La défense publique de la thèse de Madame Tiffani BOUANATI aura lieu le 11 mars à 15h30 par vidéo-conférence.

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Promoteur: Monsieur Pascal Gerbaux Co-promoteur: Monsieur Jean-Marie Raquez

Résumé de la dissertation

Algae biomass is nowadays considered as a unique reservoir of a huge variety of different molecules, such as lipids, carbohydrates, proteins and secondary metabolites, awaiting development in various fields. The ALGOTECH project has been setup with the objectives of developing and optimizing the production of microalgal biomass for energy or non-energy applications such as the production of biodiesel, bioethanol and biomethane or biobased polymers, respectively. Therefore, the study of algal biomass, with a special attention paid to the chemical profiling of the polysaccharides, proteins, lipids, pigments, is mandatory prior to the valorization of those biomolecules into products with high added value. This PhD thesis participates to the valorization of the non-energetic fraction of the macro and micro-algal biomass with a special attention paid to the polysaccharide fractions. In this project, we will focus on Arthrospira sp. PCC 8005, which are cyanobacteria that produce sulfated polysaccharides, named calcium spirulan. Sulfated oligosaccharides have been reported to present specific biological properties such as bactericide, biostimulant… properties. We also study two macroalgae, Kappaphycus alvarezii and Eucheuma spinosum, which contain carrageenan, that also are sulfated polysaccharides. The use of macroalgae is motivated by the huge availability of such a biomass and therefore these macroalgae are used in the present project to develop strategies of production and characterization of sulfated (oligo)saccharides.

The first part of the study focused on the cultivation of macroalgae and microalgae. In the case of microalgae, we implemented a pilot reactor at the lab scale to produce microalgae in high quantity under specific conditions. Doing so, we were able to modify the culture conditions to modulate the biomass composition of microalgae and therefore to produce biomolecules of interest in large quantities. In this work, we observed the large production of lipids and polysaccharides under nitrogen starvation. The second part of the work concerned the extraction and purification of polysaccharides from macroalgae and microalgae, as well as their depolymerization into oligosaccharides using a green hydrolysis approach while conserving the active sulfate functions. Beside the composition of the polysaccharides in terms of monosaccharide nature and combination thereof, the carbohydrate molecular weight, in other words the degree of polymerization (DP), has been demonstrated to strongly affect the final properties of the polysaccharides. Oligosaccharides may present enhanced or reduced biological activities when compared to polysaccharides. Only few oligosaccharides are naturally produced and therefore, many chemical-based strategies have been devised to prepare them, either by synthetic routes or by depolymerization reactions. Alternative solutions for oligosaccharide preparation are mandatory and recently the polysaccharide hydrolysis assisted by microwave activation, that is classified as physical methods, is gaining importance. The main advantage of microwave activation over conventional heating is to produce an efficient and homogeneous thermal energy source for shorter reaction times, reducing the undesirable formation of by-products by thermal decomposition due to prolonged heating. In this work, hydrolysis conditions were successfully optimized for each family of the studied polysaccharides and the reproducibility of the method was demonstrated.

In the final part, the study of the biological properties of the extracted polysaccharides and the corresponding oligosaccharides was conducted. The first study concerned the antibacterial activity of the saccharides by evaluation of the survival rate of Escherichia coli (human bacteria), Bacillus subtilis (terrestrial bacteria) and Cobetia marina (marine bacteria) in presence of these saccharides. These bacteria were selected based on their intrinsic structural differences, Bacillus subtilis being gram-positive bacteria whereas Escherichia coli and Cobetia marina are gram-negative bacteria. The second study will focus on the impact of saccharides on the growth of Fragaria ananassa « Korona », a strawberry plant of commercial relevance in the Walloon Region. The strawberry Fragaria ananassa « Korona » was selected as the model plant because, unlike some crop plants such as potatoes whose entire production is harvested, only the strawberry fruits are harvested in this plant. It is therefore interesting to take care of this plant by ensuring its growth and health. An antibacterial effect against Escherichia coli was observed in presence of polysaccharides extracted from Kappaphycus alvarezii and from Eucheuma spinosum, as well as when the corresponding oligosaccharides are tested. The data reveal that a high degree of sulfation and/or a high molecular weight promotes such an antibacterial effect. An antagonistic effect has been observed with Cobetia marina in presence of the same compounds and also in presence of hydroxymethyl furfural and spirulan. In this case, we noticed that a high degree of sulfation and a low molecular weight promote the growth of Cobetia marina. Spirulan also promotes the growth of Cobetia marina and it may be interesting to investigate enzymes produced by Cobetia marina to find which ones are able to depolymerize spirulan. Finally, for biological tests carried out on Bacillus subtilis, no effect was highlighted. The same observations were made for the growth impact assessment in presence of macroalgae compounds.