Basedonthestrongchelatingpropertyofbathophenanthrolinedisulfonicacid(BPDS)withFe(Ⅱ),rootFe(Ⅲ)chelatereductaseactivityisusuallymeasuredwithaspectrophotometerusingMES(2-morpholinoethanesulfonicacid)orHEPES(2-(4-(2-Hydroxyethyl)-1-piperazinyl)ethanesulfonicacid)bufferinthedarkbecauseofhighautoreductionrateofFe(Ⅲ)inthepresenceoflight.However,theexclusionoflightisinconvenient,especiallywhenanalyzingalargenumberofsamples.TheobjectiveofthisstudywastodevelopanewmethodfordeterminationofrootreductaseactivityundernormallaboratoryconditionsusingasuitablebuffercompositionandFe(Ⅲ)concentrationtoeliminatetheautoreductionofFe(Ⅲ).AmodifiedmethodusingaTris(2-amino-2-hydroxymethyl-1,3-propanediol)bufferatpH7.5insteadofMESorHEPESbufferandadecreasedFeEDTA(Feethylenediaminetetraaceticacid)concentrationof50μmolL-1wasdeveloped.TheautoreductionofFe(Ⅲ)usingtheTrisbufferwasundetectablefortemperaturesat4and28℃andwasalsomuchlowerthanthatusingtheotherbuffersevenwithsunlightduringmeasurementofFe(Ⅲ)reduction.Furthermore,thedifferencesinFe(Ⅲ)reductaseactivityamong5plantspeciesand14redclovercultivars(TrifoliumpratenseL.)couldbeeasilydetectedwiththemodifiedmethod.ThemethoddevelopedinthisstudytomeasurerootFechelatereductaseactivitywasnotonlyeffectiveandreliablebutalsoeasilymanagedundernormallaboratorylightconditions.
