HPLC-MS-MS法测定人血浆中石杉碱甲的浓度及人体相对生物利用度研究

目的建立人血浆中石杉碱甲浓度的HPLGM9MS测定法，研究2种石杉碱甲片在正常人体的相对生物利用度。方法以盐酸伪麻黄碱为内标，血浆样品经乙酸乙酯萃取，汉邦C18柱（4．6mm×150mm，5μm）分离后，采用HPLC-MS-MS联用法检测，18名健康男性志愿者采用双周期随机交叉试验设计，分别单剂量口服石杉碱甲片（T或R）0．2mg后测定两者的相对生物利用度。结果石杉碱甲与内标分离度好，内源性杂质不干扰测定，在0．0508～5．080μg·L^-1（r=0．9998）石杉碱甲浓度与峰面积比的线性关系良好，定量限为0．0508μg·L^-1，回收率为101．8％～109．8％（n=5），日内RSD为1．8％～3．9％（n=5）；日间RSD为2．3％～8．7％（n=15）。单次服用0．2mg石杉碱甲片后T与R的AUC0～24分别为（16．35±3．42）μg·h·L^-1和（16．38±3．61）μg·h·L^-1；AUC0-∞分别为（17．53±3．80）μg·h·L^-1和（17．70±3．97）μg·h·L^-1；Gmax分别为（2．47±0．49）μg·L^-1和（2．514-_0．51）μg·L^-1；tmax分别为（1．3±0．4）h和（1．2±0．3）h；t1／2分别为（5．9±0．8）h和（6．2±0．7）h。与R相比，T的相对生物利用度为100．5％±10．1％。结论该方法准确度高，灵敏度好，可用于石杉碱甲人体内过程研究。方差分析结果表明2种制剂的主要药动学参数之间无明显差异，双单侧t检验结果表明2种制剂为生物等效制剂。

Determination of huperzine A in human plasma by HPLC-MS-MS and its application to a relative bioavailability of huperzine A tablets in healthy volunteers

Objective To establish an HPLC-MS-MS method for the determination of huperzine A in the plasma and to study the relative bioavailability and the pharmacokinetics of huperzine A tablets. Methods With pseudoephedrine hydrochloride as the internal standard, huperzine A was extracted from the plasma sample with acetic ether and analyzed by a Han Bang C18 column （4.6 mm×150 mm, 5μm）. HPLC-MS-MS was carried out with a mobile phase consisting of methanol-0. 2% methanoic acid （85 ： 15, v/v） at a flow rate of 0. 7 mL ·min^-1. A single oral dose of 0. 2 rag huperzine A （T or R） was administered to 18 healthy volunteers in a randomized crossover study. Huperzine A concentration in the plasma was determined by the newly developed HPLC-MS-MS method, and the pharmacokinetics and bioavailability were studied. Results The calibration curve was linear within 0. 0508-5. 080 μg·L^-1 （r=0. 999 8）. The method recovery of huperzine A was 101.8%-109. 8% （n=5）, and the intraday and inter-day RSD 1.8%-4.8%（n=5）, and 2.3%-8.7% （n=15）, respectively. The AUC0-24 of the two tablets was （16.35±3.42）μg . h . L^-1 and （16.38±3.61）μg. h. L^-1, Cmax was （2.47±0.49） μg. L^-1 and （2.51±0.51） μg. L^-1, and tmax was （1.3±0.4） h and （1.2±0.3） h. The relative bioavailability of T to R was 100.5%±10. 1%. Conclusion The established HPLC-MS-MS method is accurate and sensible. It can be used to study the pharmacokineties of huperzine A. There is no significant difference between the two tablets, and they are bioequivalent.