance of both traditional plastic containers 

and an innovative material containing an oxygen scavenger was evaluated by 

monitoring the quality of bottled Cúccía during storage at 15°C, the actual storage 

condition. For comparative purposes the quality decay kinetics of Cúccía bottled 

in glass was also determined. Results showed that the containers made of PET can 

slow down the quality decay kinetics of Cúccía to that of glass containers. 

133 


alimenti funzionali 




Grazia Sepielli, tecnico laureato 

Incoronato Anna Lucia, dottoranda 

Alessandra Danza, dottoranda 








Parole chiave 

SALAMI A RIDOTTO CONTENUTO 

DI GRASSO 

PASTA GLUTEN-FREE 

COMPOSTI ATTIVI NATURALI 

VARIABILI DI PROCESSO 


Il filone di ricerca si propone di mettere a punto nuove tecnologie necessarie 

alla produzione di alimenti ad elevato valore nutrizionale. Da un lato si studiano 

sfarinati alternativi alla semola per la produzione di paste gluten-free e dall’altro 

alimenti carnei fermentati a ridotto contenuto di grasso animale. In particolare, 

gli studi sono finalizzati a valutare come vari parametri di processo, intesi sia in 

termini di materie prime della formulazione che di tecnologie di produzione, 

influiscano sulla qualità del prodotto finito. 


1 

Properties of quinoa and oat spaghetti loaded with carboxymethylcellulose 

sodium salt and pregelatinized starch as structuring agents. Chillo S., Civica 

V., Iannetti M., Suriano N., Mastromatteo M., Del Nobile M.A. Carbohydrate 

Polymers, 78, 932-937. (2009). Impact Factor: 2.644. 

Spaghetti based on quinoa or oat were manufactured using two different 

structuring agents, carboxymethylcellulose sodium salt (CMC) and pregelatinized 

starch at three different percentages (0.1%, 0.2%, 0.3% and 10%, 20%, 30%, 

respectively). The dough rheological properties were determined using a 

capillary rheometer, the mechanical characteristics of dry spaghetti by a 

dynamic mechanical analyzer and the sensorial parameters by a trained 

panel. Elongational and shear viscosity declined or increased when CMC was 

added to quinoa and oat based dough, respectively. The stress at break for dry 

non-conventional spaghetti increased for oat spaghetti added with CMC and 

pregelatinized starch, whereas decreased for quinoa spaghetti. The sensorial 

parameters of dry and cooked spaghetti in quinoa and oat base were strongly 

affected by addition of CMC and pregelatinized starch as it was not possible 

to produce spaghetti in base either quinoa or oat without using the tested 

structuring agents. 

134 



2 New 

strategies for reducing the pork back-fat content in typical Italian 

salami. Del Nobile M.A., Conte A., Incoronato A.L., Panza O., Sevi A., Marino 

R. Meat Science, 81: 263-269. (2009). Impact Factor: 2.006. 

In this work the possibility to substitute pork backfat with extra virgin olive oil 

(EVOO), adsorbed on whey protein-based crumb or white bread crumb, in typical 

Italian salami is addressed. Five types of salami were manufactured, under usual 

commercial conditions, by replacing 0 (Control), 60 and 100% of pork backfat 

with whey protein-based crumb (WP60-WP100) and white pan bread (PB60- 

PB100), respectively, soaked in EVOO. Results highlighted that pH, weight loss, 

colour parameters and microbial counts did not show statistically significant 

differences between the Control and the modified salami. On the other hand, 

malonaldehyde was slightly lower in PB100, PB60, WP100 and WP60, compared 

to the Control. Chemical composition was significantly affected by formulations. 

Modified salami presented a better fatty acid profile showing lower saturated and 

higher monounsaturated fatty acids than control. Furthermore in all modified 

salami atherogenic and thrombogenic indices displayed the lowest values. 

The Control showed the highest values for Warner-Bratzler Shear, hardness, 

cohesiveness, gumminess and chewiness. Sensory evaluation of WP60 did not 

show significant differences compared to the Control, whereas PB100 and WP100 

were unacceptable for taste. 

3 Influence 

of the addition of buckwheat flour and durum wheat bran on the 

spaghetti quality. Chillo S., Laverse J., Falcone P. M., Protopapa A., Del Nobile 

M.A. Journal of Cereal Science. 47, 144-152. (2008). Impact Factor: 1.905. 

The quality of nine spaghetti typologies, produced by using wheat durum 

semolina as a base plus the addition of buckwheat and durum wheat bran, was 

investigated. The quality of the produced spaghetti was compared with that of 

spaghetti made only of durum semolina (CTRL). Tests were run on the samples 

to determine breakage susceptibility and colour of dry spaghetti, the cooking 

135 



resistance, instrumental stickiness at optimal cooking time (OCT) and overcooking, 

the cooking loss and sensorial attributes at the optimal cooking time. Results 

suggest that the breakage susceptibility decreases with the addition of 15% and 

20% bran, the spaghetti dry colour changes with the addition of buckwheat flour 

and bran compared to the spaghetti made only of durum semolina, while the 

cooking resistance, instrumental stickiness and the cooking loss, in general, were 

equal to that of the CTRL. However, the addition of buckwheat flour and bran 

affected the sensorial attributes differently. 

4 Quality 

of spaghetti in base amaranthus wholemeal flour added with quinoa, 

broad bean and chickpea. Chillo S., Laverse J., Falcone P.M., Del Nobile M.A. 

Journal of Food Engineering. 84, 101-107. (2008). Impact Factor: 1.848. 

The quality of three gluten free spaghetti typologies, made from amaranthus flour plus 

the addition of quinoa (CQA), broad bean (CBA) or chick pea (CCA), was investigated. 

The quality of the produced spaghetti was compared to that of spaghetti made 

of durum semolina (CTRL). Tests were run on the samples to determine breakage 

susceptibility of dry spaghetti, the cooking resistance, instrumental stickiness in 

cooking and overcooking, cooking loss and some sensorial attributes at the optimal 

cooking time. The spaghetti obtained from amaranthus flour when compared to the 

CTRL, had equal or lower dry breakage susceptibility and cooking resistance, higher 

cooking loss and visibly lower instrumental stickiness. However, these spaghetti did 

not demonstrate relevant sensorial differences. 

5 Effect 

of carboxymethylcellulose and pregelatinized corn starch on the quality 

of amaranthus spaghetti. Chillo S., Laverse J., Falcone P.M., Del Nobile M.A. 

Journal of Food Engineering. 83(4), 492-500. (2007). Impact Factor: 1.848. 

Six gluten-free spaghetti typologies in base of amaranthus flour were produced, 

using two types of gluten substitutes, carboxymethylcellulose sodium salt 

(CMC) at three different percentages, 0.1%(LCMC), 0.2%(MCMC), 0.3%(HCMC) 

136 



and pregelatinized corn starch (PCS) at three different percentages, 5% (LPCS), 

7% (MPCS), 9% (HPCS). The amaranthus spaghetti were compared with a control 

spaghetti sample of durum semolina (CTRL). We ran some tests on the samples 

to determine breakage susceptibility by water sorption kinetics, cook in gloss, 

the cooking resistance by fitting the values of elastic modulus, instrumental 

stickiness and sensory analysis. The spaghetti obtained from amaranthus flour 

containing CMC and PCS showed breakage susceptibility higher than that of 

the CTRL, while they showed a cooking resistance equal or lower with respect 

to the CTRL. The spaghetti with CMC had a lower stickiness, equal values of 

cooking loss and similar values of sensory attributes with respect to that of the 

CTRL. For the spaghetti containing PCS, the cooking loss values were higher 

and the sensorial attributes were lower with respect to that of the CTRL and of 

the spaghetti containing CMC. Moreover, the stickiness value for the spaghetti 

samples with PCS increased as the pregelatinized corn-starch concentration 

increased. From the result it emerged that spaghetti samples containing 

CMC presented better performances especially in cooking with respect to the 

spaghetti samples with PCS. 

137 


caratterizzazione chimico-fisica 

Di matrici alimentari 




Pierangelo Frisullo, tecnico diplomato 








Parole chiave 

TEST MECCANICI 

TEST REOLOGICI 

MICROTOMOGRAFIA 


Questo filone di ricerca riguarda la caratterizzazione delle matrici alimentari. 

Accanto all’utilizzo di test reologici e dinamico-meccanici su vari prodotti 

alimentari si è fatto uso dell’analisi d’immagine, quale strumento utile per lo 

studio della microstruttura di alcuni sistemi alimentari. In particolare, è stata 

utilizzata la tomografia assiale computerizzata in diffrazione, per acquisire, in 

maniera non invasiva, immagini bidimensionali sulla mollica di pane. Adeguati 

algoritmi hanno poi permesso di ricostruire da queste immagini la struttura 

tridimensionale del prodotto. 


1 

Effects of additives on the rheological and mechanical properties of nonconventional 

fresh handmade tagliatelle. Chillo S., Suriano N., Lamacchia 

C., Del Nobile M.A. Journal of Cereal Science. 49, 163-170. (2009). Impact 


The effects of the following additives on the amaranth (A), quinoa (Q) and oat (O) 

dough rheological properties and the extruded tagliatelle dough mechanical 

characteristics were evaluated: carboxymethylcellulose of sodium (CMC), 

whey protein isolate (WPI), casein (CAS), chitosan (CHIT) and pregelatinized 

starch (PS). The amaranth, quinoa and oat rheological dough properties and 

amaranth, quinoa and oat tagliatelle mechanical characteristics were compared 

to those of their respective controls (ACTRL, QCTRL and OCTRL) and of the 

SEMOLINA sample. The storage modulus (G0 ) and loss modulus (G00 ) values of 

the quinoa and oat dough with PS were similar to those of the semolina dough. 

For all tagliatelle samples, WPI reduced the elastic modulus or Young’s modulus 

towards that of the semolina tagliatelle. Moreover, the additives did not 

have particular influence on the tenacity with the exception of the amaranth 

tagliatelle added with WPI. 

138 




2 Effect 

of Chitosan on the Rheological and Sensorial Characteristics of 

Apulia Spreadable Cheese. Gammariello D., Chillo S., Mastromatteo M., Di 

Giulio S., Attanasio M., Del Nobile M.A. Journal of dairy Science. 91, 4155- 

4163. (2008). Impact Factor: 2.361. 

The effect of chitosan on the rheological and sensorial properties of Apulia 

spreadable cheese during storage time was evaluated. The investigated spreadable 

cheese samples were stored at 4°C. Storage modulus (G'), loss modulus (G''), tanδ, 

and the overall sensorial quality of the spreadable cheese were monitored for 24 

d. Moreover, moisture content, pH, color, and lactic acid bacteria during storage 

time were evaluated. Results indicate that statistically significant differences in G', 

G'', and tanδ values and in the sensorial score exist between the control sample 

and the spreadable cheese samples with chitosan. In particular, chitosan improved 

the rheological and sensorial properties of the spreadable cheese, particularly its 

softness. Moreover, its addition influenced the physicochemical properties of the 

investigated spreadable cheese during storage time, without affecting the dairy 

microflora. 

3 A 

mathematical model to predict the effect of shape on pasta hydration 

kinetic during cooking and overcooking. Cafieri S., Chillo S., Mastromatteo 

M., Suriano N., Del Nobile M.A. Journal of Cereal Science. 48, 857-862. (2008). 


In this paper, a novel mathematical model to describe pasta hydration kinetic 

during cooking and overcooking is presented. The model takes into account 

the water diffusion process, the relaxation of the macromolecular matrix and 

the starch gelatinization process. The target pasta samples have three different 

geometries, specifically plane sheet, solid cylinder and hollow cylinder. The 

proposed model satisfactorily fits the experimental data, showing its ability to 

predict hydration kinetics for all the considered geometries. It is also successfully 

applied to predict the evolution of water concentration profiles. 

139 




4 Textural 

changes of Canestrello Pugliese cheese measured during storage. 

Del Nobile M.A., Chillo S., Falcone P.M., Laverse J., Pati S., Baiano A. Journal 

of Food Engineering. 83(4), 621-628. (2007). Impact Factor: 1.848. 

The changes in the fundamental textural properties of the Canestrello Pugliese 

cheese, during ripening, were studied, accounting for the internal and external 

portions of the cheese. The cheese was ripened at 12–14°C and 80% relative 

humidity and analyzed at 0, 15, 30, 45, 60, 95, and 120 days to determine the 

moisture content, water activity (Aw), pH values and some fundamental- 

mechanical and dynamic-mechanical properties such as the elastic modulus (Ec), 

storage modulus (G'), loss modulus (G'') and relaxation times distribution curve. 

Significant differences in the moisture content, Aw, pH and the elastic modulus 

were observed for the inside and outside portions of the investigated cheese. In 

particular, for the first 60 days of ripening the elastic modulus of the outer region 

was higher than that of the inner region. As the ripening process approached the 

end of the fourth month the contrary was true. Moreover, results show that, above 

all the pH and Aw play a major role in determining the mechanical properties of 

the Canestrello Pugliese cheese during ripening. 

5 Measuring 

Rheological Properties for Applications in Quality Assesment 

of Traditional Balsamic Vinegar: Description and Preliminary Evaluation of 

a Model. Falcone P.M., Chillo S., Giudici P., Del Nobile M.A. Journal of Food 

Engineering. 80, 234-240. (2007). Impact Factor: 1.848. 

In this research, a study of fundamental rheological properties of Traditional 

Balsamic Vinegar (TBV) was conducted by using a liquid food model system. 

With this purpose, aqueous solutions containing the main chemical constituents 

of TBV were prepared according to a Central Composite Design (CCD) and flow 

experiments were conducted by using a controlled-strain rotational rheometer. 

Surface response methodology was used successfully to describe effects of the 

chemical composition on the rheological behavior of the investigated TBV-like 

140 




model system (TBV-M). It was found that glucose mass fraction and glucose to 

fructose ratio strongly affected the steady shear viscosity of TBV-M and that 

the viscosity changed with temperature according to the Arrhenius’s law in 

the range of 20–60 of TBV quality before performing of sensorial analysis as 

specified by the specific production disciplinary. 

6 Use 

of the Generalized Maxwell Model for Describing the Stress 

Relaxation Behaviour of Solid-Like Foods. Del Nobile M.A., Chillo S., 

Mentana A., Baiano A. Journal of Food Engineering. 78, 978-983. (2007). 


A comprehensive study on the ability of the generalized Maxwell model to 

describe the stress relaxation behavior of food matrices is presented in this work. 

Five different food matrices were chosen as representative of a wide range of 

foods: agar gel, meat, ripened cheese, ‘‘mozzarella’’ cheese and white pan bread. 

The above foods matrices were selected on the base of their macro-structure 

and compressive stress strain behavior. Stress relaxation tests were run on the 

above food matrices at room temperature. The relaxation times distribution 

function used in this investigation to describe the viscoelastic behavior of 

the investigated foods was obtained by summing two normal distribution 

functions with the means equal to zero. Results show that the proposed model 

satisfactorily fits the experimental data. Moreover, it was also found a substantial 

difference between the relaxation times distribution curves of the investigated 

bulky and spongy foods. 

7 Influence 

of Drying Temperature on the Spaghetti Cooking Quality. Baiano 

A., Conte A., Del Nobile M.A. Journal of Food Engineering. 76, 341-347. 


The influence of protein content and drying temperature on the cooking quality 

of durum wheat spaghetti was addressed. Nine samples of spaghetti were 

141 




obtained from three wheat flours differing in protein content and the drying was 

carried out at three different temperatures (low, medium, and high). The pasta 

was submitted to sensorial analysis – in order to evaluate adhesiveness, bulkiness 

and firmness – and to physical, chemical and instrumental analyses in order to 

objectify these parameters. Mathematical equations were used to determine the 

significance of the differences among the spaghetti samples. The spaghetti with 

the highest protein content and dried at high temperature showed the highest 

firmness and the lowest stickiness. 

142 


Editore DiSA -Dipartimento di Scienze degli Alimenti, 

Università degli Studi di Foggia 

Coordinamento editoriale Artek snc 

grafica e impaginazione VogaDesign 

Stampa Shelve sas 

Finito di stampare Novembre 2010

DiSA_ttivo © DiSA - Università degli studi di Foggia - Agraria ...

Create successful ePaper yourself

Delete template?

Save as template?