谷胱甘肽测定方法研究进展

1,1,23,3,1(11;21;31,214036):HPLC,,:;;;:TQ46417,R91411:A:1004-311X(2004)01-0068-03:2003-07-02;:2003-08-05:(:JH02-101):(1978-),,,,;3:Tel:[phone],E-mail:[e-mail]1(L---L--,Glu2tathione)-,307.32,189193(),,5.93,,,,;,1921HopkinsGSH,,GSH[1],,,,,(GSH)(GSSG),,122050,,,,,,,(DSC),1GSHFig.1ChemicalStructureofGSH,[1]:,,,,[2];GSHGSSGGSSGGSH,GSSGGSH,GSHGSSGGSHGSSXProSSG[3];GSHGSHpH,,,,[4];,GSHGSSG,GSSG,GSH1/201/40,[5],,2.1,Ellman(5,5-(2-),DTNB)[2],DTNBpH8.0,4-,,412nm,DTNB400nm,DTNB,412nm,2DTNBGSHFig.2TheReactionSchemebetweenDTNBandGSHGSH,,,,,Sanger(FDNB),,ShinjiOhmori2.2(GR-DTNB)Ellman[3],1969Tietze,Ellman,(3),NADPHEllmanGSSGGSH,,4minTGSH(TotalGlu2tathione),405nm412nm,GR-DTNB,141:6820042BIOTECHNOLOGYVol114,No11:68Feb12004,,NADPH,,GSSGGSH2080,10-14mol/LGR-DTNB,[4]3GSSGFig.3TheReducingReactionofGssginGR-DTNB2.31959,Shore(OPA)1963CohnLyleGSH-OPA,1966VictorOPA,GSH(),(4),350nm,420nm,0.0632.5mol/mL,pH8.0,,15min,30min,,GSH1967ThomasGSH-OPA,,GSH,GSH1970Jocelyn[6]GSH-OPA,N-(NEM)1976,PaulGSSG,1977BeutlerGSSG,,OPAGSSG,GSSG4GSHOPA[5]Fig.4TheReactionSchemefortheFormationoftheFluorescentAdductBetweenGlutathioneandOPA,1--2,4-(CDNB)mBrB(monobromobi2mane)4-()-7--2,1,3-7--2,1,3--4-4--1-(-4-)-S-(EDTN)[6]5-()(PFB-F)[7]EDTN,,,,5min,,,,,2.4(ED),,,Hg,,Allison,Au/Hg,GSH(LCEC)[8],Au/Hg,,GSH,()(PME)(CCEs),,,,,sol/gel,,,AbdollahSalimi[9]Ru[(tpy)(bpy)Cl]PF6,,GSSG,[10]GSSGGSH,,GSHGSSG(DTT)NaBH4,,mBrBDTTNaBH4mBrB[10]mBrB,(TBP),-(2-)(TCEP),[2]GSH,GSSGNEM2-NEM,NEM,,1980Owen[13]2-,1Table1ComparerofsomeofthereportedHPLCmethodsfordetectingGlutathioneMethodLimitofdetectionReferenceHPLC-OPADEC(pH8.0)2pmol[12]CE-ECD50fmol[11]HPLC-Dithiodipyridine(DTT)3pmol[13]HPLC-OPA(pH12)100fmol[14]OPA-HPLC50fmol[15]HPLC-MS-MS200fmol[16]2Table2ExamplesfordetectionofdifferentsubstanceinbiochemicalsamplesMethodSubstanceReferenceOPA-HPLCGSH,Glu-Cys,16kindsofaminoacids[17]PFB-F-TLCGST(S-transferase),GSH[7]NPM-HPLCGSH,GSSG,Cysteine,Homocysteine&otherthiols[18]HPLC-ECDGSH,Glu-Cys,Cys-Gly,cysteine&N-acetylcysteine[19]2.5,,,,,,(HPLC)70,HPLC,HPLC-UVHPLC-OPAHPLC-GRHPLC-DECHPLC-(MS-MS)HPLC,,HPLC,C18,,HPLC,GSHGSSG,HPLC(TLC)[7](CE)[11](HPCE)59620042-(PFB-F)GS-TFB-F,TLC,10pmol[7]HPCE,,GiovanniGSHGSSG,[1]:(1),,fmol,(1);(2)(2);(3),,,,3min[5];(4),,,2.6GSH,,,GSH,GSSG,,,,(DSC)20,,,,,,,,,,,,,,3,2080,,;GSHGSSG,,,:[1]PenninckxM.Ashortreviewontheroleofglutathioneintheresponseofyeasttonutritional,environmental,andoxidativestresses[J].EnzymeMicrobTechnol,2000,26(9-10):737-742.[2]Ellman,G.L.Tissuesulfhydrylgroups[J].ArchBiochemBiophys,1959,82:70-77.[3]FTeitze1Enzymicmethordforquantitativedeterminationofnanogramamountsoftotalandoxidizedglytathione[J].Anal.Biochem,1969,27(3):502.[4]AkiraNakajima,KiyofumiYamada,Li-boZou.Interleukin-6protectspc12cellsfrom4-hydroxynonenal-inducedcytotoxicitybyincreasingintracel2lularglutathionelevels[J].FreeRadicalBiology&Medicine,2002,32(12):1324-1332.[5]PierreLeroy,AlainNicolas1Rapidliquidchromatographicassayofglu2tathioneinculturedcells[J].BiomedicalChromatography,1993(7):86-89.[6]XiaoPingChen,Regi8naldF.Cross,AlanG,etal.AnalysisofReducedGlutathioneUsingaTreactionwith2,4-Dichloro-1-(Naphthyl-4-Ethoxy)-S-Triazine(EDTN)[J].Mikrochim.Acta,1999,130:225-231.[7]MaheshK.Bhalgat,RosariaP.Haugland,ZhenjunDiwu,etal.5-(pentafluorobenzoylamino)fluorescein:Aselectivesubstrateforthedetermina2tionofglutathioneconcentrationandglutathioneS-transferaseactivityseksiriarttamangkul[J].AnalyticalBiochemistry,1999,269:410-417.[8]KristalBS,Vigneau-CallahanKE,MatsonWR.Simultaneousanalysisofthemajorityoflow-molecular-weight,redox-activecompoundsfrommito2chondria[J].Anal.Biochem.1998,263:18-25.[9]AbdollahSalimi,SimaPourbeyram.Renewablesol-/gelcarbonceramicelec2trodesmodifiedwithaRu-complexfortheamperometricdetectionofL-cys2teineandglutathione[J].Talanta,2003,60:205-214.[10]MTAnderson,JRTrudell,DWVoehringer,etal.AnImprovedMonobro2mobimaneAssayforGlutathioneUtilizingTris-(2-carboxyethyl)phosphineasthereductant[J].AnalyticalBiochemistry,1999,272:107-109.[11]WeiWang,HuaXin,HonglianShao,etal.Determinationofglutathioneinsinglehumanhepatocarcinomacellsbycapillaryelectrophoresiswithelectro2chemicaldetection[J].JournalofChromatographyB,2003,789:425-429.[12]LakritzJ,PlopperCG,BuckpittAR.Validatedhigh-performanceliquidchro2matographyelectrochemicalmethodfordeterminationofglutathioneandglutathionedisul-fideinsmalltissuesamples[J].Anal.Biochem.,1997,247:63-68.[13]K.AnderssonA,IsakssonA,Brattstrom,etal.Homocysteineandotherthi2olsdeterminedinplasmabyHPLCandthiol-specificpostcolumnderivatization[J].Clin.Chem.,1993,39:1590-1597.[14]KevinJLenton,HTherriault,JRichardWagner1Analysisofglutathioneandglutathionedisulfideinwholecellsandmitochondriabypostcolumnderivati2zationhigh-performanceliquidchromatographywithortho-Phthalaldehyde[J].AnalyticalBiochemistry,1999,274:125-130.[15]ParoniR,DeVecchiE,CighettiG,etal.HPLCwitho-phthalaldehydeprecolumnderivatizationtomeasuretotal,oxidized,andprotein-boundglu2tathioneinblood,plasma,andtissue[J].ClinChem,1995,41:448-454.[16]MichaelRMoorea,RossLNorris,GeoffreyK.E