title: High-Efficiency Microflow and Nanoflow Negative Electrospray Ionization of Peptides Induced by Gas-Phase Proton Transfer Reactions.
abstract: Liquid chromatography coupled with electrospray ionization mass spectrometry (ESI-MS) is routinely used in proteomics research. Mass spectrometry-based peptide analysis is performed de facto in positive-ion mode, except for the analysis of some post-translationally modified peptides (e.g., phosphorylation and glycosylation). Collected mass spectrometry data after peptide negative ionization analysis is scarce, because of a lack of negatively charged amino acid side-chain residues that would enable efficient ionization (i.e., on average, every 10th amino acid residue is negatively charged). Also, several phenomena linked to negative ionization, such as corona discharge, arcing, and electrospray destabilization, because of the presence of polar mobile-phase solutions or acidic mobile-phase additives (e.g., formic or trifluoroacetic acid), reduce its use. Named phenomena influence microflow and nanoflow electrospray ionization (ESI) of peptides in a way that prevents the formation of negatively charged peptide ions. In this work, we have investigated the effects of post-column addition of isopropanol solutions of formaldehyde, 2,2-dimethylpropanal, ethyl methanoate, and 2-phenyl-2-oxoethanal as the negative-ion-mode mobile-phase modifiers for the analysis of peptides. According to the obtained data, all four modifiers exhibited significant enhancement of peptide negative ionization, while ethyl methanoate showed the best results. The proposed mechanism of action of the modifiers includes proton transfer reactions through oxonium ion formation. In this way, mobile phase protons are prevented from interfering with the process of negative ionization. To the best of our knowledge, this is the first study that describes the use and reaction mechanism of aforementioned modifiers for enhancement of peptide negative ionization.
authors.affiliation: Vinča Institute of Nuclear Sciences, University of Belgrade , Belgrade, Serbia.
authors.forename: Marija
authors.initials: M
authors.lastname: Nišavić
authors.affiliation: Ruđer Bošković Institute, Bijenička cesta 54, Zagreb, Croatia.
authors.forename: Amela
authors.initials: A
authors.lastname: Hozić
authors.affiliation: Ruđer Bošković Institute, Bijenička cesta 54, Zagreb, Croatia.
authors.forename: Zdenko
authors.initials: Z
authors.lastname: Hameršak
authors.affiliation: Ruđer Bošković Institute, Bijenička cesta 54, Zagreb, Croatia.
authors.forename: Martina
authors.initials: M
authors.lastname: Radić
authors.affiliation: BIOCentre, Central Lab Services, Zagreb, Croatia.
authors.forename: Ana
authors.initials: A
authors.lastname: Butorac
authors.affiliation: Faculty of Agriculture, University of Zagreb , Zagreb, Croatia.
authors.forename: Marija
authors.initials: M
authors.lastname: Duvnjak
authors.affiliation: Ruđer Bošković Institute, Bijenička cesta 54, Zagreb, Croatia.
authors.forename: Mario
authors.identifier: 0000-0002-6545-2871
authors.initials: M
authors.lastname: Cindrić
mesh_terms: D000447:Aldehydes / Q000737:chemistry; D000804:Angiotensin II / Q000032:analysis / Q000737:chemistry; D000818:Animals; D002417:Cattle; D002853:Chromatography, Liquid; D005557:Formaldehyde / Q000737:chemistry; D005563:Formic Acid Esters / Q000737:chemistry; D010446:Peptide Fragments / Q000032:analysis* / Q000737:chemistry*; D011522:Protons; D012710:Serum Albumin, Bovine / Q000032:analysis / Q000737:chemistry; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods
chemical_list: D000447:Aldehydes; D005563:Formic Acid Esters; D010446:Peptide Fragments; D011522:Protons; C510888:ethyl formate; D000804:Angiotensin II; D005557:Formaldehyde; D012710:Serum Albumin, Bovine; C001058:pivaldehyde