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DevelopmentofanonlineSPE–LC–MS-basedassayusingendogenoussubstrateforinvestigationofsolubleepoxidehydrolaseinhibitors报告人：谢辉2011年6月29日Background1.TherapeuticpotencyofsEHIsEHinhibitorsareapromisingnewclassofpotentialdrugsfortreatmentofavarietyofdiseases,forexampleinflammation,hypertension,andpain.2.NewassaysareneededAnalyticaltechniquesareneededtoidentifyactivecompoundsandquantitativelymeasuretheirpotencies.3.Developmentofnewassayelectrosprayionizationtandemmassspectrometry(ESI–MS–MS)(电喷雾离子阱质谱)MeritsComparedtofluorescence，thismethodenablesquantitativeevaluationofpotentsEHIswithpicomolarpotencies.(13differentinhibitors/individualcompounds)Sothisassayisapromisingmethod！Procedure1.SelectionofsubstratesNaturalsubstrates:14(15)-EpETrEInternalstandards:10(11)-EpHepBaculovirus-expressedhumansEH2.SelectionofsEHIUreaderivativessynthesizedintheirlaboratoryBaculovirus-expressedhumansEHMethodMS(massspectrometry)—measurementofenzymeactivityandinhibitionLC–ESI（Liquidchromatographyelectrospray）—tomonitorconversionof14(15)-EpETrEto14,15-DiHETrE(ThemethodisanewusefortherapiddeterminationofthepotencyofsEHI.)SPE(solid-phaseextraction)—toensuresensitiveandreliabledeterminationofsmallamountsofproductformedinadifficultmatrix.methodappliedhere—short,narrowcolumnsfilledwithlargeparticles(50–60μm)—Turbulent-flowchromatography(TFC).3.Threesteps1.Preparationofstockandstandardsolutions2.Instrumentalsetup3.sEHinhibitionassayResultsSummaryAnewLC–MS-basednaturalsubstrateapproachforpotencymeasurementofsEHIhasbeendeveloped.ThisnewmethodenablesreliablemeasurementofthepotencyofsEHIwithhighprecisioninlesstime.固相萃取的简要过程1.一个样品包括分离物和干扰物通过吸附剂2.吸附剂选择性的保留分离物和一些干扰物其他干扰物通过吸附剂3.用适当的溶淋洗吸附剂，使先前保留的干扰物选择性的淋洗掉，分离物保留在吸附剂床上4.纯化、浓缩的分离物从吸附剂上淋洗下来