Microwave-Assisted 18O-Labeling of Proteins Catalyzed by Formic Acid

Oxygen exchange may occur at carboxyl groups catalyzed by acid. The reaction, however, takes at least several days at room temperature. The long-time exchanging reaction often prevents its application from protein analysis. In this study, an O-18-labeling method utilizing microwave-assisted acid hydrolysis was developed. After being dissolved in O-16/O-18 (1:1) water containing 2.5% formic acid, protein samples were exposed to microwave irradiation. LC-MS/MS analysis of the resulted peptide mixtures indicated that oxygen in the carboxyl groups from glutamic acid, aspartic acid, and the C-terminal residues could be efficiently exchanged with O-18 within less than 1.5 min. The rate of back exchange was so slow that no detectable back exchange could be found during the HPLC run.