An update for human blood plasma pretreatment for optimized recovery of low-molecular-mass peptides prior to CE–MS...

Protein precipitation and centrifugal filtration are well-established methods for concentrating and purifying peptides with a low relative molecular mass (Mr) from human blood plasma before proteomic and peptidomic studies using high-performance separation techniques, but there is little information on peptide recoveries. Here, we evaluate acetonitrile precipitation followed by a range of centrifugal filtration conditions for the analysis of low Mr peptides in human blood plasma before CE–MS and SPE coupled online to CE–MS. Three opioid peptides were used as model compounds, that is, dynorphin A 1–7, endomorphin 1, and methionine enkephalin and 3, 10, and 30 K Mr cut-off cellulose acetate filters (Amicon® Ultra-0.5) and 10 K Mr cut-off polyethersulfone filters (Vivaspin® 500) were studied. Unexpectedly, recoveries and repeatability were only optimum after passivating the 10 K Mr cut-off cellulose acetate filters with PEG to avoid peptide adsorption on the inner walls of the plastic sample reservoir.