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目的优选马兜铃酸Ⅰ-DNA加合物的体外合成条件。方法用锌活化马兜铃酸Ⅰ并分别与脱氧鸟苷(dG)及脱氧腺苷(dA)反应,合成马兜铃酸Ⅰ-DNA加合物,单因素试验及正交试验法优化反应条件,按照优选条件将北马兜铃、青木香药材提取物与dG、dA反应。液相色谱/串联质谱法对合成的马兜铃酸Ⅰ-DNA加合物进行分析。结果缓冲液pH为5.8,3倍量的dA,20倍锌粉,反应时间16 h及3倍量的dG,10倍锌粉,反应时间24 h为最佳条件。马兜铃、青木香药材提取物与dG、dA反应体系中检测到DNA加合物。结论优选方法稳定,可靠,可用于马兜铃酸Ⅰ-DNA加合物的体外合成。
Objective To optimize the in vitro synthesis conditions of aristolochic acid Ⅰ-DNA adduct. Methods Aristolochic acid I was activated by zinc and reacted with deoxyguanosine (dG) and deoxyadenosine (dA) respectively to synthesize aristolochic acid Ⅰ-DNA adduct. The reaction conditions were optimized by single factor test and orthogonal test , According to the preferred conditions Aristolochia, Aoki herbs extract with dG, dA reaction. The synthesized aristolochic acid Ⅰ-DNA adduct was analyzed by liquid chromatography / tandem mass spectrometry. Results The optimum conditions were pH3.3, dA of 20 times, zinc powder of 20 times, reaction time of 16 h, 3 times of dG, 10 times of zinc powder and reaction time of 24 h. Aristolochia, Aconitum officinalis extract and dG, dA reaction system detected DNA adduct. Conclusion The method is stable and reliable and can be used for in vitro synthesis of Ⅰ-DNA adducts of aristolochic acid.