To enhance crop yield and quality, plant cultivation in controlled-growing systems is an alternative to traditional open-field farming. The use of light-emitting diode (LED) as an adjustable light source represents a promising approach to improve plant growth, metabolism, and function. The objective of this study was to assess the impact of different light spectra (red, red/blue (3:1), blue, and white) with an emission peak of around 656, 656, 450, and 449 nm, respectively, under various replacement methods of nutrient solution (complete replacement (CR), EC-based replacement (ECBR), and replacing based on plant needs (RBPN)), on biomass, physiological traits, and macroand micronutrient contents of two best-known lettuce varieties, Lollo Rossa (LR) and Lollo Bionda (LB), in the nutrient film technique (NFT) hydroponic system. The results indicated that mix of red and blue LED spectra under RBPN method is the most effective treatment to enhance fresh and dry weights of lettuce plants. In addition, red LED spectrum under RBPN, and red and blue light under ECBR nutrient solution significantly increased leaf stomatal conductance, net photosynthesis and transpiration rate, and intercellular CO2 concentration of LR variety. Phosphorus (P), potassium (K), calcium (Ca), and magnesium (Mn) content in LR variety, and iron (Fe), zinc (Zn), copper (Cu), and manganese (Mn) content in both varieties increased upon exposure to blue and red LED light spectrum with RBPN method. Our results suggest that exposure to combination of red and blue light along with feeding plants using RBPN and ECBR methods can increase absorption of macro- and micronutrient elements and improve photosynthetic properties, and eventually increase lettuce yield with lower nitrate accumulation.