目的:建立用于测定硫酸核糖霉素有关物质及含量的高效液相色谱-电喷雾检测器(HPLC-CAD)法,优化补充2020年版《中华人民共和国药典》方法。方法:采用Agilent Zorbax SB-C18(250 mm×4.6 mm,5 μm)色谱柱,以0.15%(v/v)五氟丙酸+0.2%(v/v)三氟乙酸-甲醇(95∶5)作为流动相,流速1.0 mL·mL-1,柱温40 ℃;电喷雾检测器温度50 ℃。结果:所建立方法使核糖霉素及其有关物质实现良好分离,核糖霉素质量浓度在2~200 μg·mL-1范围内,其峰面积与浓度呈良好的线性关系(r=0.998 3),检测限(LOD)为0.8 μg·mL-1,定量限(LOQ)为2 μg·mL-1,重复性和精密度试验RSD均＜2.0%。HPLC-CAD法测定硫酸核糖霉素及有关物质含量为0.81%~1.02%、74.64%~75.22%。结论:相比于2020年版《中华人民共和国药典》硫酸核糖霉素的检测方法,该方法专属性强,灵敏度和稳定性好,为硫酸核糖霉素质量控制提供了可靠的分析手段。
Objective: To establish a high performance liquid chromatography-charged aerosol detector (HPLC-CAD) method for the determination of ribostamycin sulfate and its related substances, and optimize and supplement Chinese Pharmacopoeia (2020 edition). Methods: Agilent Zorbax SB-C18 (250 mm×4.6 mm,5 μm) column was used with 0.15% (v/v) pentafluorpropionic acid+0.2% (v/v) trifluoroacetic acid-methanol (95∶5) as the mobile phase, and the flow rate was 1.0 mL·mL-1. The column temperature was 40 ℃. The temperature of the CAD was set at 50 ℃. Results: The established method achieved good separation of ribostamycin and its related substances. The peak area and concentration of ribostamycin showed a good linear relationship in the mass concentration range of 2 μg·mL-1 to 200 μg·mL-1 (r=0.998 3), the limit of detection (LOD) was 0.8 μg·mL-1 and the limit of quantification (LOQ) was 2 μg·mL-1. The RSD of the repeatability and precision experiments were both less than 2.0%. HPLC-CAD method was used to determine the impurities and contents of ribostamycin sulfate in the range of 0.81%-1.02% and 74.64%-75.22%. Conclusion: Compared with the detection method of ribostamycin sulfate in Chinese Pharmacopoeia(2020 edition), the method has good specificity, sensitivity and stability, and provides a reliable analytical means for the quality control of ribostamycin sulfate.
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