摘要:
目的 分析比较注射用盐酸大观霉素有关物质测定的高效液相色谱脉冲安培检测法(HPLC-PAD)和高效液相色谱蒸发光散射检测器法(HPLC-ELSD),并采用两种方法测定了31批次样品.方法 两法色谱柱均为Apollo C18 (4.6 mm×250 mm,5 μm),流动相均为0.1 mol·L-1三氟乙酸溶液,柱温均为30°C,进样量均为20 μ.L.PAD检测:检测电极为金电极(3 mm),Ag-AgCl参比电极,钛合金对电极,积分安培检测器,四波形检测电位.ELSD检测:漂移管温度110°C,载气流速2.6 L·min-1,增益1.结果 两法专属性一致,检出有关物质种类和个数一致;PAD检出限为2.4 ng,ELSD检出限为72.8 ng(以S/N =3计);PAD为直接线性关系,质量浓度范围为0.15~150 μg·mL-1;ELSD为双对数线性关系,浓度范围为17~170 μg·mL-1.两法测定31批次样品有关物质,杂质D和杂质E结果一致,杂质A、4R-双氢大观霉素和总杂差异较大;PAD中杂质A采用校正因子校正后,杂质A和总杂结果均与ELSD一致.结论 两种方法均可有效检出有关物质,达到控制大观霉素有关物质的目的.HPLC-PAD需对杂质A和4R-双氢大观霉素进行响应因子校正,ELSD需采用双对数线性回归.%OBJECTIVE To compare the HPLC-PAD method and HPLC-ELSD method for determination of related substances in spectinomycin hydrochloride for injection,and determine 31 batches of samples by the two methods.METHODS The following conditions were used for both methods:the analytical column was Apollo C1s column (4.6 mm ×250 mm,5 μm),mobile phase was 0.1 mol · L-1 trifluoroacetic acid solution,the column temperature was maintained at 30 °C,and the sample volume was 20 μL For the PAD detection,gold electrode (3 mm)was used as the detection electrode,Ag-AgCl as reference electrode,titanium alloy as counter electrode,and integrated amperometric detector was employed with four waveform detection potential.For the ELSD detection,the drift tube temperature was set at 110 °C,and the carrier gas flow rate was 2.6 L · min-1,with gain of 1.RESULTS The two methods were consistent in the species and number of detected impurities.The detection limits of the PAD method and ELSD method were 2.4 and 72.8 ng,respectively (S/N =3).For the PAD method there existed a direct linear relationship in the concentration range of 0.15-150 μg · mL-1,while a double logarithmic linear relationship was shown in the concentration range of 17-170 μg · mL-1 for the ELSD method.When 31 batches of samples were determined for related substances by the two kinds of methods,the contents of impurity D and E were basically the same,but there were significant differences in the contents of impurity A and (4R)-dihydrospectinomycin.After correction,the contents of impurity A and total impurities determined by the PAD method were consistent with those by the ELSD method.CONCLUSION The two methods can both effectively detect the related substances of spectinomycin to achieve the purpose of controlling related substances.When using HPLC-PAD method,the impurity A and (4R)-dihydrospectinomycin should be adjusted by using response factor or the reference substances.The ELSD method needs to use double log linear regression.