Investigation and reduction of sub‐microgram peptide loss using molecular weight cut‐off fractionation prior to mass spectrometric analysis

This work investigates the introduction of methanol and a salt modifier to molecular weight cut‐off membrane‐based centrifugal filters (MWCO) to enrich sub‐microgram peptide quantities. Using a neuropeptide standard, bradykinin, sample loss was reduced over two orders of magnitude with and without undigested protein present. Additionally, a bovine serum albumin tryptic digestion was investigated. Twenty‐seven tryptic peptides were identified from MALDI mass spectra after enriching with methanol while only two tryptic peptides were identified after MWCO separation using H 2 O. The strategy presented here enhances recovery from MWCO separation for sub‐µg peptide samples. Copyright © 2012 John Wiley & Sons, Ltd.