The objective of this work was to statistically optimize the cultural and nutritional parameters for L-ascorbic acid by the submerged fermentation of Xanthomonas campestris MTCC 2286 using sucrose as the sole carbon source. After studying the potential parameters that influence the bioprocess by “one variable at a time” approach, we found that the Sucrose, yeast extract, and Dipotassium phosphate (K2HPO4) were the most critical parameter for the production of L-ascorbic acid. Sucrose, a cheap carbon source supports the biomass, yeast extract, an enriched nitrogen source, and K2HPO4 acts as a stimulator for growth and production. The central composite design (CCD) of the RSM was employed to evaluate the interactive effects of these three variables. This is the first report on L-ascorbic acid production by Xanthomonas campestris using statistical experimental design and RSM in submerged fermentation with sucrose as the sole source of carbon. In this study, the production and optimization of L-ascorbic acid from Xanthomonas campestris MTCC 2286 was investigated using optimized nutritional and cultural conditions in a medium containing sucrose as the carbon source in fully aerobic batch fermentation (180 rpm).Using statistical design and optimization approach, the L-ascorbic acid production was enhanced to 135.5mg/Las compared to the fermentation using non-optimized medium (95 mg/L).