An evaluation of the utility of in vacuo methylation for mass‐spectrometry‐based analyses of peptides

Abstract

In vacuo trimethylation of the N-terminus of a lyophilized peptide with methyl iodide was previously reported to enhance the peptide's signal in matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) and to suppress alkali adduct formation in electrospray ionization mass spectrometry (ESI-MS). Both the signal enhancement and alkali adduct suppression observed for methylated peptides are believed to be due to the permanent positive charge on the N-terminus of the peptide resulting from the formation of a quaternary ammonium moiety. The present work evaluates the general utility of the in vacuo methylation procedure for the MS analysis of peptides, and specifically addresses the issue of whether the methylation of nucleophilic sites other than the N-terminal amine affects the MALDI signal of modified peptides. This work establishes that, although certain side-chain modifications are inevitable using present reaction conditions, the derivatization leads to significant MALDI-MS signal improvement. The experimental results demonstrate that the N-terminal trimethylammonium derivatives of peptides exhibit MALDI signals comparable to or exceeding those of arginine-containing standards such as angiotensin I. The advantages and limitations of the in vacuo methylation procedure are discussed.

Pronounced Solvent Effect on the Hydrostannylation of Propargylic Alcohol Derivatives with nBu3SnH/Et3B at Room Temperature

“…This report describes the utility of the In Vacuo I sotope C oded A lkylation T echnique (IVICAT) as an isotopic N‐terminal label for quantitative proteomic analysis. The ability of H 9 and D 9 N‐terminal trimethylammonium‐derivatized peptides to enhance the detection signal by MALDI‐MS, as previously shown,30, 32 is demonstrated. Furthermore, we describe the fragmentation characteristics of IVICAT‐labeled peptides in collision‐induced dissociation (CID) ESI tandem mass spectrometry (MS/MS) modes.…”

“…Recently, the in vacuo trimethylation of peptides with methyl iodide was shown to be a promising approach in labeling of all peptides at their N‐termini for MS 30–32. It was demonstrated that the permanent positive charge induced by the quaternary methylated ammonium group enhanced the detection signal of the labeled versus the non‐labeled peptides in MALDI‐MS 30.…”

1-(3-Aminopropyl)-3-butylimidazolium bromide for carboxyl group derivatization: potential applications in high sensitivity peptide identification by mass spectrometry

“…Currently, mass spectrometry (MS) based detection techniques have emerged as indispensable tools for peptide analysis [11]. To increase the ionization efficiency of pep-tides (especially for those with low abundance and poor ionization efficiency) by MS, a variety of chemical derivatization reagents [12] that target labeling of the amino group [13][14][15], thiol group [16][17][18] and hydroxyl group [19,20] have been developed. Smith et al applied the aldehydebased derivatization reagent hexanal and butanal for the alkylation of primary amines in peptides to provide an increase in ionization efficiency and concomitant signal enhancement.…”

Recent Advancement of Chemical Derivatization and Its Applications to High Sensitive Analysis of Peptide in Mass Spectrometry

“…Moreover, nucleophilic side chain modifications could occur, disturbing subsequent MS identification. Choma et al [25] further evaluated the general utility of iodomethane derivatization for MS-based peptide identification. With the present reaction conditions, although side chain modifications were inevitable, the signal intensity of peptides, especially for those lacking lysine and arginine residues, could be improved via MALDI-TOF MS.…”