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目的:建立高效液相色谱(HPLC)分析方法研究珊瑚菜内酯在大鼠肝微粒体生物转化的时程曲线,并对转化产物E-5-甲氧基紫罗绒苔素乙酸酯(M3)和Z-5-甲氧基紫罗绒苔素乙酸酯(M4)进行朔源。方法:应用HPLC分析法测定珊瑚菜内酯的大鼠肝微粒体生物转化中的原形及其转化产物白当归素的含量。色谱柱为DiamonsilTMODS C18(250 mm×4.6 mm,5μm);流动相为甲醇-水,梯度洗脱,流速为1.0 m L·min-1;检测波长为315 nm;柱温为25.0℃。通过转化产物不同萃取方法的比较,朔源M3和M4。结果:绘制了珊瑚菜内酯在大鼠肝微粒体体外的生物转化时程曲线,在60 min内下降较快,在60~240 min内基本稳定。其转化产物白当归素在60 min时达最大值,然后开始下降,到120 min时基本稳定。在大鼠肝微粒体温孵培养物中,珊瑚菜内酯和白当归素分别在600~1 800和0.2~2.4μg·m L-1的浓度范围内线性关系良好(r2>0.999),方法的精密度和稳定性RSD均小于15%;回收率分别为88.6%~92.9%和79.0%~89.5%,RSD均小于15%。结论:本研究建立的分析方法经方法学验证,可对大鼠肝微粒体温孵培养物中珊瑚菜内酯和白当归素含量进行测定,快速、准确、灵敏;珊瑚菜内酯在大鼠肝微粒体体外的生物转化缓慢;M3和M4是“人工”转化产物。
OBJECTIVE: To establish a time-course curve for the bioconversion of coral lactone in rat liver microsomes by high performance liquid chromatography (HPLC). The conversion of E-5-methoxyproslonidine acetate M3) and Z-5-methoxy pperone acetate (M4). Methods: HPLC method was used to determine the protoplast and the content of white angelica in the liver microsomes of Colactone. The column was DiamonsilTMODS C18 (250 mm × 4.6 mm, 5 μm). The mobile phase was methanol-water with a gradient of 1.0 mL · min-1. The detection wavelength was 315 nm. The column temperature was 25.0 ℃. Through the transformation of different extraction methods of comparison, Schomburg M3 and M4. Results: The time course of bioconversion of coral lactone in rat liver microsomes was plotted, which decreased rapidly within 60 min and basically stabilized within 60 min to 240 min. The conversion product of white Angelica reached its maximum at 60 min, then began to decline, to 120 min when the basic stability. In the rat liver microsomal incubation cultures, there was a good linear relationship (r2> 0.999) between the coral vegetable lactone and the white angelica at concentrations of 600-1 800 and 0.2-2.4 μg · mL-1, respectively The RSDs of precision and stability were less than 15%. The recoveries were 88.6% -92.9% and 79.0% -89.5%, respectively. The RSDs were less than 15%. Conclusion: The analytical method established in this study is validated by methodology, which can be used to determine the content of coral lactone and white Angelica in rat liver microsomal incubation culture rapidly, accurately and sensitively. The content of coral lactone in rat liver Microsomes have slow biotransformation in vitro; M3 and M4 are “artificially” transformed products.