Identification of recombinant human insulin and biosynthetic insulin analogues by multiplexed targeted unlabeled mass spectrometry of proteotypic tryptic peptides

Publication date: Available online 1 May 2019Source: Journal of Pharmaceutical and Biomedical AnalysisAuthor(s): Rani J. Qasem, Anas S. Aldawsari, Faisal E. Almutairi, Abdullah S AlsadoonAbstractDirect qualitative methods that allow the rapid screening and identification of insulin products during early stages of the drug development process and those already in the market can be of great utility for manufacturers and regulatory agencies and the recent scientific literature describes several methods. Herein, a qualitative proteomic method is presented for the identification of recombinant human insulin and all marketed biosynthetic analogues -insulin lispro, aspart, glulisine, glargine, detemir and degludec- via tryptic digestion and identification of proteotypic peptides for each insulin. Individual insulins were first denatured under reducing conditions and the cysteine residues blocked by iodoacetamide. The proteins were then digested with trypsin and the peptide products separated by reversed phase liquid chromatography on an Ascentis® Express ES-C18 column and detected by positive polarity ESI-MS/MS. The digestion peptides were characterized using a multiplexed MRM approach that monitors the fragmentation of the doubly charged unlabeled precursor ion of each peptide into a collection of signature y and b ions. The MRM transitions for the individual peptides were optimized to allow maximal ionization on a standard triple quadrupole mass spectrometer. All products of the ...
Source: Journal of Pharmaceutical and Biomedical Analysis - Category: Drugs & Pharmacology Source Type: research