大鼠体内淫羊藿素葡萄糖醛酸代谢物的定量研究
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摘要
目的建立超高效液相色谱-串联质谱法(UHPLC-MS/MS)同时测定大鼠血浆中淫羊藿素(ICT)及其葡萄糖醛酸代谢物(GICT)浓度,并用于定量研究ICT在大鼠体内的代谢。方法采用乙腈提取血浆中ICT和GICT,以香豆雌酚(CMT)为内标,在C18色谱柱上以乙腈-水-甲酸铵-甲酸梯度洗脱分离;在电喷雾离子化电离源上以选择反应监测方式进行负离子检测,用于定量分析的离子反应分别为m/z 367.1→297.1(ICT),m/z 543.3→367.1(GICT)和m/z 267.0→211.1(CMT)。大鼠腹腔注射10 mg/kg ICT,采集不同时刻血样并测定ICT和GICT浓度。结果 ICT和GICT的线性范围分别为0.2~20 ng/ml和2~200 ng/ml,定量下限分别为0.2 ng/ml和2ng/ml。ICT和GICT的精密度(RSD)在10.3%以内,准确度在95.1%~103.7%,提取回收率为89.1%~92.4%,基质效应为93.2%~104.2%。结论 UHPLC-MS/MS法专属性好、准确度高,适用于ICT在大鼠体内代谢研究,2 h时GICT达到最大浓度。ICT在大鼠体内迅速代谢成葡萄糖醛酸Ⅱ相代谢物GICT。
Objective To establish an ultra-high performance liquid chromatographytandem mass spectrometry( UHPLC-MS / MS) method for the simultaneous quantification of icairitin( ICT) and its major metabolite glucuronidated icaritin( GICT),and investigate metabolism of ICT in rats following a single intraperitoneal administration. Methods ICT,GICT and an internal standard coumestrol( CMT) were extracted from rat plasma using acetonitrile and separated on a C18 column using a gradient mobile phase of acetonitrile,water,ammonium formate and formic acid. In the negative electrospray ionization mode,selected reaction monitoring of the precursor-product ion transitions of m / z 367. 1→297. 1for ICT,543. 3 → 367. 1 for GICT and 267. 0 → 211. 1 for CMT was used for the quantification. Plasma was collected for the determination of ICT and GICT after rats were intraperitoneally administered with ICT at a single dose of 10 mg / kg. Results The calibration curves were linear over the concentration range of 0. 2- 20 ng / ml for ICT and2- 200 ng / ml for GICT with the respective lower limit of quantification at 0. 2 and 2 ng / ml.The intra- and inter-day accuracy values fell in the range of 95. 1- 103. 7%,andprecisions were within 10. 3%. Recovery and matrix effect were 89. 1- 92. 4% and 93. 2-104. 2%, respectively. Conclusion The UHPLC-MS / MS method was specific and accurate,suitable for the metabolism study of ICT. ICT was rapidly metabolized to GICT in rats.
Objective To establish an ultra-high performance liquid chromatographytandem mass spectrometry( UHPLC-MS / MS) method for the simultaneous quantification of icairitin( ICT) and its major metabolite glucuronidated icaritin( GICT),and investigate metabolism of ICT in rats following a single intraperitoneal administration. Methods ICT,GICT and an internal standard coumestrol( CMT) were extracted from rat plasma using acetonitrile and separated on a C18 column using a gradient mobile phase of acetonitrile,water,ammonium formate and formic acid. In the negative electrospray ionization mode,selected reaction monitoring of the precursor-product ion transitions of m / z 367. 1→297. 1for ICT,543. 3 → 367. 1 for GICT and 267. 0 → 211. 1 for CMT was used for the quantification. Plasma was collected for the determination of ICT and GICT after rats were intraperitoneally administered with ICT at a single dose of 10 mg / kg. Results The calibration curves were linear over the concentration range of 0. 2- 20 ng / ml for ICT and2- 200 ng / ml for GICT with the respective lower limit of quantification at 0. 2 and 2 ng / ml.The intra- and inter-day accuracy values fell in the range of 95. 1- 103. 7%,andprecisions were within 10. 3%. Recovery and matrix effect were 89. 1- 92. 4% and 93. 2-104. 2%, respectively. Conclusion The UHPLC-MS / MS method was specific and accurate,suitable for the metabolism study of ICT. ICT was rapidly metabolized to GICT in rats.
引文
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[2]HUANG X,ZHU D,LOU Y.A novel anticancer agent,icaritin,induced cell growth inhibition,G1arrest and mitochondrial transmembrane potential drop in human prostate carcinoma PC-3 cells[J].Eur J Pharmacol,2007,564(1-3):26-36.
[3]LI S,PRICEMAN S J,XIN H,et al.Icaritin inhibits JAK/STAT3 signaling and growth of renal cell carcinoma[J].PLo S One,2013,8(12):e81657.
[4]SUN L,CHEN W,QU L,et al.Icaritin reverses multidrug resistance of Hep G2/ADR human hepatoma cells via downregulation of MDR1 and P-glycoprotein expression[J].Mol Med Rep,2013,8(6):1883-1887.
[5]LAI X,YE Y,SUN C,et al.Icaritin exhibits antiinflammatory effects in the mouse peritoneal macrophages and peritonitis model[J].Int Immunopharmacol,2013,16(1):41-49.
[6]WANG Z,ZHANG X,WANG H,et al.Neuroprotective effects of icaritin against beta amyloid-induced neurotoxicity in primary cultured rat neuronal cells via estrogen-dependent pathway[J].Neuroscience,2007,145(3):911-922.
[7]HUANG J,YUAN L,WANG X,et al.Icaritin and its glycosides enhance osteoblastic,but suppress osteoclastic,differentiation and activity in vitro[J].Life Sci,2007,81(10):832-840.
[8]CHANG Q,WANG GN,LI Y,et al.Oral absorption and excretion of icaritin,an aglycone and also active metabolite of prenylflavonoids from the Chinese medicine Herba Epimedii in rats[J].Phytomedicine,2012,19(11):1024-1028.
[9]刘海培,孟繁华,郭继芬,等.HPLC-MS/MS法分析大鼠体内淫羊藿素葡糖醛酸结合物[J].质谱学报,2010,31(10):376-379.
[10]WONG S P,SHEN P,LEE L,et al.Pharmacokinetics of prenylflavonoids and correlations with the dynamics of estrogen action in sera following ingestion of a standardized Epimedium extract[J].J Pharm Biomed Anal,2009,50(2):216-223.