Approche intégrée couplant procédé microbiologique et procédé séparatif in situ pour la production d’acide 3-hydroxypropionique par Lactobacillus reuteri

Abstract : Within the development of bioeconomy, there is an increasing drive towards the production of chemicals from renewable biomass-derived feedstock. In this context, interest in the sustainable production of biobased building blocks, such as the bifunctional 3-hydroxypropionic acid (3-HP), is growing. Among its various applications, the 3-HP can be used for the synthesis of biobased polymers and acrylic acid. The development of a biotechnological process to produce this platform chemical is a key issue. Lactobacillus reuteri is known as a natural producer of 3-HP from glycerol (by-product of biodiesel manufacturing industries) although at low yield and productivity, due to the inhibitory effects of the product and of its metabolic intermediate 3-hydroxypropionaldehyde (3-HPA), as well as the formation of 1,3-propanediol (1,3-PDO) as co-product. The integration of an in situ 3-HP extraction process to the bioconversion system could lead to a decrease in inhibition phenomena and to the continuous recovery of the targeted molecule. To this aim, the reactive liquid-liquid extraction assisted by a membrane contactor was considered in the present study. This technique shows a high specific surface area, unrivaled when compared to usual dispersive processes, without creating emulsion and limits the direct contact between bacteria and the organic phase. First, three L. reuteri strains, representative of the intra-species diversity, were studied in flasks and compared in terms of growth performance on glucose and of 3-HP production using glycerol as substrate. The DSM 17938 strain, a good glycerol consumer and 3-HP producer, was then selected for the project. During bioconversion, a total loss in cultivability concomitant with a decrease of the cellular enzymatic activity was observed. Flow cytometry analyses revealed a great impact of 3-HPA and, to a lesser extent, of 3-HP only in acidic conditions. To optimize 3-HP production performances and further understand its metabolic determinants, bioconversion was set up in bioreactor with a controlled feeding in glycerol (fed-batch mode). Concerning the 3-HP extraction, the use of a mixed extractants (tri-octyl-amine and Aliquat 336 in n-decanol) showed high performances over a wide range of experimental conditions (pH, 3-HP concentration), highlighting a synergistic effect. 3-HP was predominantly extracted over all the molecules of the bioconversion medium, confirming the selectivity of the extraction process studied. Nevertheless, when extraction was performed on a real bioconversion broth, some cell-originating compounds were suspected to adsorb to the interface between the two phases near the pores of the membrane. This effect, combined to the possible competition between 3-HP and other compounds able to react with the extractants, such as salts, led to limitations in the extraction performances and selectivity. To our best knowledge, this work is the first one dedicated to the study of 3-HP extractive bioconversion. However, this new integrated process currently displays low performances (productivity, final product concentration, production duration) due to bacterial inhibition by the bioconversion metabolites and the extractant organic phase. Several optimization tracks have been suggested to overcome the identified locks.
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Submitted on : Monday, May 7, 2018 - 5:23:59 PM
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Grégoire Burgé. Approche intégrée couplant procédé microbiologique et procédé séparatif in situ pour la production d’acide 3-hydroxypropionique par Lactobacillus reuteri. Sciences du Vivant [q-bio]. Ecole doctorale ABIES (Agriculture, Alimentation, Biologie, Environnement, Santé), 2015. Français. ⟨tel-01787709⟩

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