2,3-Butanediol plus acetoin obtention by <i>Enterobacter aerogenes</i> ATCC 13048: inhibition by target products and cells reuse during fed-batch cultivation

2,3-Butanediol (2,3-BD) is a highly valued building block, and optimizing its production by fermentation, particularly with crude glycerol, is crucial. <i>Enterobacter aerogenes</i> is a key microorganism for this process; however, there are limited studies addressing the inhibition effects of products and by-products on 2,3-BD production. This study investigates these inhibition effects to maximize 2,3-BD production. Final concentrations of 2,3-BD plus acetoin reached 89.3, 92.7, and 71.1 g.L^-1 with productivities of 1.22, 1.69, and 0.99 g.L^-1.h^-1 in pure glycerol, glucose, and crude glycerol media, respectively. Acetic acid was the main by-product, with concentrations ranging from 10 to 15 g.L^-1. The reinoculation of <i>E. aerogenes</i> cells highlighted the strong effect of 2,3-BD and acetic acid on microbial growth and metabolism, with the cultivation environment exerting selective pressure. Notably, cells reuse enhanced performance in crude glycerol media, achieving a specific productivity in relation to biomass (Y_P/X) of 9.18 g.g^-1; about 25% higher than in fed-batch without cells reuse. By combining results from two fed-batch cycles, the total final concentration of 2,3-BD plus acetoin reached 99.4 g.L^-1, alongside a 33% reduction in total acetic acid production with reused cells.