低温生物膜中微生物脱氢酶活性分析韩晓云

　　:2006-03-10:(11511287):(1970-),,,,.:1671-2021(2006)05-0817-04韩晓云1,2,姜安玺1,贲　岳1,姜　明2(1.,150090;2.,150080)　:对低温生物膜的生物活性进行快速、准确的评价.以硫化钠作还原剂,甲苯作提取剂,用氯化三苯基四氮唑(TTC)法测定低温生物膜脱氢酶活性,探讨酶反应过程中温度、pH值、反应时间等因素对生物膜活性的影响.生物膜脱氢酶的反应温度范围为2～50℃,其中最适温度为30℃;适宜的pH值范围是7.5～9.0;反应速率随温度的下降而降低,在最适温度下的适宜时间是3h,而4℃时为8h.活细菌数(ABN)的TTC-还原染色试验证明组成生物膜的耐冷细菌都能够还原TTC而变色,但是不同种的微生物脱氢酶活性也不同.TTC-脱氢酶活性检测的方法可以准确、快速地反映低温生物膜中的生物活性.在低温4℃下,生物膜仍具有一定的生化活性,比常温下的活性污泥催化温度低约10℃.:;TTC-;;:X703　　　:A　　　　　[1].,[2].、[3-5].,,,,,.,,.,,.TTC(2,3,5-),,TTC(TF),[4].TTC-,、.1　1.1　1.0.5t/d,,66.15L.,,,.1.2　,1.200609225　()JournalofShenyangJianzhuUniversity(NaturalScience)　Sep.　2006Vol.22,No.51.3　1.3.1　TTC-[6].1.　2.　3.　4.5.　6.　7.　8.9.10.11.12.1　1　ρ(COD)/(mg·L-1)ρ(BOD5)/(mg·L-1)pHρ(DO)/(mg·L-1)144～37550～3007～8.24～81.3.2　50ml,25ml,(200r/min)15min,(3000r/min)15min,3,50ml,.,,[7].1.3.3　(ABN)TTC-DHA[8].1.3.2,LB,4℃48h,,,3%TTC1%1∶1,.2　2.1　20041120054,,,0～10℃.[9].,,25dCOD70%90%,BOD92%.2.2　,,.,pH8.0,3h,.2,30℃,10℃,19.2%;30℃,,50℃,.,,10℃,10℃.2　2.3　pHpH,3pH8.5,pH7.5～9.0,pH.2.4　4℃30℃.(45):,,TF,,;30℃,,1h,,3h;4℃,818　　()22,,8h.3　pH2.5　(ABN)TTC-(ABN),.,,,.,.6,4℃48h,TTC.6～8.,,,().TTC,,,TTC,,.,2.4　30℃5　4℃　　2,10min,,6　7　30min8　120min,30min88.2%,2h100%..,,,、.3　　TTC-,22:819　　、、.2　TTC-DHAt/min/(cfu·mL-1)/(cfu·mL-1)/%101273930.73012711288.26012711590.5120127127100　　2～50℃,30℃;pH7.5～9.0;,3h,4℃8h.:[1]　NedwellDB.Effectoflowtemperatureonmicrobialgrowth:loweredaffinityforsubstrateslimitsgrowthatlowtemperature[J].FEMSMicrobiologyEcology,1999(30):101-111.[2]　,,,.[J].,2003,35(4):412-415.[3]　,,,.[J].,2006,23(1):1-5.[4]　KlapwiJA,Drent,SteenvoordenJHAM.AmodifiedprocedurefortheTTC-dehydrogenasetestinactivat-edsludge[J].WaterResearch,1974,8(2):121-125.[5]　,,,.[J].,2005,25(2):31-34.[6]　ChrysiSL,BruceER.Modelingthedevelopmentofbiofilmdensityincludingactivebacteria,inertbiomass,andextracellularpolymericsubstances[J].WaterRe-search,2004(38):3349-3361.[7]　,,,.TTC[J].,1996,14(2):3-5.[8]　Praveen-KumarJC.Tarafdar.2,3,5-Triphenylte-trazoliumchloride(TTC)aselectronacceptorofcul-turablesoilbacteria,fungiandactinomycetes[J].BiolFertilSoils,2003(38):186-189.[9]　,,,.[J].,2002,34(4):563-565.TTC-DehydrogenaseActivityinLowTemperatureBiofilmsHANXiaoyun1,2,JIANGAnxi1,BENYue1,JIANGMing2(1.SchoolofMunicipal&EnvironmentalEngineering,HarbinInstituteofTechnology,Harbin150090,China;　2.LaboratoryofMicrobiology,CollegeofLifeSciences,HeilongjiangUniversity,Harbin,150080,China)Abstract:Inordertoinvestigatemicrobialactivityinlowtemperaturebiofilms,Dehydrogenaseactivity(DHA)wasmeasuredwithTTCmethodusingsodiumsulfideasreductantandtolueneasextractant.Theresultsshowedthatthereactiontemperaturewererangedfrom2℃to50℃,theoptimumreactiontemper-aturewas30℃,andsuitablepHrangewasfrom7.5to9.0.Thereactionspeeddecreasedwithtemperaturedrops,theproperreactingtimeswere3hoursat30℃and8hoursat4℃separately.BysprayingTTConculturablemicrobialcolonies,weobservedthatallspeciesofbacteriainbiofilmwerecapableofusingTTCtodevelopredcolours,thecoloursweredevelopeddeeperoncolonieswhichgrewfaster.Theresearchconclud-edthatTTC-dehydrogenasemethodcouldbeusedforfast,accuratemeasurementoflowtemperaturebiofilmactivity.At4℃,thebiofilmswerestillabletocatalysebiochemicalreaction,whichsecuredthehighefficiencyoflowtemperaturewastewatertreatment.KeyWords:low-temperaturebiofilm;TTC-dehydrogenase;psychrotrophs;microbialactivity820　　()22