黄嘌呤氧化酶抑制剂筛选体系的建立

目的建立黄嘌呤氧化酶(xanthine oxidase,XOD)抑制剂筛选体系。方法应用理化法,测定体外XOD活性、血尿酸、血清XOD活性、组织XOD活性及肝肾功能有关血液指标;病理分析肝脏肾脏的损伤情况。结果选择牛奶来源XOD3U·L^-1,黄嘌呤(Xanthine,XA)50μmol·L^-1,37℃,pH7.4,20min为XOD抑制剂体外高通量筛选的最适条件。单次灌胃次黄嘌呤联合皮下注射氧嗪酸钾诱导ICR小鼠,血尿酸水平一过性升高;以血尿酸变化曲线和血尿酸-时间曲线下面积评价急性高尿酸血症小鼠模型。ICR小鼠连续皮下注射氧嗪酸钾血尿酸平稳升高,成模率约70%;以血尿酸水平评价慢性高尿酸血症小鼠模型。上述2种模型动物的血清和组织XOD活性均未见明显变化,肝脏肾脏均未见明显损伤。结论XOD抑制剂体外筛选方法与急性高尿酸血症小鼠模型、慢性高尿酸血症小鼠模型等体内实验方法相互验证,形成基于分子靶点XOD的抗高尿酸血症药物研发的实验体系。

Establishment of a screening system for xanthine oxidase (XOD) inhibitors

Aim To establish a screening system for xanthine oxidase (XOD) inhibitors.Methods The XOD activity in vitro,serum uric acid level,serum XOD activity,tissue XOD activity and the blood indexes related to liver and kidney function were determined by biochemical method,respectively.Pathological analysis was used to observe liver and kidney.Results The optimized reaction consists of 3 U·L^-1 XOD and 50 μmol·L^-1 XA under pH7.4,37 ℃ for 20 min.The high throughput screening method was established for XOD inhibitors in vitro.The acute hyperuricemia models were induced by signle dose of hypoxanthine and oteracil potassium in ICR mice.In this case,the serum uric acid level had a transient elevation after inducer administered.The changes of serum uric acid levels and the area under the uric acid-time curve were used to evaluate the acute hyperuricemia mice.The chronic hyperuricemia models were selected from the ICR mice stimulated with oteracil potassium consecutively.No significant changes of XOD activities in serum and tissue,of function and pathological structure in liver and kidney were observed in both acute hyperuricemia mice and chronic hyperuricemia mice.Conclusions The high throughput screening method for XOD inhibitors in vitro and the evaluation in vivo,in acute or chronic hyperuricemia mice,respectively,are mutually verified,forming an experimental system for developing the anti-hyperuricemia drug targeting at XOD.