Combining anxiolytic activity of thymol and trans-ferulic acid: a potential GABAergic intervention

Trans-ferulic acid (TFA), a phenolic acid abundant in fruits and cereals, and thymol (THY), a monoterpenoid phenol from thyme oils, are bioactive natural compounds with notable neuropharmacological potential. This study evaluated the combined anxiolytic effects of THY and TFA in Swiss albino mice, focusing on GABAergic mechanisms and pharmacokinetic profiles through in vivo and in silico methods. Mice received vehicle, diazepam (DZP, 2 mg/kg), TFA (50 mg/kg), THY (50 mg/kg), or their combination (THY + TFA; 50 + 50 mg/kg, p.o.). Behavioral assessments in open-field, swing, hole-cross, and light-dark tests indicated that the combined treatment produced more pronounced effects, markedly reducing locomotor and exploratory activity (e.g., square crosses: 25.8 ± 3.7 vs. 94.6 ± 6.2 in control; hole crosses: 3.4 ± 1.8 vs. 15.6 ± 1.9) and increasing time in the light chamber, consistent with anxiolytic-like activity comparable to DZP. Molecular docking targeted gamma-aminobutyric acid A (GABA_A) receptor subunits (α2, α3), and ADME/toxicity profiles were analyzed in silico. Docking analysis revealed moderate binding affinities (BAs) to GABA_A receptor subunits AF-P26048-F1 (α2 subunit) (- 5.8 kcal/mol) and 8G4X (α3 subunit) (- 5.3 kcal/mol) for TFA and α2 (- 4.7 kcal/mol) and α3 (- 5.6 kcal/mol) for THY, relative to DZP (- 6.9 kcal/mol). These findings suggest that the THY + TFA combination elicits enhanced GABAergic modulation, offering a potential natural therapeutic strategy for anxiety management with reduced toxicity risk. However, formal interaction analyses (e.g., isobologram or two-way ANOVA) are warranted to confirm the combining effect.