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:2003207230:,,1936,,LC/MS,Tel:(025)3271560,E2mail:shenglsh@sohu.com.//1,2,1,3,1,1(1.,210009;2.,200020;3.,210029)://(LC/DAD/MS)C;,,ABC,KL;N2N2:;;;;;;:O658:A:100028713(2003)0620549205ApplicationsofLiquidChromatography/PhotodiodeArrayDetection/MassSpectrometryforAnalysisofChineseTraditionalMedicinesSHENGLongsheng1,WANGYing2,MARenling1,DINGGang3,LIQian1,ZHANGZunjian1(1.CentreforInstrumentAnalysis,ChinaPharmaceuticalUniversity,Nanjing210009,China;2.AgilentTechnologicalCo.Ltd.,Shanghai200020,China;3.CentreforChineseMedicineModernizationofJiangsuProvince,Nanjing210029,China)Abstract:Abasicmethodbasedonliquidchromatography/photodiodearraydetection/massspectrome2try(LC/DAD/MS)techniqueswasestablishedfortheanalysisofChinesetraditionalmedicines.Sal2vianicA,protocatechuicaldehyde,caffeicacid,lithospermicacid,salvianicacidCandsalvianolicacidCweredetectedinwaterextractfromDanshen.BesidesbilobalideandgingkolideA,BandC,newcompoundswerefoundfromgingkolidesextract.Thestructuresoftwonewcompoundswerecon2firmedandnamedasgingkolideKandL.IntheextractofKushen,matrine,oxymatrine,sopho2carpineorisosophocarpine,N2oxysophocarpineandsophoranolN2oxidewereidentified.Keywords:liquidchromatography;photodiodearraydetection;massspectrometry;phenolicacids;ginkgolides;alkaloides;Chinesetraditionalmedicine,,,2,,,,//(LC/DAD/MS)1,,,,;1%;,C18,//,;,DAD200311November2003ChineseJournalofChromatographyVol.21No.6549553©1994-2006ChinaAcademicJournalElectronicPublishingHouse.Allrightsreserved.(ESI)(APCI),,pH,(1HPLC/UV(a)HPLC/MS(b)Fig.1HPLC/UV(a)andHPLC/MS(b)fingerprintsofDanshenForpeakidentification,seeTable1.)2[1]2.1(,S.mil2tiorrhizaBge.),604h,,802g,100mL,40mL,2h,20mL,20mL,12h,20mL,100mL,,0145m2.2:AlltimaC18(416mmi1d1250mm,5m):A,1%();B,1%:055min,10%90%B;5565min,90%B:110mL/min;,012mL/min(UV):281nm,DADUV200400nm2.3:ESI(-)ESI(+):m/z80800:10L/min:350:320kPa:4kV:70V2.4LCESI,,,LC/ESI(-)/MS,LC/ESI(+)/MSUVMS(TIC)1,LC/MS1,(salvianicA)(pro2tocatechuicaldehyde)(caffeicacid)(lithospermicacid)(salvianicacidC)C(salvianolicacidC)1LC/MSTable1LC/MSdataofmainwatersolubleconstituentsofDanshentR/minm/zAssignment9.7197,219[M-H]-,[M+Na-2H]-salvianicA14.2137[M-H]-protocatechuicaldehyde19.2179,201[M-H]-,[M+Na-2H]-caffeicacid25.227.6537,559[M-H]-,[M+Na-2H]-lithospermicacidoritsisomer28.5717,739[M-H]-,[M+Na-2H]-salvianicacidC31.6493,515[M-H]-,[M+Na-2H]-salvianolicacidC3[2]3.1:Shim2PackCLC2ODS(6mmi1d1150mm,5m):35:A,;B,/(31):020min,05521©1994-2006ChinaAcademicJournalElectronicPublishingHouse.Allrightsreserved.:115mL/min;,013mL/min:UV(2208)nm,DADUV190400nm3.2LC/MS,,(in2sourceCID),250V3.33.3.1UVMSTIC2,2UV/MS,tR1612,1914,34114013minCAB,4UV,220nm,n33.3.2UV,tR4219,56175718min3UV,ABCUV220nm,240nm;3GCGAGB3GCGAGB18,3UV240nm,;,3ABC18,,2UV(a)MSTIC(b)Fig.2ChromatogramwithUVdetection(a)andMSTICchromatogram(b)ofgingkolidesForpeakidentification,seeTable2andTable3.2LC/MS(m/z)Table2LC/MSdata(m/z)ofgingkolidesCompound3tR/min[M-H]-[M+Cl]-[M+NO3]-[M+Na]+[2M+Na]+MrGA34.1407443502431839408GB40.3423459486447871424GC19.4439475470463925440BD16.23253613883496753263GA,GB,GC:gingkolideA,B,C,respectively;BD:bilobalide.3LC/MS(m/z)Table3LC/MSdata(m/z)ofunknowncompoundstR/min[M-H]-[M+Cl]-[M+NO3]-[M+Na]+[2M+Na]+Mr42.8421457484--42256.8389425452--39057.84054414684298354061556://©1994-2006ChinaAcademicJournalElectronicPublishingHouse.Allrightsreserved.(CID),,BtR5718minCID,[M-H-28]-[M-H-56]-,,COC2H4CO3,3.3.3tR5618min5718min(IR)(NMR)(1H2NMR13C2NMR1H21HCOSYHMQCHMBC)10223,1421,10223,142,LK,[3]44.14.1.1(,SophoraFlavescensAit.)(40)110g,150mL,40mL,1h(),,,0145m,50%(),50%,,,,4.1.2()(),A(4.2)210g/L,0145m,4.1.30145m,4.2:ZorbaxExtendC18(211mmi1d150mm,5m):ZorbaxExtendC18(211mm1215mm,5m):25:A,10mmol/L,pH810;B,815mmol/L(91);:030min,40%B;3050min,40%100%B;5min:015mL/minDAD220nm60min,,,,;,:HypersilAPS2(416mmi1d1150mm,5m),ZorbaxExtendC18,211mmi1d150mm,5m,4.3LC/MS4.44115L,LC/DAD/MS10min,,,,3HPLC/UV1,2,3,4,5,3,LC/ESI/MS,,,[4],35N2(sophoranolN2oxide)(oxymatrine)N2(N2oxysophocarpine)(matrine)(sopho2carpine)(isosophocarpine)(4)43Table4IdentificationofmainpeaksinFig.3PeakNo.MrAssignment1280sophoranolN2oxide2264oxymatrine3262N2oxysophocarpine4248matrine5246sophocarpineorisosophocarpine25521©1994-2006ChinaAcademicJournalElectronicPublishingHouse.Allrightsreserved.(a)(b)(c)Fig.3FingerprintsofcrudematerialofSophoraFlavescensAit.(a),intermediate(b)andinjection(c)Forpeakidentification,seeTable4.5,,/,/(2D2GC/MS)[5]/(LC/MSn)[6]/(2D2LC/MS),[7]2D2LC/MSpHC18,ZorbaxExtendC18,,,LC/DAD/MS:[1]LiQian.[MSThesis].Nanjing:ChinaPharmaceuticalUniversi2ty,2003..:,2003[2]WangYing,ShengLongsheng,LouFengchang.ActaPharma2ceuticaSinica,2001,36(8):606,,.,2001,36(8):606[3]WangYing,ShengLongsheng,LouFengchang.JournalofChinaPharmaceuticalUniversity,2001,32(5):371,,.,2001,32(5):371[4]MiaoKangli,ZhangJianzhong,DongYing,XiYufei.NaturalProductResearchandDevelopment,2001,13(2):69,,,.,2001,13(2):69[5]XuGuowang,RuanChunhai,LuXin,KongHongwei.JournalofInstrume