【作者】 刘婷；

【导师】 葛志强；

【作者基本信息】 天津大学 ， 制药工程， 2018， 硕士

【摘要】 为了探究不同pK_a的药物经口服给药后与消化道不同部位消化酶的化学作用,本文选取两种具有不同pK_a的药物乙酰唑胺(ACZ,pK_a 7.2)和氨苯砜(DDS,pK_a 2.4)作为模型药物,使用多光谱法结合分子对接技术研究在消化道不同生理pH值条件下两种药物对胃蛋白酶和胰蛋白酶的酶学性质及构象的影响。主要内容包括:(1)ACZ在肠道pH 6.8条件下对胰蛋白酶活性的抑制作用大于在胃pH 2.0条件下对胃蛋白酶的抑制。酶动力学结果显示ACZ对胃蛋白酶为非竞争性抑制,对胰蛋白酶为竞争性抑制,K_i值分别为74.8μM/L和19.0μM/L。光谱法分析表明ACZ与胰蛋白酶的结合力大于胃蛋白酶。ACZ与胃蛋白酶的结合作用力为氢键和范德华力,与胰蛋白酶结合作用力主要为静电力。此外,ACZ对胰蛋白酶的蛋白二级、三级构象影响大于胃蛋白酶。(2)DDS在胃pH 2.0条件下对胃蛋白酶活性的抑制大于在肠道pH 6.8条件下对胰蛋白酶的抑制。DDS对胃蛋白酶为竞争性和非竞争性结合的混合型抑制,对胰蛋白酶为非竞争性抑制,K_i值分别为15.4μM/L和64.2μM/L。光谱法分析表明DDS与胃蛋白酶的结合力大于胰蛋白酶。DDS与胃蛋白酶的结合作用力主要为静电力,与胰蛋白酶结合作用力为氢键和范德华力。另外,DDS对胃蛋白酶二级、三级构象的影响大于胰蛋白酶。(3)利用Autodock软件模拟了两种电荷状态下ACZ和DDS与胃蛋白酶和胰蛋白酶相互作用,模拟结果说明药物分子不同的电荷状态将导致药物与蛋白酶结合位点及结合作用力的差异,分子对接模拟结果支持上述实验结果。以上结果表明,不同pK_a的药物在不同生理pH下所形成的不同离子状态对该生理条件下的消化酶会产生不同的化学作用。该结果对临床用药及制剂开发具有重要的指导意义。更多还原

【Abstract】 In order to explore the chemical interaction between digestive enzymes in different part of the digestive tract and drug molecules with different p K_a after oral administration,two drugs acetazolamides(pK_a 7.2)and dapsone(pK_a 2.4)with different p K_a were selected as model drugs.Using multispectral methods combined with molecular docking techniques,explore the effects of two drugs on the enzymatic properties and conformation of pepsin and trypsin at physiological pH of the digestive tract.The main contents include:(1)The effect of acetazolamide on the enzymatic properties and conformation of pepsin and trypsin under physiological pH conditions in the digestive tract(stomach:pH 2.0;intestinal:pH 6.8)was studied.Acetazolamide inhibits trypsin activity more than pepsin.The results of enzyme kinetics showed that acetazolamide was non-competitive to pepsin and competitive to trypsin,with K_i values of 74.8μM/L and 19.0μM/L,respectively.Spectroscopic analysis indicated that acetazolamide binds trypsin more strongly than pepsin.The binding forces of acetazolamide and pepsin are hydrogen bonds and van der Waals forces,and the binding force with trypsin is electrostatic.In addition,acetazolamide has a greater effect on the secondary and tertiary conformation of trypsin than pepsin.(2)The effect of dapsone on the enzymatic properties and conformation of pepsin and trypsin at the physiological pH of digestive tract was studied.The results showed that dapsone had greater inhibition of pepsin activity than trypsin.Dapsone was a mixed inhibition of pepsin for competitive and non-competitive,and non-competitive inhibition for trypsin,with K_i values of 15.4μM/L and 64.2μM/L,respectively.Spectroscopic analysis showed that the binding force of dapsone to pepsin was greater than trypsin.The binding force of dapsone and pepsin is mainly electrostatic force,and the interaction force with trypsin is hydrogen bond and van der Waals force.In addition,the effect of dapsone on the secondary and tertiary conformations of pepsin was greater than that of trypsin.(3)Using Autodock software to simulate the interaction of acetazolamide and dapsone with pepsin and trypsin under two kinds of charge states.The simulation results show that the different charge states of drug molecules will lead differences to the binding site and binding of drug and protease.The simulation results are consistent with the above experimental results.All results indicate that the different ionic states of drugs with different pK_a will have different chemical effects on the digestive enzymes under the physiological pH conditions.This result has important guiding significance for clinical use and formulation development of drugs.更多还原