## 3 Results
### Fed-Batch Fermentation for Lycopene Production  
Based on the results above, the fermentation medium was modified as follows: 3 g/L yeast extract, 400 mg/L proline and 2 g/L glycerol (medium A) or 2 g/L glucose (medium B), MgSO\({}_{4}\) 7H\({}_{2}\)O 0.5 g/L, NH\({}_{4}\)Cl 0.5 g/L, NaCl 0.5 g/L, K\({}_{2}\)HPO\({}_{4}\) 0.5 g/L, pH 7.0. The lycopene-producing capacity for the \(\Delta\)_crtC_ mutant strains was evaluated in a 500 mL fed-batch cultivation under aerobic dark conditions. The results show that, during the initial stage following the inoculation of the \(\Delta\)_crtC_ mutant strain to medium A and the subsequent two rounds of fed-batch cultivation, both the growth of the \(\Delta\)_crtC_ mutant strain and the production of lycopene continued to increase, while the intracellular lycopene content remained relatively stable. During the two batches of fermentation period, there was an increasing trend toward bacterial proliferation and lycopene accumulation, which confirmed the feasibility of lycopene production using fed-batch cultivation of the \(\Delta\)_crtC_ mutant strain. However, further increase in the \(OD_{660}\) and lycopene yield was not observed in the third batch of fermentation. After three rounds of fed-batch cultivation, the yield of lycopene reached 151.10 \(\pm\) 0.13 mg/L (Figure 8A), which is comparable to the yield 150.15 mg/L reported by [PERSON] et al. [14].  
In contrast, the fed-batch cultivation of the \(\Delta\)_crtC_ mutant strain proved to be less effective in medium B with glucose as a carbon source. In fact, active growth and lycopene accumulation merely occurred in the initial stage, with the second stage receiving the first round of fresh medium addition. The subsequent addition of fresh medium did not lead to a further increase in either bacterial biomass or lycopene production. This was because the fermentation of glucose by this bacterium led to the progressive acidification of the medium B until the lowest value of pH 6.0 was reached, creating an environment condition unfavorable for bacterial growth. At the end of the three rounds of fed-batch cultivation, the maximum lycopene yield harvested was 66.11 \(\pm\) 0.12 mg/L, which was far below that obtained in the medium A (Figure 8B). Therefore, glycerol can be used as a carbon source that is superior to glucose, in that it is a non-ferentative substrate, precluding the possibility of medium acidification during lycopene production using the \(\Delta\)_crtC_ mutant  
Figure 8: Lycopene production in fed-batch fermentation. Initial medium: 3 g/L yeast extract, 400 mg/L proline and 2 g/L glycerol (**A**), 2 g/L glucose (**B**), MgSO\({}_{4}\) 7H\({}_{2}\)O 0.5 g/L, NH\({}_{4}\)Cl 0.5 g/L, NaCl 0.5 g/L, K\({}_{2}\)HPO\({}_{4}\) 0.5 g/L, pH 7.0. The arrowheads indicated that addition of fresh medium at 4 N, 8 N and 16 N concentrations, respectively. The data with different lower-case letters in the figures are statistically different in the content of lycopene (\(p\) < 0.05).  
strain. As a by-product during oleochemical and biodiesel fuel production, it can be used as a more cost-effective, sustainable and environmentally friendly substrate for industrial applications in the future [13].