Production of primary metabolites by Pseudomonas reptilivora B-6bs at the flask level using a full factorial experimental design

The metabolites gluconic acid, 5-ketogluconic acid, proline, and glutamic acid, produced by Pseudomonas reptilivora B-6bs, are industrially important, particularly in food and pharmaceutical sectors. However, producing these metabolites involves biotin supplementation to enhance yields, which is an expensive additive, and reducing its use can significantly lower production costs. Thus, This study aimed to enhance the production of gluconic acid, 5-ketogluconic acid, proline, and glutamic acid without biotin supplementation. To achieve this, a full factorial design was employed, varying agitation speed, glucose concentration, and temperature to determine the optimal conditions for metabolite production. Metabolite concentration was measured using spectrophotometric analysis and thin-layer chromatography (TLC), and the results were statistically analyzed using Minitab^® 18. The findings demonstrate that Pseudomonas reptilivora B-6bs effectively produce gluconic acid (50.51 ± 0.035 g/L, Y_P/S: 0.917 g/g) and 5-ketogluconic acid (44.46 ± 0.23 g/L, Y_P/S: 0.947 g/g), along with proline (0.1727 ± 0.00085 g/L, Y_P/S: 0.00004 g/g) and glutamic acid (0.853 ± 0.142 g/L, Y_P/S: 0.013 g/g) without biotin supplementation. Optimal production was observed with a glucose concentration of 55 g/L. These findings provide a viable biotin-independent strategy for high-value metabolite production. This study contributes novel insights into cost-effective production processes, making it relevant to industrial applications.