醋酸曲普瑞林缓释注射剂中杂质的检测分析  

Analysis of impurities in the sustained-release injection of triptorelin acetate

作　　者：孙悦[1] 郭宁子 梁成罡[1] 杨化新[1] SUN Yue;GUO Ning-zi;LIANG Cheng-gang;YANG Hua-xin(National Institutes for Food and Drug Control,Beijing 102629,China)

机构地区：[1]中国食品药品检定研究院

出　　处：《药物分析杂志》2019年第10期1870-1881,共12页Chinese Journal of Pharmaceutical Analysis

基　　金：国家“重大新药创制”科技重大专项（2015ZX09303001）;“十三五”国家重大新药创制专项(2017ZX09101001-7)

摘　　要：目的:对醋酸曲普瑞林缓释注射剂中的未知杂质进行检测和结构确证。方法:采用高效液相色谱法进行杂质分离检测,使用XBridge?Peptide BEH C18色谱柱(4.6 mm×250 nm,3.5μm,300?),以磷酸-三乙胺缓冲液及乙腈-正丙醇为流动相体系,流速1 mL·min^-1,柱温35℃,检测波长210 nm;采用高效液相色谱-线性离子阱串联质谱法(HPLC-LTQ-MS/MS法)对未知杂质进行结构确证,采用Kromasil C18预柱(2.1mm),柱温30℃,检测波长210 nm,纯水(含0.05%甲酸)为流动相A,甲醇为流动相B,梯度洗脱,流速0.2m L·min^-1;电喷雾电离源(ESI),正离子模式,扫描范围m/z 150~1 500,毛细管电压35 V,毛细管温度275℃,喷雾电压4 kV,鞘气速度35 arb,辅助气流速10 arb。结果:对3种来源的缓释制剂进行了检测,发现并确证了25种未知杂质的结构,并进行了来源归属,其中工艺杂质有缺失肽、插入肽、光学异构体,降解杂质有氧化杂质、还原杂质、酰化杂质等。结论:不同企业醋酸曲普瑞林缓释注射剂中的杂质与制剂工艺有关,曲普瑞林与辅料聚乳酸聚乙醇酸(PLGA)相互作用生成的酰化杂质,酰化位点为肽链4位丝氨酸残基上的游离羟基。Objective:To study and confirm the structure of the impurity profiles in the sustained-release injection of triptorelin acetate. Methods:Impurities were detected and separated by high performance liquid chromatography(HPLC). The HPLC separation was carried out on XBridge? Peptide BEH C18 column(4.6 mm×250 nm,3.5 μm,300 ?),with a mobile phase consisting phosphate-triethylamine buffer and acetonitrile n-propanol system at a flow rate of 1 mL·min^-1,a column temperature of 35 ℃,a detection wavelength of 210 nm. The high performance liquid chromatography-multiple stage Orbitrap mass spectrometric method(HPLC-LTQ-MS/MS method) was developed to confirm the structure of its unknown impurify. The desalination of impurities was carried out on Kromasil C18 pre-column(2.1 mm),with a mobile phase consisting of 0.05% formic acid aqueous solution(A) and methanol(B) at a flow rate of 0.2 mL·min^-1 by gradient elution. The MS settings were as follows:scan range,m/z 150^-1,500;capillary voltage,35 V;capillary temperature,275 ℃;spray voltage,4 kV;sheath gas, 35 units;sweep gas,10 units. Results:The structures of 25 kinds of unknown impurities in the sustained-release injection of triptorelin acetate from three different sources were determined by the information of HPLC-MS-MS,and its origin was attributed to be the drug-excipient adducts related to the manufacturing processes. Processrelated impurities were found in the production process including missing peptide and insert peptide ligands,optical isomers and degradation of impurities(oxide impurities,reduced impurities and acylated impurities). Conclusion:The impurities of the sustained-release injection of triptorelin acetate from different enterprises are all related to the manufacturing processes. The formation mechanism is identified to be the pharmaceutic adjuvant[poly(lactic-co-glycolic acid),PLGA] interaction with the 4’-OH Ser of triptorelin to generate acylated impurities.

关 键 词：醋酸曲普瑞林 缓释注射剂 杂质谱 酰化杂质 聚乳酸聚乙醇酸 高效液相色谱-线性离子阱串联质谱法 

分 类 号：R917[医药卫生—药物分析学]