LC-MS/MS法测定大鼠血浆中5-羟基-7-(4-羟基-3-甲氧基苯基)-1-苯基-3-庚酮浓度

Development of LC-MS/MS Method for Determination of 5-Hydroxy-7-(4-Hydroxy-3-Methoxyphenyl)-1-Phenyl-3-Heptanone in Rat Plasma

目的:建立快速、灵敏测定大鼠血浆中5-羟基-7-(4-羟基-3-甲氧基苯基)-1-苯基-3-庚酮(DPHA)的LCMS/MS方法。方法:血浆样品经正己烷萃取后进行分析,采用Kinetex XB-C18色谱柱(2.10 mm×50 mm,2.6μm),柱温40℃,以水(含0.01%甲酸)-甲醇(含0.01%甲酸)为流动相梯度洗脱,流速0.30 m L/min,ESI离子源,多反应离子监测(MRM),用于定量分析的离子对为m/z 329.2→163.0,内标化合物益智酮甲为313.1→137.0。结果:DPHA的线性范围为1.0～2000.0 ng/mL(r=0.9996),最低定量限为1.0 ng/m L;提取回收率为73.53%～85.77%,基质效应为99.63%～110.50%;日内和日间精密度RSD均低于15%,重复性好。用该法测定静脉注射给药DPHA(1.0 mg/kg)后0.5 h大鼠血药浓度为36.2±5.1 ng/m L(n=4,RSD=14.0%)。结论:本法经方法学验证,适用于大鼠血浆中DPHA浓度的测定,适合药代动力学研究。

Objective:To develop a rapid sensitive LC-MS/MS method for determination of 5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)-1-phenyl-3-heptanone(DPHA)in rat plasma.Methods:The plasma samples were extracted by n-hexane,and subjected to chromatographic analysis running with Kinetex XB-C18 column(2.10 mm×50 mm,2.6μm)at the column temperature of 40℃.The mobile phase consisted of water(contain 0.01%formic acid)and methanol(contain 0.01%formic acid)with gradient elution at a flow rate of 0.3 mL/min.Electrospray ionization(ESI)with a positive-ion and multiple reaction monitoring mode(MRM)was adopted.Yakucinone A was used as the internal standard,the precursor ion→product ion transitions m/z 329.2→163.0 for DPHA and m/z 313.1→137.0 for internal standard were monitored.Results:The plasma standard curve of DPHA was linear in the range of 1.0~2000.0 ng/mL(r=0.9996).The limit of quantification was 1.0 ng/mL.The plasma extraction recovery of DPHA was 73.53%~85.77%.Matrix effects were 99.63%~110.50%.The intra-day and inter-day RSDs were less than 15.0%and the repeatability was good.The concentration of DPHA in rat plasma was 36.2±5.1 ng/mL(n=4,RSD=14.0%)0.5 h after intravenous injection.Conclusion:The established method is proved to be suitable for determination of DPHA in rat plasma,and it can be used as an effective method for the pharmacokinetics studies.