The Optimization of the Nutrient Medium Composition for the Submerged Cultivation of the Mycolicibacterium neoaurum Strain VKM Ac-3067D in a 100 L Bioreactor Under Controlled Conditions by Mathematical Planning

The biotechnological production of carotenoids offers a promising alternative to their chemical synthesis or extraction from plants. Mycolicibacterium species have shown potential as pigment-producing microorganisms. However, bacterial strains typically exhibit lower productivity compared to fungal and yeast strains. Earlier, we enhanced the β-carotene biosynthesis in M. neoaurum strain VKM Ac-3067D by modifying the cultivation medium. Key changes included replacing glucose with glycerol and soybean meal with skimmed milk powder (SMP) and increasing the urea content from 0.5 to 1.0 g/L. To further optimize β-carotene yield, a steepest ascent method was applied combining factorial design with a gradient-based optimization (Wilson–Box method). The resulting regression model showed that the most influential factors were the glycerol concentration and SPM use. The in-flask fermentation of the Ac-3067D strain in a medium containing 25.5 g/L of glycerol (carbon source) and 12.80 g/L of SMP (nitrogen source) increased β-carotene yield to 318.4 ± 8.3 mg/kg. In a 15 L bioreactor, β-carotene yield increased to 432.3 ± 10.4 mg/kg, while the biomass concentration reached 23.2 ± 1.2 g/L. The further scaling up to a 100 L bioreactor increased both β-carotene yield (450.4 ± 8.2 mg/kg) and biomass concentration (25.2 ± 1.1 g/L). Thus, β-carotene production technology using the M. neoaurum strain AC-3067D was successfully scaled up from 750 mL flasks to a 100 L bioreactor, confirming its potential for industrial-scale application.