Background: Due to increasing domestic and industrial demand, edible oil production is not keeping up with demand. To fill this gap, the productivity of oilseeds can be increased by applying adequate nutrients, particularly sulphur (S), at the crucial growth stage. Purpose: The present study aims to explore the best concentration of S for its foliar application on various cultivars of mustard. Methods: A factorial randomized pot experiment was conducted to investigate the role of leaf-applied S on growth, physiobiochemistry, yield and quality traits of three cultivars of Brassica juncea L. (mustard). Five levels of S viz. 0 (water), 15, 30, 45 and 60 ppm S constituted one variant, and the three cultivars (Chutki, Nath Sona and Rohini) were the other variants. The various levels of S were sprayed at 50 and 70 days after sowing (DAS). The growth and physio-biochemical characteristics were studied at 90 DAS, and yield and quality attributes at 120 DAS (harvest). Results: The data indicated that increasing S levels up to 45 ppm S improved all parameters of mustard and thereafter (at the level above 45 ppm S) decreased. Cultivar Nath Sona, followed by Rohini and Chutki, performed best. Among the foliar spray treatment of different levels of S, the application of 45 ppm S increased plant dry weight by 40.21, 35.65 and 30.96%, photosynthetic rate by 28.27, 27.44 and 36.29%, pods of a plant by 15.23, 12.12 and 10.80%, seed yield of a plant by 7.54, 3.89 and 4.91%, oil content by 48.70, 46.31 and 43.15% and oil yield of a plant by 24.56, 23.93 and 22.35% in cultivar Nath Sona, Rohini and Chutki, respectively, compared with their respective water-treated plants. Moreover, the oil was examined by GC-MS technique for its various components. The analysis revealed that there were 36 compounds in the oil of the non-treated plants and 44 compounds in the oil of plants treated with 45 ppm S. The extra compounds resulted from the application of 45 ppm S. Conclusion: It may be concluded that two sprays of 45 ppm S proved effective in improving the growth, physio-biochemical characteristics, yield and quality of cultivars of mustard, particularly Nath Sona.