Abstarct: Abstract Salinity stress is one of agricultural production's most significant limiting factors, adversely affecting plant growth and yield. Due to its medicinal and nutritional properties, fenugreek holds great importance, but its growth and performance decline under salinity stress. Dopamine, an organic compound from the catecholamine family, acts as an elicitor and plays a crucial role in regulating plants' physiological and biochemical responses to environmental stresses. This study aimed to investigate the effects of different dopamine concentrations (0, 100, and 200 µM) on the physiological, biochemical, and molecular responses of fenugreek under salinity stress conditions (0, 150, and 300 mM). Salinity had multiple negative impacts on the physiological and biochemical traits of fenugreek. It reduced relative water content, photosynthetic pigments (chlorophyll and carotenoids), endogenous auxin content, and potassium levels, while increasing electrolyte leakage, malondialdehyde, hydrogen peroxide, nitric oxide, sodium, chloride, and the sodium-to-potassium ratio. The results of this experiment showed that dopamine application significantly mitigated the adverse effects of salinity. Dopamine enhanced relative water content, photosynthetic pigments, total protein, flavonoids, phenols, and potassium levels. Additionally, the content of antioxidant enzymes (catalase, ascorbate peroxidase, superoxide dismutase, and guaiacol peroxidase) and abscisic acid increased under salinity stress and dopamine application. Gene exxpression analysis revealed that salinity upregulated most genes except for the C7 gene. Furthermore, dopamine application up to a concentration of 200 µM increased the exxpression of all genes and the endogenous diosgenin content. Overall, this study demonstrated that salinity negatively affects the physiological and biochemical characteristics of fenugreek. However, dopamine application, particularly at 200 µM, can serve as an effective stimulant in reducing these effects and improving plant tolerance to salinity. This study highlights that dopamine, by regulating defense mechanisms and enhancing physiological and biochemical traits, can be a sustainable strategy for managing salinity stress in agricultural systems.