There is an increasing emphasis on the critical evaluation of interbatch purity and physical stability of therapeutic peptides. This is due to concerns over the impact that product- and process-related impurities may have on safety and efficacy of this class of drug. Aspartic acid isomerization to isoaspartic acid is a common isobaric impurity that can be very difficult to identify without first synthesizing isoAsp peptide standards for comparison by chromatography. As such, analytical tools that can determine if an Asp residue has isomerized, as well as the site of isomerization within the peptide sequence, are highly sought after. Ion mobility-mass spectrometry is a conformation-selective method that has developed rapidly in recent years particularly with the commercialization of traveling wave ion mobility instruments. This study employed a cyclic ion mobility (cIMS) mass spectrometry system to investigate the conformational characteristics of a therapeutic peptide and three synthetic isomeric forms, each with a single Asp residue isomerized to isoAsp. cIMS was able to not only show distinct conformational differences between each peptide but crucially, in conjunction with a simple workflow for comparing ion mobility data, it correctly located which Asp residue in each peptide had isomerized to isoAsp. This work highlights the value of cIMS as a potential screening tool in the analysis of therapeutic peptides prone to the formation of isoAsp impurities.

中文翻译：

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使用循环离子淌度质谱法在治疗性肽中快速定位天冬氨酸异构体

人们越来越重视对治疗性肽的批次间纯度和物理稳定性进行严格评估。这是由于担心与产品和工艺相关的杂质可能对此类药物的安全性和有效性产生影响。天冬氨酸异构化为异天冬氨酸是一种常见的同量异位素杂质，如果不首先合成 isoAsp 肽标准品进行色谱比较，就很难识别。因此，可以确定 Asp 残基是否已异构化以及肽序列内异构化位点的分析工具受到高度追捧。离子淌度-质谱法是一种构象选择方法，近年来发展迅速，尤其是随着行波离子淌度仪器的商业化。本研究采用循环离子淌度 (cIMS) 质谱系统来研究治疗性肽和三种合成异构体的构象特征，每种异构体都有一个 Asp 残基异构化为 isoAsp。cIMS 不仅能够显示每个肽之间明显的构象差异，而且至关重要的是，结合用于比较离子淌度数据的简单工作流程，它可以正确定位每个肽中的哪个 Asp 残基已异构化为 isoAsp。这项工作突出了 cIMS 作为潜在筛选工具在分析易于形成 isoAsp 杂质的治疗性肽方面的价值。cIMS 不仅能够显示每个肽之间明显的构象差异，而且至关重要的是，结合用于比较离子淌度数据的简单工作流程，它可以正确定位每个肽中的哪个 Asp 残基已异构化为 isoAsp。这项工作突出了 cIMS 作为潜在筛选工具在分析易于形成 isoAsp 杂质的治疗性肽方面的价值。cIMS 不仅能够显示每个肽之间明显的构象差异，而且至关重要的是，结合用于比较离子淌度数据的简单工作流程，它可以正确定位每个肽中的哪个 Asp 残基已异构化为 isoAsp。这项工作突出了 cIMS 作为潜在筛选工具在分析易于形成 isoAsp 杂质的治疗性肽方面的价值。