UPLC法测定度鲁特韦有关物质含量

目的:建立测定度鲁特韦原料药有关物质含量的超高效液相色谱方法。方法:色谱柱为Waters BEH C 18(2.1 mm×150 mm×1.7μm),流动相:10 mmol·L-1甲酸铵溶液(pH=2.5)为流动相A,乙腈为流动相B,梯度洗脱:0~15 min,流动相B 20%~80%,柱温30℃,流速0.28 mL·min-1,进样量1.0μL,检测波长246 nm。结果:度鲁特韦与各杂质之间的分离度均>1.5;杂质A~D的检测限和定量限分别为8 ng·mL-1和25 ng·mL-1;度鲁特韦浓度和杂质A~D浓度在相应浓度范围内与峰面积线性关系良好,相关系数r均>0.998;杂质A~D加样回收率均在80.0%~120.0%之间,RSD<5.0%(n=9),度鲁特韦含量准确度结果在99.0%~101.0%范围内,RSD<0.2%(n=9),3批原料药含量结果分别为99.79%,99.77%,100.03%,杂质A分别为0.10%,0.10%,0.11%,杂质B、C均未检出,未知单杂均小于0.05%,总杂分别为0.15%,0.13%,0.14%。结论:本研究建立的UPLC法,专属性强,重复性好,灵敏度和准确度高,可作为度鲁特韦原料药有关物质和含量的控制方法。

Determination the Assay of Related Substances in Dolutegravir by UPLC

Objective:To establish UPLC method for determination of related substances and assay in Dolutegravir drug substance.Methods:The determination was carried out on Waters BEH C 18(2.1 mm×150 mm×1.7μm)with 10 mmol·L-1 ammonium formate buffer pH=2.5 adjusted with formic acid(mobile phase A)and acetonitrile(mobile phase B)as mobile phase.Linearity gradient elution was run by mobile phase B changed from 20%to 80%in the range of 0~15 min,with the column temperature of 30℃,and the injection volume of 1.0 uL at the flow rate of 0.28 mL(min),monitored at 246 nm.Results:The resolutions between Dolutegravir peak and other impurities peaks were not less than 1.5.The LOD and LOQ for impurity A,B,C and D all were 8 ng·mL-1 and 25 ng·mL-1,respectively.The calibration curves for Dolutegravir and Impurity A,B,C and D were all linear in the range of the corresponding concentration of them with r>0.998.The recovery of Impurity A,B,C and D all were within the range 80%~120%,and RSD were less than 5.0%(n=9).The accuracy results of Dolutegravir was within the range of 99.0%~101.0%,and RSD was not more than 0.2%(n=9).The assay results of 3 batches of Dolutegravir drug substance were 99.79%,99.77%and 100.03%respectively,and the content of Impurity A were 0.10%,0.10%and 0.11%respectively.Impurity B and C were both no detection,the results of unknown impurity all were less than 0.05%,and the results of total impurities were 0.15%,0.13%and 0.14%respectively.Conclusion:The UPLC method is specific,repeatable,sensitive and accurate,and is effective for quality control in Dolutegravir.