RP-HPLC测定注射用硫普罗宁的有关物质

Development and validation of a novel and simple RP-HPLC method for the determination of related substances of tiopronin for injection

建立一种高效液相色谱法测定注射用硫普罗宁有关物质的方法。色谱柱为C18柱,流动相为乙腈-0.1%磷酸(8∶92),流速为1.0mL/min。光电二极管阵列检测器的波长为210nm。采用该方法共检测到硫普罗宁中7种有关物质,并用质谱法对其进行了结构推测。该方法具有良好的系统适用性、专属性,在0.3~50μg/mL质量浓度范围内具有良好的线性(r≥0.999),检测限和定量限分别为0.10和0.31μg/mL,准确度在98.7%~103.7%之间,该方法的日内、日间精密度较好(RSD≤4.4%),耐用性好(RSD≤6.4%)。该方法成功地应用于注射用硫普罗宁中有关物质的测定,揭示了巯基化合物和甘氨酸类似物在高效液相色谱上的保留行为以及流动相pH对待测物色谱行为的影响。

In this paper,a novel and simple RP-HPLC method for the determination of related substances of tiopronin for injection was described.The RP-HPLC analysis was performed on a C 18 column,with acetonitrile-0.1% phosphoric acid (8 ∶ 92),mobile phase in isocratic mode at a rate of 1.0 mL/min.The photodiode array detector was set at 210 nm.Seven related substances were detected and the structures were characterized by mass spectrometry.The method showed great suitability,specificity and excellent linearity over the concentration range of 0.3 to 50 μg/mL ( r ≥0.999),and the limits of detection and quantitation were found to be 0.10 and 0.31 μg/mL,respectively.The accuracy of the method determined by the entire mean recovery ranged from 98.7% to 103.7%.The intra-and inter-day precision was satisfactory (RSD≤4.4%) and robust (RSD≤6.4%).And this method was successfully applied for the determination of related substances of tiopronin for injection,which revealed the retention of sulfhydryl compounds and glycine analogues on the RP-HPLC and the effect of the pH value of the mobile phase on the chromatographic behavior of the analytes.