【作者】 刘中博；

【导师】 毕开顺； 陈晓辉；

【作者基本信息】 沈阳药科大学 ， 药物分析学， 2011， 博士

【摘要】 盐酸艾咪朵尔(imidol hydrochloride),化学名为1-甲基-4-(1-咪唑基)甲基-2-(苯基亚磺酰甲基)-5-羟基-6-溴-1H-吲哚-3-羧酸乙酯盐酸盐,英文名为1-methyl-4-(1一imidazole)methyl-2-(phenyl-sulfinylmethyl)-5-hydroxy-6-bromo-1H-indole-3-carboxylic acid ethyl ester hydrochloride,是我国自主研制的新型抗乙肝病毒药,属化学药分类1.1。无致畸、致突变作用,长毒和生殖毒性研究亦未见明显毒性,并且合成成本低,具有非常好的开发前景。本文研究结果如下：1.盐酸艾咪朵尔体外稳定性考察建立了测定盐酸艾咪朵尔含量的RP-HPLC方法,考察了温度、溶液pH值和光照对盐酸艾咪朵尔稳定性的影响和其在尿液、胆汁和血浆中的稳定性。引入化学动力学研究方法,得到不同pH条件下(pH 1.5,3.0,5.2,7.4,9.0,11.0)的降解动力学方程分别为：lnCA=-0.02570t+4.279,r=0.9338；ln CA=-0.02790 t+1.156,r=0.9337；ln CA=-0.03050 t+0.07820,r=0.9348；ln CA=-0.04750 t+3.622,r=0.9621；ln CA=-0.2077t+1.828,r=0.9718和ln CA=-0.4143 t+2.004,,r=0.9431；t1/2分别为27.0 h,24.8 h,22.7 h,14.6 h,3.3 h,1.7 h。不同温度下(0℃,20℃,37℃,70℃,100℃)的降解动力学方程分别为:ln CA=-0.02920 t+4.4025,r=0.9380；ln CA=-0.04750 t+3.6223,r =0.9621；ln CA=-0.05540 t+2.558,r=0.9156；ln CA=-03980 t+1.3182,r=0.9016；lnCA=-0.4844 t+2.6208,r=0.9653；t1/2分别为23.7 h,14.6 h,12.5 h,1.7 h,1.4 h。光照下的降解动力学方程为ln CA=-0.01350 t+4.012,r=0.9014,t1/2=5.1h。在生物样品中(胆汁,尿液,血浆)的降解动力学方程分别为：ln CA=-0.04680 t+6.022，r=0.9753；1nCA=-0.02059 t+4.513,r=0.9775；ln CA=-0.03070 t+4.416,r=0.9585；t1/2分别为14.8h,33.6 h,22.6 h。结果证明,盐酸艾咪朵尔的稳定性受温度、溶液pH值、光照和溶液组成的影响。因此在生物样品的采集和保存过程中必须避免盐酸艾咪朵尔发生降解,尽可能避免接触碱性溶液、高温和强光照射。2.盐酸艾咪朵尔在大鼠体内的吸收动力学考察采用UPLC-MS-MS技术,建立了测定大鼠血浆中盐酸艾咪朵尔浓度的分析方法,该方法快速、准确、灵敏度高,定量下限为2.518ng·mL-1。应用已建立的分析方法对盐酸艾咪朵尔在大鼠体内的动力学行为进行考察。以10.02,30.25,70.55,320.3mg·mL-1分别单剂量灌胃给予大鼠盐酸艾咪朵尔后,其在动物体内的Cmax分别为64.79±6.44,188.1±29.4,660.2±174.9,4430±1288ng·mL-1；Tmax分别为0.4±0.1,0.4±0.1,0.4±0.1,0.4±0.1h；AUC0-t分别为80.1±7.5,269.3±53.4,769.4±77.8,6130±2063 ng·h·mL-1；AUCo-∞分别为93.8±11.1,287.0±62.2,827±97,6410±2022ng·h·mL-1；t1/2分别5.4±1.2,3.5±0.6,4.0±1.0,4.1±1.4 h。以10.02 mg·kg-1尾静脉注射给予大鼠盐酸艾咪朵尔后,其在动物体内的Cmax为1896±634ng·mL-1,AUCo-t和AUCo-∞分别为500.8±83.2和534.5±71.76ng·h·mL-1,t1/2为4.1±1.5 h。以上数据经统计学分析后结果表明,在10.02～70.55mg·.kg-1剂量范围内,灌胃给予大鼠盐酸艾咪朵尔后,其在大鼠体内的动力学行为具有线性特征。随着剂量的进一步加大(320.3mg·kg-1),出现较明显的蓄积和饱和现象。由10.02mg·kg-1剂量下灌胃给药组和静脉注射给药组中的AUCo-∞。数据比值计算得到药物的绝对生物利用度为17.55%。3.盐酸艾咪朵尔在大鼠体内外代谢初步考察采用直接进样的方式,利用UPLC-MS-MS技术对盐酸艾咪朵尔的甲醇溶液在不同碰撞能量下产生的离子碎片信息进行全扫描分析,推断了盐酸艾咪朵尔在ESI源实验条件下的质谱裂解规律,为其体内代谢产物结构的推断提供参考。系统的研究了盐酸艾咪朵尔在大鼠体内外的代谢情况。在体内样品中共发现8种代谢产物,其中I相代谢产物3种,II相代谢产物5种；体外代谢共发现3种代谢产物,全部为I相代谢产物。确证出药物主要发生N-去甲基化、S-氧化、O-葡萄糖醛酸化轭合和O-硫酸酯化轭合等反应。4.盐酸艾咪朵尔大鼠在体内排泄考察对70.55 mg·kg-1单剂量灌胃给予大鼠盐酸艾咪朵尔后,原型药物在大鼠体内的排泄情况进行了考察。在给药后的0～72 h,盐酸艾咪朵尔在大鼠粪便和尿液中的累积排泄量为2808±856μg和8.14±2.62 ng,分别相当于给药量的20.06%和0.065%,提示大部分药物已被代谢掉,以代谢产物的形式排出体外；其在胆汁中的排泄平稳而缓慢,给药后0～12 h累计排泄量为18.79±3.78 ng,仅相当于给药量的0.13%。说明其在大鼠体内的肝肠循环现象十分微弱,主要以原形药物的形式由粪便中排出。更多还原

【Abstract】 Imidol hydrochloride was a new excellent anti-Hepatitis B Virus drug with its chemical name of 1-methyl-4-(1-imidazole)methyl-2-(phenyl-sulfinylmethyl)-5-hydroxy-6-bromo-lH-indole-3-carboxylic acid ethyl ester hydrochloride. Imidol hydrochloride is a new anti-viral drug appealed as a class I new drug, which was investigated and developed in our country. With perfect development prospect, imidol has not been seen teratogenesis, mutagenesis, obvious toxicity in long-term toxicity and reproductive toxicity study, and it has a low synthetic cost.1. The stability study of imidolIn this paper, a RP-HPLC method was developed to determine the concentration of imidol hydrochloride. The effect of temperature, pH and light on the stability of imidol and the effect of different biological matrix were assessed. The concept of chemical kinetics and Arrhenius equations were introduced to investigate the degradation kinetics of imidol in aqueous solutions at different pH (pH 1.5,3.0,5.2,7.4,9.0,11.0) with 37℃. The results were In CA=-0.02570t+4.279, r=0.9338; In CA=-0.02790t+1.156, r=0.9337; In CA=-0.030501+0.07820, r=0.9348; In CA=-0.047501+3.622, r=0.9621; In CA=-0.20771 +1.828, r=0.9718 and In CA=-0.4143 t+2.004, r=0.9431; t12 were 27.0,24.8,22.7, 14.6,3.3,1.7 h respectively. The degradation kinetics equations at different temperature(0℃,20℃,37℃,70℃,100℃)were:In CA=-0.02920 t+4.402, r=0.9380, In CA=-0.04750 t+3.622, r=0.9621; In CA=-0.05540 t+2.557, r=0.9156; In CA=-0.3980 t+1.318, r=0.9016; In CA=-0.4844 t+2.621, r=0.9653 and the corresponding values of t12 were 23.7,14.6,12.5,1.7,1.4 h, respectively. The degradation kinetics equation on light was, In CA=-0.01350 t+4.012, r=0.9014;In different biological matrix (urine, bile, plasma) the degradation kinetics equations were, In CA=-0.02059 t+4.513, r = 0.9775; In CA=-0.04680 t+6.022, r=0.9753; In CA=-0.03070 t+4.416, r=0.9585, respectively; and the t12 were 33.6,14.8,22.6 h, respectively.This study revealed that the degradation of imidol is effected by temperature, pH and light. So in the biological sample collection and preservation process of imidol in order to prevent the degradation of imidol, we should avoid too high pH solution, high temperature and contact of glare as far as possible.2. The pharmacokinetic study of imidol in ratsA rapid, accurate and sensitive UPLC-MS-MS method, with an LLOQ (Low Limit of Quantitation) of 2.518 ng·mL-1, was developed and fully validated for the pharmacokinetic study of imidol in rats. The pharmacokinetic processes of imidol in rats after intravenous injection (10.02 mg·kg-1) and oral administration with different doses (10.02,30.25, 70.55 and 320.3 mg·kg-1) were investigated. With the oral doses of 10.02,30.25,70.55 and 320.3 mg·kg-1, Tmax were 0.4±0.1,0.4±0.1,0.4±0.1 and 0.4±0.1 h; Cmax were 64.79±6.44,188.1±29.4,660.2±174.9,4430±1288 ng·mL-1; AUC0-t were 80.1±7.5, 269.3±53.4,769.4±77.8 and 6130±2063 ng·h·mL-1; AUC0-∞were 93.8±11.1, 287.0±62.2,827±97 and 6410±2022ng·h·mL-1; T1/2 were 5.4±1.2,3.4±0.6,4.0±1.0 and 4.1±1.4 h, respectively. After intravenous injection of 10.02 mg·kg-1, the drug reached its maximum concentration of 1896±634 ng·mL-1; AUC0-t and AUC0-∞, were 500.8±83.2 and 534.5±71.76 ng·h·mL-1, respectively; t1/2 was 4.1±1.5 h. The parameters obtained were then compared and analyzed. Dose-dependent linear relationships of AUCo-t and Cmax for imidol was found in the range of 10.02 to 70.55 mg·kg-1 after oral administration to rats. t1/2 from each group showed no significant difference by rank sum test. Bioavailability of this drug was determined as 17.55%.3. The metabolite study of imidol in rats and liver microsomesThe MS fragmentation pathways of imidol by ESI source were studied and summarized with UPLC-MS-MS method. The result Was quite essential and helpful for the further drug metabolism investigation.The UPLC-MS-MS system was used for the identification of imidol metabolites from rat feces, urine and plasma samples. There were altogether 8 metabilites detected and identified in vivo bio transformation products, including 3 phase I ones and 5 phase II ones. There were altogether 3 metabilites detected and identified in vitro biotransformation products. They are all phase I ones. It is concluded that imidol metabolism was mainly accomplished such as N-demethyl,S-oxidation and O-conjugation with Glu or SO3.4. The excretion study of imidol in ratsThe excretion of imidol from rats after an oral administration of 70.55 mg·kg-1 was investigated. The cumulative amount for imidol was 2808±856μgμg in feces within 72 h after administration, which amounted to 20.06% of the given dosage. In urine, the cumulative amount was only 8.14±2.62 ng, indicating a minor urinary excretion of the parent compound. The excretion of imidol from bile was slight and constant, with the cumulative amount of 18.79±3.78 ng. The data above suggested that most of the drug administrated has been metabolized, and enteropatic recycling of imidol is weak in rats.更多还原