FLEISCHWIRTSCHAFT international_04_2018

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Fleischwirtschaft international 4_2018
43
Research & Development
Calorific value
Two grams of sample was electrically burnt in excess of oxygen in a bomb
calorimeter. The maximum temperature rise of the bomb calorimeter was
measured with a thermocouple and a galvanometer system. By comparing this
rise with that obtained when a sample of known calorific value is burnt, the
calorific value of the sample material could be determined. Gross energy of
samples was determined by a Gallenkamp and Ballistic Bomb Calorimeter (HAQUE
andMURALI LAL, 1999).
Water activity (aw)
The water activity of functional mutton patties was measured by an Aqua LAB
dew point water activity meter 4TE.
Total phenolics
The total phenolic content in the phyto-ingredients powder and mutton patties
were quantified using the Folin-Ciocalteu colorimetric method as described by
MAKKAR (2000). Suitable aliquots of the extracts were taken in test tubes, and
the volume was made upto 0.5 ml with distilled water and 0.25 ml Folin-Ciocalteu
(1N) reagent was added and then the reaction was neutralized by addition
with 1.25 ml sodium carbonate solution (20%). The tubes were vortexed and the
absorbance of the resulting blue color was measured using a Beckman DU-640
UV/Vis spectrophotometer at 760 nm against blank after incubation for 40min
at room temperature. From the standard calibration curve equation y= f(x) the
quantification of phenolics was done and expressed as mg tannic acid equivalents
per gram of sample.
Texture profile analysis
The texture profile of mutton patties was measured with the help of instrumental
texture profile analyzer (TAHD Plus Texture analyser). The procedure used for
instrumental texture profile analysis was similar to those described by BOURNE
(1978). Chilled samples were tempered to bring to room temperature and were
cut into 1cm squares. The samples were placed on a platform in a fixture and
compressed twice to 85% of their original height by a compression probe (P75)
at a cross head speed of 10 mm/s through a two cycle sequence, using a 50 kg
load cell.
Antioxidant capacity
Total Phenolics by F-C Method
Dried (finely ground) phyto-ingredients powder (0.2g) was taken in a centrifuge
tube of 25 ml capacity. 10 ml of aqueous acetone (70%) was added and subjected
to centrifugation for 10 min at 3000 g in a REMI research centrifuge and
then the contents were cooled by keeping the centrifuge tube in the refrigerator
at 4±1°C (MAKKAR, 2000). Five gram of mutton patties was taken in a beaker
of 100ml capacity. 25 ml of aqueous ice cold 70% acetone was added and
subjected to homogenization for 60 s in an Ultra Turrax T25 tissue homogenizer
(Janke and Kenkel IKA Labortechnik, Germany) and then the beaker was
wrapped with aluminium foil and kept overnight for extraction at a refrigeration
temperature of 4±1°C (NAVEENA et al., 2008).
Preparation of a calibration curve using Standard Tannic Acid (TA)
Table 1 shows the tannic acid stock standard concentration. The calibration
curve was drawn and the equation was calculated in a Microsoft Excel 2007
spread sheet. The linear correlation between standard concentration and
absorbance was expressed with the equation y= f(x) and r 2 value. Where y=
absorbance, x= standard concentration (µg/ml) and r 2 = correlation coefficient.
Reducing power assay
Suitable aliquots of the extracts containing 50 to 100 µg phenolics from
phyto-ingredients and mutton patties were taken in test tubes, and the
volume was made equal with acetone (70%) and mixed with 2.5 ml phosphate
buffer (200 mM, pH 6.6) and 2.5 ml potassium ferricynide (1% w/v).
This mixture was kept at 50 o C in a water bath for 20 min. After cooling,
2.5 ml of 10% trichloro acetic acid was added and centrifuged at 5000 rpm
for 10 min in a REMI research centrifuge. The upper layer of the solution
(2.5 ml) was mixed with (2.5 ml) distilled water and 0.5 ml of freshly prepared
ferric chloride (0.1% w/v) solution. The absorbance was measured
using Beckman DU-640 UV/Vis spectrophotometer at 700 nm against
blank without any extracts and 0.1% ferric chloride. An increase in absorbance
of the reaction mixture indicated the reducing power of the
sample.
DPPH radical scavenging activity
The ability to scavenge the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical by
phyto-ingredients and mutton patties was estimated by the method of
SINGH et al. (2002). Different concentrations (50 and 100 µL equivalent to
50 and 100ppm of total phenolics) of phyto-ingredients and mutton
patties were taken in different test tubes. The volume was adjusted to
100 µL by adding MeOH. 5ml of 0.1mM methanolic solution of DPPH was
added to these tubes and shaken vigorously. The tubes were allowed to
stand at 27 °C for 20 min. The control was prepared as above without any
extract, and MeOH was used for the baseline correction. Changes in the
absorbance of the samples were measured at 517 nm using a Beckman
DU-640 UV-VIS spectrophotometer. The radical scavenging activity was
expressed as the inhibition percentage and was calculated using the
following formula:
Calibration
Tab. 1: Preparation of the calibration curve using
standard Tannic Acid (TA)
Test
tube
Final
conc. of
TA
(µg/ml)
Stock TA
0.1 mg/
ml
(µl)
Distilled
water
(ml)
F-C
reagent
(ml)
Sodium
carbonate
solution
(ml)
Final
volume
(ml)
Blank 0 0 0.50 0.25 1.25 2.0
T1 2 20 0.48 0.25 1.25 2.0
T2 4 40 0.46 0.25 1.25 2.0
T3 6 60 0.44 0.25 1.25 2.0
T4 8 80 0.42 0.25 1.25 2.0
T5 10 100 0.40 0.25 1.25 2.0
Stock standard (tannic acid) concentration – 0.1mg/ml;
Vortexed and kept at room temp for 40 min then recorded at 760 nm
Source: MALAV et al. FLEISCHWIRTSCHAFT international 4_2018
TBARS value
The TBARS value of mutton patties was determined by using the distillation
method described by TARLADGIS et al. (1960).
Instrumental color analysis
The color of mutton patties was measured using a Lovibond Tintometer
(Model F, Greenwich, UK). Samples were cut with the help of scissors to
the inner diameter of the sample holder and secured against the viewing
aperture. The sample color was matched by adjusting the red (a*) and
yellow (b*) units, while keeping the blue unit fixed at 0.1. The corresponding
color units were recorded. The hue and chroma values were determined
by using the formulae tan -1 (b/a) (LITTLE, 1975) and (a 2 +b 2 ) 1/2
(FROEHLICH et al., 1983), respectively, where a= red unit and b= yellow unit.
Free fatty acids
The free fatty acids content were determined by the procedure as described
by KONIECKO (1979):