The Effects of Baicalin in Combination with Cefotaxime on the Biofilm and Metabolic Reprogramming of Multidrug-Resistant Pseudomonas aeruginosa

Baicalin, a natural plant-derived compound, holds promise in addressing clinical bacterial resistance when combined with antibiotics. This study evaluated the antibacterial activity of the combination of baicalin and cefotaxime and explored its mechanism of action on the cell wall and biofilm of multidrug-resistant Pseudomonas aeruginosa (MRPA). The results showed that the combination of baicalin and cefotaxime exerted a synergistic inhibitory effect on the growth of MRPA, with a fractional inhibitory concentration index (FICI) of 0.28. Mechanistically, compared with cefotaxime alone, the combination of baicalin and cefotaxime enhanced the permeability of the cell membrane and cell wall of MRPA, thereby increasing cell damage. It also exhibited stronger antibiofilm activity by inhibiting numerous virulence factors (pyocyanin, elastase, lectin), reducing cellular metabolic activity, and downregulating the expression of biofilm genes (pslA, pelA, algD) and quorum-sensing genes (lasl, lasR, rhll, rhlR, pqsA, pqsR). The molecular docking results revealed that baicalin could stably bind to wbpE, LasR, and RhlR. Therefore, this interaction may indirectly influence the processes related to antibiotic resistance and biofilm formation in bacterial cells. Metabolomic analysis revealed that the combination of baicalin and cefotaxime upregulated 863 metabolites and downregulated 587 metabolites. These metabolites mainly included amino acids, lipids, nucleotides, carbohydrates, and secondary metabolites. The combination primarily enriched key pathways such as amino acid metabolism, lipid metabolism (sphingolipid metabolism) and secondary metabolite biosynthesis. Through these pathways, it triggers significant metabolic reprogramming, thereby interfering with the supply of cell wall synthesis precursors, membrane structural stability, and the generation of biomembrane matrix. Ultimately, it synergistically enhances the effects of cell wall damage and biomembrane inhibition. In conclusion, this study confirms that the combination of baicalin and cefotaxime exerts significant synergistic antibacterial activity against MRPA. It also reveals the mechanism of action of the combination on the cell wall and biofilm of MRPA at the metabolic level, providing theoretical support for the development of novel strategies to combat MRPA.