Radi R, Bush K M, Cosgrove T P, Freeman B A
Department of Anesthesiology, University of Alabama, Birmingham 35233.
Arch Biochem Biophys. 1991 Apr;286(1):117-25. doi: 10.1016/0003-9861(91)90016-c.
Xanthine oxidase and purines have recently been detected in the circulation during acute viral infection and following hepatotoxicity and shock. Reactions of xanthine oxidase-generated oxidants with human plasma or bovine serum albumin (BSA) and egg phosphatidylcholine (PC) liposomes have been studied by measuring protein sulfhydryl oxidation and two markers of free radical-mediated lipid peroxidation, thiobarbituric acid reactive substances (TBARS) and conjugated dienes. Plasma incubated with 5 mU/ml xanthine oxidase (XO) and 0.5 mM hypoxanthine (Hx) for 2 h at 37 degrees C had 25-53% oxidation of sulfhydryl groups, with greater than 80% of the oxidation occurring during the first 20 min of the reaction. Concentrations of BSA similar to those present in serum, when exposed to XO/Hx-mediated oxidative stress, showed an even greater decrease in sulfhydryl concentration than that of plasma. No significant increase in plasma TBARS and conjugated dienes was observed during the 2-h incubation period in the presence of XO. Egg PC liposomes, suspended to a plasma phospholipid-equivalent concentration, showed a minor increase in TBARS and conjugated dienes under similar XO/Hx incubation conditions. In the presence of 0.23 mM BSA, lipid peroxidation was completely inhibited. A similar inhibition of lipid peroxidation was induced by cysteine but not by uric acid. Electrophoretic and arsenite-mediated sulfur reduction analysis revealed that BSA was oxidized beyond the disulfide form, with sulfenic acid formed during the initial period of oxidation. Protein sulfhydryls served as sacrificial antioxidants, preventing plasma lipid peroxidation, as well as being targets for oxidative damage. Plasma protein thiol oxidation was determined to be a more sensitive and specific indication of oxidant stress to the vascular compartment than assessment of lipid oxidation byproducts.
最近在急性病毒感染期间以及肝毒性和休克后，已在循环系统中检测到黄嘌呤氧化酶和嘌呤。通过测量蛋白质巯基氧化以及自由基介导的脂质过氧化的两个标志物——硫代巴比妥酸反应性物质（TBARS）和共轭二烯，研究了黄嘌呤氧化酶产生的氧化剂与人类血浆、牛血清白蛋白（BSA）和卵磷脂（PC）脂质体的反应。在37℃下，将血浆与5 mU/ml黄嘌呤氧化酶（XO）和0.5 mM次黄嘌呤（Hx）孵育2小时，巯基基团有25%-53%被氧化，其中超过80%的氧化发生在反应的前20分钟内。与血清中浓度相似的BSA，在暴露于XO/Hx介导的氧化应激时，其巯基浓度的下降甚至比血浆中的更大。在XO存在的情况下，2小时孵育期内血浆TBARS和共轭二烯未观察到显著增加。悬浮至与血浆磷脂等效浓度的鸡蛋PC脂质体，在类似的XO/Hx孵育条件下，TBARS和共轭二烯略有增加。在存在0.23 mM BSA的情况下，脂质过氧化被完全抑制。半胱氨酸可诱导类似的脂质过氧化抑制作用，而尿酸则不能。电泳和亚砷酸盐介导的硫还原分析表明，BSA被氧化至二硫键形式以上，在氧化初期形成了亚磺酸。蛋白质巯基作为牺牲性抗氧化剂，可防止血浆脂质过氧化，同时也是氧化损伤的靶点。与通过评估脂质氧化产物来评估相比，血浆蛋白巯基氧化被确定为血管腔室氧化应激更敏感和特异的指标。
