高分辨液质联用技术推定泊马度胺胶囊有关物质

目的：采用高分辨液质联用技术快速分析和推定泊马度胺胶囊中的降解产物。方法：采用AgilentZORBAX SB-CN色谱柱（150 mm×4.6 mm，5 mm），以质谱兼容性0.1%甲酸水溶液-甲醇为流动相，梯度洗脱，流速1.0 mL·min^-1，检测波长为240 nm，对泊马度胺胶囊经酸、碱、高温、氧化及光照破坏产生的有关物质进行分离。通过电喷雾正离子化四极杆-飞行时间质谱（Q-TOF/MS）测定各有关物质的精确分子质量及分子式，再根据二级质谱（MS/MS）碎裂信息推定未知降解产物结构；喷雾电压3.5 kV，雾化器流量11 L·min^-1，去溶剂气温度350℃，离子源气压310 kPa，驱簇电压120 V，MS扫描范围m/z 100～1 000，MS/MS扫描范围m/z 50～800，MS/MS碰撞能量10～20 eV。结果：建立的HPLC分析方法下各有关物质与主成分分离良好，在强制降解供试品溶液中共检测到14个主要有关物质，经液质初步分析推定了它们的结构分别为泊马度胺的水解产物（有关物质1、2、3、7和8），3-氨基-1，2-苯二甲酸（有关物质4），2-硝基-6-[[（2，6-二氧代-3-哌啶基）氨基]羰基]-苯甲酸（有关物质5），2-亚硝基-6-[[（2，6-二氧代-3-哌啶基）氨基]羰基]-苯甲酸（有关物质6），N’-[2-（2，6-二氧代哌啶-3-基）-1，3-二氧代-4-异二氢吲哚]-乙酰肼（有关物质9），泊马度胺羟基化产物（有关物质10和12），3-硝基-N-（2，6-二氧-3-哌啶基）苯邻二甲酰亚胺（有关物质11），3-甲基氨基-N-（2，6-二氧-3-哌啶基）苯邻二甲酰亚胺（有关物质13），泊马度胺的羟基化二聚体产物（有关物质14）。结论：新建立的液质联用方法适用于泊马度胺胶囊中有关物质的检查，为泊马度胺制剂质量控制和工艺优化提供重要参考。

Objective: To develop a rapid and sensitive method by high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry(HPLC-Q-TOF/MS) for screening and identification of the degradation products in pomalidomide capsules.Methods: LC was performed on a ZORBAX SB-CN column (150 mm×4.6 mm,5 mm) by linear gradient elution using a mobile phase containing 0.1% formic acid solution and methanol at the flow rate of 1.0 mL·min^-1.The pomalidomide capsules test solution was treated with acid, alkaline,oxidation,thermal and photolysis to obtain the degradation products.They were characterized by high resolution Q-TOF/MS with positive ionization as well as the product MS/MS spectra determination.The conditions were optimized as follow:capillary voltage:3.5 kV;drying gas:11 L·min^-1;gas temperature:350℃;nebulizer:310 kPa;fragmentor:120 V;MS scan range:m/z 100-1 000;MS/MS scan range:m/z 50-800;MS/MS collision energy was set at 10-20 eV.Results: The related substances and polimadomide was completely separated using the established HPLC method.A total of 14 related substances were tentatively identified or characterized by their high resolution mass spectrometry(HRMS) and the feature fragmentation from MS/MS experiments in degradation solution samples.They were hydrolysates of pomalidomide(impurity 1,2,3,7 and 8),3-amino-1, 2-phthalate(impurity 4),2-nitro-6-[[(2,6-dioxo-3-piperidinyl) amino]carbonyl]-benzoic acid(impurity 5),2-nitroso-6-[[(2,6-dioxo-3-piperidinyl) amino]carbonyl]-benzoic acid(impurity 6),N'-[2-(2,6-dioxo-3-yl)-1,3-dioxo-isoindoline]-acetohydrazide(impurity 9),hydroxylated of pomalidomide(impurity 10 and 12),3-nitro-N-(2,6-dioxo-3-piperidin-yl) phthalimide(impurity 11),3-methyl-amino-N-(2,6-dioxo-3-piperidin-yl) phthalimide(imputiy 13),and hydroxylated dimer of pomalidomide(impurity 14),respectively. Conclusion: The established LC-MS method is useful for the qualitative analysis of the unknown degradation products in pomalidomide capsules.The related substances identified in pomalidomide capsules are valuable for its quality control and manufacturing process optimization.