Formic acid and Acetic acid for peptide LC-MS

ABSTRACT: Despite the general acceptance of formic acid as the additive of choice for peptide reversed-phase LC-MS/MS applications, some still argue that the selection of acetic acid represents a better option. To settle this debate, we investigated both the difference in MS sensitivity and chromatographic behaviour of peptides between these two systems. This interlaboratory study was performed using different MS setups and C18 separation media employing both 0.1% formic and 0.5% acetic acid as ion pairing modifier. Relative to formic acid, we find an overall ~2.2-2.5x increase in MS signal and slight decrease in RP LC retention (-0.7% acetonitrile on average) for acetic acid conditions. While these two features have opposing effects on peptide detectability, we find that acetic acid produces up to 60% higher peptide ID output depending on the type of sample. The drop in RPLC retention increases with peptide net charge at acidic pH. MS signal is dependent on the difference between charge of the precursor ion and charge of peptide in solution, favoring species with low pI. Lower peptide retention under acetic acid conditions demonstrates its higher hydrophilicity and as expected, leads to composition and sequence-dependent character of the observed retention shift. A custom version of SSRCalc retention prediction model has been developed to accommodate these charges, with formic and acetic acid models showing identical results independent of the origin of data collection.

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Oleg Krokhin 

PROVIDER: PXD029416 | JPOST Repository | Sat Dec 10 00:00:00 GMT 2022

REPOSITORIES: jPOST