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LCMSMS＊王 松1,2,3＊＊ 孙铁珩1,2,4 RichardStuetz3 孙丽娜2 刘家女5(1,110004;2,110044;3,2052;4,110016;5,300071) 以固相萃取为预处理手段,用高效液相色谱-串联四极杆质谱联用技术,针对澳大利亚南威尔士州畜牧业废水中的丙酸睾酮等13种类固醇化合物含量建立了分析方法.采用大气压化学电离源,在正离子模式下,对色谱条件和质谱条件进行优化,其中,丙酸睾酮等7种化合物以质子化的准分子离子峰[M+H]+、另6种化合物以产生了脱去水的离子峰[M+H-H2O]+为母离子进行二级质谱扫描,以最大丰度确定定量离子对.结果表明:该方法所建立的13种化合物的9点标准曲线的线性相关范围为1～1000ng·ml-1,在该范围内,相关系数均0.9990;各化合物的平均回收率在83.75%～111.50%,相对标准差2.02%～14.21%;除美雌醇和雌三醇的灵敏度相对较低,检测限高于15ng·ml-1外,其余物质的检测限均低于5ng·ml-1;实际样品测定时,不同处理流程中各化合物的浓度均能得到较好体现,该方法能满足检测要求. 畜牧业废水 类固醇 液相色谱-串联质谱 大气压化学电离 检测＊(FLOT.333).＊＊.E-mail:ws0506@163.com2009-05-13,2009-10-13. 1001-9332(2009)12-3027-07 X8 AOptimizationofLCMSMSdeterminationmethodforsteroidcompoundsinwastewateranditsapplicationinfeedlotwastewater.WANGSong1,2,3,SUNTie-heng1,2,4,RichardStuetz3,SUNLi-na2,LIUJia-nǜ5(1CollegeofResourcesCivilEngineering,NortheasternUniversity,Shen-yang110004,China;2ShenyangKeyLaboratoryofEnvironmentEngineering,ShenyangUniversity,Shenyang110044,China;3CentreforWaterandWasteTechnology,UniversityofNewSouthWales,Sydney2052,Australia;4InstituteofAppliedEcology,ChineseAcademyofSciences,Shenyang110016,China;5CollegeofEnvironmentalScienceandEngineering,NankaiUniversity,Tianjin300071,China).-Chin.J.Appl.Ecol.,2009,20(12):3027-3033.Abstract:TakingSolid-PhaseExtraction(SPE)aspre-treatmentprocess,alaboratoryexperimentwasconductedbyadoptingthetechnologyofHighPerformanceLiquidChromatography-TandemMassSpectrometry(HPLC-MSMS)todevelopamethodtodeterminetheconcentrationsof13ster-oidsinfeedlotwastewater.AtmosphericPressChemicalionization(APCI)sourcewasappliedandoperatedinnegativeionmodel,withtheparametersofChromatographyandSpectrometrybeingop-timized.Quasi-molecularionpeak[M+H]+appearedinthefullscanofthefirstquadrupoleofmassspectrometer(Q1)ofTestosteronepropionateandother6steroidcompounds,whilequasi-mo-lecularionpeak[M+H-H2O]+appearedinthefullscanofQ1ofrest6steroidcompounds.Thelinearrangeofthe9pointscalibrationcurveforthe13targetcompoundswasdeterminedtobefrom1to1000ng·ml-1,andthecalibrationcurveregressioncorrelationcoefficients(R)werealwaysabove0.9990forallsamplebatches.Theaveragerecoveryrateofalltargetcompoundswas83.75%-111.50%,andthemethodologicalstabilitywasdeterminedtobeacceptable,withtherel-ativestandarddeviationsbetween2.02%-14.21%(n=6).Exceptthatthelimitofdetection 200912 20 12 ChineseJournalofAppliedEcology,Dec.2009,20(12):3027-3033DOI:10.13287/j.1001-9332.2009.0433(LOD)ofMestradiolandOestriolwashigherthan15ng·ml-1,alltheothertargetcompoundshadaLODlowerthan5ng·ml-1.Inthedeterminationofrealsamplesfromfeedlotwastewater,thedevelopedmethodrepresentedagoodprofiletoalltargetcompoundsatdifferentconcentrationsineachstageoftreatment.Keywords:feedlotwastewater;steroids;HPLC-TandemMass;APCI;determination. 、,.,,[1-2],.,.,,[3].[4][5-7],[8-9].,,.,.,、、,.,APCI、Positive13(、、、、、、17β-、、、、、17α-),,.1 1.1 1.1.1 :Agilent1200se-riesLCsystem.(G1379BDegas-ser)、(G1312ABinPump)、(G1329AALS)、(G1330BFC/ALSTherm)、(G1316ATCC)10μl.:PhenomenexLunaC18(2),100A,5μm,150mm×4.6mm.(BiolabC18,5μm,4mm×2mm,100Acolumnguard).:AppliedBiosystems4000QTrapLC/MS/MSsystem.(PeakScientificNitroGenN300DRGasSta-tion).:VarianHS-602rotaryvanepump.:ThermoSavantSPD121PSpeedVacsystem.(ThermoSavantRVT400-230VaporTrap).:Ther-moSavantVLP80ValuPump.:MilliporeDirect-Q5UltrapureWaterSystems.:Thermoelec-tronModulyod230FreezeDryer.:SupelcoVisiprepDLSPE.:WatersOasisHLB6cc(500mg)LPExtractionCar-tridge.1.1.2 :(4-Andro-sten-17β-ol-3-one17-propionate)solid,Sigma;(Estriol)≥97.0%(HPLC)Fluka;(Trenbolo-ne)≥95%,Fluka;(4-Androstene-3,17-di-one)VETRANAL,Fluka;(Etiocholan-3α-ol-17-one)Sigma;(Estrone)≥99%,Sigma;17β-(β-Estradiol)≥98%,Sig-ma;(17α-Ethynylestradiol)≥98%,Sigma;(Androsterone)≥97.0%(TLC),Sigma;(Tes-tosterone)≥99.0%(HPLC),Fluka;(5α-Androstan-17β-ol-3-one)VETRANAL,Fluka;17α-(α-Estradiol)powder,≥98%(TLC),Sigma;(Mestranol)Aldrich.:HPLC(≥99.9%),Sigma-Aldich;HPLC(≥99.7%),AjaxFinechem;(≥99%),AjaxFinechem;(MilliQwater).1.1.3 ,TareeMidcoastWater,4L,3,.1.2 1.2.1 0.1%(A)、0.1%(B).:40%(3028 20AA、B,)A5min;15min,40%A0%,0%A3min;2min,0%A60%,25min.700μl·min-1.20μl;30℃.1.2.2 :(AP-CI);:(Pos);(CUR):20kPa;(CAD):Medium;(IS):5000V;1(GS1):40kPa;2(GS2):0kPa;(Ihe):On;(DP):91V;(EP):10V;(CE):39V;(CXP):10V;(TEM):500℃.1.2.3 10mg13,40ml,20ml500μg·ml-113;,500μl,40ml,18.5mg,10μg·ml-1.,1000、500、100、50、25、10、5、2.5、1ng·ml-1,.1.2.4 ,[10-13],.,.、、,4ml、、1%、pH=3.12(1L),0.25～0.5L·h-1;4ml2,10ml,1.5h,;0.5ml,2,1.5ml,,.1.2.5 .,,,、,.1.3 Analyst,SPSS10.0.2 2.1 ,2(ESI)(AP-CI).APCI、,[14],APCI.,、.,,,[15];,.,,[16],,0.1%,、,,,,0.1%.,40%A5min.、,、0%A2min.,30℃.2.2 1320μl·min-1,(MS1),;,4Da,(MS2),8,2;(DP)、(CE)(CXP),MS2.7,.(FIA),13,(MRM)(CUR)、(CAD)、(IS)、(TEM)、1(GS1)2(GS2),3.302912 :LCMSMS 13100ng·ml-11,2613(2).1,,,,.2.3 ,、,1325min.1 100ng·ml-113Fig.1 SIMofstandardsample13steroidcompoundsat100ng·ml-1concentration.1 13、Tab.1 Retentiontime,precursor/productiontransitions,andmasschromatographyparametersoptimizedof13steroidcompoundsSteroidcompoundRetentiontime(min)Molecularmass(g·mol