Microbiological Production of Citric and Isocitric Acids from ...

54 S.V. KAMZOLOVA et al.: Production of Citric and Isocitric Acids from Sunflower Oil, Food Technol. Biotechnol. 46 (1) 51–59 (2008)where X 2 and X 1 are the biomass at the moment of timet 2 and t 1 , respectively.The biomass yield was calculated as follows:Y X/S =X/S /2/where X is the total amount of biomass in the cultureliquid at the end of fermentation (g/L) and S is theamount of the consumed oil (g/L).The specific rates of CA and ICA production (q CA orq ICA , respectively) were calculated using the followingequation:q=(P/X) ⋅ t /3/where P is the total amount of CA or ICA in the cultureliquid at the end of fermentation (g), X is biomass contentin a fermentor at the end of fermentation (g), and tis fermentation time (h).The mass yield coefficients of CA or ICA productionwere calculated as follows:Y=P/S /4/where P is the total amount of CA or ICA in the cultureliquid at the end of fermentation (g), and S is theamount of consumed oil (g).The volumetric productivity of the process was calculatedusing the following equation:C=(P/V) ⋅ t /5/where P is the total amount of CA or ICA in the cultureliquid at the end of fermentation (g), V is the volume offermentor (L), and t is fermentation time (h).Results and DiscussionProperties of wild type strain Y. lipolytica VKMY-2373 and the development of regime of oil feedingThe wild type strain Y. lipolytica was cultivated inthe complete medium containing 10 g/L of sunfloweroil and 5.0 g/L of (NH 4 ) 2 SO 4 in a 10-litre fermentor withinitial working volume of 5 L; pH=5.0 was adjusted automaticallywith 10–15 % NaOH. Fig. 1 illustrates a typicaltime course of growth of Y. lipolytica. Cell growth wasaccompanied by a decrease in the content of nitrogenand oil in the medium; 10 g/L of biomass were accumulatedat the end of fermentation (27 h) (Fig. 1a).The maximum specific growth rate (m max ) calculatedfrom the linear segment of the growth curve (Fig. 1)amounted to 0.230 h –1 ; a value comparable with literaturedata obtained for Y. lipolytica grown on fatty acids(8,9). The biomass yield from the consumed oil (Y X/S )was 1.1 g/L. The above findings support the potential ofY. lipolytica for the production of single-cell protein fromfatty material. High Y X/S of (1.1±0.3) g/L has been reportedfor different strains of Y. lipolytica cultivated onvarious types of crude fat (8,9,30,31). Recently, in experimentswith Y. lipolytica grown on completely saturatedfatty acid mixture (9), it has been demonstrated that anincrease in dissolved oxygen concentration from 5–15 to60–70 % of air saturation resulted in the increase of Y X/Sfrom 1.1 to 1.6, whereas the lipid production decreasedat the same time. Moreover, the growth parameters appearedto be critically influenced by fatty acid compositionof the carbon source. It was indicated that yeaststrains (Saccharomycopsis lipolytica and Apiotrichum curvatum)did not grow sufficiently well on saturated fat dueto inadequate dispersion of these substrates into the liquidmedium; solid fat required considerable agitation(1200 rpm) for dispersal in the growth medium (32,33).In other reports, it was observed that Y. lipolytica demonstratedsignificant growth without differentiations, regardlessof the fatty acid composition of the culture medium(saturated free fatty acids, rapeseed oil and mixture ofthese two substrates) (8).The utilization of sunflower oil involved its hydrolysisby lipase that resulted in the formation of di- andmonoglycerides, glycerol, and free fatty acids. The glycerolconcentration rapidly increased during the first 9 hof cultivation (up to 0.125 g/L) (Fig. 1b) and then remainedat this level. It should be noted that residual glycerolremained at the trace level as sunflower oil wasconsumed and no accumulation of glycerol was observed.Also, that a ratio among di-, monoglycerides, andfree fatty acids in the culture broth remained at the constantlevel in the course of cultivation (data not shown).Analysis of free fatty acid composition revealed that inthe course of cultivation, the fractions of oleic acid(D 9 C 18:1 ) and linoleic acid (D 9,12 C 18:2 ) decreased from 36 to27 % and from 52.1 to 35.0 % (by mass), respectively,while the amount of stearic acid (C 18:0 ) increased from4.0 to 26.0 % (by mass). It may be considered that thestrong fatty acid specificity of Y. lipolytica was probablyg(biomass, oil)/(g/L)g(glycerol)/(g/L)141210864200.50.40.30.20.10OilNH 4+(a)Biomass0 3 6 9 12 15 18 21 24 27t/hC 18:2(b)C 18:1C 18:0C 16:0Glycerol0 3 6 9 12 15 18 21 24 271200t/hFig. 1. The growth of Y. lipolytica VKM Y-2373. Variation of sunfloweroil, biomass, nitrogen concentrations (a), glycerol and freefatty acids (b) with time. Cultivation conditions: 10 g/L of sunflowerand 5.0 g/L of (NH 4 ) 2 SO 4 ; pH=5.09006003000706050403020100g(NH )/(mg/L)4 +w(fatty acids)/%

HEALTHTALK TRINITY HEALTH 5 • NOVEMBER 2007Trinity Health Goes Tobacco-Free November 15thCome November 15,Trinity Health will be atobacco-free zone –both inside and out.The Trinity HealthBoard of Directorsvoted unanimouslyto adopt atobacco-freecampus policy totake effect duringthe Great AmericanSmoke-Out, a daywhen Americanstraditionally try to kickthe habit.President and CEO TerryHoff says the new policymeans that patients,visitors, employees,volunteers and medicalstaff will be asked torefrain from using tobaccoproducts on allTrinity-owned and leasedproperty, includingoutdoor areas.“We know that tobaccouse around our facilitiesposes health and safetyrisks to those mostvulnerable to its harmfuleffects,” Hoff states. “As ahealthcare organization, wehave an obligation toprovide a healthy,tobacco-free environmentfor our patients, visitorsand staff.”The decision to eliminateboth smoking andsmokeless tobacco onTrinity campuses wasrecommended by anemployee committeecomprised of smokers andnonsmokers. CommitteeChair Ron Meier says thepolicy keeps faith withTrinity’s mission and alignsthe organization withregulatory initiativesunderway to require thathealthcare facilitiesprovide tobacco-freegrounds as well asbuildings.“We felt it was importantto set an example as healthprofessionals,” Meier says.“We realized that thischange would be difficultfor people who smoke, andthat’s why we announcedit months ago to givepeople plenty of time toprepare.”By implementing atobacco-free campuspolicy, Trinity Health joinsa growing number ofhealthcare organizations inNorth Dakota that areprohibiting tobacco use ontheir grounds.“Tobacco consumptionremains the leadingpreventable cause of deathand disability in NorthDakota,” says TrinityHospital Chief of StaffJeffrey Verhey, MD.“As apulmonaryspecialist, Ihave witnessedthe humancost associatedwith tobaccouse. By providinga smoke-freecampus we candemonstrate ourcommitment to thehealth of our patients,visitors and employees.”Meier says the policy isn’tintended to force anyoneto quit smoking, althoughstudies show thattobacco-free policies doencourage smokers to quit.“In the months that ourcommittee has beenmeeting one of ourmembers has stoppedsmoking,” he notes. Twoexceptions to thetobacco-free campus policywill be allowed. Residentsof Trinity Homes (thefacility serves as theirhome, after all) willcontinue to have access toa designated smoking area;and ambulatory patients inTrinity’s addictionsprogram will also use adesignated area. “In consultingwith our addictionTobacco’s Toll on North DakotaHealth Impacts* Approximately 11,000 North Dakota youth areprojected to die prematurely due to smoking.* Disease-specific, smoking-attributable death ratesper 100,000 people:- Lung cancer - 64.1- Coronary heart disease - 53.1- Chronic obstructive pulmonary disease - 50.9Economic Impacts* Smoking-attributable direct medical expenditures:Total - $247,000,000Annual cost per capita - $388* Smoking-attributable productivity costs:Total - $190,000,000Annual cost per capita - $299* Medicaid expenditures for smoking-relatedillnesses and diseases: $47,000,000 annuallyprofessionals the consensuswas that it’s best to dealwith one addiction at atime,” Meier explains.“Even so, we do providepatients with informationabout smoking andsmoking cessationprograms.”He notes smokers whohave family members inthe ETC or ICU at TrinityHospital will most likelyhave the biggest challengerefraining from lighting upoutside. Says Mr. Hoff:“We understand andrespect every person’s rightto make choices related totheir health. We also havean obligation to protectour patients, employeesand visitors from theeffects of tobacco use. Weknow that people don’twant to harm our patientsor other vistors, so wedon’t anticipate havingproblems,” he adds.Ready to quit?Tobacco cessation assistance is available throughMinot’s First District Health Unit. Call 852-1376 formore information. Additional help is also available bycalling the ND Tobacco Quitline at 1-866-388-7848.Dear Babydoc,Why is my doctor always running late? I am tired of waiting!Busy ParentPlease sendyour questions toMarketing DepartmentAttn: Kids’ KornerTrinity Hospital – St. Joseph’s407 3rd Street SEMinot, ND 58701Dear BP,Your doctor regrets running late! Unfortunately we are not always able topredict how much time each child will need. Sometimes 10 minutes ismore than enough, other times 30 minutes is NOT enough for acomplicated problem. And there are emergencies or sick newborns whichcan never be on a schedule. We do try to stay on schedule because werespect your time. Please be patient and remember that some day yourchild may be the one who needs the extra time.BDDear Babydoc,Why doesn’t my child’s doctor take the time to see my other child too? We are already there and he probably has the same illness ashis sister!Don’t Want Extra CopaysDear DWEC,You are right, both your children may have the same illness. However, extra time spent with your child takes away time from other patientswho have appointments on the schedule. And as every child handles illness slightly differently, the ‘added on’ patient may be sicker than thesibling and thus take more time than you realize. If you have concerns about your other children, please schedule time for them also.BDDear Babydoc,Why is the nighttime doctor always so crabby on the phone? I only called for a dosage of cold medicine!Frustrated DadDear FD,Doctors don’t mean to be crabby on the phone. Sometimes parents abuse the after-hours phone call to ask questions that should be askedduring the office hours. After 5 pm, all phone calls should be emergencies. You wouldn’t want to be disturbed for non-urgent matters either.Remember: most doctors are parents also and evenings are family time for them too!BDDear Babydoc,I like to give my daughter Tylenol and Motrin, she seems to feel better faster. Why doesn’t my doctor let me do this?Concerned MomDear CM,Unfortunately, as both those medicines are digested by the liver, using them together can cause liver problems later. Ask your doctor whichhe/she prefers and use that instead. And remember, fever is not a problem by itself. It is the body’s way of fighting illness, use the medicinesonly to treat your child’s fussiness or pain.BD

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FTB 46 (1) 51-59.Mikrobna proizvodnja limunske i izolimunske kiseline na podlozi odsuncokretovog uljaSažetakU radu je ispitan rast divljeg soja kvasca Yarrowia lipolytica VKM Y-2373 imutantnog soja Yarrowia lipolytica N 15, te biosinteza limunske i izolimunske kiseline napodlozi od suncokretovog ulja. Utvrđeno je da je rast stanica povezan s istodobnimiskorištenjem glicerola i slobodnih masnih kiselina nastalih tijekom hidrolize ulja. Usporednoje ispitana aktivnost enzima koji sudjeluju u metabolizmu glicerola (glicerol kinaze),glioksilatnom ciklusu asimilacije masnih kiselina (izocitrat liaze i malat sintaze) i ciklusulimunske kiseline u kvascu Y. lipolytica uzgojenom na podlozi od suncokretovog ulja,glicerola i oleinske kiseline. Glicerol kinaza i enzimi glioksilatnog ciklusa bili su aktivnicijelo vrijeme uzgoja stanica na podlozi od suncokretovog ulja. Količina proizvedenelimunske kiseline i omjer limunske i izolimunske kiseline ovise o upotrijebljenom soju isastavu podloge. Otkriveno je da je divlji soj Y. lipolytica VKM Y-2373 proizveo skorojednaku količinu limunske i izolimunske kiseline pri pH=4,5, a pretežno izolimunsku kiselinupri pH=6.0. Mutantni soj Y. lipolytica N 15 proizveo je samo limunsku kiselinu (150 g/L, smasenim stupnjem pretvorbe Y CA =1,32 g/g). U radu su također razmotrene biokemijskeznačajke mutantnog soja Y. lipolytica N 15.