Biochar enhances nitrogen use efficiency in lettuce by promoting its metabolic assimilation.

Background and aims: Peat replacement with biochar (BC) offers a sustainable strategy in horticultural substrates but its effects on plant nitrogen (N) metabolism and N use efficiency (NUE) remain unclear. This study tested whether vineyard-pruning-derived BC can boost NUE and metabolic activity in lettuce, providing a pathway toward more productive and sustainable horticulture. Methods: Plant substrates (BC, peat and vermiculite) were prepared in the following proportions (v:v:v): B0 (0:70:30), B15 (15:55:30) and B30 (30:40:30) for growing lettuce (Lactuca sativa L. var. Batavia) under greenhouse conditions for 31 days. We assessed plant growth and physiological traits, quantified N species and calculated NUE parameters and the activities of key N assimilation enzymes. Results: B30 plants produced 44.2% more biomass and 23.2% larger leaf area than B0, resulting in lower specific leaf area and greater succulence. BC addition decreased available NO₃⁻ and NH₄⁺ in substrate and roots without causing any plant stress symptoms, as chlorophyll content and PSII efficiency remained stable. B30 increased N uptake flux, N utilization efficiency, partial N balance, and N productivity by 31.8%, 34.8%, 27.8%, and 13.8%, respectively, relative to B0, coinciding with enhanced N-assimilation enzymatic activity. Despite lower total N in roots and shoots, protein accumulation increased, indicating more efficient N conversion into organic compounds. Conclusion: These findings demonstrate the potential of BC-based substrates (especially 30% BC) to enhance lettuce productivity by improving NUE through the stimulation of N assimilation pathway, offering a promising strategy to optimize N-fertilizer needs to support more sustainable agriculture and soil management practices. Supplementary Information: The online version contains supplementary material available at 10.1007/s11104-025-07997-0.