凝血酶时间法测定家兔血浆中重组水蛭素  

Determination of r-Hirudin Concentration in the Plasma of Rabbits by Thrombin Time Method

作　　者：任洪灿[1] 芮明 韩国柱[1] 吕勇[1] 李颖[1] 肖述[1] 叶红伟[1] 

机构地区：[1]大连医科大学药理教研室,大连116027 [2]哈尔滨博莱制药有限公司,哈尔滨150525

出　　处：《分析化学》2006年第4期547-550,共4页Chinese Journal of Analytical Chemistry

基　　金：国家"九五"科技攻关计划基金资助(No.96-CO2-01-05A)

摘　　要：建立了凝血酶时间（thrombin time,TT）法测定国产重组水蛭素血药浓度的方法,以便为进一步研究该药在家兔体内的药物代谢动力学提供方法学基础.重组水蛭素血浆水平依其抗凝血酶活性（TT延长）予以测定.对该法的影响因素进行了探讨,最适测定条件确定为5 U/mL的凝血酶溶液,0.1 mol/L pH 7.4的Tris-HCL缓冲液及血浆各50 μL;结果发现对凝血酶时间延长百分率的对数与浓度呈线性相关,r为0.98～0.99;线性范围为0.12～0.60 mg/L,定量限为0.12 mg/L.日内和日间精密度（RSD）分别为5.6%～9.1%和9.2%～11.5%;方法回收率97.5%～99.2%;超出线性范围的高浓度血浆样品作2～100倍稀释,其稀释回收率为92.4%～94.8%.血浆样品在-20℃冰冻保存,至少1星期内稳定.应用本方法测定家兔iv重组水蛭素2.0 mg/kg后不同时间血药浓度,表明该药符合二室模型、一级动力学过程.A bioassay called thrombin time（TT） method was developed for the determination of r-hirudin （rH） concentration in plasma of rabbits and thereby for further study of pharmacokinetics of rH. The rH plasma level was measured ex vivo by antithrombin activity with TT prolongation as index. The TT influencing factors were explored. The most suitable conditions for the measurement were established as 50 mL thrombin solution of 5 U/mL, 0.1 mol/L pH 7.4 Tris-HCL buffur and plasma each. It was found that the log （TT） prolongation rate was linearly correlated to the concentration of rH in plasma （ r = 0.98 - 0.99 ） with a linear range of 0. 12 -0. 60 mg/L and a quantitation limit of 0.12 mg/L. The intra- and inter-day precision （RSD） were 5.6% -9.1% and was 9.2% -11.5%, respectively. The method recovery was 97.5% -99.2% and dilution recovery was 92.4% - 94.8% when dilution fold of 2 - 100 was made for high concentration samples. The stability was 1 week when storage temperature of plasma samples was -20℃. Application of the method to the determination of rH in plasma of rabbits receiving iv 2 mg/kg showed that this drug pharmacokinetically behaved as the two-compartment model and the first-order kinetics.

关 键 词：重组水蛭素 生物测定法 凝血酶时间 家兔 药物代谢动力学 

分 类 号：R285[医药卫生—中药学]