结合代谢的抗疟药溶血毒性体外检测法

以苯巴比妥i.p.60mg/kg×3d及单剂苯并黄酮i.p.80mg/kg诱导雄性Sprague-Dawley大鼠,摘肝制微粒体,加氧化型辅酶Ⅱ(NADP)等辅助因子,组成体外代谢系统。选取6-磷酸葡萄糖脱氢酶(G6PD)偏低的兔血,制成1％红细胞悬液。不同剂量的伯喹、氯喹或三氟乙酰伯喹(M8506)分别于体外和肝微粒体代谢系统在37℃共同温育后,再取其上清液与1％红细胞悬液共同温育,测定温育后各管上清液的OD值。结果表明,在3×10~1～3×10~3μmol/L的浓度范围内,伯喹和M8506均可测出明显的溶血毒性,并呈一定的剂量一反应关系;而氯喹则无溶血毒性。提示本方法可用作抗疟药溶血毒性的快速初筛试验。

IN VITRO ASSAY FOR DETECTING HEMOLYTIC TOXICITY OF ANTIMALARIALS INCORP RATED WITH MICROSOMAL METABOLIC SYSTEM

The in vitro metabolic system comprised NADP co-factors and liver microsomes isolated from male rats pre-treated with phenobarbital, 60 mg/kg, i.p. for 3d combined with a single i.p. dose of 80 mg/kg of naphtholflavone. The 1% RBC suspension was made up from G6PD-deficient rabbit blood. Primaquine, chloroquine and M 8506 at various dosages were incubated at 37 degrees C with the microsomal metabolic system in vitro respectively. The supernatants were incubated with 1% RBC suspension. The OD635nm values of the supernatants were detected after incubation and centrifugation. The results showed that both primaquine and M 8506 exhibited potent hemolytic toxicity at the dose-range of 1.5-3 x (10(1)-10(3) mumol/L, with certain dose-effect relationship, while chloroquine exhibited no hemolytic toxicity. It is suggested that the in vitro assay incorporated with microsomal metabolic system might be a useful preliminary screening method for testing hemolytic toxicity of various antimalarials.