高效液相色谱-电喷雾源-质谱联用法测定紫杉醇热敏脂质体及其大鼠体内药动学研究

Quantification of Paclitaxel in Rat Plasma by LC-MS/MS and Pharmacokinetics of Paclitaxel Thermosensitive Liposomes in Rats

目的建立测定大鼠血浆中紫杉醇的高效液相色谱-电喷雾源-质谱联用(LC-MS/MS)方法,并进行大鼠体内药动学研究。方法血浆经NaHCO3溶液(pH 8.0)碱化后,用叔丁基甲醚提取。液相分离采用Zorbax SB C18(2.1 mm×50 mm,3.5μm)分析柱,柱温25℃,以甲醇-水(含0.5 mmol.L-1乙酸铵)(70∶30,V/V)为流动相,流速为0.3 mL.min-1,进样量5μL;采用四级杆质谱检测器,电喷雾源(ESI),正离子方式检测,在选择离子监测(SIM)模式下检测离子对m/z 876.3→308.1(紫杉醇)和m/z 830.2→549.2(多西他赛)。两组SD大鼠分别尾静脉推注7 mg·kg-1紫杉醇热敏脂质体与紫杉醇注射液,采集血样后采用高效液相色谱-电喷雾源-质谱联用法测定血药浓度,计算主要药物动力学参数。结果血浆中无干扰测定的内源性物质,每个样品的分析时间为3.5 min;紫杉醇在2～1 000 ng.mL-1内呈良好的线性关系(r>0.998 5),定量限为2 ng.mL-1,日内、日间精密度RSD均小于15%。紫杉醇热敏脂质体的t1/2β、MRT、AUC和CL与紫杉醇注射液均具有显著性差异。结论本方法操作简便,特异性强,灵敏度高,可用于紫杉醇的药动学研究。与紫杉醇注射液相比,紫杉醇热敏脂质体能控制药物的释放,在一定程度上减轻了急性毒性,提高机体耐受性。

OBJECTIVE To develop and validate a liquid chromatography-tandem mass spectrometry （LC-MS/MS） method for the quantification of paclitaxel in rat plasma and to study the pharmacokinetics of paclitaxel thermosensitive liposomes in rats. METH- ODS The plasma was alkalized with sodium bicarbonate （ pH 8.0） and extracted with tea-butyl methyl ether （TBME）. Paclitaxel and IS（docetaxel） were separated on an Agilent Zorbax SB C18 column （2. 1 mm ×50 mm,3.5 μm） with the mobile phase consisting of methanol-0. 5 mmol·L-1 ammonium acetate （70： 30, V/V） at a flow rate of 300 μL·min-1. The injection volume was 5 μL. An Agilent 6410A QQQ triple-quadrople mass spectrometer equipped with an electrospray ionization（ESI） source was used as detector and was operated in positive ion mode. Selected ion monitoring （SIM） was performed and the m/z of ions selected for quantitation were 876. 3→308. 1 （paclitaxel） and 830. 2→549. 2（ internal standard, docetaxel）. SD rats were injected paclitaxel thermosensitive lipo- somes and paclitaxel injection via tail vein at the dose of 7 mg·kg-1, respectively. Pharmacokinetic parameters were calculated. RE- SULTS The chromatograms showed no endogenous interfering peaks in blank rat plasma. Each analysis was completed within 3.5 min. The linear range of paclitaxel in plasma was 2 - 1 000 ng·mL-1. The lower limit of quantification was 2 ng·mL-1. The intra- run and inter-run relative standard deviations （RSDs） were less than 15%. The differences of t1/2β, MRT, AUC and CL were signifi- cant between the two preparations. CONCLUSION The method is sensitive, accurate, and convenient for the pharmacokinetic study of paclitaxel thermosensitive liposomes. Compared with paclitaxel injection, paclitaxel thermosensitive liposomes has a controlled re- lease rate, which can alleviate the acute toxicity and improve the tolerance to a certain extent.