摘要
目的 建立测定鲑降钙素鼻用喷雾剂中有关物质的HPLC方法.方法 使用Macherey-Nagel Nucleosil 100-5 C18AB色谱柱(250 mm×4.0 mm,5 μm);以 0.04 mol/L四甲基氢氧化铵溶液(用磷酸调pH 2.5)-乙腈(9∶1)为流动相A,以0.04 mol/L四甲基氢氧化铵溶液(用磷酸调pH 2.5)-乙腈(4∶6)为流动相B,梯度洗脱;检测波长为 220 nm;体积流量为 1.0 mL/min;进样量为 100 μL;柱温为 65℃;样品室温度:10℃.按加校正因子的主成分自身对照法计算降钙素C、N-乙酰半胱氨酰鲑降钙素(杂质A)、9-D-亮氨酸鲑降钙素(杂质B)、去-22-酪氨酸鲑降钙素(杂质C).结果 降钙素C和鲑降钙素杂质A、B、C在 0.20～7.50 μg/mL线性关系良好,平均回收率分别为 98.1%、98.9%、100.5%、99.7%,RSD值分别为 1.6%、1.5%、1.0%、1.1%.杂质A、B、C的校正因子均超出 0.90～1.10.结论 方法简便、快速,可作为鲑降钙素鼻用喷雾剂中有关物质的检测方法.
Abstract
Objective To establish an HPLC method for determination of related substance in Salmon Calcitonin Nasal Spray.Methods The separation was performed on Macherey-Nagel Nucleosil 100-5 C18AB column(250 mm×4.0 mm,5 μm).The mobile phase A was 0.04 mol/L tetramethyl ammonium hydroxide buffer(adjusted with phosphoric acid to pH 2.5)-acetonitrile(9∶1),and the mobile phase B was 0.04 mol/L tetramethyl ammonium hydroxide buffer(adjusted with phosphoric acid to pH 2.5)-acetonitrile(4∶6),with gradient elution.The detection wavelength was 220 nm,the flow rate was 1.0 mL/min,injection volume was 100 μL,column temperature was 65℃,and sample temperature was 10℃.The contents of calcitonin C and N-acetylcysteine salmon calcitonin(impurity A),9-D-leucine salmon calcitonin(impurity B),and de-22-tyrosine salmon calcitonin(impurity C)were determined by principal component self-control with correction factor.Results The linear ranges of calcitonin C and impurity A,B,and C were all 0.20—7.50 μg/mL.The average recoveries were 98.1%,98.9%,100.5%,and 99.7% with RSD values of 1.6%,1.5%,1.0%,and 1.1%,respectively.The correction factors of impurity A,B,and C were all above 0.90—1.10.Conclusion The method is simple,rapid,and can be used for the determination of related substances in Salmon Calcitonin Nasal Spray.
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