Pharmaceutical Candidate Optimization-Bioanalytical and Discovery Analytical Sciences, Princeton, NJ 08543-4000, USA.
Anal Bioanal Chem. 2013 May;405(12):4283-7. doi: 10.1007/s00216-013-6795-3. Epub 2013 Feb 21.
PEGylation has been widely used to improve the biopharmaceutical properties of therapeutic proteins and peptides. Previous studies have used multiple analytical techniques to determine the fate of both the therapeutic molecule and unconjugated poly(ethylene glycol) (PEG) after drug administration. A straightforward strategy utilizing liquid chromatography-mass spectrometry (LC-MS) to characterize high-molecular weight PEG in biologic matrices without a need for complex sample preparation is presented. The method is capable of determining whether high-MW PEG is cleaved in vivo to lower-molecular weight PEG species. Reversed-phase chromatographic separation is used to take advantage of the retention principles of polymeric materials whereby elution order correlates with PEG molecular weight. In-source collision-induced dissociation (CID) combined with selected reaction monitoring (SRM) or selected ion monitoring (SIM) mass spectrometry (MS) is then used to monitor characteristic PEG fragment ions in biological samples. MS provides high sensitivity and specificity for PEG and the observed retention times in reversed-phase LC enable estimation of molecular weight. This method was successfully used to characterize PEG molecular weight in mouse serum samples. No change in molecular weight was observed for 48 h after dosing.
聚乙二醇化（PEGylation）已广泛用于改善治疗性蛋白质和肽类的生物制药特性。以前的研究使用了多种分析技术来确定治疗分子和未缀合的聚（乙二醇）（PEG）在给药后的命运。本文提出了一种利用液相色谱-质谱（LC-MS）在生物基质中对高分子量 PEG 进行特征分析而无需复杂样品制备的简单策略。该方法能够确定体内是否将高分子量 PEG 切割成低分子量 PEG 物质。反相色谱分离用于利用聚合物材料的保留原理，其中洗脱顺序与 PEG 分子量相关。然后，在源碰撞诱导解离（CID）与选择反应监测（SRM）或选择离子监测（SIM）质谱（MS）结合使用，以监测生物样品中的特征 PEG 片段离子。MS 对 PEG 具有高灵敏度和特异性，并且在反相 LC 中的观察保留时间可用于估计分子量。该方法成功地用于在小鼠血清样品中对 PEG 分子量进行特征分析。给药后 48 小时内未观察到分子量变化。
