<p>Vitamins exhibit biostimulant potential to enhance soybean grain yield by promoting plant growth and/or mitigating abiotic stress. Therefore, this study aimed to evaluate the effects of foliar application of thiamine and nicotinamide on soybean growth and yield traits under two different cultivation environments. The experiments were conducted at the Mato Grosso do Sul State University (UEMS) in Cassilândia, MS, and at the Federal University of Mato Grosso do Sul (UFMS) in Chapadão do Sul, MS. A randomized block design was used, with five treatments and four replicates. The treatments included a control, individual applications of thiamine and nicotinamide at 100 mg L^− 1 of water, and combined applications of both vitamins at 50 and 100 mg L^− 1 concentrations. The vitamins were applied exogenously at the V6 growth stage. Gas exchange parameters, grain yield, and its components were evaluated. Regarding gas exchange, vitamin applications resulted in an average increase of 11.42% for the net photosynthetic rate. For yield components, based on the average across both cultivation sites, the combined application of thiamine and nicotinamide at 50 mg L^− 1 led to increases of 17.43%, 14.89%, 7.04%, 11.24%, and 15.13% in the number of grains and pods per plant, grains per pod, 1,000-grain weight, and grain yield, respectively. Vitamin application influenced physiological behavior related to gas exchange activity, yield components, and grain yield in soybean plants across different environments, with the combined application of thiamine and nicotinamide at 50 mg L^− 1 showing the most promising results.</p>