HPLC-ELSD和HPLC-MSn分析硫酸奈替米星及其注射液

 目的建立高效液相色谱-蒸发光散射检测法(HPLC-ELSD)测定硫酸奈替米星及注射液的含量和有关物质,并采用高效液相色谱-电喷雾离子阱质谱联用(HPLC-MSn),鉴定硫酸奈替米星及其有关物质。方法采用Agilent SB-C₁₈(4.6 mm×250mm,5μm)色谱柱,流动相为0.05 mol·L^-1三氟乙酸溶液-丙酮(90∶10),流速为1.0 mL·min^-1,漂移管温度为50℃,雾化气体压力为3.5×10⁵Pa,在正离子检测方式下,用电喷雾离子阱质谱法对奈替米星和有关物质进行多级质谱分析。结果奈替米星和有关物质分离良好。奈替米星在28～870 mg·L^-1内呈较好的线性关系(r=0.999 9),重复性试验RSD=1.1%,回收率为99.9%,检测限为5.6 mg·L^-1,结合多级质谱裂解初步推断奈替米星中主要一个未知有关物质是奈替米星分子中发生B环与C环之间的糖苷键断裂,生成的脱去C环(氨基葡萄糖)的降解产物。结论本方法快速、简便,结果准确可靠,重现性好。

Determination of Netilmicin Sulfate and Its Related Substances by HPLC-ELSD and HPLC-MSn

OBJECTIVE To develop a new and simple HPLC-ELSD method for the determination of netilmicin sulfate and its related substances.To build a HPLC-MSⁿ method for the structural identification of netilmicin sulfate and its related substances.METHODS The column was a Agilent SB-C₁₈(4.6 mm×250 mm,5 μm).The mobile phase was 0.05 mol·L^-1trifluoroacetic acid-acetone(90∶10).The drift tube temperature was 50 ℃.The pressure of nebulizing gas was 3.5×10⁵ Pa.In positive mode,netilmicin sulfate and its related substances were elucidated by use of electrospray ion trap mass spectrometry in the multi-stage MS full scan mode.RESULTS Good separation of netilmicin and main relate substances were achieved.The standard curve was linear in the range of 28～870 mg·L^-1(r=0.999 9).The reproducibility was RSD 1.1%.The average recoveries was 99.9%.The limit of detection was 5.6 mg·L^-1.The possible structure of an unknown related substance in netilmicin was deduced based on the HPLC-MSⁿ data.CONCLUSION The method is simple and rapid,the results are accurate and reproducible.