【作者】 拜秀玲；

【导师】 冯霞；

【作者基本信息】 天津大学 ， 物理化学， 2007， 硕士

【摘要】 本论文研究了有机溶剂、温度、pH值对工业上广泛应用的青霉素酰化酶活性的影响,优化反应条件,如反应介质、温度、pH值、底物浓度等因素,研究了青霉素酰化酶在催化合成阿莫西林中的应用。主要进行了如下工作:1.根据LogP规律筛选出12种有机溶剂,考察了它们对青霉素酰化酶活性的影响。在pH值等于7的缓冲溶液和有机溶剂各占50%(V/V)的体系中,通过测定酶在此体系中活性的变化,选出对酶活性影响较小的有机溶剂。发现丙三醇、乙二醇、二甲基亚砜等有机溶剂对酶活性影响较小;研究了温度及pH值对青霉素酰化酶活性及稳定性的影响,发现在20℃~55℃的范围内,随着温度升高,酶的活力增大,但酶的稳定性逐渐降低,综合得出温度为40℃时酶的活性及稳定性较好;pH值在6~8的范围内变化时,酶的活力先升高后下降,pH值等于7.5时,活力达到最大,而酶稳定性却随着pH值增大而下降,综合得出pH值为6.5或7时酶的活性及稳定性较好。2.从对羟基苯甘氨酸邓钾盐出发制备了对羟基苯甘氨酸甲酯(p-OH-PGME),红外和熔点分析表明,制备的产物纯度较高。测定了对羟基苯甘氨酸甲酯(p-OH-PGME)在乙二醇,1,4-丁二醇,环己烷,正丁醇,丙三醇,叔丁醇等几种溶剂中的表观溶解度,发现对羟基苯甘氨酸甲酯(p-OH-PGME)在正丁醇,乙二醇等溶剂中的表观溶解度较好,底物的溶解性好在酶催化合成β-内酰胺抗生素反应中有利于向合成方向进行。3.优化了青霉素酰化酶催化合成阿莫西林的反应条件,分别考察了温度、pH值、底物浓度以及不同反应介质时,6-APA的转化率和对羟基苯甘氨酸甲酯(p-OH-PGME)的水解率,得出较理想的反应条件为:乙二醇-缓冲溶液体系、温度为20℃、溶液pH值为6.5、底物6-APA浓度为75 mmol/L、原料浓度配比:6-APA: p-OH-PGME为1:2。更多还原

【Abstract】 The effect of organic solvent, temperature and pH value on the activity of the industrial widely used penicillin G acylase(PGA) were studied in this thesis. The reaction conditions, including the reaction medium, temperature, pH value and the concentration of substrate, were optimized and used in the synthesis of amoxicillion with PGA as the catalytist. The following works have been done:1. Some organic solvents were selected according to the law of logP for the further investigation about their effect on the activity of PGA. The preferable organic solvent which has slightly effect on the activity of PGA was chosen out by monitoring the change of the activity of PGA. Glycerin, ethylene glycol and dimethyl sulphoxide(DMSO) showed slightly effect on the activity of PGA. The effects of temperature and pH value on the activity and stability of PGA were also investigated. It was found that the activity of PGA increased and the stability of PGA decreased as the temperature increased from 20℃to 55℃. The most suitable temperature was 40℃. The activity of PGA increased firstly and then decreased when the pH changed from 6 to 8. However, the stability of PGA decreased as the pH value increasing. The activity and stability of PGA reached to the best point at pH 6.5 or 7.2. The Methyl p-hydroxyphenylglycinate (p-OH-PGME) was synthesized from the p-hydroxyphenylglycinate dane salt. The IR and melting point analysis were used to characterized the synthesized p-OH-PGME. The apparent solubility of p-OH-PGME in some solvents was measured. The n-butyl alcohol and ethylene glycol are better solvents for p-OH-PGME. The better solubility of substrate is propitious to the synthesis ofβ-lactam antibiotic.3. The reaction conditions in the synthesis of amoxicillion using PGA as catalyst were optimized. The conversion ratio of 6-APA and hydrolytic ratio of p-OH-PGME are investigated under different conditions. As a result, the favorable reaction conditions including the reaction medium, temperature, pH value of solution, concentration of 6-APA , the ratio of 6-APA and p-OH-PGME are ethylene glycol-buffer solution, 20℃, 6.5, 75 mmol/L, 1:2, respectively.更多还原