The isolation of pentose-assimilating yeasts and their xylose fermentation potential

h ttp : / / w w w . b j m i c r o b i o l . c o m . b r /

Biotechnology

and

Industrial

Microbiology

The

isolation

of

pentose-assimilating

yeasts

and

their

xylose

fermentation

potential

Gisele

Marta

Martins

a

_,

_Daniela

_Alonso

_{Bocchini-Martins}

b

_,

_Carolina

_{Bezzerra-Bussoli}

a

_,

Fernando

Carlos

Pagnocca

c

_,

_Maurício

_Boscolo

a

_,

_Diego

_Alves

_Monteiro

a

_,

Roberto

da

Silva

a

_,

_Eleni

_Gomes

a,∗

a_{UniversidadeEstadualPaulista(UNESP),InstitutodePesquisaemBioenergia-IPBen,LaboratóriodeMicrobiologiaaplicada,SãoJosédo}

RioPreto,SP,Brazil

b_{UniversidadeEstadualPaulista(UNESP),InstitutodeQuímica,Araraquara,SP,Brazil}

c_{UniversidadeEstadualPaulista-UNESP,CentrodeEstudosdeInsetosSociais-Ceis,CampusofRioClaro,SP,Brazil}

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Articlehistory:

Received5May2016 Accepted13November2016 Availableonline26August2017 AssociateEditor:SolangeI. Mussatto Keywords: Xylose Yeasts Ethanol

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Fortheimplementationofcellulosicethanoltechnology,themaximumuseoflignocellulosic materialsisimportanttoincreaseefficiencyandtoreducecosts.Inthiscontext,appropriate useofthepentosereleasedbyhemicellulosehydrolysiscouldimprovedeeconomicviability ofthisprocess.SincetheSaccharomycescerevisiaeisunabletofermentthepentose,thesearch forpentose-fermentingmicroorganismscouldbeanalternative.Inthiswork,theisolation ofyeaststrainsfromdecayingvegetalmaterials,flowers,fruitsandinsectsandtheir appli-cationforassimilationandalcoholicfermentationofxylosewerecarriedout.Fromatotal of30isolatedstrains,12wereabletoassimilate30gL−1ofxylosein120h.ThestrainCandida tropicalisS4produced6gL−1ofethanolfrom56gL−1ofxylose,whilethestrainC.tropicalisE2 produced22gL−1_{ofxylitol.ThestrainsCandidaoleophilaG10.1andMetschnikowiakoreensis}

G18consumedsignificantamountofxyloseinaerobiccultivationreleasingnon-identified metabolites.Thedifferentmaterialsinenvironmentweresourceforpentose-assimilating yeastwithvariablemetabolicprofile.

©2017PublishedbyElsevierEditoraLtda.onbehalfofSociedadeBrasileirade Microbiologia.ThisisanopenaccessarticleundertheCCBY-NC-NDlicense(http://

creativecommons.org/licenses/by-nc-nd/4.0/).

Introduction

Bioethanol or first generation ethanolin Brazilis obtained byfermentingglucosefrom sugarcanejuice andmolasses (aresiduefromsugarmaking)usingSaccharomycescerevisiae.

While this technologyisa consolidated industrial process,

∗ _{Correspondingauthor.}

E-mail:eleni@ibilce.unesp.br(E.Gomes).

productionofsecondgenerationethanolremainsachallenge. Oneofthebottlenecksistheutilizationofpentosesreleased byhydrolysisofhemicellulosethatcorrespondtoaround30% ofthelignocellulosicfeedstock.Yeastsabletoconvertxylose intoethanolhavebeendescribedlikeasScheffersomyces(Pichia) stipitis,S.(Candida)shehataeandPachysolentannophilus.1,2

How-evertheyieldhasnotreachedasatisfactorylevelforindustrial

https://doi.org/10.1016/j.bjm.2016.11.014

1517-8382/©2017PublishedbyElsevierEditoraLtda.onbehalfofSociedadeBrasileiradeMicrobiologia.Thisisanopenaccessarticle undertheCCBY-NC-NDlicense(http://creativecommons.org/licenses/by-nc-nd/4.0/).

applicationsOntheotherhand,pentose-assimilatingyeasts can beused for obtainmentofhigh aggregate value prod-ucts.Thepentose-assimilatingcapacityofyeastsPseudozyma antarcticaPYCC5048T_{,P. aphidisPYCC5535}T _{and P.rugulosa} PYCC5537T_{andproductionofaglycolipidbiosurfactantwas} demonstrate3_{aswellastheproductionofxylitolandarabitol}

byDebaryomyceshanseniifrompentose.4

A number of recent studies have been focused on the geneticengineeringofS.cerevisiae, aimedatmakingitable toproduceethanolfromglucoseandxylose.5_{Nevertheless,}

theyieldachievedresemblesthexylose-fermentingspecies, the xylose utilization is slow and occurs only after glu-cose exhaustion.3,4,6,7 _{The search for genetically modified}

strains is the focus of several studies. Thus, the study of microorganismspentose-fermenting and isolationofgenes involvedinpentosefermentationcanaffordsupportforthese studies.

Anotherresearchtendencyistheuseofpentosetoproduce alternativecompoundslikeorganicacids,8–12_xylitol,13_lipids

forbiodieselproduction,14–16_isobutanol17_andhydrogen18_all

usingmainlyyeastandbacteria.Besidescompoundsalready cited,newalternativeproductswithhighaggregatevalue,are stillappearing,increasing theoptions fortheutilization of thesesugarsinbiorefineriessystem.19

Consideringtheimportanceoftheuseofpentosesfrom lignocellulosic materials, the present work aimed at iso-lating yeast strains with the ability to assimilate and/or ferment xylose for future use for obtainment of products withbiotechnological interest.Thus,this studycontributes byincreasingthe informationonxylose-fermentingstrains andoffersgenomesforfuturestudiesofthemetabolismof microorganisms.

Materials

and

methods

Isolationandidentificationofstrains

Thestrains were isolated from fruits, flowers, insects and graperesidues.About0.5gofmaterialwassuspendedin4mL ofYEPXmedium(1%ofyeastextract,2%ofpeptoneandof 2%xylose) and incubated at30◦C.A loop of the homoge-nizedculturewasthenstreakedonthesolidYEPXmedium inPetridishestoisolatethecolonies.Thestrain identifica-tionwasmadeusingsequencesoftheD1/D2domainsofthe rDNA.20

Xyloseassimilationassay

Toevaluatethepotentialofxylose assimilation,thestrains werepre-cultivated inaYEP medium(1% yeastextract, 2% peptoneand2%glucose)and thebiomasswascentrifuged, watchedtwicewithsteriledistilledwaterandasuspensionof themusedasinoculumat1gofdrybiomassperliterofculture mediumcomposedofKH2PO4(2.0gL−1),(NH4)2SO4(2.0gL−1), MgSO4·7H2O(1.0gL−1),urea(0.3gL−1),CaCl2 (0.3gL−1)and xylose(30.0gL−1).Theincubationwascarriedoutat30◦Cand 150rpm.Theassaysweredoneinduplicateandwiththree repetitions.

Ethanolproductionassay

The alcoholic fermentation was assayed in 125mL Erlen-meyerflasksadaptedforalcoholicfermentation,closedwith avalvecontainingsodiummetabisulfitesolutionat1gL−1to ensurethatnooxygengot,andcontaining60mLofmedium describedabovebutwithyeastextractat10gL−1andxyloseat 100.0gL−1.Theincubationwasat30◦C.Theassaysweredone induplicatewiththreerepetitions.

Analyticalmethods

Toevaluatethedrycellmass,samplesoffermentedmedium werecentrifugedat10,000×g,by15min,thesupernatantwas discardedandtheprecipitatedcellsweredriedat60◦Cuntil theyreachedconstantweight.

Thereducingsugarconcentrationwasdeterminedusing the dinitrosalicylic (DNS) acid method, based on Miller (1959).21

The ethanol concentration was measured by a gas chromatograph (HP 5890) with an FFAP capillary column (polyethyleneglycol–30mm×0.22mm×0.3␮m)andaflame ionizationdetector,onesplit/splitlessinjector.Nitrogenwas utilizedasacarriergasat30mLmin−1.Temperaturesat injec-toranddetectorwere250◦C.

The xylitol quantification was performed by high-performance anion-exchange chromatography with pulsed amperometric detection(HPAEC-PAD).All sampleswere fil-tered(0.22␮mmembrane)andinjected(20␮L)inHPAEC-PAD System(ICS,DionexCorporation,USA)equippedwith auto-matic sampler AS40. The form of wave pattern used was the standard quadruple with the followingpotential pulse anddurations:E1=0.10V(t1=0.40s);E2=−2.00V(t2=0.02s); E3=0.60V(t3=0.01s);E4=−0.10V(t4=0.06s).AnIsocraticrun wasperformedwith10mMNaOHataflowrateof1mLmin−1 and35◦C.

Results

Isolation,identificationofyeastsandevaluationofxylose assimilation

Atotalof30 strainswereisolated andable togrowon the liquidYEPXmedium,but,amongthose,fourwerenotableto growinliquidmediumwithxyloseasthesolecarbonsource

(Table1).Inaerobiccultivation,ninestrainsconsumedallthe

xylosepresentinthemedium(30gL−1)in120h.

ThestrainPichiaguilliermondiiG1.2wasthemostefficient inthexyloseconsumption(maximumat48h)whilestrains

CandidatropicalisFP,C. tropicalisS4andP. guilliermondiiG4.2 exhaustedthesugarin72h.

Speciesorgeneraisolatedinthiswork,suchas Aureobasid-iumpullulans, Issatchenkiaterricola,I. orientalis,Metschnikowia bicuspidata,M.chrysoperlae,M.zobellii,P.guilliermondii,P. stipi-tis,RhodotorulaminutaandR.mucilaginosaarementionedinthe literatureasbeingabletoproduceethanolfromxylose.22–29

ThegeneraHanseniasporaandthespecieI.terricolaare glu-cosefermenting,butthereiscontradictinginformationabout

Table1–Yeastsisolatedandxyloseconsumptionduringaerobiccultivationusingabasalmediumandxylose(30gL−1)

asthesolecarbonsource.

Isolatedstrains Samples Xyloseconsumption Maximumbiomass production

Yield(biomass/xylose)

(gL−1) Time(h) (gL−1) Time(h) (gg−1)

AureobasidiumpullulansG19 Flor(Nyctaginaceae) 9.9 120 9.7 120 0.9

CandidatropicalisE2 Graperesidue 30.0 120 8.0 120 0.2

C.tropicalisFP Anthill 30.0 72 7.2 120 0.2

C.tropicalisS2 Grapeleaf 10.0 120 3.3 96 0.3

C.tropicalisS3 Grape 10.0 120 3.1 120 0.3

C.tropicalisS4 Grapeleaf 30.0 72 7.3 120 0.2

C.tropicalisT Winemust 20.1 120 7.1 120 0.3

C.oleophilaG10.1 Leaf(Asteraceae) 30.0 96 8.2 120 0.3

HanseniasporaguilliermondiiG9.1 Leaf(Asteraceae) 7.6 120 2.6 96 0.3

Hanseniasporasp.G1.3 Fruit(Anarcadiaceae) 19.4 120 3.7 72 0.2

Hanseniasporasp.G11.1 Fruit(Rutaceae) 7.6 120 3.5 120 0.5

Hanseniasporasp.G13 Fruit(Rutaceae) 10.4 120 2.7 72 0.2

Hanseniasporasp.G14 Flower(Myrtaceae) 9.9 120 2.0 120 0.2

Hanseniasporasp.G15 Fruit(Malpighiaceae) 3.5 120 2.2 96 0.6

Hanseniasporasp.G17 Flower(Rubiaceae) 5.4 120 2.8 48 0.5

Hanseniasporasp.G2 Flower(Amaryllidaceae) 10.2 120 5.6 48 0.5

Hanseniasporasp.G4.1 Flower(Nyctaginaceae) 23.0 120 8.2 120 0.3

Hanseniasporasp.G7.1 Fruit(Myrtaceae) 23.9 120 8.1 120 0.3

IssatchenkiaterricolaG20 Fruit(Myrtaceae) 5.1 120 5.0 96 0.9

MetschnikowiakoreensisG18 Flower(Amaryllidaceae) 30.0 120 6.7 96 0.2

PichiaguilliermondiiG1.2 Fruit(Anarcadiaceae) 30.0 48 10.8 120 0.3

P.guilliermondiiG4.2 Fruit(Rosaceae) 30.0 72 10.6 120 0.3

P.guilliermondiiB1 Winemust nda _{– nd – –}

P.guilliermondiiB2 Grape nd – nd – –

P.kluyveriG1.1 Fruit(Anarcadiaceae) 20.5 120 7.2 120 0.3

RhodotorulamucilaginosaG7.2 Flower(Euphorbiaceae) 29.4 120 11.5 120 0.4

R.mucilaginosaG11.2 Flower(Plantaginaceae) 17.3 120 6.7 120 0.4

Rhodotorulasp.G10.2 Flower(Plantaginaceae) 27.8 120 8.1 120 0.3

YarrowialipoliticaCO1 Grape nd – nd – –

YarrowialipolíticaCO2 Grape nd – nd – –

IdentificationbasedonsequencesoftheD1/D2domainsoftherDNA(atleast98%similarity,withreferencetohomologoussequencesfrom GenBank).

a _{Notquantified,thestrainswerenotabletogrowwithxyloseasthesolecarbonsource.}

thexyloseassimilationbytheseyeasts.22,25_{Themostefficient}

xylose-assimilatingyeastswerechosentocontinuethe exper-imentsandusedforfermentationforethanolproductionin subsequentassays.

Ethanolproductionbyyeasts

The ethanol production was evaluated for 30 strains and four of them were able to produce ethanol from xylose (from3to6gL−1)(datanotshown).ThethreestrainsofC. tropicalisshoweddifferentprofilesofconsumptionofxylose andethanolproduction.Thehighestethanolproductionwas observedwithC.tropicalisS4with6gL−1ofethanolandwhose xyloseassimilationwas56gL−1resultinginayieldof0.1gg−1, followingbyC.tropicalisFPthatproduced4.6gL−1ofethanol from84gL−1ofxylose(yieldof0.05gg−1).

ThestrainsRhodotorulasp.G10.2andC.tropicalisE2showed similarproductionofethanol(around3gL−1)toeachotherbut

Rhodotorulasp.G10.2consumed73gL−1ofxylosewithyieldof 0.04gg−1andC.tropicalisE2consumed53gL−1withyieldof 0.05gg−1(Fig.1A).

Fig. 1Bshows thexylose assimilation and biomass pro-ductionbystrainsthatnotwereabletoproduceofethanol.

P. guilliermondii G4.2,C.oleophilaG10.1 and M.koreensis G18 showed yieldsof biomassof0.7gg−1 meanwhileP. guillier-mondii G1.2 and R. mucilaginosa G7.2 had yields of 1.1 up to1.3gg−1, suggestingthat P. guilliermondiiG4.2, C.oleophila

G10.1andM.koreensisG18producednotidentifiedmetabolite. SinceC.tropicalishasbeenreportedasaxylitol-producing species30_{andtakingintoaccountitsconsumptionofxylose,}

thexylitolquantificationinthefermentationmediawas

car-riedout(Table2).

ThestrainC.tropicalisE2reachedaproductionof22gL−1 ofxylitolfrom100gL−1ofxylose(yieldof0.2gg−1)whileS4 produced15.5gL−1ofxylitol(0.15gg−1)andC.tropicalisFP pro-duced4.5gL−1ofxylitol(0.04gg−1).Thecomparisonbetween xylitolandethanolfermentativeparametersshowsthatthe strainC.tropicalisE2exhibitsmorepotentialforxylitol pro-ductionwhileC.tropicalisFPproducedsimilarconcentrations ofbothalcoholsbut withlowyield.Inthe culturemedium ofstrainC.tropicalisS4ahigherconcentrationofxylitolthan ethanolwasmeasured.

A

B

Ethanol (g.l -1) 7 6 5 4 3 2 1 0 0 12 24 36 48 60 72 84 96 108 120 0 20 40 60 80 100 Xylose (g.l -1) Xylose / biomass (g.l -1) Time (h) Time (h) 60 50 40 30 10 5 0 0 12 24 36 48 60 72 84 96 108 120

Fig.1–Alcoholicfermentationassayinabasalmediumwithxylose(100gL−1)andyeastextract(10gL−1).(A)Ethanol

productionandxyloseconsumptionbyC.tropicalisE2(squares),C.tropicalisS4(circles),C.tropicalisFP(triangle),and

Rhodotorulasp.G10.2(diamond);Opensymbols:xylose;Fullsymbols:ethanol.(B)ConsumptionofxyloseandbiomassbyP. guilliermondiiG1.2(squareshalfup)andG4.2(circlehalfright),C.oleophilaG10.1(star),M.koreensisG18(diamondhalfleft),

R.mucilaginosaG7.2(cross).

Table2–XylitolproductivitybyC.tropicalisFPstrain.Fermentationrealizedinabasalmediumwithxylose(100gL−1)

andyeastextract(10gL−1)at30◦Cin120h.

Parametersevaluated Xylitol Ethanol

E2 S4 FP E2 S4 FP Sugarconsumed(gL−1) 100 100 100 100 100 100 Production(gL−1) 22 15.5 4.5 3.0 5.9 4.6 Yield–Yp/s(gg−1) 0.22 0.15 0.04 0.03 0.06 0.04 Volumetricproductivity –Qp(gL−1h−1) 0.18 0.12 0.03 0.12 0.24 0.05 Conversionefficiency– Á(%)a 24 16 4 6 12 9

Adecreasingintheethanolconcentrationduringthe fer-mentativeroutewasobserved(Fig.1A),probablybecausethe yeastcanassimilate ethanol and,inthis case,theethanol determinedinthemediumis,perhaps,notreal.

Discussion

Ingeneral,theyeastsstrainspresentedxyloseassimilation butlowethanolproduction.C.tropicalisS4producedthe high-estamountofethanol(6gL−1),withayieldof0.1gg−1.This strainshowedgoodsugarconsumptionandthespeciesisnot amongthemostpromisingyeastsforethanolproduction.In fact,theliteratureshowsthatthemostpromisingspeciesfor ethanolproductionareScheffersomycesstipitis,S.shehataeand

Pachysolentannophilus,whileC.tropicalisisrelatedtoxylitol production.

ThestrainsC.oleophilaG10.1,M.koreensisG18andP. guil-liermondiiG4.2showedsmalleryieldsofbiomassproduction thanP.guilliermondiiG1.2andRmucilaginosaG7.2.Thissugar consumptionwithminorgrowthsuggeststheproductionof metaboliteswhich werenot ethanolorxylitol. Thus, these strains need further investigation about alternative com-poundslikeorganicacids.

P.guilliermondiiisfrequentlycitedforproductionofxylitol and/orethanol,butC.oleophilaandM.koreensishavenotbeen reportedintheliteratureforthefermentationofhydrolysate lignocellulosicmaterial.C.oleophilahasbeenreportedasan agent ofbiologic control due toits abilityto suppress the growth ofsomefungithat causefruit spoilagelike Botrytis cinereaand,Penicilliumdigitatum.32,33_{M.koreensiswasdescribed}

morerecently27_{andwasreportedtobeacarbonylreductase}34

andanL-talitolproducer.35

ThespecieA.pulluluans,ayeast-like,isabletoassimilate glucoseandxylosebutisreportedasunabletoferment glu-cose.Thisfeaturemakesthisyeastinterestinginapossible co-cultivation withS. cerevisiae. Furthermore,it isreported asbeingproducerofhighxylanaseactivity22,28 _allowingthe

hydrolysisofderivedfromhemicelluloseintheproductionof ethanolfromlignocellulosicmaterial.

ThestrainC.tropicalisS4showedslowerxylose assimila-tionbutconsumedallthesugarofthemediumin120hand producedthemostethanol.Furthermore,thedecreaseinthe ethanolinthefermentationmediaoverthe courseoftime indicatesthat, perhaps,itisabletoassimilatetheethanol, whichwouldjustifytheslowxyloseutilization.36

Undertheconditionstested,C.tropicalisFPconsumedall the xylose.Nevertheless, it presenteda low yieldfor both ethanolandxylitol.Also,duringthefermentationprocess,the straindidnotpresentalargeincreaseinthebiomass (maxi-mumbiomasswas7gL−1),thatsuggeststhepresenceofother coproducts.Meanwhile,C.tropicalisE2presentedhighxylose assimilationwithahigheryieldforxylitolthanforethanol, showingitspromiseforresearchintoxylitolproduction.

The amount of xylitol produced by C. tropicalis strains shows the importanceofthe individual study ofthe char-acteristics of each strain, since differences may occur in theproductionofcompounds, althoughbelongingtosame species. While the strain E2produced a small quantity of

ethanolandalargerquantityofxylitol,thestrainFPproduced thesamequantityofbothcompounds.

Theethanolandxylitolproductionisstronglyinfluenced bythefermentationconditionssuchaspH,temperature, oxy-genandnutrientswhichwerenotstudiedinthiswork.Maybe productivitycouldbeincreasedindifferentfermentation con-ditions,mainlycontrollingtheoxygensupply.

Themissingofethanolproductionfromxylosebythemost ofyeastsismarkedbyaredoximbalancethatoccursbetween cofactorsNADandNADPduringtheformationofxylitoland xylulose.ThexylosereductaseenzymereducesD-xyloseto xylitolusingNADPH,whereasxylitoldehydrogenaserequires NAD+_{mainly,whichhamperstherecyclingofcofactors.This} imbalancecausesxylitolaccumulationinthemediumandthe pathwaydoesnotproceedtoproduceethanol.Therecycling ofNADP+_{canberelievedthroughrespiratorychain.However,} dependingontheoxygenlevel,cellgrowthmayoccurinstead ofethanolproduction.37,38

Xylitolisindustriallyinterestingforitsapplicabilityasa sweetenerfordiabeticsandisutilizedinthefood, pharma-ceutical, oral health and cosmetics industries. In addition, itsproductionfromseverallignocellulosicmaterials,suchas corncobandcottonstalk,hasbeentested.30,39,40_Furthermore,

to increase the xylitol yield from lignocellulosic materials, somestudieslookedatstrategiessuchashydrolysate detox-ificationoryeastadaptation,cellimmobilizationandvarying theprocessconditions.41–46

Inconclusion,theassimilatingyeastsarecommoninthe environment but the understanding of the absorption and assimilationofthesugarisneededsothatbiotechnological utilizationofthispathwayispossible.Ourresultsshowedthe importanceoftheindividualstudyofeachstrain,sincethey havedifferentcharacteristics concerningresponseto exter-nalfactors,evenifthestrainsbelongtothesamespecies.The studyoftheproductionofalternativecompoundstoethanol isimportantbecausetheapplicationofthesestrainsin indus-trialprocesseshasnotbeenwellstudied.

Conflicts

of

interest

Theauthorsdeclarenoconflictsofinterest.

Acknowledgments

The authors would like to thanksFundac¸ão de Amparo à PesquisadoEstadodeSãoPaulo(Procn◦2010/12624-0), Con-selhoNacionaldeDesenvolvimentoCientíficoeTecnológico and Coordenac¸ão de Aperfeic¸oamento dePessoal emNível Superior fortheirfinancial supportandDaniela Correiade OliveiraLisboaforthetechniciansupport.

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c

e

s

1.BajwaPK,PhaenarkC,GrantN,etal.Ethanolproduction

fromselectedlignocellulosichydrolysatesbygenome

shuffledstrainsofScheffersomycesstipitis.BioresourTechnol.

2.WebbSR,LeeH.Regulationofd-xyloseutilizationby

hexosesinpentose-fermentingyeasts.BiotechnolAdv.

1990;8(4):685–697.

3.FariaNT,SantosMV,FernandesP,FonsecaLL,FonsecaC,

FerreiraFC.Productionofglycolipidbiosurfactants,

mannosylerythritollipids,frompentosesand

d-glucose/d-xylosemixturesbyPseudozymayeaststrains.

ProcessBiochem.2014;49(11):1790–1799.

4.GírioFM,AmaroC,AzinheiraH,PelicaF,Amaral-Collac¸oMT.

Polyolsproductionduringsingleandmixedsubstrate

fermentationsinDebaryomyceshansenii.BioresourTechnol.

2000;71(3):245–251.

5.MoonJ,LewisLiuZ,MaM,SliningerPJ.Newgenotypesof

industrialyeastSaccharomycescerevisiaeengineeredwithYXI

andheterologousxylosetransportersimprovexylose

utilizationandethanolproduction.BiocatalAgricBiotechnol.

2013;2(3):247–254.

6.HectorRE,DienBS,CottaMA,MertensJA.Growthand

fermentationofD-xylosebySaccharomycescerevisiae

expressinganovelD-xyloseisomeraseoriginatingfromthe

bacteriumPrevotellaruminicolaTC2-24.BiotechnolBiofuels.

2013;6(1):84.

7.GuoC,JiangN.Physiologicalandenzymaticcomparison

betweenPichiastipitisandrecombinantSaccharomyces

cerevisiaeonxylosefermentation.WorldJMicrobiolBiotechnol.

2013;29(3):541–547.

8.KatoH,MatsudaF,YamadaR,etal.Cocktail␦-integrationof

xyloseassimilationgenesforefficientethanolproduction

fromxyloseinSaccharomycescerevisiae.JBiosciBioeng.

2013;116(3):333–336.

9.WenS,LiuL,NieKL,DengL,TanTW,FangW.Enhanced fumaricacidproductionbyfermentationofxyloseusinga modifiedstrainofRhizopusarrhizus.BioResources.

2013;8(2):2186–2194.

https://ojs.cnr.ncsu.edu/index.php/BioRes/article/view/Bio

Res0822186WenFumaricAcidFermentationAccessed

01.05.16.

10.ZhaoJ,XuL,WangY,etal.Homofermentativeproductionof

opticallypureL-lacticacidfromxylosebygenetically

engineeredEscherichiacoliB.MicrobCellFact.2013;12

(1):57.

11.YeL,ZhouX,HudariMS,LiZ,WuJC.Highlyefficient

productionofL-lacticacidfromxylosebynewlyisolated

BacilluscoagulansC106.BioresourTechnol.2013;132:38–44.

12.ChenT,ZhuN,XiaH.Aerobicproductionofsuccinatefrom

arabinosebymetabolicallyengineeredCorynebacterium

glutamicum.BioresourTechnol.2014;151:411–414.

13.JarosAM,RovaU,BerglundKA.Acetateadaptationof

clostridiatyrobutyricumforimprovedfermentation

productionofbutyrate.SpringerPlus.2013;2(1):47.

14.IversonA,GarzaE,ZhaoJ,etal.Increasingreducingpower

output(NADH)ofglucosecatabolismforreductionofxylose

toxylitolbygeneticallyengineeredEscherichiacoliAI05.World

JMicrobiolBiotechnol.2013;29(7):1225–1232.

15.BabauM,CescutJ,AlloucheY,etal.Towardsamicrobial

productionoffattyacidsasprecursorsofbiokerosenefrom

glucoseandxylose.OilGasSciTechnol–Revd’IFPEnergies

Nouv.2013;68(5):899–911.

16.GaoD,ZengJ,ZhengY,YuX,ChenS.Microbiallipid

productionfromxylosebyMortierellaisabellina.Bioresour

Technol.2013;133:315–321.

17.MondalaA,HernandezR,HolmesW,etal.Enhanced

microbialoilproductionbyactivatedsludgemicroorganisms

viaco-fermentationofglucoseandxylose.AIChEJ.

2013;59(11):4036–4044.

18.BratD,BolesE.IsobutanolproductionfromD-xyloseby

recombinantSaccharomycescerevisiae.FEMSYeastRes.

2013;13(2):241–244.

19.ZhaoC,LuW,WangH.Simultaneoushydrogenandethanol

productionfromamixtureofglucoseandxyloseusing

extremethermophiles.II.Effectofhydraulicretentiontime.

IntJHydrogenEnergy.2013;38(22):9131–9136.

20.IsernNG,XueJ,RaoJV,CortJR,AhringBK.Novel

monosaccharidefermentationproductsinCaldicellulosiruptor

saccharolyticusidentifiedusingNMRspectroscopy.Biotechnol Biofuels.2013;6(1):47.

21.MeloWGP,ArcuriSL,RodriguesA,MoraisPB,MeirellesLA,

PagnoccaFC.Starmerellaacetif.a.,sp.nov.,anascomycetous

yeastspeciesisolatedfromfungusgardenoftheleafcutter

antAcromyrmexbalzani.IntJSystEvolMicrobiol.2014;64(PART 4):1428–1433.

22.MillerGL.Useofdinitrosalicylicacidreagentfor

feterminationofreducingsugar.AnalChem.

1959;31(3):426–428.

23.KurtzmanCP,FellJW.In:KurtzmanCP,FellJW,eds.The Yeasts.5thed.London:Elsevier;2011,

http://dx.doi.org/10.1016/B978-0-444-52149-1.00168-3.

24.GongC,ChenL,FlickingerMC,ChiangL,TsaoGT.Production

ofethanolfromD-xylosebyusingD-xyloseisomeraseand

yeasts.ApplEnvironMicrobiol.1981;41(2):430–436.

25.ToivolaA,YarrowD,vandenBoschE,vanDijkenJP,

ScheffersWA.Alcoholicfermentationofd-xylosebyyeasts.

ApplEnvironMicrobiol.1984;47(6):1221–1223.

26.NigamJN,IrelandRS,MargaritisA,LachanceMA.Isolation andscreeningofyeaststhatfermentd-xylosedirectlyto ethanol.ApplEnvironMicrobiol.1985;50(6):1486–1489.

http://aem.asm.org/content/50/6/1486.abstractAccessed

01.05.16.

27.SreenathH,JeffriesT.Productionofethanolfromwood

hydrolyzatebyyeasts.BioresourTechnol.2000;72(3):253–260.

28.HongSG,ChunJ,OhHW,BaeKS.Metschnikowiakoreensissp

nov.,anovelyeastspeciesisolatedfromflowersinKorea.IntJ

SystEvolMicrobiol.2001;51:1927–1931.WOS:000171162800039.

29.BhadraB,RaoRS,SinghPK,SarkarPK,ShivajiS.Yeastsand

yeast-likefungiassociatedwithtreebark:diversityand

identificationofyeastsproducingextracellular

endoxylanases.CurrMicrobiol.2008;56(5):489–494.

30.RaoRS,BhadraB,ShivajiS.Isolationandcharacterizationof

ethanol-producingyeastsfromfruitsandtreebarks.Lett

ApplMicrobiol.2008;47(1):19–24.

31.RafiqulISM,SakinahAMM.Processesfortheproductionof

xylitol—areview.FoodRevInt.2013;29(2):127–156.

32.GulatiM,KohlmannK,LadischMR,HespellR,BothastRJ.

Assessmentofethanolproductionoptionsforcornproducts.

BioresourTechnol.1996;58(3):253–264.

33.Bar-ShimonM,YehudaH,CohenL,etal.Characterizationof

extracellularlyticenzymesproducedbytheyeastbiocontrol

agentCandidaoleophila.CurrGenet.2004;45(3):

140–148.

34.SpadaroD,GullinoML.Stateoftheartandfutureprospects

ofthebiologicalcontrolofpostharvestfruitdiseases.IntJ

FoodMicrobiol.2004;91(2):185–194.

35.SinghA,ChistiY,BanerjeeUC.Stereoselectivebiocatalytic

hydridetransfertosubstitutedacetophenonesbytheyeast

Metschnikowiakoreensis.ProcessBiochem.

2012;47(12):2398–2404.

36.SasaharaH,IzumoriK.ProductionofL-talitolfromL-psicose

byMetschnikowiakoreensisLA1isolatedfromsoysaucemash.

JBiosciBioeng.2005;100(3):335–338.

37.CarolanG,CatleyBJ,KellyPJ.Ethanolproductionduringthe

growthcycleofAureobasidiumpullulans.BiochemSocTrans.

1976;4(6):1021–1022.

38.LiangM,HeQP,JeffriesTW,WangJ.Elucidatingxylose

metabolismofScheffersomycesstipitisbyintegratingprincipal

componentanalysiswithfluxbalanceanalysis.In:2013Am

39.BarbosaMFS,MedeirosMB,MancilhaIM,SchneiderH,LeeH.

Screeningofyeastsforproductionofxylitolfromd-xylose

andsomefactorswhichaffectxylitolyieldinCandida

guilliermondii.JIndMicrobiol.1988;3(4):241–251.

40.VanVleetJH,JeffriesTW.Yeastmetabolicengineeringfor

hemicellulosicethanolproduction.CurrOpinBiotechnol.

2009;20(3):300–306.

41.PingY,LingH-Z,SongG,GeJ-P.Xylitolproductionfrom

non-detoxifiedcorncobhemicelluloseacidhydrolysateby

Candidatropicalis.BiochemEngJ.2013;75:86–91.

42.DengLH,TangY,LiuY.Detoxificationofcorncobacid hydrolysatewithSAApretreatmentandxylitolproduction byimmobilizedCandidatropicalis.SciWorldJ.2014;2014,

http://dx.doi.org/10.1155/2014/214632.

43.MohamadNL,KamalSMM,AbdullahN,IsmailI.Evaluation

offermentationconditionsbyCandidatropicalisforxylitol

productionfromsagotrunkcortex.BioResources.

2013;8(2):2499–2509.

44.SoleimaniM,TabilL.Evaluationofbiocomposite-based

supportsforimmobilized-cellxylitolproductioncompared

withafree-cellsystem.BiochemEngJ.2014;82:

166–173.

45.ZhangQ,LiY,XiaL,LiuZ,PuY.Enhancedxylitolproduction

fromstatisticallyoptimizedfermentationofcottonstalk

hydrolysatebyimmobilizedCandidatropicalis.ChemBiochem

Eng.2014;28(1):87–93.

46.LiZ,GuoX,FengX,LiC.Anenvironmentfriendlyand

efficientprocessforxylitolbioconversionfromenzymatic

corncobhydrolysatebyadaptedCandidatropicalis.ChemEngJ.