Background <p>Salinity stress, as one of the most significant and widespread abiotic stresses, affects a large portion of the world’s arable lands. Developing salt-tolerant plants through classical breeding, biotechnology, and molecular methods is time-consuming and expensive. Considerable research attention has been directed toward the application of exogenous biostimulants as potential strategies to mitigate the adverse effects of salinity. The current research sought to investigate the impacts of varying silymarin (0, 250, and 500 µM) and serotonin (0, 50, and 100 µM) concentrations on the physiological, biochemical, and molecular responses of fenugreek under salt stress conditions (200 mM).</p> Results <p>Salinity-induced oxidative stress adversely affected physiological parameters, including photosynthetic pigments, K⁺ concentration, relative water content, and auxin levels. Conversely, it triggered an upregulation in abscisic acid and nitric oxide levels, along with increased activity of both enzymatic and non-enzymatic antioxidants, accumulation of compatible osmolytes, and elevated cellular Na⁺ content. The enhancement of fenugreek tolerance to salt stress by these two compounds was associated with their ability to modulate key physiological and biochemical pathways. These findings are likely attributable to the reduction in Na⁺ content, as well as decreased hydrogen peroxide concentrations, which contribute to maintaining membrane integrity and cellular turgor, and reducing lipid peroxidation and ELI. Moreover, these included the regulation of photosynthetic pigment biosynthesis, abscisic acid and auxin levels, and the signaling molecules, including nitric oxide and hydrogen peroxide. Furthermore, the utilization of these two elicitors significantly enhanced the expression of genes associated with the diosgenin biosynthesis pathway, resulting in increased diosgenin content.</p> Conclusions <p>Our results lay the groundwork for further investigation into the molecular mechanisms of plant defense mediated by these compounds, providing valuable perspectives on novel approaches to crop cultivation that leverage these two plant growth regulators.</p>