骨刺片中士的宁和马钱子碱含量测定方法的研究

目的：拟建立骨刺片中士的宁和马钱子碱的高效液相色谱（HPLC）含量测定方法，并考察马钱子生物碱对照提取物（简称对照提取物）和马钱子生物碱混合对照溶液（简称混合对照溶液）替代单体对照品，测定骨刺片中士的宁和马钱子碱含量的可行性。方法：采用Symmetry Shield TM RP C₁₈（4.6 mm&#215;250 mm，5 μm）色谱柱，以乙腈-0.01 mol&#183;L^-1庚烷磺酸钠与0.02 mol&#183;L^-1磷酸二氢钾等量混合溶液（21：79）为流动相，流速1 mL&#183;min^-1，检测波长260 nm，柱温30℃。分别采用士的宁、马钱子碱的对照品，对照提取物和混合对照溶液为对照，测定骨刺片中士的宁和马钱子碱的含量。结果：士的宁和马钱子碱进样量分别在0.14~10.00 μg和0.10~10.00 μg范围内线性关系良好；以对照品、对照提取物和混合对照溶液为对照，士的宁的平均回收率（n=6）分别为95.7%、98.8%和97.5%，RSD分别为1.6%、1.2%和1.4%；马钱子碱的平均回收率（n=6）分别为97.7%、99.5%和96.9%，RSD分别为1.3%、0.92%和2.0%。采用建立的HPLC方法测定了10个厂家22批次骨刺片样品，以对照品、对照提取物和混合对照溶液为对照，士的宁含量范围分别为0.09~0.69、0.10~0.68和0.10~0.69 mg&#183;g^-1，马钱子碱含量范围分别为0.05~0.62、0.05~0.61和0.05~0.62 mg&#183;g^-1。结论：本研究建立的HPLC方法，可快速、准确地测定骨刺片中士的宁和马钱子碱的含量。本文采用士的宁和马钱子碱单体对照品、对照提取物和混合对照溶液为对照，测定骨刺片中士的宁和马钱子碱的含量，结果无显著差异（P<0.05），表明对照提取物和混合对照溶液可替代单体对照品测定骨刺片中士的宁和马钱子碱的含量。

Objective:To establish an HPLC assay method for the quality control of strychnine and brucine in Guci tablets and explore the suitability for the reference extracts of Strychni Semen(RE) and the mixture reference solution of Strychni alkaloids strychnine and brucine (MRS) to replace reference substances of strychnine and brucine (RS) for assay. Methods:The HPLC analysis was performed on a Waters Symmetry Shield TM RP C₁₈ column (4.6 mm&#215;250 mm, 5 μm) with a mixture of acetonitrile and a mixture solution of equal quantities of 0.01 mol&#183;L^-1 sodium heptanesulfonate solution and 0.02 mol&#183;L^-1 potassium dihydrogen phosphate solution (21:79) as the mobile phase at a flow rate of 1 mL&#183;min^-1. The spectrophotometer was set at 260 nm and the column temperature was 30℃. The contents of strychnine and brucine in Guci tablets were determined by RS, RE and MRS. Results:The results showed good linearity of strychnine and brucine in the ranges of 0.14-10.00 μg and 0.10-10.00 μg, respectively. Assay by using RS, RE and MRS, the average recoveries (n=6) of strychnine were 95.7%, 98.8% and 97.5% with RSDs of 1.6%, 1.2% and 1.4%. The average recoveries (n=6) of brucine were 97.7%, 99.5% and 96.9% with RSDs of 1.3%, 0.92% and 2.0%. The HPLC method was evaluated by 22 batches of Guci tablets from 10 manufacturers. The contents were determined by RS, RE and MRS. The contents of strychnine were 0.09-0.69, 0.10-0.68 and 0.10-0.69 mg&#183;g^-1 and those of brucine were 0.05-0.62, 0.05-0.61 and 0.05-0.62 mg&#183;g^-1. Conclusion:The proposed HPLC method can be used for the determination of strychnine and brucine in Guci tablets rapidly and accurately. The concentrations of strychnine and brucine in Guci tablets were detected with RS, RE and MRS. No obvious difference was observed between the detection results obtained with difference calibration methods (P<0.05), which suggested that RE or RMS can be used as the alternative for RS in the determination of strychnine and brucine in Guci tablets.