高效液相色谱法测定大鼠组织中乌头碱的含量

Determination of Aconitine in Tissues of Rats by HPLC

目的:建立高效液相色谱法测定口服乌头碱后大鼠组织中的含量。方法:采用Waters 2690-996PAD色谱系统。色谱条件:用HAlSIL100 C18(5μm,250 mm×4.6 mm),流动相为甲醇∶水∶二乙胺(75∶25∶0.1),流速0.9 mL/min,检测波长240 nm。脏器样品按重量体积比(1∶5)加入1.0 mmoL/L HCl,匀浆离心取上清液,冷冻干燥,有机溶剂提取,减压蒸干,用适量甲醇溶解残渣,30μL进样。结果:大鼠脏器心、肝、脾、肺、肾均检测出乌头碱,脑与脊髓中未检出。心、脾、肺、肾线性范围为8.0 ng^2.0μg,相关系数(r)分别为0.9972,0.9986,0.9993,0.9994;肝脏线性范围为0.04μg^4.0μg,相关系数(r)为0.9990。最低检测限为0.4μg/mL。样品回收率均大于88%,精密度RSD均小于10%。结论:该法是一种准确高效的检测方法,为临床检测乌头碱中毒提供了依据。

Objective：To develop a HPLC method for determination of aconitine in tissues of rats. Methods：The Waters 2690-996 PAD system was adopted,the analytical column was a HAISIL 100 ODS column （5 μm,250 ×4.6 mm）. The mobile phase was methanol,water, and diethylamine in the ratio of 75：25：0.1. The flow rate was 0.9 mL· min^-1. The wavelength of detector was 240 nm. The tissue samples were added to 1.0 mmol/L HCI with weight and volumn in the ratio of 1：5, the above solution was collected after the samples were homogenized and centrifugalized. The solution was lyophilized and extracted by organic solvent. After evaporation of the solvent under reduced pressure, the residue was redissolved into methanol, 30 μL was injected to the sampler. Results： Aconitine was detected in heart, liver, spleen,lung, kidney of rat, none in brain and spinal cord. The linear range of heart, spleen, lung and kidney was 8.0 ng-2.0 μg, the relation coefficients were 0.997 2,0. 998 6,0. 999 3 and 0.999 4 respectively. The linear range of liver was 0.4 μg- 4.0μg and teh relation coefficient was 0.999 0. The detection limit was 0.4 μg/mL, the extracted recovery was 88%, the RSD of precision was less than 10% .Conclusion： It appears to be an accurate and effective method that can offer scientific basis for toxication of aconitine clinically.