高效液相色谱法测定甲钴胺片的有关物质

Content determination of related substances in Mecobalamin tablet by HPLC

目的 建立检测甲钴胺片有关物质的高效液相色谱法。方法 采用HPLC法,色谱柱为Agilent Poroshell EC-C18 (4.6×250 mm, 4μm);以含己烷磺酸钠0.47%的0.02 mol·L^(-1)磷酸二氢钾溶液(用稀磷酸调节pH至3.5)为流动相A,乙腈为流动相B,梯度洗脱;检测波长:266 nm;进样盘:4℃;流速:1.0 mL·min^(-1);柱温:40℃;进样量:40μL;按照加校正因子的自身对照品法计算杂质含量。结果 主成分与有关物质能完全分离,各杂质峰分离良好;各已知杂质不同质量浓度的平均回收率均在95.56%~108.04%之间,RSD均小于2.4%;各批次依甲钴胺片总杂质含量均不大于1.0%。结论 该方法专属性强、灵敏度高、重复性好,可用于甲钴胺片中有关物质的检测。

Objective To establish an HPLC method for the content determination of related substances in Mecobalamin tablet. Methods The chromatographic column was Agilent Poroshell EC-C18(4.6 ×250 mm, 4 μm). The mobile phase A was consisted of 0.02 mol·L^(-1)monopotassium phosphate solution containing 0.47% sodium hexanesulfonate(pH adjusted to 3.5 by dilute phosphoric acid), and the mobile phase B was consisted of acetonitrile with gradient elution. The detection wavelength was 266 nm, the temperature of sample plate was 4 ℃, the flow rate was 1.0 mL·min^(-1), the column temperature was 40 ℃ and the injection volume was 40 μL. The impurity content was calculated by self-reference method with correction factor. Results The main components and related substances were completely separated, and the impurity peaks were separated well from each other. The average recoveries of all known impurities in different concentrations ranged from 95.56% to 108.04%, and RSD were less than 2.4%. The content of total impurities in Mecobalamin tablets in each batch was not more than 1.0%. Conclusion: The method is specific, sensitive and reproducible, which can be used for the content determination of related substances in Mecobalamin tablets.