化学试剂

建立高效液相色谱法(HPLC-DAD)同时检测布美他尼原料药中的8种杂质含量。采用Thermo Hypersil GOLD C18(4.6 mm×250 mm, 5μm,孔径120?)为色谱柱，以0.1%(体积分数)三氟乙酸溶液(取1 mL三氟乙酸溶于1 000 mL水中)为流动相A,甲醇为流动相B,梯度洗脱(0～5 min 60%A;5～10 min 60%A→15%A;10～20 min 15%A;20～25 min 15%A→60%A;25～30 min 60%A),流速为1.0 mL/min,柱温为30℃,进样量为20μL。空白溶剂不干扰布美他尼及其8种杂质的测定，杂质A、B、C、D、E、G、I、J质量浓度分别在0.020 1～4.028 0、0.028 0～3.730 1、0.045 8～3.661 8、0.050 9～4.069 2、0.048 5～3.877 3、0.101 1～4.042 8、0.050 9～4.070 0和0.051 8～4.147 7μg/mL范围内与相应峰面积呈良好的线性关系，相关系数r分别为0.999 3、0.999 1、0.999 9、0.999 4、0.999 6、0.999 2、0.999 9、0.998 8,检测限(LOD)为7.05～35.37 ng/mL,定量限(LOQ)为20.14～101.07 ng/mL,杂质A、B、C、D、E、G、I、J的平均回收率分别为95.4%、93.0%、101.1%、98.7%、100.0%、101.0%、101.2%、98.1%,RSD分别为1.26%、0.63%、1.97%、0.40%、3.08%、0.79%、1.18%、0.85%。所建立的高效液相色谱法(HPLC)高效、稳健、准确，可同时测定布美他尼原料药中8种杂质(工艺杂质与降解产物)的含量。

A high-performance liquid chromatography method with diode-array detection(HPLC-DAD) was established for the simultaneous determination of eight impurities in bumetanide active pharmaceutical ingredient(API).A Thermo Hypersil GOLD C18(4.6 mm×250 mm, 5 μ m, pore size120 ?) was used, with 0.1% volume fraction trifluoroacetic acid solution(prepared by dissolving 1 mL trifluoroacetic acid in 1 000 mL water) as mobile phase A and methanol as mobile phase B.A gradient elution was employed(0～5 min 60% A;5～10 min 60% A→15% A;10～20 min 15% A;20～25 min 15% A→60% A;25～30 min 60% A).The flow rate was 1.0 mL/min, the column temperature was 30 ℃,and the injection volume was 20 μL.The blank solvent showed no interference with the detection of bumetanide or its eight impurities.The mass concentrations of impurities A,B,C,D,E,G,I,and J showed good linear relationships with their corresponding peak areas in the range of 0.020 1～4.028 0,0.028 0～3.730 1,0.045 8～3.661 8,0.050 9～4.069 2,0.048 5～3.877 3,0.101 1～4.042 8,0.050 9～4.070 0,and 0.051 8～4.147 7 μg/mL,respectively.The correlation coefficients r was 0.999 3,0.999 1,0.999 9,0.999 4,0.999 6,0.999 2,0.999 9,and 0.998 8.The limits of detection(LOD) ranged from 7.05 to 35.37 ng/mL,and the limits of quantification(LOQ) ranged from 20.14 to 101.07 ng/mL.The average recoveries of impurities A,B,C,D,E,G,I,and J were 95.4%,93.0%,101.1%,98.7%,100.0%,101.0%,101.2%,and 98.1%,respectively, with corresponding relative standard deviations(RSDs) of 1.26%,0.63%,1.97%,0.40%,3.08%,0.79%,1.18%,and 0.85%.The established HPLC method is efficient, robust and accurate, and is suitable for the simultaneous determination of eight impurities(both process-related impurities and degradation products) in bumetanide API.