高效液相色谱法测定大鼠组织样品中马兜铃酸A的含量

目的建立用高效液相色谱法（HPLC）测定大鼠脾虚模型组织与血浆样品中马兜铃酸A含量的方法。方法色谱条件为HypersilODS2柱（4．6mm×250mm，5μm），柱温30℃，流动相为甲醇：乙腈：水（0．6％冰醋酸，pH=3．60）（30：30：40），流速为1ml／min，检测波长为250nm，20μl进样。结果空白血浆中马兜铃酸A在0．05～25μg／ml范围内与峰面积线性关系良好（Y=57672X-10065，R^2=0．9994），提取回收率为83．46％～94．13％，方法回收率为99．67％～106．43％。低、中、高3个浓度的日内精密度RSD分别为6．28％、3．36％、1．88％，日间精密度RSD分别为9．67％、4．27％、4．56％。血浆中马兜铃酸A的最低检测浓度为0．05μg／ml，组织中最低检测浓度为0．31ng／g。结论建立的HPLC法准确、稳定、重复性好、灵敏度高，可应用于大鼠脾虚模型组织和血浆样品中马兜铃酸A的含量测定。

Determination of aristolochic acid A in the issure samples of rat by HPLC

Objective To establish a method for the determination of aristolochic acid A in the issure samples of spleen dificiency syndrome＇s rat by high performance liquid chromatography（HPLC）. Methods A Hypersil ODS2 column （4.6 mm × 250 mm, 5 μm）, at the column temperature of 30℃, was used. The mobile phase was Methanol-Acetonitrile-0.6%Eisessig solution （30：30：40; pH=3.6） and the flow rate was 1 ml/min. The UV wave length was sd.at 250 nm and the sample volume used for the analysis was 20 μl. Results In the blood plasma sample, the calibration curve was linear at the range of 0.05 - 25 μg/ml （Y=57672X-10065 ,R^2=0.999 4） and the minimum detectable concentration was 0.05μg/ml （S/N 〉13）. The abstraction recoveries were from 83.46% to 94.13% and the relative recoveries were from 99.67% to 106.43%. The Inter-day RSD and Intra-day RSD of low, middle and high concentrations was 9.67%, 4.27%, 4.56% and 6.28%, 3.36%, 1.88%, respectively. The lowest detectable concentration of aristolochic acid A in blood plasma and tissues was 0.05 μg/ml and 0.31 ng/g, respectively. Conclusion The HPLC method is simple, accurate, stable and reproducible. It can be used for the determination of aristolochic acid A in issues and blood plasma from spleen dificiency syndrome＇s rat.