The extensive development in light-emitting diodes (LEDs) in recent years provides an opportunity to positively influence plant growth and biomass accumulation and to optimize biochemical composition and nutritional quality. This study aimed to assess how different light spectra affect the growth, photosynthesis and biochemical properties of Eruca sativa. Therefore two LED lighting modes - red:blue (RB, 1:1) and red:green:blue (RGB, 2:1:2) were compared to the conventional white light fluorescent tubes (WL). Plant biomass, photosynthetic performance, several antioxidants, polyamines and nitrates contents were analyzed across different treatments. The plant growth was affected by the light quality - the presence of green light in the spectrum resulted in smaller plants and leaves, and correspondingly less biomass. RB spectral mode enhanced the total antioxidant and guaiacol peroxidase activity, pigments, flavonoids, polyphenols, ascorbate and polyamines contents. This effect under RB was combined with better leaf development compared to RGB and less nitrate in the leaves among all treatments. The RB light generated modifications in polyamines, which are interrelated with the nitrate content, further induce important metabolite and antioxidant changes. Both RB and RGB enhanced photosynthesis. The afterglow thermoluminescence band varied according to leaves development, being higher in RB and WL as a consequence of their faster growth. The RB light spectrum was found to be the most efficient for promoting the growth, biochemical composition, and overall quality of Eruca sativa compared to RGB and WL. These findings suggest that RB LEDs can be an effective tool for improving crop production in controlled environments.