外文文献.docx
- 文档编号:25964716
- 上传时间:2023-06-16
- 格式:DOCX
- 页数:15
- 大小:54.08KB
外文文献.docx
《外文文献.docx》由会员分享,可在线阅读,更多相关《外文文献.docx(15页珍藏版)》请在冰豆网上搜索。
外文文献
MediumoptimizationforglutathioneproductionbySaccharomycescerevisiae
Chi-HsienLiu,
Chin-FaHwang,
Chii-CherngLiao
FoodIndustryResearchandDevelopmentInstitute.P.O.Box246,Hsinchu,Taiwan,ROC
Received28April1997.Revised25March1998.Accepted28March1998.Availableonline19August1999.
http:
//dx.doi.org/10.1016/S0032-9592(98)00055-7,HowtoCiteorLinkUsingDOI
Permissions&Reprints
Abstract
Glucose,peptoneandmagnesiumsulphatewerefoundtobesuitablecomponentsforthecellgrowthandglutathione(GSH)productionintheyeaststrain.SaccharomycescerevisiaeCCRC21727.TheBox–Behnkendesignandresponsesurfacemethodologywereemployedtoderiveastatisticalmodeltoinvestigatetheeffectsofglucose,peptoneandmagnesiumsulphateconcentrationsonGSHproduction.Neuralnetworkswerecomparedwithasecond-order-polynomialmodelinpredictingtheeffectsofcomponentconcentrationsontheproductionofGSHanddrycellweight(DCW).NeuralnetworkmodelscanpredictcellgrowthandGSHproductionmorepreciselythansecond-order-response-surfacemodels.
Keywords
mediumoptimization;
glutathione;
neuralnetworkprediction;
response-surfacemodel
1.Introduction
Glutathione(GSH)isatripeptideconsistingofl-glutamate,l-cysteineandglycine.In1921,HopkinsisolatedGSHfromyeasts,animalliversandmuscle[1].GSHwidelyaffectsoxidationandreductioninvivo.Itisusedasamedicineforliverandasascavengeroftoxigen[2].HighlevelsofGSHcontentexistinsomestrainsofyeastswhicharethemicroorganismscommonlyusedforcommercialGSHproduction[3] and [4].
Neuralnetworks(NNs),computermodelsystems,matchthefunctionalityofthebraininafundamentalmanner.NNshaverecentlybeenusedinmanyareasoffermentationsuchasbrewing[5],[6] and [7]andprocessstatevariableestimate[8],[9] and [10].Kennedyetal.comparedtheuseofNNstofactorialdesignastoolsformediumdesignandreportedthatNNscouldhetrainedonlessdataandpredictedtheexperimentswithaccuracyformediumdesign[11].
Responsesurfacemethodology(RSM),anexperimentalstrategyforseekingtheoptimumconditionsforamultivariablesystem,isamuchmoreefficienttoolforoptimizationthantheone-factor-at-a-timeapproach[12].RSMhasbeensuccessfullyappliedforoptimizingmediagradientsinshake-flasksystems[13],[14],[15] and [16].Thesteepestascentmethod(SAM)wasusedtoevaluatetheinfluenceofcomponentconcentrationsontheproductviatheregressionmodel[12],[13] and [17].Inthisstudy,aGSH-producingstrainofSaccharomycescerevisiae,wasused.TheauthorsadoptedBox–Behnkendesign[12]andRSMtodeterminetheoptimalmediumforGSHproductionandcellgrowth.Neural-networkmodelswerecomparedwithsecond-order-response-surfacemodelstopredictGSHproductionanddrycellweight(DCW)alongthesteepestascentpaths.
2.Materialsandmethods
2.1.Organismandcultivationconditions
SaccharomycescerevisiaeCCRC21727(ATCC7754)fromtheCultureCollectionandResearchCenterofFoodIndustryResearchandDevelopmentInstituteinTaiwanwasmaintaincdonYMAgar(DifcoLaboratories,MI,USA)andgrowninYMBroth(DifcoLaboratories)at30°Cand150rpmfor18hastheinoculum.Forallexperiments,a4%inoculumwasusedina50mlitreHinton-flaskcontaining10mlitreculturemediumandincubatedat30°Cand150rpmfor24h.Eachtrialwasconductedintriplicate.
2.2.Experimentaldesignandstatisticalanalysis
Variouscarbon,nitrogenandsaltsourcesweretestedtodeterminethebestcomponentsforGSHproductionandcellgrowth.Themostimportantcomponentswerethenchosenforoptimizationofconcentrations.Thethree-variableBox–Behnkendesignwith13combinationsandthreereplicationsofthecentrepointwasused[18].Eachtrialwasconductedintriplicate.TheaveragedataofGSHproductionandDCWwereanalysedbySASsoftware[19]toconstructthesecond-ordermodelsandusedtotrainNNssoftware[20].AsetofexperimentsdesignedbySAMwasusedtoevaluatetheaccuracyofthesecond-ordermodelsandthetrainedNNs.
TheartificialNNhasbeeninspiredbybiologicalmodelsandthebuildingblocksofartificialNNareprocessingelements(PEs).PEsarecombinedintolayers,theinputisreceivedandweightedaccordingtoweightingfactorsandtheresultingquantitiesaresummed.PEsthenexecuteatransferfunctionontheweightedsum,andpassthevalueontothenextPE.ThetransferfunctionmapsthepossiblyinfinitesummationofPEinputtoapredefinedrange,theoutput.ThelearningortrainingphaseofaNNrequirespairedinput-outputdata.Theinputisfedintothenetwork,transferredthroughthenetworklayers.Ultimately,thenetworkcalculatesapredictedoutput.Thispredictedoutputissubsequentlycomparedwiththeactualoutput,andtheconnectionweightsbetweenthePEsaremodifiedtominimizethedeviationbetweenthepredictedoutputandtheactualone.Thelearningrulespecifieshowconnectionweightsarechangedduringthelearningprocess.Theprocedurecontinuesuntiladefinedaccuracyhasbeenreached.Thisistheconceptofback-propagation[24].
Inthisstudy,twoback-propagationNNsconsisting,offourlayers,oneinputlayer,twohiddenlayersandoneoutputlayer,wereused.ThenumbersofPEsinthesetwohiddenlayersweresixandthree,respectively.Thehyperbolictangentfunctionanddeltarulewerechosenforthetransferfunctionandlearningrule.
2.3.Assays
Afterfermentationthebrothwasboiledfor5min,ice-cooledandthencentrifugedat12 000rpmfor3min,theGSHcontentinthesupernatantwasdeterminedaccordingtothepublishedmethods[21] and [22]bymeasuringtheabsorbanceofreactionsolutionsat412nmusingaspectrophotometer(BeckmanInc.,DU7400Spectrophotometer,CA,USA).CellgrowthwasdeterminedbymeasuringtheDCW.DCWwasdeterminedafteryeastcellswerecentrifugedat12 000rpmfor3minandwashedtwicewithdeionizedwaterbeforedryingat70°Cfor24h.
3.Resultsanddiscussion
TheeffectsofvariouscarbonsourcesonGSHproductionandcellgrowthinS.cerevisiaecultureareillustratedinTable1.GlucosewasfoundtobethebestcarbonsourceforGSHproductionwhilefructoseandsucrosewerethetwomostsuitablecarbonsourcesforgrowthofS.cerevisiae.Shimizuandcoworkersreportedthatwhenglucoseratherthanethanolwasusedassolecarbonsource,bothspecificgrowthrateandGSHspecificproductionratewerehigher[3].CellgrowthwassimilarbutGSHproductionwas92%higherwhenglucosewasthecarbonsourceascomparedtoethanol.Withglycerolascarbonsource,theratioofGSHtoDCWwasthehighestamongallthecarbonsourcestestedbutGSHandDCWproductionswas56·9and25·8%ofthosewhenglucosewasusedascarbonsource.Whenlactoseormannitolwasused,GSHandDCWproductionwaspoor.Glucosewasusedasthecarbonsourceforfurtherexperiments.
Table1.EffectsofvariouscarbonsourcesonrelativeGSHproductionandrelativeDCWinS.cerevisiaeculturea
Carbonsource
RelativeGSHproduction(%)
RelativeDCW(%)
RatioofGSHtoDCW(%)
Glucose
100·0
100·0
0·48
Lactose
22·8
21·6
0·51
Fructose
77·1
117·3
0·32
Sucrose
87·9
115·3
0·37
Glycerol
56·9
25·8
1·06
Mannitol
24·0
28·9
0·40
Ethanol
51·7
100·0
0·25
a
Themediumcontained1%yeastextract,andthecarbonsourceswereaddedataconcentrationof1%(w/v).GSHproductionandDCWwhichequateto100%are28·26mglitre−1and5·88glitre−1,respectively.
Full-sizetable
TheeffectsofnitrogensourcesonGSHproductionandcellgrowthareshowninTable2.ThehighestGSHcontentperunitofDCWwasobtainedwhentryptonewasusedasnitrogensource,althoughtheamountsofGSHandDCWwere61·4and57·6%ofthosewhenpeptonewasused.PeptoneandyeastextractwerethetwobestnitrogensourcesforGSHproductionbyS.cerevisiae.Therewasatwo-foldincreaseingrowthwhenyeastextractwasusedinsteadofpeptoneasnitrogensource,resultinginlowerGSHcontentperunitofDCW.Peptonewasthuschosenasthenitrogensourceforfurtherexperiments.
Table2.EffectsofvariousnitrogensourcesonrelativeGSHproductionandrelativeDCWinS.cerevisiaeculturea
Nitrogensource
RelativeGSHproduction(%)
RelativeDCW(%)
RatioGSHtoDCW(%)
Peptone
100·0
100·0
0·97
Tryptone
61·4
57·6
1·04
Meatextract
80·7
112·8
0·70
Yeastextract
100·0
202·8
0·48
Maltextract
10·7
42·8
0·24
a
Themediumcontained1%glucose,andthenitrogensourceswereaddedataconcentrationof1%(w/v).GSHproductionandDCWwhichequateto100%are28·26mglitre−1and2·90glitre−1,respectively.
Full-sizetable
MetalionssuchasK+,Mg+2andCa+2directlyinfluencefermentativemetabolisminyeasts[25]and(NH4)2SO4andNH4ClstimulateethanolproductionbyS.cerevisiae[27].Khanetal.reportedthat(NH4)2SO4wasasuitablenitrogensourcefortheproductionofyeastbiomass[26].TheeffectsofvarioussaltsourcesonGSHproductionandcellgrowthareshowninTable3.AllsaltsexceptMgSO4showedinhibitiononbothGSHproductionandcellgrowth.TherewasnopreviousreportontheeffectofMgSO4onGSHproductioninyeast.Millsetal.reportedthatmagnesiumdeficientdietinhibitsbiosynthesisofbloodGSHinrat[23].AsMgSO4couldstimulateGSHproductionandcellgrowth,itwaschosenforfurtherstudy.
Table3.EffectsofvarioussaltsonrelativeGSHproductionandrelativeDCWinS.cerevisiaeculturea
Saltsource
RelativeGSHproduction(%)
RelativeDCW(%)
RatioofGSHtoDCW(%)
Controlb
100·0
100·0
0·97
CaCl2
39·0
87·2
0·44
KH2PO4
46·2
75·2
0·60
(NH4)2SO4
43·7
73·4
0·58
NH4CL
41·4
43·1
0·94
- 配套讲稿:
如PPT文件的首页显示word图标,表示该PPT已包含配套word讲稿。双击word图标可打开word文档。
- 特殊限制:
部分文档作品中含有的国旗、国徽等图片,仅作为作品整体效果示例展示,禁止商用。设计者仅对作品中独创性部分享有著作权。
- 关 键 词:
- 外文 文献