高效液相色谱法测定马兜铃酸A在细胞培养基中含量变化*

目的]建立细胞培养基中马兜铃酸A高效液相色谱(HPLC)法的测定方法,考察在细胞培养条件下马兜铃酸A在培养基中的稳定性。方法]样品经甲醇沉淀蛋白,10 000 r/min,离心5 min,取上清液进样。采用HPLC法,Waters C18柱(150 nm×4.6 nm,5μm),甲醇-水(含3%的冰醋酸)(70∶30,V/V)为流动相;检测波长260 nm,柱温30℃,流速1 mL/min。结果]马兜铃酸A在0.5~50 mg/L范围内线性良好,相关系数r=0.999 8;日内、日间精密度相对标准偏差(RSD)分别为0.96%、1.92%;回收率为94.01%~104.34%。结论]马兜铃酸A在培养基中3 d内含量稳定,该法简便、灵敏、特异性强,适用于细胞培养基中马兜铃酸A含量的测定。

Determination of the concentration of aristolochic acid A in cell medium with HPLC-UV

Objective] To establish the HPLC-UV method for determination of the concentration of aristolochic acid A.Methods] Samples was precipitated by methanol and centrifugated for 5 min(10 000 r/min).Using HPLC-UV method the samples was determined,and the chromatographic system was consisted of Waters C18 column(150 nm×4.6 nm,5 μm),with 3% acetic acid water solution-methanol(30∶70) at the flow rate of 1.0 mL/min.The detection wavelength was 260 nm and column temperature was 30 ℃.Results] The aristolochic acid A showed a good linear relationship at the range of 0.5~50 mg/L(r=0.999 8).The intra-run and inter-run validation were 1.92%,0.96%.The recovery rate were 94.01%~104.34%.Conclusion] The samples are stabile in 3 days.The method is high specific and sensitive,simple and reliable,so it can be used to study the concentration of aristolochic acid A in culture medium.