The present study aims at comparing the performances of three Lactobacillus reuteri strains (DSM 20016, DSM 17938, and ATCC 53608) in producing 3-hydroxypropionic acid (3-HP) from glycerol and at exploring inhibition phenomena during this bioconversion. Differences were highlighted between the three strains in terms of 3-HP production yield, kinetics of substrate consumption, and metabolite production. With a maximal productivity in non-optimal conditions (free pH) around 2 g.L(-1).h(-1) of 3-HP and 4 g.L(-1).h(-1) of 3-hydroxypropionaldehyde (3-HPA) depending on the strain, this study confirmed the potential of L. reuteri for the biotechnological production of 3-HP. Moreover, the molar ratios of 3-HP to 1,3-propanediol (1,3-PDO) obtained for the three strains (comprised between 1.25 and 1.65) showed systematically a higher 3-HP production. From these results, the DSM 17938 strain appeared to be the most promising strain. The impact of glycerol bioconversion on the bacteria's physiological state (a decrease of around 40 % in DSM 17938 cells showing an enzymatic activity after 3 h) and survival (total loss of cultivability after 2 or 3 h depending on the strains) was revealed and discussed. The effect of each metabolite on L. reuteri DSM 17938 was further investigated, displaying a drastic inhibition caused by 3-HPA, while 3-HP induced lower impact and only at acidic pH.

BACKGROUND: Bioconversion is a promising route to produce bio-based building blocks such as 3-hydroxypropionic acid (3-HP). Reactive extraction can be used for 3-HP recovery, and ultimately integrated to the bioconversion process. To the best of our knowledge, there is no published experimental data about the reactive extraction of 3-HP. This work aimed to study the extraction of 3-HP using tri-n-octylamine and Aliquat 336 as extractants in n-decanol. Comparison was also made with its positional isomer, lactic acid. Finally, the extraction of 3-HP from model and real bioconversion broths was examined.

BACKGROUND: Reactive liquid-liquid extraction is a promising technique for the direct recovery of carboxylic acids from bioconversion media. This work focused on the optimization of 3-hydroxypropionic acid (3-HP) reactive extraction assisted by hollow-fiber membrane contactor (HFMC), using tri-n-octylamine (TOA) and Aliquat 336 as extractants in n-decanol, and on its practical application for the recovery of 3-HP obtained from glycerol bioconversion by Lactobacillus reuteri.

The aims of this study is to compare the growth and glucose metabolism of three Lactobacillus reuteri strains (i.e. DSM 20016, DSM 17938, and ATCC 53608) which are lactic acid bacteria of interest used for diverse applications such as probiotics implying the production of biomass, or for the production of valuable chemicals (3-hydroxypropionaldehyde, 3-hydroxypropionic acid, 1,3-propanediol). However, the physiological diversity inside the species, even for basic metabolisms, like its capacity of acidification or glucose metabolism, has not been studied yet. In the present work, the growth and metabolism of three strains representative of the species diversity have been studied in batch mode. The strains were compared through characterization of growth kinetics and evaluation of acidification kinetics, substrate consumption and product formation. The results showed significant differences between the three strains which may be explained, at least in part, by variations in the distribution of carbon source between two glycolytic pathways during the bacterial growth: the phosphoketolase or heterolactic pathway (PKP) and the Embden-Meyerhof pathway (EMP). It was also shown that, in the context of obtaining a large amount of biomass, DSM 20016 and DSM 17938 strains were the most effective in terms of growth kinetics. The DSM 17938 strain, which shows the more significant metabolic shift from EMP to PKP when the pH decreases, is more effective for lactate production.

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BACKGROUND: 3-hydroxypropionic acid (3-HP) microbial synthesis through glycerol bioconversion byLactobacillus reuteri is at the moment characterized by too low performances to consider production at industrial scale. To avoid its toxic accumulation in the medium and to recover this molecule of interest, 3-HP in situ reactive extraction from bioconversion broth was investigated using a hollow fiber membrane contactor (HFMC) in order to intensify its production. The so-called integrated system was compared with the conventional bioconversion system. The impact of the extractive bioconversion on the overall production performance and on cell physiological state was studied. RESULTS: Results underlined drastic inhibitory effects on the producing bacteria, especially under extractive bioconversion conditions despite the use of a HFMC supposed to avoid direct contact between organic phase and bacteria. Indeed, the extractant phase components (trioctylamine in n-decanol) were found to be toxic for the cells (due to solubility and by direct contact). These phenomena were increased by the presence of 3-hydroxypropionaldehyde (3-HPA) and 3-HP produced during glycerol bioconversion. These cumulative effects induced complete loss of the cell membrane integrity and esterase activity after 1.5 h of extractive bioconversion. When the bioconversion was conducted alone, the bacterial inhibition was lower, as around 50% of L. reuteri cells remained active with unaltered membrane after 3 h. Hypotheses concerning the mechanisms of action of the observed inhibitions were proposed and discussed.CONCLUSIONS: This work highlighted the importance of performing such an integrative approach to address specific questions prior to the optimization of the whole process.

HPLC-based quantification of 3-HPA using a synthetic acrolein-free 3-HPA standard obtained from commercially available 1,2,4-butanetriol through a straightforward and easy synthetic process.

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