Vervoort L M, Ronden J E, Thijssen H H
Department of Pharmacology, Cardiovascular Research Institute, University of Maastricht, The Netherlands.
Biochem Pharmacol. 1997 Oct 15;54(8):871-6. doi: 10.1016/s0006-2952(97)00254-2.
In the vitamin K cycle, vitamin K-hydroquinone, the active cofactor for gamma-glutamylcarboxylase, is continuously regenerated. The successive pathways contain oxidation of the hydroquinone to the epoxide, followed by reduction to the quinone and reduction to the hydroquinone. Vitamin K-hydroquinone is a potent radical scavenging species (Mukai et al., J Biol Chem 267: 22277-22281, 1992). We tested the potential antioxidant activity of the vitamin K cycle in lipid peroxidation reactions (thiobarbituric acid reactive substances, TBARS) in rat liver microsomes. As prooxidant we used Fe2+/ascorbate, NADPH-Fe3+/ATP, and NADPH/CCl4. Vitamin K (< or = 50 microM) on its own did not influence the formation of TBARS. In combination with 1 mM dithiothreitol (DTT), the reductive cofactor for the microsomal enzyme vitamin K epoxide reductase, vitamin K suppressed lipid peroxidation with a concentration that blocked the maximal response by 50% (IC50) of ca. 0.2 microM. Vitamin K1 (phylloquinone) and vitamin K2 (menaquinone-4) were equally active. Warfarin (5 microM) and chloro-vitamin K (50 microM), inhibitors of vitamin K epoxide reductase and gamma-glutamylcarboxylase, respectively, were able to completely abolish the antioxidant effect. Lipid peroxidation was inversely related to the amount of vitamin K hydroquinone in the reaction. Vitamin K epoxide reductase seemed sensitive to lipid peroxidation, with half of the activity being lost within 10 min during oxidation with NADPH/CCl4. The inactivation could be attenuated by antioxidants such as vitamin E, reduced glutathione, and menadione and also by a K vitamin in combination with DTT, but not by superoxide dismutase and catalase. The results show that the vitamin K cycle could act as a potent antioxidant, that the active species in all probability is vitamin K-hydroquinone, and that the primary reaction product is the semiquinone. The results also show that the reaction product is processed in the vitamin K cycle to regenerate vitamin K-hydroquinone.
在维生素K循环中，γ-谷氨酰羧化酶的活性辅因子维生素K氢醌会持续再生。后续途径包括氢醌氧化为环氧化物，接着还原为醌，再还原为氢醌。维生素K氢醌是一种有效的自由基清除剂（Mukai等人，《生物化学杂志》267: 22277 - 22281, 1992）。我们在大鼠肝微粒体的脂质过氧化反应（硫代巴比妥酸反应性物质，TBARS）中测试了维生素K循环的潜在抗氧化活性。作为促氧化剂，我们使用了Fe2+/抗坏血酸、NADPH - Fe3+/ATP和NADPH/CCl4。单独的维生素K（≤50微摩尔）不会影响TBARS的形成。与1毫摩尔二硫苏糖醇（DTT）（微粒体酶维生素K环氧化物还原酶的还原辅因子）联合使用时，维生素K抑制脂质过氧化，其浓度能使最大反应被阻断50%（IC50），约为0.2微摩尔。维生素K1（叶绿醌）和维生素K2（甲萘醌 - 4）具有同等活性。华法林（5微摩尔）和氯维生素K（50微摩尔）分别是维生素K环氧化物还原酶和γ-谷氨酰羧化酶的抑制剂，它们能够完全消除抗氧化作用。脂质过氧化与反应中维生素K氢醌的量呈负相关。维生素K环氧化物还原酶似乎对脂质过氧化敏感，在用NADPH/CCl4氧化过程中，10分钟内其活性会丧失一半。抗氧化剂如维生素E、还原型谷胱甘肽和甲萘醌以及维生素K与DTT联合使用可减弱这种失活，但超氧化物歧化酶和过氧化氢酶则不能。结果表明，维生素K循环可作为一种有效的抗氧化剂，其活性物质很可能是维生素K氢醌，主要反应产物是半醌。结果还表明，反应产物在维生素K循环中被处理以再生维生素K氢醌。
