Itiswithoutdoubtthattestingplaysapivotalrolethroughoutthewholeofthevinificationprocess.ToproducethebestpossIBLequalitywineandtominimiseprocessproblemssuchas“stuck”fermentationortroublesomeinfections,itisnowrecognisedthatifpossibletestingshouldbeginpriortoharvestingofthegrapesandcontinuethroughtobottling.TrADItionalmethodsofwineanalysisareoftenexpensive,timeconsuming,requireeitherelaborateequipmentorspecialistexpertiseandfrequentlylackaccuracy.However,enzymaticbio-analysisenablestheaccuratemeasurementofthevastmajorityofanalytesofinteresttothewinemaker,usingjustonepieceofapparatus,thespectrophotometer(seepreviousissueNo.116foradetailedtechnicalreview).Grapejuiceandwineareamenabletoenzymatictestingasbeingliquidstheyarehomogenous,easytomanipulate,andcangenerallybeanalysedwithoutanysamplepreparation.

ManyoftheenzymatictestkitsareofficialmethodsofprestigiousorganisationssuchastheAssociationofOfficialAnalyticalChemicals(AOAC)andtheAmericanAssociationofCerealChemists(AACC)inresponsetotheinterestfromoenologists.Megazymedecidedtouseitslonghistoryofenzymaticbio-analysistomakeasignificantcontributiontothewineindustry,bythedevelopmentofarangeofadvancedenzymatictestkits.Thistaskhasnowbeensuccessfullycompletedthroughthestrategicandcomprehensiveprocessofidentifyinglimitationsofexistingenzymaticbio-analysistestkitswheretheyoccurred,andthenusingadvancedtechniques,suchasmolecularBIOLOGy(photo1),torapidlyovercomethem.Noveltestkitshavealsobeendevelopedforanalytesofemerginginteresttotheoenologist,suchasyeastavailablenitrogen(YAN;seepages2-3ofissue117article),orwherepreviouslyenzymesweresimplyeithernotavailable,orweretooexpensivetoemploy,suchasforD-mannitolanalysis.

Thispaperfocusesonfermentationoflactosefromamodelsystem(blacktea)andfromtwotypesofmilk(0.9%w/wand2.2%w/woffat)byapplicationofKombucha.QuantitiesoftheappliedKombuchastarterwere10%v/vand15%v/v.Allfermentationswereperformedat42°C.TheprocesstoachieveadesirablepH=4.5wasslowerinthemodelsystem(16h)thaninmilks(9-10h).Regardingstarterquantity,10%v/vprovedtheoptimal.Regardingtypesofmilk,higherfatcontentguaranteesshorterfermentationandhigheryieldofmetabolites.Utilizationoflactosewasfoundatalevelof≈20%and≈30%inmilkswith0.9%w/wand2.2%w/woffat,respectively.Thiswascorrelatedwithanappearanceofintermediatesand/orproducts.Glucoseunderwentfurthertransformationsalmostentirely,whilegalactoseshowedmuchlowerreactivity.Seventotwelvetimeshighercontentsoflacticacidwerefoundcomparedtoaceticacid.Milk-basedbeveragefromthereducedfatsample,inoculatedwith10%v/vofKombuchastarter,hasthebestphysicalcharacteristics(syneresisandwaterholdingcapacity).Italsodevelopedagoodtexture(especiallycohesivenessandindexofviscosity).Milklactosefermentationwasaprocessthatcouldhavebeenusedforobtainingnewmilk-basedproducts.

Wineisacomplexconsumerproductproducedpredominatelybytheactionofyeastupongrapejuice.Modelmustsystemshaveproventobeidealforstudiesintotheeffectsoffermentationconditionsontheproductionofcertainwinevolatiles.Toclarifythecontributionofgrapejuicetotheproductionofwinevolatiles,wehaveemployedamodelmustsystemspikedwithincreasingamountsofgrapejuice(RieslingorCabernetSauvignon).TheresultingfermentedwineswereanalyzedbySPME-GC-MSandthedataobtainedgroupedusingANOVAandclusteranalysestorevealthosecompoundsthatvariedinconcentrationwithreproducibletrendsrelativetojuiceconcentration.Suchgroupinghighlightsthosecompoundsthataregrape-dependentorforwhichproductionismodulatedbygrapecomposition.Insomecases,increasingtheproportionofgrapejuiceinthefermentationsstimulatedtheproductionofcertainesterstolevelsbetween2-and140-foldhigherthanthoseseeninfermentationsmadewithmodelgrapejuicemediaalone.Theidentificationofthegrapecomponentsresponsiblefortheincreasedproductionofthesewinevolatileswillhaveimplicationsfortheimpactofgrapeproductionandenologyonwineflavorandaroma.

Saccharomycescerevisiaemetabolizesarginine,oneofthemajoraminoacidsingrapemusts,toornithineandureaduringwinefermentations.WineyeaststrainsofS.cerevisiaedonotfullymetabolizeureaduringgrapemustfermentation.Ureaissecretedbyyeastcellsanditreactsspontaneouslywithethanolinwinetoformethylcarbamate,apotentialcarcinogenicagentforhumans.ThelackofureacatabolismbyyeastinwinemaybeascribedtothetranscriptionalrepressionoftheDUR1,2genebygoodnitrogensourcespresentinthegrapemust.WeexpressedtheDUR1,2geneundercontroloftheS.cerevisiaePGK1promoterandterminatorsignalsandintegratedthisDUR1,2expressioncassette,flankedbyura3sequences,intotheURA3-locusoftheindustrialwineyeastUCDavis522.InvivoassaysshowedthatthemetabolicallyengineeredindustrialstrainreducedethylcarbamateinChardonnaywineby89.1%.Analysesofthegenotype,phenotype,andtranscriptomerevealedthattheengineeredyeast522^EC−issubstantiallyequivalenttotheparental522strain.

Theobjectiveofthisresearchwastodeterminetheconversionefficiencyofprosomillettoethanolcomparedtocorninabench-scaledry-grindprocedure.SevenprosomilletcultivarsandsixadvancedbreedinglinescontainingwaxystarchwerefermentedwithSaccharomycescerevisiaeandethanolproductionwascomparedwithnormalcornand“highlyfermentable”corn.Thehighlyfermentablecornexhibitedthehighestfermentationefficiency(97.0±1.4%).Amongprosomilletlines,thosewiththehighestfermentationefficiencieswere:Huntsman(85.9±0.6%),172-2-9(90.8±0.2%),172-2-13(85.1±2.5%),and182-4-24(84.7±2.1).Waxyprosomilletlinesresultedinhigherfermentationefficienciesthanthenon-waxyprosomilletvarietiescontainingnormalstarch(82.4±5.5%vs.75.5±7.4%,respectively,p=0.01).Prosomilletdistiller"sdriedgrainswithsolubles(DDGS)containedmoreprotein(26.6–33.4%)thantheDDGSfromcorn(17.2–23.4%).Thesedataindicatethatprosomilletexhibitspromiseasafeedstockforethanolproduction,especiallyifbreedingprogramsfocusonselecting“highlyfermentable”linesforadvancement.

Productionofhighvalueproductsfromglycerolviaanaerobicfermentationisofutmostimportanceforthebiodieselindustry.ThemicroorganismEscherichiacoli(E.coli)K12wasusedforfermentationofglycerol.Theeffectsofglycerolconcentrationandheadspaceconditionsonthecellgrowth,ethanolandhydrogenproductionwereinvestigated.Afullfactorialexperimentaldesignwith3replicateswasconductedinordertotestthesefactors.Underthethreeheadspaceconditionstested,theincreaseofglycerolconcentrationacceleratedglycerolfermentation.Theyieldsofhydrogenandethanolwerethelowestwhenglycerolconcentrationof10g/Lwasused.Themaximumproductionofhydrogenwasobservedwithaninitialglycerolconcentrationof25g/Latafinalconcentrationofhydrogenwas32.15mmol/L.Thisstudydemonstratedthathydrogenproductionnegativelyaffectscellgrowth.Maximumethanolyieldwasobtainedwithaglycerolconcentrationof10g/Landwasupto0.40g/gglycerolundermembraneconditionheadspace.Statisticaloptimizationshowedthatoptimalconditionsforhydrogenproductionare20g/Linitialglycerolwithinitialspargingofthereactorheadspace.Theoptimalconditionsforethanolproductionare10g/Linitialglycerolwithmembrane.

AdifferentialpHmeasurementdevicewasusedtoachieveoperationconditionsofalcoholdehydrogenasereaction.Optimumoperatingconditionsweretemperatureof30°C,10μlofalcoholdehydrogenaseenzymevolume(withafinalactivityof563.75unitsml^-1)per50μlofsample,NAD⁺concentrationof0.05mMand20mMglycine-pyrophosphatebuffersolutionofpH9.1.Inthismethodarangeofethanolconcentrationsfrom0-99,985%,whichmeans0.000001714-17.14M,wereused.ThemaximumobtainedchangeinpH,deltapH,was(-33)mpH.AcalibrationcurveoflogarithmicvaluesofethanolconcentrationsagainstchangeinpHforstandardethanolsampleswasdone.Sincethiscalibrationcurveisalinearwithacorrelationcoefficient(R)of0.998,thiscalibrationcurvecanbeusedinquantificationofethanolconcentration.Endpointofequilibriumconcentrationsofreactantsandproductsofethanoloxidationreactionwasmeasuredwithinspectrophotometer.Theresultsindicated100secondsofprocesstimeisrequiredtoreachtheendpointforallethanolstandardsamples.ThisrequiredtimewassatisfiedwithresultsofmeasuringchangeinpHwithindifferentialpHanalyzersystem.

Thethermophilicbacteriareceiveconsiderablyinterestnowadaysbecauseofacurrentchallengeofincreasingglobaltemperature.Particularlyforethanolproduction,thethermo-ethanologenicbacteriapossessadvantagesduetolowercontaminationrisk,costsavinginindustrialscale,andthewiderangeofsugarsutilization.Inthisstudy,13bacterialisolatesobtainedfromthepreviousisolationstudyweretestedfortheirfermentativecapacityandethanoltoleranceathightemperatures.FivebacterialisolatesHM2,M2,MC3,MR1andRDwerefoundtobetolerantupto12%ethanol.OfwhichHM2,M2andMR1couldfermentglucosewellat30,35and40°C,particularlyisolatesHM2andMR1couldperformthefermentativecapacityat45°Candeven50°C.Inthepresenceof12,16,and20%w/vglucose,isolatesHM2,M2,andMR1showedthehighfermentationratebygivinghighgasproduction;however,therateslightlydecreasedinthepresenceof24%w/vglucose.ThefermentativeperformancebythesethreeisolatescouldhappenatdifferentpHlevelsof4.0,5.0and6.0.Thefavourableconditionsofethanolfermentationwerefoundat18.5%glucose,pH5.0,and33°CforisolateHM2andat14%glucose,pH5.5,and40°CforisolateMR1.Theresultsofsequencinganalysisofpartial16SrRNAgeneshowedthatthegenesequencesoftheselectedisolateHM2shared99%similaritywithBacillussubtilis.

Inthisstudy,variousyeasts(Kluyveromycesmarxianus,Saccharomycesturicensis,Pichiafermentans)andlacticacidbacteria(Lactobacilluskefiranofaciens,Lactobacilluskefiri,Leuconostocmesenteroides)wereentrappedin2differentmicrospheresusinganentrapmentratioforthestrainsthatwasbasedonthedistributionratiooftheseorganismsinkefirgrains.Thepurposeofthisstudywastodevelopanewtechniquetoproducekefirusingimmobilizedstarterculturesisolatedfromkefirgrains.Anincreaseincellcountswithfermentationcycleswasobservedforboththelacticacidbacteria(LAB)andyeasts,whereasthecellcountsofkefirgrainswereverystableduringcultivation.Scanningelectronmicroscopyshowedthattheshort-chainlactobacilliandlactococcioccupiedthesurfaceoftheLABmicrospheres,whereasthelong-chainlactobacilliwereinsidethemicrospheres.Whentheyeastswereanalyzed,cellsatahighdensitywereentrappedincracksonthesurfaceandwithinthemicrospheres,wheretheyweresurroundedbytheshort-chainlactobacilli.ThedistributionoftheLABandyeastspeciesinkefirproducedfromgrainsandmicrospheresshowedthattherewasnosignificantdifferencebetweenthekefirsproducedbythe2methods;moreover,Leu.mesenteroidesandK.marxianuswerethepredominatingmicroflorainbothtypesofkefir.Therewasnosignificantdifferenceintheethanolandexopolysaccharidecontentsbetweenthe2kefirs,althoughtheaciditywasdifferent.

Thepresentstudy,taraxerone(D-friedoolean-14-en-3-one)wasisolatedfromSedumsarmentosumwithpurity96.383%,anditsenhancingeffectsonalcoholdehydrogenase(ADH)andacetaldehydedehydrogenase(ALDH)activitiesweredetermined:EC₅₀valueswere512.42±3.12and500.16±3.23μMforADHandALDH,respectively.Inordertoobtainmoreinformationontaraxeronerelatedwiththealcoholmetabolism,40%ethanol(5mL/kgbodyweight)with0.5–1mMoftaraxeronewereadmiNISTeredtomice.Theplasmaalcoholandacetaldehydeconcentrationsoftaraxerone-treatedgroupsweresignificantlyloweredthanthoseofthecontrolgroup(p<0.01):=""approximately=""20–67%=""and=""7–57%=""lowered=""for=""plasma=""alcohol=""and=""acetaldehyde,=""respectively.=""compare=""to=""the=""control=""group,=""the=""adh=""and=""aldh=""expressions=""in=""the=""liver=""tissues=""were=""abruptly=""increased=""in=""the=""taraxerone-treated=""groups=""after=""ethanol=""exposure.=""in=""addition,=""taraxerone=""prevented=""catalase,=""superoxide=""dismutase,=""and=""reduced=""glutathione=""concentrations=""from=""the=""decrease=""induced=""by=""ethanol=""administration=""with=""the=""concentration=""dependent=""manner.="">

InSaccharomycescerevisiae,genesencodingmaltosepermeasesandmaltasesarelocatedinthetelomericregionsofdifferentchromosomes.TheCOMPASSmethylationcomplex,whichmethylateslysine4onhistoneH3,controlsthesilencingoftelomericregions.YeaststrainsdeletedforSWD1,SWD3,SDC1,SET1,BRE2,orSPP1,encodingcomponentsoftheCOMPASScomplex,fermentedamediumcontaining22%maltosewithnoticeablyhigherattenuationthandidthewildtype,resultinginproductionofupto29%moreethanol.Theleasteffectivestrainwasspp1.AbsenceofCOMPASScomponentshadnoeffectonthefermentationofmediawith20%glucose,20%sucrose,or16%maltose.DeletionofSWD3resultedinlargeramountsofMAL12transcript,encodingmaltase,atthelatestagesoffermentationof22%maltose.Asimilareffectonmaltaseactivityandmaltoseuptakecapabilitywasseen.Thelysine4residueofhistoneH3wastrimethylatedinwild-typecellsatthelatestages,whileonlysmallamountsofthedimethylatedformweredetected.TrimethylationanddimethylationofthisresiduewerenotdetectedinstrainsdeletedforSWD1,SWD3,SET1,BRE2,orSDC1.Trimethylatedlysine4wasapparentonlyattheearlystages(48and96h)offermentationinanspp1strain.ThisworkindicatesthattheCOMPASScomplexrepressestheexpressionofmaltoseutilizationgenesduringthelatestagesoffermentationofahighconcentrationofmaltose.

Ethanolproductionfromnon-pretreatedlignocellulosewascarriedoutinaconsolidatedbioprocesswithwood-decayfungiofphlebioidPolyporales.Ethanolproductionwasattemptedonglucose,sprucewoodsawdustandwastecoreboard.Substantialquantitiesofethanolwereachieved,andisolatePhlebiaradiata0043produced5.9 g/Lofethanolreachingtheyieldof10.4%ethanolfromcoreboardlignocellulosesubstrate.Acidicinitialcultureconditions(pH3)inducedethanolfermentationcomparedtothemoreneutralenvironment.Togetherwithbioethanol,thefungiwereabletoproduceorganicacidssuchasoxalateandfumarate,thusbroadeningtheircapacityandapplicabilityasefficientorganismstobeutilizedforbioconversionofvariouslignocelluloses.Inconclusion,fungiofPhlebiagrowon,convertandsaccharifysolidlignocellulosewastematerialswithoutpre-treatmentsresultinginaccumulationofethanolandorganicacids.Thesefindingswillaidinapplyingfungalbiotechnologyforproductionofbiofuelsandbiocompounds.

Thepreparationofcompositesoflivingfunctionalcellsandpolymersisamajorchallenge.Wehavefabricatedsuch“livingcomposites”bypreparationofpolymericmicrotubesthatentrapyeastcells.Ourapproachwastheprocessofcoaxialelectrospinninginwhichacorecontainingtheyeastwas“spun”withinashellofnonbiodegradablepolymer.WeutilizedtheyeastCandidatropicalis,whichwasisolatedfromolivewaterwaste.Itisparticularlyusefulsinceitdegradesphenolandothernaturalpolyphenols,anditiscapableofaccumulatingethanol.Theelectrospunyeastcellsshowedsignificantactivityofbioremediationofphenolandproducedethanol,and,inaddition,themetabolicprocessesremainedactiveforaprolongedperiod.Comparisonofelectrospuncellstoplanktoniccellsshoweddecreasedcellactivity;however,theolivewaterwasteaftertreatmentbytheyeastwasnolongertoxicforEscherichiacoli,suggestingthatdetoxificationandprolongedviabilityandactivitymayoutweighthereductionofefficiency.

LargedeformationmechanicaltestingandFourier-transforminfraredspectroscopywereusedtogainfurtherinsightsintothemechanismofheatresistanceinethylcellulose-(EC)stabilizedchocolatepreparedusingthesolventsubstitutionmethod.HereweshowthatthepresenceoflecithinatthesurfaceofsucrosereducedheatresistancebyimpedinginteractionsbetweenECandsucrose.ThesetechniquesalongwithfluorescencemicroscopyalsoshowedthattheEtOHusedinsolventsubstitutionchocolatewasabletoremovetheEtOHsolublelecithinphospholipidsfromthesurfaceofthesucrose.RemovalofthelecithinandtheslightsolubilityofsucroseinEtOHbothhavepositiveimpactsonheatresistance.ItwasalsofoundthatEtOHmayreduceheatresistancebydestabilizingthecaseinmicelleinsamplesmadewithskimmilkpowder.Finally,resultshaveindicatedthatECislikelyabletointeractwiththelactoseinskimmilkpowderandthestarchincocoapowderleadingtogreaterheatresistance.Thesefindingswillbeusefulindevelopingtheidealheatresistantchocolateformula.

Fishsolidwaste(faeces)producedinrecirculatedaquaculturesystems(RAS)mightbeusedforon-farm,single-sludgedenitrificationiftransformedintosolubleorganiccarbonsubstances.Thecurrentstudyinvestigatedtheeffectoffeedingdietswithincreasingproteintoenergyratios(P:E_15,17,19,21and23 g/MJ)torainbowtrout(Oncorhynchusmykiss)ontheproductionofvolatilefattyacids(VFAs)andethanolduring7 daysfermentationoftheproducedfishfaeces.ThetotalyieldsofVFAsandethanolobtained(expressedaschemicaloxygendemand(COD))rangedbetween0.21–0.24 gCOD/gTCOD,showingnodifferencesbetweentreatments.However,thetypeandquantitiesofindividualVFAsandethanolchangedaccordingtothedietarytreatment.LowerP:Eratiodietsresultedinhigherproductionofbutyricacidandethanol,whereashigherP:Eratiodietsresultedinanincreasedproductionofaceticandvalericacid.Changingthedietcompositionthusaffectsthecompositionofreadilyavailablecarbonthatcanbederivedfromthefaeces.Thiscanbeappliedtoenhanceon-farmsinglesludgedenitrificationandreducetheneedforaddingexternalcarbonsourcessuchase.g.methanol.

Toinvestigatetheanti-hangovereffectsofCudraniatricuspidatarootextract(CTE),thebloodalcoholmetabolismandbloodgasimbalanceofCTEinratstreatedwith10ml/kgalcoholwereexamined.CTE(500mg/kgand750mg/kg)wasadministratedafter30minutesofalcoholconsumption(10ml/kg).Bloodcollectionwasimplementedfromthetailsoftheanimalsafter1,3,and5hourspostalcoholconsumption.TheConditiondrink(acommercialanti-hangoverbeverage)wasusedasapositivecontrol.Singleadministrationbytheoralroutewasperformed.TheconsumptionofCTE(500mg/kgand750mg/kg)decreasedtheserumalcoholconcentrationbyincreasingandmaintainingboththealcoholdehydrogenase(ADH)andacetaldehydedehydrogenase(ALDH)enzymeactivitylevelsinthebloodandliver.Inaddition,CTEledtorecoveryfromtheimbalancesinthebloodgaslevels,includingcarbondioxide(CO₂^-)andchangesinpH,bicarbonate(HCO₃^-)andlacticacidlevelsduetoalcoholingestion.Inconclusion,CTEexertedamorepronouncedanti-hangovereffectthanacommercialanti-hangoverdrink.Therefore,CTEcanbeanovelandsafeanti-hangovernaturalproductagentforthepreventionortreatmentofsymptomscausedbyexcessivealcoholconsumption.

Alcoholicfermentationswereperformedadaptingthetechnologytoexploittheresidualthermalenergy(hotwaterat83-85°C)ofacogenerationplantandtovalorizeagriculturalwastes.Substrateswereapple,kiwifruitandpeacheswastesandCornThreshingResidue(CTR).Saccharomycesbayanuswaschosenasbiocatalyst.Thefruits,freshorblanched,weremashed;CTRwasgelatinizedandliquefiedbyaddingLiquozyme^®SCDS(Novozyme);saccharificationsimultaneoustofermentationwascarriedoutusingtheenzymeSpirizyme^®Ultra(Novozyme).Lab-scalestaticfermentationswerecarriedoutat28°Cand35°C,usingrawfruits,blanchedfruitsandCTR,monitoringtheethanolproduction.ThehighestethanolproductionwasreachedwithCTR(10,22%9andamongfruitswithapple(8,71%).Distillationsatlowtemperaturesandundervacuum,toexploitwarmwaterfromcogenerationplant,weretested;distillationat80°Cand200mbaror400mbarallowedtorecover93,35and89,59%ofethanolrespectively.Theseresultssupportafermentationprocesscoupledtoacogenerationplant,fedwithapplewastesandwithCTRwhenapplewastesarenotavailable,wherehotwaterfromcogenerationplantisusedinblanchinganddistillationphases.Thescaleupinapilotplantwasalsocarriedout.

Resveratrol(RSV)isanaturallyoccurringpolyphenoliccompoundendowedwithinterestingbiologicalproperties/functionsamongstwhichareitsactivityasanantioxidantandasSirtuinactivatingcompoundtowardsSIRT1inmammals.SirtuinscompriseafamilyofNAD⁺-dependentproteindeacetylasesthatareinvolvedinmanyphysiologicalandpathologicalprocessesincludingagingandage-relateddiseases.TheseenzymesareconservedacrossspeciesandSIRT1istheclosestmammalianorthologueofSir2ofSaccharomycescerevisiae.Inthefieldofagingresearches,itiswellknownthatSir2isapositiveregulatorofreplicativeLifespanand,inthiscontext,theRSVeffectshavebeenalreadyexamined.Here,weanalyzedRSVeffectsduringchronologicalaging,inwhichSir2actsasanegativeregulatorofchronologicallifespan(CLS).Chronologicalagingreferstoquiescentcellsinstationaryphase;thesecellsdisplayasurvival-basedmetabolismcharacterizedbyanincreaseinoxidativestress.WefoundthatRSVsupplementationattheonsetofchronologicalaging,namelyatthediauxicshift,increasesoxidativestressandsignificantlyreducesCLS.CLSreductionisdependentonSir2presencebothinexpiredmediumandinextremeCalorieRestriction.Inaddition,alldatapointtoanenhancementofSir2activity,inparticularSir2-mediateddeacetylationofthekeygluconeogenicenzymephosphoenolpyruvatecarboxykinase(Pck1).ThisleadstoareductionintheamountoftheacetylatedactiveformofPck1,whoseenzymaticactivityisessentialforgluconeogenesisandCLSextension.

Productionofbiofuelsfromfarmanimalwasterepresentsapromisingapproachtodiversifyinggreenenergyproductionandreducingcompetitionforcultivablelandsbetweenfuelandfood-orientedcrops.Thisworkwasaimedtodefinethetechnicalfeasibilityandthespecificsuitabilityofcattle,swineandpoultrymanuretointegratingbioethanolandbiomethaneproduction,usingthebiorefineryconcept.Saccharificationobtainedbydiluteacidpretreatment(3.5%H₂SO₄,121°C,30min)followedbyenzymatichydrolysisresultedintotalsugarrecoveryof230.16,160.40,and98.40mgg^-1(ofdrymatter)forcattle,pig,andpoultrymanurerespectively.Thesugarwasthenfermentedusingfreeyeastco-cultures.Theethanolobtainedwas56.32mgg^-1ofdrymatterforcattle(about52.59%ofthetheoreticalethanolyield);27.98mgg^-1forswine(about88.66%ofthetheoreticalethanolyield);12.69mgg^-1forpoultry(about31.32%ofthetheoreticalethanolyield).Methaneproductionfromdistillationwastewas72.95mgg−1fromdryrawfaecesforcattle,126.48mgg^-1forswineand119.03mgg^-1forpoultry.Cattlemanureshowedthebestenergybalanceintermsofethanolproductionwithabout824.16kJkg^-1ofdryfaeces,butthetwointegratedprocessesgeneratedanetenergybalanceof1.28MJkg^-1forcattle,4.57MJkg^-1forswineand4.79MJkg^-1forpoultry.