脱氧核苷酸钠制剂的有关物质研究

Study on related substances and stability of sodium deoxyribonucleotide formulations

目的建立HPLC法测定注射用脱氧核苷酸钠和脱氧核苷酸钠注射剂中的有关物质,并应用LC-MS/MS法推测有关物质结构。方法采用HPLC法测定2种脱氧核苷酸钠制剂中的有关物质。色谱柱为Agilent Zorbax SAX柱(250 mm×4.6 mm,5μm),流动相为浓度50 mmol·L-1的磷酸二氢钾溶液(取6.80 g磷酸二氢钾,加水800 m L,用磷酸调p H值至3.3,再加水稀释至1 000 m L),流速为0.6 m L·min-1,柱温为30℃,进样量为20μL,检测波长为258 nm。结果 2种脱氧核苷酸钠制剂中脱氧核糖胞嘧啶核苷酸钠(d CMP-Na)、脱氧核糖腺嘌呤核苷酸钠(d AMP-Na)、脱氧核糖胸腺嘧啶核苷酸钠(d TMP-Na)和脱氧核糖鸟嘌呤核苷酸钠(d GMP-Na)分别与其相邻色谱峰完全分离,测得的注射用脱氧核苷酸钠中最大单个有关物质百分含量均不大于2.0%,有关物质总量均不大于4.0%。脱氧核苷酸钠注射剂中最大单个有关物质百分含量平均大于30%,而有关物质总量更是达到79%以上。经质谱分析,推测注射用脱氧核苷酸钠有关物质为5-甲基胞嘧啶脱氧核糖核苷酸,脱氧核苷酸钠注射剂有关物质为胞嘧啶核糖核苷酸、尿嘧啶核糖核苷酸和鸟嘌呤核糖核苷酸。结论该方法简便,准确,专属性较好,适用于脱氧核苷酸钠制剂的质量分析,为脱氧核苷酸钠制剂临床用药的安全性改善提供参考。

Objective To develop a method for the related substances analysis of sodium deoxyribonucleotide by HPLC and apply LC-MS / MS method to predict the structures of the related substances. Methods The analysis was performed by using a Agilent Zorbax SAX（ 250 mm × 4. 6 mm,5 μm） at elution of50 mmol·L- 1KH2PO4 with Ultraviolet Detector（ 258 nm）,the flowrate was 0. 6 mL·min^- 1and the column temperature was maintained at 30 ℃. Results The dCMP-Na,d AMP-Na,dTMP-Na and d GMP-Na of the two formulations were completely separated with the adjacent artifactual bands. The percentage content of the largest single related substance of deoxynucleotide sodium for injection was less than 2. 0%,and the total related substances were less than 4. 0%. While the percentage content of the largest single related substance of sodium deoxyribonucleotide injection was more than 30%,and the total related substances were more than79%. Conclusions This method is simple,precise and of effective separation. It is suitable for the quality analysis of sodium deoxyribonucleotide formulations,and providing reference for its clinical medication safety.