【作者】 张丽娟；

【导师】 陈清西；

【作者基本信息】 厦门大学 ， 生物化学与分子生物学， 2006， 硕士

【摘要】 酪氨酸酶(EC.1.14.18.1)是一种含铜的氧化还原酶,它具有单酚酶活性和二酚酶活性,是生物体合成黑色素的限速酶[1-3],广泛存在于自然界中,其抑制剂具有多方面的应用包括皮肤美白、果蔬保鲜、防治虫害等方面。本论文围绕两大部分进行,第一部分是探讨蘑菇酪氨酸酶的抑制剂及其对单酚酶和二酚酶催化反应的抑制机理。研究了苯甲酸的不同取代物对蘑菇酪氨酸酶的单酚酶和二酚酶效应。研究它们的抑制作用机理,并测定其抑制常数。第二部分是对苯甲酸类衍生物的抑菌作用进行研究。(1)苯甲酸的单羟基取代物对蘑菇酪氨酸酶的单酚酶和二酚酶的抑制作用:研究结果表明,2-羟基苯甲酸、3-羟基苯甲酸和4-羟基苯甲酸对蘑菇酪氨酸酶的单酚酶和二酚酶活性均有抑制作用。其抑制活性由强到弱的顺序是:4-羟基苯甲酸、3-羟基苯甲酸、2-羟基苯甲酸。它们均是酪氨酸酶的可逆竞争型抑制剂。(2)苯甲酸的二羟基取代物对蘑菇酪氨酸酶的单酚酶和二酚酶的抑制作用。2,3-二羟基苯甲酸、2,4-二羟基苯甲酸和2,5-二羟基苯甲酸对酪氨酸酶二酚酶活力均没有影响,而2,4-二羟基苯甲酸和2,5-二羟基苯甲酸对酪氨酸酶单酚酶活力却有着明显的抑制活性。2,3-二羟基苯甲酸对单酚酶仍无抑制活性。3,4-二羟基苯甲酸和3,5-二羟基苯甲酸仅对二酚酶有一定的抑制作用,但是抑制活性很低,当浓度达到5 mmol/L时,其抑制率均未达到50%。(3)苯甲酸的甲基取代物对蘑菇酪氨酸酶的单酚酶和二酚酶的抑制作用。2-甲基苯甲酸、3-甲基苯甲酸和4-甲基苯甲酸对蘑菇酪氨酸酶的单酚酶和二酚酶均具有明显的抑制作用。4-甲基苯甲酸的抑制活性最强,3-甲基苯甲酸次之,2-甲基苯甲酸最差。它们对二酚酶的抑制类型略有不同,4-甲基苯甲酸是反竞争抑制剂,而3-甲基苯甲酸和2-甲基苯甲酸是混合型抑制剂。(4) 4-卤代苯甲酸对蘑菇酪氨酸酶的单酚酶和二酚酶的抑制作用。4-氟苯甲酸、4-氯苯甲酸和4-溴苯甲酸对酪氨酸酶的单酚酶和二酚酶均具有明显的抑制作用。4-溴苯甲酸的抑制活性均最强,4-氯苯甲酸次之,4-氟苯甲酸最弱。并且,三个化合物均是酪氨酸酶二酚酶的可逆非竞争型抑制剂。(5) 4-甲氧基苯甲酸和4-甲氧基水杨酸对蘑菇酪氨酸酶的单酚酶和二酚酶的抑制作用。4-甲氧基苯甲酸和4-甲氧基水杨酸均可以抑制酪氨酸酶的单酚酶和二酚酶活性。4-甲氧基苯甲酸比4-甲氧基水杨酸的抑制活性高。虽然,两者的取代基更多还原

【Abstract】 Tyrosinase (EC 1.14.18.1) is a kind of redoxidase containing copper, which is of the activity of monophenolase and diphenolase, widely distributed in nature. It is a key enzyme during the melanin biosynthesis. Its inhibitors can be used widely in many fields including whitening agents, keeping fruits and vegetables fresh, insecticides.In this paper, it mainly is of two parts. One part is study the inhibitory effect of the tyrosinase inhibitors. It studied the inhibitory mechanism of analogs of benzoic acid on the activity of monophenolase and diphenolase, and their kinetic constants were determined. The effects of o-hydroxy benzoic acid, m-hydroxy benzoic acid and p-hydroxy benzoic acid on mushroom tyrosinase were studied. o-Hydroxy benzoic acid, m-hydroxy benzoic acid and p-hydroxy benzoic acid can inhibit the monophenolase activity and diphenolase activity.The inhibitory capacity of p-hydroxy benzoic acid was the best. They were all competitive inhibitors. The effects of five dihydroxy benzoic acids on mushroom tyrosinase were studied. 2,3-Dihydroxy benzoic acid, 2,4-dihydroxy benzoic acid and 2,5-dihydroxy benzoic acid can’t inhibit the diphenolase activity. But, 2,4-Dihydroxy benzoic acid and 2,5-dihydroxy benzoic acid can inhibit apparently the monophenolase activity, and 2,3-dihydroxy benzoic acid can’t inhibit the monophenolase activity. When the concentrations of 3,4-dihydroxy benzoic acid and 3,5-dihydroxy benzoic acid were less than 5 mmol/L, the inhibitory rates were less than 50%. 3,4-Dihydroxy benzoic acid and 3,5-dihydroxy benzoic acid had a little inhibitory effect on the diphenolase activity.It studied the inhibitory effects of toluic acids on the activity of monophenolase and diphenolase. They all can inhibit the activity of monophenolase and diphenolase. p-Toluic acid was of most strong inhibitory effect of the three compounds. p-Toluic acid was a uncompetitive inhibitor, while o-toluic acids and m-toluic acids were mixed-type inhibitors.The inhibitory effects of 4-halobenzoic acids on mushroom tyrosinase were elucidated, too. 4-Fluorobenzoic acid, 4-chlorobenzoic acid and 4-bromobenzoic acid all can inhibit the activity of monophenolase and diphenolase. The inhibitory effect of 4-bromobenzoic acid was more strong than 4-fluorobenzoic acid and 4-chlorobenzoic acid. Furthermore, they were all non-competitive inhibitors of diphenolase.更多还原