目的 解析合成醋酸西曲瑞克的两种难解析杂质氨基酸序列及修饰,探讨氨甲酰基中性丢失效应在质谱氨基酸序列分析中的影响。方法 在合成工艺分析的基础上,利用静电场轨道阱质谱碰撞诱导解离,结合高能碰撞解离多级碎裂,解析无法合成纯品并且单一高能碰撞解离无法解析的两种杂质。结果 精氨酸侧链无修饰的杂质形成氢键,避免了氨甲酰基中性丢失。氨甲酰基中性丢失效应和空间位阻效应导致杂质3理论碎片与实测碎片不匹配。在杂质2的结构中,磺酸基与瓜氨酸羰基氧形成的氢键避免氨甲酰基中性丢失,但由于磺酸基位阻较小,发生N-S键断裂。结论 氨甲酰基中性丢失效应可能导致单一高能碰撞解离碎片与理论碎片不匹配,空间位阻效应、形成氢键也可能影响氨甲酰基中性丢失,因此当数据解析引擎显示不匹配时,还应深入分析推测氨基酸侧链的低能键,才能实现充分的杂质质控。
Abstract
OBJECTIVE To analyze the amino acid sequences and modification of two perplexing impurities in cetrorelix acetate, and discuss the effect of carbamyl neutral loss on the amino acid sequencing using mass spectrometry.CONCLUSION Based on the analysis of synthesis process, collision-induced dissociation combined with high-energy collision dissociation was used to analyze two impurities which can neither be synthesized nor solved by high-energy collision dissociation alone.RESULTS The hydrogen bond avoids the neutral loss of carbamyl group in the unmodified impurities of arginine side chain. The effects of carbamyl neutral loss and steric hindrance make the theoretical fragments of impurity 3 inconsistent with the measured fragments. In the structure of impurity 2, the hydrogen bond between sulfonic acid group and citrulline carbonyl oxygen avoids the neutral loss of carbamoyl group. However, due to the small steric hindrance of sulfonic acid group, the N-S bond is broken.CONCLUSION The neutral loss effect of carbamyl groups may lead to a mismatch between the dissociated fragments of a single high-energy collision and the theoretical fragments. Steric hindrance and hydrogen bonding may also affect the neutral loss of carbamyl groups. Therefore, when the data analysis engine shows a mismatch, we should analyze and speculate the low-energy bonds of the amino acid side chains in depth to achieve sufficient impurity quality control.
关键词
氨甲酰基中性丢失效应 /
静电场轨道阱质谱 /
碰撞诱导解离 /
高能碰撞解离 /
多级碎裂
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Key words
carbamyl neutral loss effect /
Orbitrap-MS /
collision-induced dissociation /
high-energy collision dissociation /
MSn
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中图分类号：
R917
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