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Accelerated Tryptic Digestion for the Analysis of Biopharmaceutical Monoclonal Antibodies in Plasma by Liquid Chromatography with Tandem Mass Spectrometric Detection by Antoine Lesur; Emmanuel Varesio; Gérard Hopfgartner is a Chemistry article available to read on EtoBox.

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Accelerated tryptic digestion of a therapeutic protein including microwave irradiation and thermal transfer by convection at 60 • C and 37 • C was investigated. An analytical setup was devised to follow the protein digestion rate using 1D gel electrophoresis and liquid chromatography coupled a triple quadrupole linear ion trap mass spectrometer. The formation kinetic of its tryptic peptides was monitored in the selected monitoring mode (LC-SRM/MS). Different digestion end points (e.g. 2, 5, 10, 15, 30 and 60 min) as well as an overnight digestion were tested using a therapeutic human monoclonal antibody (mAb) with the goal of its LC-SRM/MS quantification in human plasma. The peptides from the human mAb were generated at different rates and were classified into three categories: (1) the fast forming peptides, (2) the slow forming peptides and (3) the peptides degrading over time. For many monitored peptides, a heating temperature of 37 • C with a 750 rpm mixing applied for at least 30 min provided equivalent results to microwave-assisted digestion and generally allowed the achievement of an equivalent peptide concentration as an overnight digestion carried out at 37 • C. The disappe

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Author

Antoine Lesur; Emmanuel Varesio; Gérard Hopfgartner

Publisher

Elsevier Science; Elsevier ; Elsevier BV (ISSN 1873-3778)

Published

2010

Language

EN

Field

Chemistry (Physical Sciences)