摘要
建立了基于在线二维高效液相色谱-四极杆飞行时间质谱(2D HPLC-Q TOF/MS)的两性霉素B杂质谱分析方法。第一维液相色谱(^(1)D-LC)以XBridge Shield C18为色谱柱、甲醇-乙腈-4.2 g/L柠檬酸水溶液为流动相。第二维液相色谱(^(2)D-LC)以Xtimate C8为捕集柱,以含0.1%甲酸的10 mmol/L甲酸铵水溶液-乙腈(95∶5, v/v)为流动相进行捕集和脱盐;以Xtimate C8为色谱柱,以含0.1%甲酸的10 mmol/L甲酸铵水溶液-乙腈为流动相进行洗脱。该方法结合了在线稀释和多针捕集装置,实现了10个组分/杂质的高效分离,并通过四极杆飞行时间质谱实现了对不稳定组分/杂质的结构推断,解决了2D HPLC溶剂兼容性的难题,增加了分析通量,提高了2D HPLC的自动化能力,避免了两性霉素B与部分杂质在色谱分离过程中相互转化的问题,提高了检测灵敏度。利用该方法可以灵敏地对两性霉素B的杂质谱进行分析,所得到的研究结果对两性霉素B的生产工艺改进具有指导意义。
Amphotericin B(AmB)is a polyene-macrolide antimicrobial drug with a broad antibacterial spectrum and remarkable efficacy against deep fungal infections.It binds to ergosterol on the fungal cell membrane and alters its permeability,thereby destroying the membrane.AmB is a multicomponent antimicrobial medication that contains a wide range of impurities,rendering quality analysis extremely difficult.In the current Chinese Pharmacopoeia(Edition 2020)and European Pharmacopoeia(EP10.3),high performance liquid chromatography(HPLC)is applied to examine related substances in AmB.However,this technique presents a number of issues.For instance,the mobile phases used in the HPLC method described in both references contain nonvolatile inorganic salts,which cannot be coupled with a mass spectrometry(MS)detector.In addition,because the mobile phases used have a low pH,the component/impurities of AmB drug can easily be degraded or interconverted during the analytical process,leading to reduced analytical accuracy.Therefore,the accuracy and sensitivity of this method must be improved.In this study,a method based on on-line two-dimensional high performance liquid chromatography-quadrupole time-of-flight mass spectrometry(2D HPLC-Q TOF/MS)was developed to analyze the impurity profile of AmB in accordance with the Chinese Pharmacopoeia(Edition 2020)and European Pharmacopoeia(EP10.3).The method combines on-line dilution and a multiple-capture HPLC system to achieve the efficient separation of AmB component/impurities.It also resolves the issue of poor solvent compatibility in 2D HPLC,increases the analytical flux,enhances the automation capability,reduces the mutual conversion of AmB and its impurities during the analytical process,and increases the detection sensitivity of the method.MS was also used to determine the structural inference of unstable components and impurities.An XBridge Shield C18 column(250 mm×4.6 mm,3μm)was used for first-dimensional-liquid chromatography with gradient elution using methanol-acetonitrile-4.2 g/L
作者
翁荣文
王轩堂
闻宏亮
刘浩
WENG Rongwen;WANG Xuantang;WEN Hongliang;LIU Hao(College of Pharmacy,Fudan University,Shanghai 201203,China;Shanghai Institute for Food and Drug Control,Shanghai 201203,China;China State Institute of Pharmaceutical Industry,Shanghai 201203,China)
出处
《色谱》
CAS
CSCD
北大核心
2024年第5期432-444,共13页
Chinese Journal of Chromatography
关键词
在线二维高效液相色谱-四极杆飞行时间质谱
两性霉素B
杂质谱
on-line two-dimensional high performance liquid chromatography-quadrupole time-of-flight mass spectrometry(2D HPLC-Q TOF/MS)
amphotericin B
impurity profile
