硫酸西索米星注射液有关物质分析方法的比较

Comparison of Analytical Methods for Related Substances of Sisomicin Sulfate Injection

目的建立HPLC-PAD测定硫酸西索米星注射液有关物质的方法并与法定方法进行比较。方法采用IonPac AMG C_(18)(4.0 mm×150 mm,3μm)色谱柱,以0.1 mol·L^(-1)三氟乙酸(含体积分数0.025%五氟丙酸,不含碳酸盐的质量浓度50%氢氧化钠溶液5 mL,用质量浓度50%氢氧化钠溶液调节pH值至2.3)-乙腈(95∶5)为流动相,流速0.7 mL·min^(-1);柱后碱液为0.76 mol·L^(-1)氢氧化钠,流速0.35 mL·min^(-1);柱温30℃,用脉冲安培检测器检测,检测池温度35℃,工作电极金电极(直径3 mm),参比电极Ag/AgCl电极,检测电位为四电位,测定硫酸西索米星有关物质,并将测定结果与法定方法测定结果进行比较。结果西索米星峰能与庆大霉素C_(1a)峰和奈替米星峰完全分开,其他各杂质峰之间能有效分离,空白无干扰,方法的LOD(S/N=3)为2 ng,LOQ(S/N=10)为6 ng,进样精密度RSD(n=6)为0.9%。法定方法与HPLC-PAD法测定得到的硫酸西索米星注射液总杂(%)、最大单杂(%)和含量(%)进行配对样品的t检验,得到P=0.034、P=0.364和P=0.605。结论与法定方法比较,该法专属性好,灵敏度高,精密度好,测定结果准确可靠,可用于硫酸西索米星注射液有关物质测定。

OBJECTIVE To establish an HPLC-PAD method to determine the related substances of sisomicin sulfate injection and compare with the statutory method.METHODS IonPac AMG C_（18）（4.0 mm×150 mm,3 μm） chromatographic column was used with acetonitrile-0.1 mol·L^-1 trifluoroacetic acid（containing 0.025% of pentafluoropropionic acid,5 mL of 50% NaOHsolution without carbonate,pH of the aqueous solution adjusted to 2.3 with 50% NaOHsolution.） as mobile phase at a flow rate of 0.7 mL·min^-1.NaOHsolution of 0.76 mol·L^-1 was added post column at a flow rate of 0.35 mL·min^-1.The column temperature was maintaine at 30℃.PAD detector was operated with the cell temperature set at 35℃.The working electrode was a gold electrode（diameter of 3 mm） and a quadruple-potential waveform was selected as detection waveform.The reference electrode was Ag/AgCl,the detection potential was four potential.The determination result of the related substances of sisomicin sulfate injection was compared with that of the statutory method.RESULTS The peaks of sisomicin sulfate,gentamicin C1a and netilmicin could be completely separated,and other impurities could also be effectively separated.The blank sample had no interferences.The LOD and LOQ of etimicin were found to be 2 and 6 ng respectively,and the RSD of precision test（n=6） was 0.9%.Paired-samples t-test showed significance levels of P=0.034,P=0.364 and P=0.605 for total amount of impurities（%）,the biggest single impurity（%） and content（%） respectively between the statutory method and the method of HPLC-PAD.CONCLUSION Compared with the statutory method,this HPLC-PAD method shows higher sensitivity,and is accurate and reliable.It can be applied to the determination of related substances in sisomicin sulfate injection.