Ataturk University, Science Faculty, Department of Chemistry, Erzurum, Turkey.
Int J Vitam Nutr Res. 2009 May;79(3):188-94. doi: 10.1024/0300-9831.79.3.188.
The present paper investigates the in vitro effect of L-ascorbic acid (vitamin C), menadione sodium bisulfate (vitamin K3), and folic acid on purified lactoperoxidase (LPO).
This enzyme was purified from bovine milk by Amberlite CG 50 resin, CM Sephadex C-50 ion-exchange chromatography, and Sephadex G-100 gel filtration chromatography.
Rz (A412/A280) value for the purified LPO was found to be 0.8. Lactoperoxidase was purified 20.45-fold with a yield of 28.8 %. Purity of enzyme was checked by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) method and a single band was observed. All tested vitamins caused inhibition of the enzyme activity and displayed a competitive type of inhibition mechanism. IC(50) values of these three vitamins were 2.03 microM, 0.025 mM, and 0.0925 mM, and the K(i) constants were 0.508+/-0.257 microM, 0.0107+/-0.0044 mM, and 0.0218+/-0.0019 mM respectively.
The vitamins discussed here displayed inhibition-type competition with LPO enzyme at varying concentrations. Our study showed that L-ascorbic acid exhibited a much higher inhibitory effect at lower concentrations, so it was evidently a more potent inhibitor than other vitamins tested.
本研究旨在探讨抗坏血酸（维生素 C）、甲萘醌亚硫酸氢钠（维生素 K3）和叶酸对纯化乳过氧化物酶（LPO）的体外影响。
本实验通过 Amberlite CG 50 树脂、CM Sephadex C-50 离子交换层析和 Sephadex G-100 凝胶过滤层析从牛初乳中纯化 LPO。
发现纯化 LPO 的 Rz（A412/A280）值为 0.8。乳过氧化物酶经 Amberlite CG 50 树脂、CM Sephadex C-50 离子交换层析和 Sephadex G-100 凝胶过滤层析纯化 20.45 倍，收率为 28.8%。酶的纯度通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）方法进行检查，观察到单一带。所有测试的维生素均抑制了酶的活性，并表现出竞争性抑制机制。这三种维生素的 IC50 值分别为 2.03 μM、0.025 mM 和 0.0925 mM，K i 常数分别为 0.508±0.257 μM、0.0107±0.0044 mM 和 0.0218±0.0019 mM。
在所讨论的维生素中，它们在不同浓度下以抑制型竞争与 LPO 酶相互作用。我们的研究表明，抗坏血酸在较低浓度下表现出更高的抑制作用，因此它显然是比其他测试的维生素更有效的抑制剂。
