斑马鱼毒性-代谢关联分析法评价补骨脂香豆素及其糖苷成分安全性

Safety Evaluation of Coumarins of Psoralea corylifolia L. and Their Glycosides by Toxicity-Metabolism Correlation Analysis Using Zebrafish

目的用斑马鱼毒性-代谢关联分析法评价补骨脂香豆素及其糖苷成分安全性。方法将受精后1 d(1 dpf)的健康斑马鱼胚胎置于不同浓度补骨脂素(PS)、异补骨脂素(IPS)、补骨脂苷(PSS)和异补骨脂苷(IPSS)的供试液中,观察记录给药后1~5 d(2~6 dpf)的鱼脏器形态和死亡情况,显微检视3~6 dpf斑马幼鱼形态,用SPSS16.0计算不同化合物6 dpf鱼的半数致死浓度(LC50);此外,各化合物分别选择一个浓度,同法给药,每天取药液,分析各成分的动态代谢变化。结果 PS和IPS以原型成分致3 dpf斑马鱼中毒;PSS和IPSS在鱼3 dpf时以原型成分为主,未见致鱼脏器形态明显变化,但经4~6 dpf斑马鱼分别动态转化成PS和IPS后致鱼中毒。结论 PSS和IPSS对斑马鱼的毒性与体内转化紧密相关。PS和IPS既是药材中的原型成分,也是其糖苷在体内转化的代谢成分,是补骨脂致毒的相关成分。斑马鱼毒性-代谢关联分析法可高效辩识微量成分的在体毒性及其与代谢转化的相关性。

OBJECTIVE To evaluate the safety of coumarins from Psoralea corylifolia L. and their glycosides by toxicity-metabolism correlation analysis using zebrafish. METHODS Healthy zebrafish embryos 1 d post fertilization(1 dpf) were exposed to different concentrations of psoralen(PS), isopsoralen(IPS), psoralenoside(PSS) and isopsoralenoside(IPSS). Death numbers of the embryos or larvals were counted from 1 to 5 d after dosing(2-6 dpf);embryonic micro-morphology of zebrafish(3 dpf or 3-6 dpf) was observed and pictures were taken;LC50 value of 6 dpf zebrafish was calculated by SPSS16.0. In addition, a relatively safe concentration of each sample was exposed to 1-6 dpf zebrafish as above and sampled every day to analyze the dynamic metabolites changes. RESULTS Psoralen and isopsoralen original form caused 3 dpf zebrafish poisoning;while psoralenoside and isopsoralenoside were mainly in original form when zebrafish was 3 dpf, and no obvious morphology changes were observed. But they were transformed by 4-6 dpf zebrafish into psoralen and isopsoralen, respectively, and caused zebrafish poisoning. CONCLUSION The toxicity of psoralenoside and isopsoralenoside to zebrafish is closely related to the transformation in zebrafish. Psoralen and isopsoralen are not only the original components in Psoralea corylifolia L., but also the metabolic components converted in the body. They are components related to toxicity. The zebrafish toxicity-metabolism correlation method can efficiently identify the in vivo toxicity of micro-components and their correlation with metabolic transformation.