目的 建立高效液相色谱-脉冲安培检测法(HPLC-PAD)测定硫酸核糖霉素的含量和有关物质。方法 采用Thermo AcclaimTMAmG C18(4.6 mm×150 mm,3 μm)色谱柱,以0.2%(V/V)五氟丙酸溶液-乙腈(99∶1)为流动相,脉冲安培检测器,四电位检测波形,检测器温度35 ℃,进样体积为25 μL。结果 所建方法使核糖霉素及其有关物质良好分离,核糖霉素的检出限为0.15 μg·mL-1(3.75 ng),定量限为0.375 μg·mL-1(9.38 ng),在0.15～40 μg·mL-1内,核糖霉素的浓度与峰面积呈良好的线性关系(r=0.999 3),含量测定重复性RSD(n=6)为0.33%,总杂质测定重复性RSD(n=6)为1.10%。结论 该方法专属性强,灵敏度高,稳定好,检验成本较现行Ch.P 2015方法显著降低,有望实现方法替代。
Abstract
OBJECTIVE To establish an HPLC method combined with pulsed amperometric detection for the analysis of ribostamycin sulfate and related substance. METHODS The HPLC was performed on Thermo AcclaimTMAmG C18 column (4.6 mm×150 mm,3 μm). The mobile phase consisted of acetonitrile and 0.2%(V/V) pentafluoropropionic acid aqueous solution containing 0.15%(V/V) trifluoroacetic acid (1∶99, V/V). The pH of the aqueous solution was adjusted to 1.5 with 50%(m/m) sodium hydroxide solution. The pulsed amperometricdetector was operated with aquadruple-potential wave form at 35 ℃ and the injection volume was 25 μL. RESULTS Ribostamycin and its related substances were adequately separated under the established HPLC conditions. The LOD and LOQ of ribostamycin were 0.15 μg·mL-1(3.75 ng injected) and 0.375 μg·mL-1(9.38 ng injected), respectively. The linearity of ribostamycin ranged from 0.15 to 40.0 μg·mL-1 with a correlation coefficient of 0.999 3.The repeatability RSDs(n=6)for method validation of the content assay and total impurities test were 0.33% and 1.10%, respectively. CONCLUSION The established method is characterized by high specificity, sensitivity and good stability. The established method has much lower test cost than the current Ch.P 2015 method and is hopeful to replace it.
关键词
反相高效液相色谱-脉冲安培检测法 /
硫酸核糖霉素 /
含量测定 /
有关物质
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Key words
HPLC-PAD /
ribostamycin sulfate /
content determination /
related substance
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中图分类号：
R917
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参考文献
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脚注
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基金
“重大新药创制”科技重大专项课题“化药制剂质量评价关键技术研究”(课题编号:2015ZX09303001)
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