Kyselova Z, Rackova L, Stefek M
Institute of Experimental Pharmacology, Slovak Academy of Sciences, Dubravska cesta 9, 842 16, Bratislava, Slovakia.
Arch Gerontol Geriatr. 2003 May-Jun;36(3):221-9. doi: 10.1016/s0167-4943(02)00167-x.
On exposure to free radicals generated by the Fenton reaction system of Fe(2+)/EDTA/H(2)O(2)/ascorbate, bovine serum albumin (BSA), used as a model of water-soluble protein, was losing its water solubility depending on the concentration of the chelated iron. The precipitate was found irreversibly insoluble even in concentrated urea. In the soluble fraction, SDS-PAGE analysis proved the presence of dimers and trimers of BSA, accompanied by enhanced bityrosine fluorescence. The pyridoindole antioxidant stobadine inhibited the process of albumin insolubilization in a concentration-dependent manner, the protective effect being more efficient than that of 2-keto-4-methiolbutyric acid (KMBA). Stobadine was, however, less effective than trolox. The inhibitory effect of the antioxidants, expressed as IC(50), correlated well with the reciprocal values of corresponding second order rate constants for scavenging OHz.rad; radicals. The results indicated that the insolubilization of BSA induced by the Fenton system of Fe(2+)/EDTA/H(2)O(2)/ascorbate was caused by OHz.rad; radical mediated cross-linking of the albumin. The model system proved to be suitable for convenient testing of OHz.rad; radical scavenging ability of new antioxidants in a non-lipid environment.
在暴露于由Fe(2+)/EDTA/H(2)O(2)/抗坏血酸的芬顿反应体系产生的自由基时，用作水溶性蛋白质模型的牛血清白蛋白（BSA），其水溶性会根据螯合铁的浓度而丧失。即使在浓尿素中，沉淀也被发现是不可逆的不溶性。在可溶部分，SDS-PAGE分析证明存在BSA的二聚体和三聚体，并伴有双酪氨酸荧光增强。吡啶并吲哚抗氧化剂司他丁以浓度依赖的方式抑制白蛋白不溶性化过程，其保护作用比2-酮-4-甲硫基丁酸（KMBA）更有效。然而，司他丁的效果不如曲洛司坦。抗氧化剂的抑制作用以IC(50)表示，与相应的清除OHz.rad;自由基的二级速率常数的倒数密切相关。结果表明，Fe(2+)/EDTA/H(2)O(2)/抗坏血酸的芬顿体系诱导的BSA不溶性化是由OHz.rad;自由基介导的白蛋白交联引起的。该模型体系被证明适用于在非脂质环境中方便地测试新型抗氧化剂的OHz.rad;自由基清除能力。
