TMT-based quantitative proteomics and non-targeted metabolomics reveal the antibacterial mechanism of rhein against Bacillus cereus and its potential application as a food preservative

The study aims to investigate the antibacterial effect and mechanism of rhein against Bacillus cereus using Tandem Mass Tag (TMT) quantitative proteomics combined with non-targeted metabolomics. The results showed that rhein exhibited strong antimicrobial effect against B. cereus , with a minimum inhibitory concentration (MIC) and a minimum bactericidal concentration (MBC) of 1.2 μg/mL and 2.4 μg/mL, respectively. Additionally, rhein treatment increased membrane permeability, induced reactive oxygen species generation, and disrupted the cell morphology of B. cereus cells. Notably, rhein could effectively inhibit the proliferation of B. cereus in cooked rice and skimmed milk during the storage at 4 °C. Proteomics and metabolomics analyses identified 288 differentially expressed proteins (DEPs) and 402 differential metabolites (DMs) in rhein-treated B . cereus . These molecules were involved in multiple biological pathways related to primary metabolism, carbon metabolism, amino acid biosynthesis and energy production, as well as the synthesis of secondary metabolites and stress response-related signaling pathways. Bioinformatics analysis further revealed that rhein disrupted key cellular processes, including amino acid metabolism, nucleotide metabolism, glycolysis/gluconeogenesis and flagellar structure. These disruptions significantly impaired bacterial metabolic functions and motility, ultimately inhibiting bacterial growth and leading to cell death. This research provides valuable understanding into the mechanism of rhein's action against B . cereus and lays theoretical foundations for its potential use as an antibacterial agent in the food industry. • Rhein presented strong antibacterial activity against B. cereus with a MIC value of 1.2 μg/mL. • Rhein disrupted B. cereus cell membrane and metabolism, impairing bacterial growth and survival. • Rhein showed potential as a natural antibacterial agent for food preservation.