中成药中氰苷及其差向异构体的成分分析

目的: 建立HPLC 法测定中成药中D-和L-苦杏仁苷的含量,比较D-和L-苦杏仁苷体外酶解差异,为进一步建立中成药中的苦杏仁安全限量标准奠定基础科学数据。方法: 采用Agilent ZORBAX EclipsePlus C₁₈ 色谱柱(4.6 mm×250 mm,5 μm),以乙腈-10 mmol·L^-1 磷酸二氢钠水溶液为流动相进行梯度洗脱,流速为1 mL·min^-1,检测波长为210 nm,柱温为25 ℃。结果: 中成药中的D-和L-苦杏仁苷得到良好分离,在质量浓度2~500 mg·L^-1(r=0.999 9)范围内均呈良好线性关系。D-和L-苦杏仁苷在体外β-葡萄糖苷酶的作用下,酶解速率存在明显差异,提示L-苦杏仁苷可能具有不同的药理毒理作用。对于通宣理肺丸这一氰苷类中成药的代表,不同厂家的通宣理肺丸和不同剂型的通宣理肺方中的D-、L-苦杏仁苷含量范围以及异构化比例分别为0.015%~0.117%、0.004%~0.021% 和5.06%~48.28%,均存在较大差异。结论: 该HPLC 方法灵敏简便,可用于中成药中苦杏仁苷的质量控制,所得含量及异构化比例等数据可为氰苷类中成药中苦杏仁的安全限量标准制定提供参考。

Objective: To establish an HPLC method for determination of D-amygdalin and L-amygdalin in Chinese patent medicine,and compare in vitro enzymatic differences of D-and L-amygdalin,so as to provide fundamental scientific data for the establishment of standard limit of safety for amygdalin in Chinese patent medicine. Methods: The HPLC separation was performed on an Agilent ZORBAX Eclipse Plus C₁₈ column (4.6 mm×250 mm,5 μm)using acetonitrile-10 mmol·L^-1 sodium dihydrogen phosphate solution as the mobile phase by gradient elution. The flow rate was 1 mL·min^-1,the detection wavelength was 210 nm,and the column temperature was 25 ℃. Results: Good separation was obtained for D-and L-amygdalin in Chinese patent medicine, with a good linearity relationship ranged from 2 to 500 mg·L^-1(r=0.999 9). The in vitro β-glucosidase induced hydrolytic rates of D-and L-amygdalin are significantly different. It is suggested that L-amygdalin might have different pharmacological and toxicological effect from D-amygdalin. Next,the Tongxuanlifei pill was chosen as a representative of cyanide glycoside-containing Chinese patent medicine. The D-,L-amygdalin content and epimerization ratio ranges of Tongxuanlifei from different manufacturers and formulations were 0.015%-0.117%, 0.004%-0.021%,and 5.06%-48.28%,respectively,which were quite different. Conclusion: The established HPLC method is simple and sensitive for quality control,the obtained content and epimerization ratio can provide good reference data for drafting the safety limit standard of amygdalin in Chinese patent medicine.