离子色谱法测定反义寡核苷酸抗流感新药中铵残留量

Content Determination of Residual Ammonia in Antisense Oligonucleotide Antiviral Drugs by Ion Chromatography

目的:建立测定反义寡核苷酸抗流感新药Flutide中铵残留量的方法。方法:用阴离子交换树脂前处理样品去除核酸干扰(去除效果用紫外分光光度法进行监测),用阳离子色谱法测定铵含量。阳离子交换色谱柱为IonPacCS12,淋洗液为12mmol/L甲烷磺酸,电导检测,流速为1ml/min,抑制器抑制电流为59mA。结果:铵离子检测质量浓度线性范围为0.6～30μg/m(lr=0.9995),平均回收率为99.44%(RSD=1.3%,n=6),检测限为0.12ng。结论:该方法结果准确,可用于生物大分子水溶性寡核苷酸原料药中杂质铵的控制。

OBJECTIVE： To establish the method for the determination of residual ammonia in antisense oligonucleotide antiviral drugs （Flutide）. METHODS： The sample was pre-treated by anion-exchange resin and removed interference from nucleic acid （monitored with UV spectrophotometry）. The content of ammonia was determined by cation exchange chromatography. IonPac CS12 column was used with elutriant consisted of 12 mmol/L loprazolam at the flow rate of 1 ml/min. The current of suppressed conductivity detector was 59 mA. RESULTS： The linear range of ammonia ion were 0.6-30 μg/mL （r=0.999 5） with average recovery rate of 99.44% （RSD=1.3%, n=6）. The detection limit was 0.12 ng. CONCLUSIONS： The method is accurate and suitable for the residual ammonia control in macromolecule water-solubile antisense oligonucletide.