HPLC法测定吡嗪酰胺的含量及有关物质

Determination of pyrazinamide and its related substances by HPLC

建立用于吡嗪酰胺含量及其有关物质测定的HPLC法，为吡嗪酰胺原料药制定质量控制标准，并采用LCMS—IT-TOF对检出主要杂质的结构进行推定。色谱柱为TSK—gelODS一100V（250mm×4．6mm，5μm），流动相为甲醇-水（冰醋酸调节pH3．0）（8：92），检测波长为268nm，柱温为35cC，流速为1．0mL/Mmin。对方法进行了系统的验证：主峰能与相邻杂质峰较好地分离；在0．4～250μg／mL范围内，吡嗪酰胺的质量浓度与峰面积呈良好的线性关系（r：1．00）；最低检测限和最低定量限分别为6．5和40．5ng／mL；9批样品含量测定结果与英国药典的非水电位滴定法结果一致。所建方法专属、耐用，操作简便，可用于吡嗪酰胺原料的含量及有关物质测定。已建议中国药典和国际药典收载。

An HPLC method was established for the determination of pyrazinamide and its related sub-stances. The method was carried out on a TSK-gel ODS-gel C18 column（250 mm ×4. 6 mm, 5μm）with a mobile phase consisting of methanol-water （pH 3.0 adjusted with glacial acetic acid） （8 ： 92） at the flow rate of 1.0 mL/min. The detection wavelength was set at 268 nm and the column temperature was kept at 35 ℃. There was fair resolution between pyrazinamide and its related substance. The calibration curves were linear over the range of0. 4 μg/mL and 250 μg/mL （ r = 1.00）. The limits of detection and quantitation were 6.5 ng/mL and 40.5 ng/mL, respectively. The contents data of nine batches of pyrazinamide determined by the developed method were consistent with those measured by non-aqueous potentiometric titration （ BP 2012）. The established HPLC method is specific, accurate, durable and easy to operate, has been proposed to be used in the Chinese Pharmaco- poeia and the International Pharmacopoeia. It could be applied for the determination of pyrazinamide and its related substances.