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目的建立梯度洗脱高效液相色谱法测定利格列汀原料杂质的方法。方法色谱柱为Agilent C18柱,流动相A为磷酸盐缓冲液(取磷酸二氢钠2.0 g,加水1 000 m L溶解,并用磷酸调节p H值至2.5±0.1),流动相B为甲醇-乙腈(55∶45),以1.0 m L·min?1流速;检测波长为226 nm;柱温为30℃,进样量为10μL。结果 8-[(3R)-哌啶-3-氨基]-7-(2-丁炔基)-3,7-二氢-3-甲基-1-[(4-甲基-2-喹唑啉基)甲基]-1H-嘌呤-2,6-二酮(杂质A)、8-[(3R)-3-氨基-1-哌啶基]-7-(3-溴-2-丁烯基)-3,7-二氢-3-甲基-1-[(4-甲基-2-喹唑啉基)甲基]-1H-嘌呤-2,6-二酮(杂质B)、8-[(3R)-3-甲酰胺基-1-哌啶基]-(7-(2-丁炔基)-3,7-二氢-3-甲基-1-[(4-甲基-2-喹唑啉基)甲基]-1H-嘌呤-2,6-二酮(杂质C)和利格列汀及其他未知杂质均能达到很好的分离,且杂质A、杂质B、杂质C与利格列汀分别在0.59~5.91μg·m L?1(r=0.999 9),0.59~5.86μg·m L?1(r=0.999 8)、0.58~5.79μg·m L?1(r=0.999 5)和1.32~13.22μg·m L?1(r=0.999 7)内具有良好的线性关系。杂质A、杂质B、杂质C平均回收率(n=9)分别为99.12%(RSD=2.9%)、99.35%(RSD=2.4%)和98.52%(RSD=1.1%)。结论本方法灵敏快速、准确、可靠,专属性强,可作为利格列汀的杂质检查。
Objective To establish a gradient elution HPLC method for the determination of impurity of linagliptin. Methods The column was Agilent C18 column. The mobile phase A was phosphate buffer solution (taking 2.0 g sodium dihydrogen phosphate, dissolving with 1000 ml water and adjusting pH value to 2.5 ± 0.1 with phosphoric acid). The mobile phase B was methanol- Acetonitrile (55:45) at a flow rate of 1.0 m L · min -1. The detection wavelength was 226 nm. The column temperature was 30 ℃ and the injection volume was 10 μL. Results 8 - [(3R) -Piperidin-3-amino] -7- (2-butynyl) -3,7-dihydro-3- (Impurity A), 8 - [(3R) -3-amino-1 -piperidinyl] -7- (3-bromo-2- Butenyl) -3,7-dihydro-3-methyl-1 - [(4-methyl-2- quinazolinyl) methyl] -1 H -purine-2,6-dione (Impurity B ), 8 - [(3R) -3-carboxamido- 1 -piperidinyl] - (7- (2-butynyl) -3,7 -dihydro-3- methyl- 1 - [(4 -quinazolinyl) methyl] -1H-purine-2,6-dione (impurity C) and linagliptin and other unknown impurities, and impurities A, Impurity B, impurity C and linagliptin were in the range of 0.59-5.91μg · m L -1 (r = 0.999 9), 0.59-5.86μg · m L -1 (r = 0.999 8), 0.58-5.79μg · m L (r = 0.999 5) and 1.32-13.22 μg · m L · 1 (r = 0.999 7) respectively.The average recoveries of impurity A, impurity B and impurity C (n = 9) were 99.12% (RSD = 2.9%), 99.35% (RSD = 2.4%) and 98.52% (RSD = 1.1%) respectively.Conclusion The method is sensitive, rapid, accurate, reliable and specific and can be used as impurity of linagliptin .