柱前衍生HPLC法测定硫酸阿米卡星及其制剂中基因毒性杂质肼的研究

摘要：目的 建立柱前衍生HPLC法测定硫酸阿米卡星及其制剂中基因毒性杂质肼的含量。方法 以5%苯甲醛甲醇溶 液为衍生化试剂，采用Agilent Zobax Eclipse XDB-C18色谱柱(2.1 mm×150 mm，5 μm)，流动相A为0.1%甲酸溶液，流动相B 为乙腈，进行线性梯度洗脱，流速1.0 mL/min，DAD紫外检测器，检测波长300 nm，柱温25℃，进样量20 μL。结果 肼在 0.5022~12.5543 μg/mL浓度范围与峰面积的线性关系良好(r=0.9994)，硫酸阿米卡星、注射用硫酸阿米卡星、硫酸阿米卡星注射 液的平均回收率分别为94.8%、94.1%和94.3%。国内企业硫酸阿米卡星、注射用硫酸阿米卡星、硫酸阿米卡星注射液中的肼分 别为未检出~1.8 μg/g、未检出~0.2 μg/g、未检出~3.1 μg/g；日本Nichiiko公司的注射用硫酸阿米卡星和硫酸阿米卡星注射液均未 检出肼。结论

Determination of genotoxic impurity hydrazine in amikacin sulfate and its preparations by HPLC with precolumn derivatization

Abstract Objective To establish a pre-column derivatization HPLC method for the determination of hydrazine in amikacin sulfate and its preparations. Methods 5% benzaldehyde methanol solution was used as the derivatization reagent. Agilent Zobax Eclipse XDB-C18 column (2.1 mm×150 mm, 5 μm) was used. The mobile phase A was 0.1% formic acid solution, the mobile phase B was acetonitrile, and linear gradient elution was carried out according to the attached table. The flow rate was 1.0 mL/min. DAD UV detector was used, and the detection wavelength was 300 nm. The column temperature was 25℃, and the injection volume was 20 μL. Results The linear relationship of hydrazine was found in the concentration range of 0.5022~12.5543 μg/mL(r=0.9994). The recovery values of amikacin sulfate, amikacin sulfate for injection, and amikacin sulfate injection were 94.8%, 94.1%, and 94.3%, respectively. The content of hydrazine in amikacin sulfate, amikacin sulfate for injection, and amikacin sulfate injection in domestic enterprises were undetected~1.8 μg/g, undetected~0.2 μg/g, and undetected~3.1 μg/g, respectively. No hydrazine was detected in amikacin sulfate for injection and amikacin sulfate injection in nichiiko pharmaceutical Co. Ltd. Conclusion The new precolumn derivatization HPLC method was accurate, simple, and specific. It can be used to control the genotoxic impurity hydrazine in amikacin sulfate and its