Increased chemical acetylation of peptides and proteins in rats after daily ingestion of diacetyl analyzed by Nano-Lc-MS/MS

Abstract

Background: Acetylation alters several protein properties including molecular weight, stability, enzymatic activity, protein-protein interactions, and other biological functions. Our previous findings demonstrating that diacetyl/peroxynitrite can acetylate L-lysine, L-histidine, and albumin in vitro led us to investigate whether diacetyl-treated rats suffer protein acetylation as well. Methods: Wistar rats were administered diacetyl daily for 4 weeks, after which they were sacrificed, and their lung proteins were extracted to be analysed by Nano-LC-MS/MS (Q-TOF). A C18 reversed-phase column and gradient elution with formic acid/acetonitrile solutions from 2 to 50% over 150 min were used to separate the proteins. Protein detection was performed using a microTOF-Q II (QTOF) equipped with captive source and an electrospray-ionization source. The data from mass spectrometry were processed using a Compass 1.7 and analyzed using Protein Scape, software that uses Mascot algorithms to perform protein searches. Results: A set of 3162 acetylated peptides derived from 351 acetylated proteins in the diacetyl-treated group was identified. Among them, 23 targeted proteins were significantly more acetylated in the diacetyl-treated group than in the PBS control. Protein acetylation of the group treated with 540 mg/kg/day of diacetyl was corroborated by Western blotting analysis. Conclusions: These data support our hypothesis that diacetyl exposure in animals may lead to the generation of acetyl radicals, compounds that attach to proteins, affecting their functions and triggering adverse health problems.

Effects of Diacetyl Flavoring Exposure in Mice Metabolism

“…We previously demonstrated the generation of acetyl radicals from the reaction between diacetyl and peroxynitrite [ 12 – 16 ] and also demonstrated the increase in protein acetylation in vivo after diacetyl consumption [ 17 ]. However, the broader effects of this flavoring compound have not been clearly studied.…”