Bioanalytical Studies in the Brain of Wistar Rats (Rattus norvegicus) Exposed to 2,3-Butanedione

2,3-Butanedione is widely used as a flavoring, preservative, and fragrance additive, and increasing evidence suggests that chronic exposure may induce molecular alterations in biological systems. This exploratory study investigated the impact of prolonged 2,3-butanedione exposure on the brain proteomic profile of Wistar rats. Thirty-three animals were assigned to three groups: a vehicle-treated control and two exposed groups receiving 46.6 or 99.1 µg kg–1 day–1 via intragastric administration for 90 days. Following euthanasia under deep anesthesia, brain tissues were analyzed by nano-liquid chromatography-mass spectrometry (nano-LC-MS), and the resulting data were evaluated using chemometric approaches. Data processing in Skyline identified 9,296 proteins, of which 2,500 were retained for multivariate analysis. Clear discrimination among experimental groups was observed, even in the absence of sex stratification, indicating that exposure influenced the global brain proteomic profile. Ontological and pathway analyses revealed 78 affected protein classes, with several dysregulated proteins annotated to biological pathways previously implicated in neurological processes. Importantly, these proteomic alterations represent pathway-level associations and do not constitute evidence of neurological disease or pathology. Overall, the findings demonstrate that chronic exposure to 2,3-butanedione modulates brain proteomic patterns and associated molecular pathways, supporting its use as a hypothesis-generating framework for future mechanistic and functional studies.