UHPLC-Q-Orbitrap HRMS法鉴定盐酸阿扎司琼中的有关物质

Identification of related substances in azasetron hydrochloride by UHPLC-Q-Orbitrap HRMS

目的:利用超高效液相色谱-四极杆/静电场轨道阱高分辨质谱技术鉴定盐酸阿扎司琼氯化钠注射液有关物质的化学结构。方法:采用ACQUITY UPLC?HSS C18色谱柱(2.1 mm×50 mm,1.7μm),以含0.1%甲酸的水(A)-乙腈(B)为流动相,梯度洗脱,离子源采用HESI(heated ESI),碰撞能梯度为20、50、100 eV。对盐酸阿扎司琼氯化钠注射液分别进行加盐酸、氢氧化钠溶液、过氧化氢溶液、紫外光照射和90℃水浴等强制降解试验,利用超高效液相色谱-四极杆/静电场轨道阱高分辨质谱检测盐酸阿扎司琼氯化钠注射液及其强制降解溶液中的有关物质,获得阿扎司琼及其有关物质的质谱数据,根据碎片离子裂解规律,结合阿扎司琼的碎片信息和化学结构,推测有关物质可能的化学结构。结果:在盐酸阿扎司琼氯化钠注射液及其强制降解溶液中发现了15种有关物质,鉴定了其可能的化学结构,并根据其来源分为4类:合成中间体(9)、合成副产物(2和13)、■嗪环开环降解产物(1、3、4和11)和酰胺键降解产物(5、6、7、8、10、12、14和15)。结论:本文建立的超高效液相色谱-四极杆/静电场轨道阱高分辨质谱分析方法能快速有效地鉴定盐酸阿扎司琼有关物质的化学结构,对其生产工艺优化、质量标准完善和药品质量控制具有重要意义。

Objective:To identify the chemical structure of the related substances in azasetron hydrochloride and sodium chloride injection by ultra-high performance liquid chromatography-quadrupole/orbitrap high resolution mass spectrometry(UHPLC-Q-Orbitrap HRMS).Methods:An ACQUITY UPLC?HSS C18 column(2.1 mm×50 mm,1.7μm)was used for the separation of the related substances in azasetron hydrochloride and a mixture of water with 0.1%formic acid and acetonitrile was employed as the mobile phase by gradient elution.An HESI(heated ESI)ion source was used and collision energy was set to 20,50 and 100 e V.Forced degradation test was carried out at 1 m L a zasetron hydrochloride and sodium chloride injection,which was added with 1 m L HCl solution(1 mol·L-1),1 mL NaOH solution(1 mol·L-1),or 1 mL H2O2 solution(3%)at room temperature for 6 h,respectively.Besides,1 m L azasetron injection was exposed under the ultraviolet lamp(254 nm)at room temperature for 24 h,or was heated in the 90℃water bath for 5 h.Mass spectrum data of related substances in azasetron hydrochloride and sodium chloride injection and its forced degradation solution was obtained by UHPLCQ-Orbitrap HRMS.The probable chemical structures of related substances were speculated according to the fragmentation pattern of fragment ions,combined with the fragment information and chemical structure of azasetron.Results:Fifteen related substances were found in this study and divided into four categories according to the source:synthetic intermediate(9),synthetic by-product(2 and 13),degradation product of oxazine ring(1,3,4 and 11),and degradation product of amido bond(5,6,7,8,10,12,14 and 15).Conclusion:The established UHPLC-Q-Orbitrap HRMS analysis method can identify the chemical structure of related substances in azasetron hydrochloride rapidly and efficiently.It is significant for manufacturing process optimization,quality standards improvement and drug quality control.