Mechanistic Study of 3-Methoxy-5-hydroxy-2-(3-methyl-2-butenyl) bibenzyl from Pigeon Pea Against MRSA and Biofilms: A Highly Promising Natural Inhibitor

Aims The study aimed to investigate the anti-MRSA and anti-biofilm activity of 3-methoxy-5-hydroxy-2-(3-methyl-2-butenyl) bibenzyl (DELC) isolated from Cajanus cajan (L.) Millsp (pigeon pea) leaves, while evaluating its biocompatibility and exploring its potential as an antibiotic alternative. Methods and Results DELC exhibited potent anti-MRSA activity, with a MIC of 3.12 μg/mL and MBC of 6.25 μg/mL. At 2×MIC, it achieved complete eradication of 10⁶ CFU/mL MRSA within 120 minutes, demonstrating faster bactericidal action than vancomycin. Antibiofilm studies:2×MIC DELC inhibited biofilm formation as effectively as 4×MIC vancomycin. In mature biofilms, merely1/2×MIC DELC penetrated and eliminated a portion of embedded viable bacteria—a feat unattainable even with 8×MIC vancomycin. Mechanistic studies: DELC induced oxidative stress, generating ROS that damaged the bacterial membrane, causing nucleic acid leakage. It also suppressed α-hemolysin secretion. RNA-seq analysis revealed: At 1×MIC, DELC blocked ATP synthesis and repair systems, leading to metabolic inhibition and virulence attenuation. At ≥2×MIC, it directly lysed bacterial cells. Biocompatibility: DELC showed no hemolysis or cytotoxicity, indicating excellent safety. Conclusions DELC demonstrates potent anti-MRSA activity characterized by rapid bactericidal effects, a concentration-dependent dual mechanism - metabolic inhibition at low concentrations progressing to direct cellular lysis at higher concentrations, and superior antibiofilm capacity. Its outstanding biocompatibility strongly supports its clinical translation potential.