【作者】 韩莹；

【导师】 钟大放； 陈笑艳；

【作者基本信息】 沈阳药科大学 ， 药物分析学， 2003， 硕士

【摘要】 临床前药代动力学研究是新药评价的重要组成部分，进行药代动力学研究首先必须建立灵敏和特异的定量分析方法。本文建立了测定血浆中艾瑞昔布浓度的液相色谱-串联质谱（LC／MS／MS）法，并应用此方法对艾瑞昔布的临床前药代动力学及毒代动力学进行了研究。 一、血浆中艾瑞昔布浓度测定的LC／MS／MS分析方法的建立 建立了快速、灵敏、专属的LC／MS／MS法，测定动物血浆中艾瑞昔布的浓度。以其结构类似物BAP910为内标，100μL血浆样品经液-液萃取后，以乙腈-水-甲酸（80：20：1）为流动相，采用YMC-pack Pro C₁₈柱进行分离，通过液相色谱-串联质谱法以选择离子反应监测（SRM）方式进行检测。用于定量分析的离子反应分别为m/z 370→236（艾瑞昔布）和m/z 374→236（内标BAP910）。艾瑞昔布的线性范围为2.0-5000.0ng·mL^-1，定量下限为2.0ng·mL^-1，每个样品测试时间仅为2.5min。样品预处理的回收率大于89％，日内、日间精密度（RSD）小于9％，准确度（RE）在±3％范围内。 二、LC／MS／MS法研究艾瑞昔布的临床前药代动力学 本文采用上述的LC／MS／MS法，对艾瑞昔布的临床前药代动力学进行了系统的研究。 艾瑞昔布的吸收动力学研究 分别以三个剂量对大鼠灌胃和对犬口服给药，用LC／MS／MS法测定不同时刻的血药浓度。用Topfit非室模型计算药代动力学参数，大鼠给药后3个剂量组C_max、AUC_0-t和AUC_0-∞与给药剂量线性相关（r＞0.8217，P＜0.01）。以一个剂量对大鼠和犬静脉给药。血药浓度数据经Topfit拟合，药物在大鼠和犬体内的动力学过程均符合二室模型。比较口服和静脉给药后的AUC_0-t值，大鼠灌胃和犬口服给药后的平均绝对生物利用 摘 要2度分别为37．2O和23．9O。 艾瑞昔布在大鼠体内的药物动力学性质，表现出一定程度的性别差异。静脉或灌胃给予艾瑞昔布后，在雄性大鼠体内的消除较雌性大鼠快o＜0．05h雌性大鼠药时曲线下面积A UCUC。值明显高于雄性大鼠汐＜o．1卜雌性大鼠的绝对生物利用度为41．8％，雄性大鼠为31．5％。 艾瑞昔布在犬体内的药物动力学性质未表现出性别差异(＞0刀5卜 艾瑞昔布的组织分布研究 大鼠灌胃给予艾瑞昔布后，受试化合物自各组织清除较快，在各组织内不易蓄积，主要在肝脏发生代谢转化，易在脂肪组织分布。受试化合物在脑中含量较低，说明该药不容易通过血脑屏障。另外，试验中还发现，给药后的前3h受试化合物艾瑞昔布在大鼠各组织中的含量未表现出性别差异，但给药3h后，艾瑞昔布在雌性大鼠的某些组织内含量明显高于雄性大鼠，表明艾瑞昔布在大鼠体内吸收过程不存在性别差异，可能在分布及代谢过程存在性别差异。 艾瑞昔布的血浆蛋白结合率研究艾瑞昔布在大鼠血浆中浓度分别为400，1000，4000 ng·mL”‘时，血浆蛋白结合率分别为 94．4土 0．5、94．0土 0．8和 92．4士1．3％。三、LC／MS／MS法研究艾瑞昔布的临床前毒代动力学 以三个剂量对犬连续口服给药，用己建立的LC／MS／MS法测定给药后不同时刻的血药浓度。结果表明，多剂量口服给药时，艾瑞昔布在犬体内并无蓄积。更多还原

【Abstract】 Preclinical pharmacokinetic study is important in evaluating a novel drug, and the basis of the study is a sensitive and selective method with good accuracy and precision. A new LC/MS/MS method for determination of imrecoxib in biological fluids was developed and validated and successfully used in the preclinical pharmacokinetic and toxicokinetic study.1. The development of a LC/MS/MS method for determination of imrecoxib in animal plasmaA highly sensitive and rapid method using reversed-phase liquid chromatography combined with tandem mass spectrometry has been developed for the quantitative analysis of imrecoxib in animal plasma. BAP910, a structurally related analog of imrecoxib, was used as the internal standard. A small volume of plasma (100μL) was processed using liquid-liquid extraction. The extract was chromatographed on a YMC-pack ProC18 column. The mobile phase consisted of acetonitrile-water-formic acid (80: 20: 1, v/v), at a flow rate of 0.6 mL·min-1. A Finnigan TSQ tandem mass spectrometer equipped with atmospheric pressure chemical ionization source was used as detector and was operated in the positive ion mode. Selected reaction monitoring (SRM) using the precursor/product ion combinations of m/z 370- 236 (imrecoxib) and m/z 374→236 (internal standard, BAP910) was used to quantify imrecoxib. The assay was reproducible and linear for imrecoxib in the range of 2.0 - 5000.0 ng·mL-1. The lower limit of quantification was 2.0 ng·mL-1. Each sample was chromatographed only 2.5 min. The precision was within 9% and RE value was within the range of \3%, respectively. The extraction recovery was greater than 89%.2. The preclinical pharmacokinetics of imrecoxib studied by LC/MS/MSThe preclinical pharmacokinetics of imrecoxib was systematically studied by the validated LC/MS/MS method.Investigation of imrecoxib absorptionFollowing an oral administration of imrecoxib to rats and dogs in three doses, respectively, the plasma concentrations of imrecoxib were determined by the validated LC/MS/MS method. The pharmacokinetic parameters were assessed by non-compartmental method using TOPFIT. Cmax, AUCo.t andAUCo.x were all dose proportional (r>0.8217, p<0.01) in rats. The concentration in plasma-time curves after an i.v. administration to rats and dogs were all fitted to two-compartment models. Comparison between AUCo.t after an oral administration and an i.v. administration indicated that the average absolute bioavailabilities of imrecoxib in rats and dogs were 37.2% and 23.9%, respectively.There were gender differences in imrecoxib pharmacokinetics in rats. After an oral administration of imrecoxib, there’s higher elimination rate in male rats (p<0.05). Estimated AUCo-t values in male and female rats showed that AUCo.t value in female was much higher than that in male (p<0.1). The absolute bioavailabilities of imrecoxib in female and male rats were 41.8% and 31.5%, respectively.. There was no gender difference observed in imrecoxib pharmacokinetics in dogs.Investigation of imrecoxib distributionAfter oral administration of imrecoxib to rats, the elimination rates of imrecoxib in all tissues were relatively fast and the drug was unlikely to accumulate. Relatively high concentration presented in liver and fat showed that imrecoxib was easily metabolized in hepar and accumulated in fatty tissues due to its highly lipophilic character. The lowest concentration existed in brain showed a poor penetration into central nervous system. Within 3 h after i.g. administration, the concentrations in the tissues showed no gender difference, while after 3 hpostdose, concentrations of imrecoxib in some tissues of female were significantly higher than those of male. This indicated that gender difference may exist in distribution and metablism phases rather than absorption phase in rats.Investigation of the plasma protein binding of imrecoxibAt rat plasma concentrations of 400 ng·mL-1, 1000 ng·mL-1 and 4000 ng·mL-1, the plasma protein bindings of imrecoxib were measured 94.4±更多还原