Production d'exopolysaccharides par fermentation avec des cellules immobilisées de LB. Rhamnosus RW-9595M d'un milieu à base de perméat de lactosérum

Bergmaier, Dirk
Philosophiæ doctor (Ph.D.)
Université Laval
Doctorat en sciences et technologie des aliments

Direction de recherche : Lacroix, Christophe

Codirection de recherche : Champagne, Claude-P.

2002-11

Dédicace

Für meine Großeltern, Eltern und Schwester,deren Nähe mir oft fehlte,meine Familie in Québec,die mich mit offenen Armen empfang,Sophie und Victor,die ich von Herzen liebe.

Table des matières

Introduction générale
Chapitre 1 Revue de littérature
Chapitre 2 New method for exopolysaccharide determination in culture broth using stirred ultrafiltration cells
Chapitre 3 Exopolysaccharide production during batch cultures with free and immobilized Lactobacillus rhamnosus RW-9595M
Chapitre 4 Exopolysaccharide production during chemostat cultures with free and immobilized Lactobacillus rhamnosus RW-9595M
Chapitre 5 Aggregate formation during chemostat culture with immobilized Lactobacillus rhamnosus RW-9595M
Conclusion générale
Bibliographie
Annexe 1 Concentration en ATP pendant une fermentation avec des cellules libres de Lb. rhamnosus RW-9595M
Annexe 2 Cell counts and lactic acid, soluble EPS and immobilized cell concentration during a 32 day continuous culture in a two stage bioreactor system.
Annexe 3 Analysis of variance and parameter estimates for lactic acid concentration in the effluent of the ICB and the FCB.

Liste des illustrations

Fig. 1.1 Structure du dextrane, un homopolymère.
Fig. 1.2 Structure de l'EPS de Lb. rhamnosus RW-9595M (Van Calsteren et al. 2002).
Fig. 1.3 Schéma des voies métaboliques impliquées dans le catabolisme du lactose/galactose et la biosynthèse des EPS par Lactococcus lactis Gal⁺ (via les systèmes phosphoénolpyruvate PEP et phospho-transférase PTS) et par Lactobacillus delbrueckii ssp. bulgaricus et Streptococcus thermophilus Gal^- (via l'antiport lactose/galactose) (De Vuyst et al. 2001).
Fig. 1.4 Modèle de la biosynthèse d'EPS par Lactococcus lactis ssp. lactis NIZO B40 (Van Kranenburg et al. 1998)
Fig. 1.5 Organisation des gènes codant pour la production d'EPS chez Lactococcus lactis NIZO B40 (plasmidique) et chez Streptococcus thermophilus Sfi6 (adapté de De Vuyst et al., 2001).
Fig. 1.6 Classification des systèmes d’immobilisation selon quatre classes (adapté de Willaert et Baron 1996).
Fig. 2.1 Schematic diagram of the laboratory device used for ultrafiltration experiments in stirred cells.
Fig. 2.2 Change in viable cell count (●), lactose (■) and lactic acid (◆) concentrations, and EPS production measured by the UF (▲) and conventional (▼) methods during pH-controlled batch cultures with Lb. rhamnosus RW-9595M in BMM. Means and standard deviations for three fermentations.
Fig. 2.3 Change in viable cell count (●), volume of base consumed (―), and EPS production measured by the UF (▲) and conventional (▼) methods during a pH-controlled batch culture with Lb. rhamnosus RW-9595M in WP. Means and standard deviations for duplicate analyses by the UF method.
Fig. 2.4 Change in viable cell count (●), lactose (■) and lactic acid (◆) concentrations, and EPS production measured by the UF (▲) and conventional (▼) methods during pH-controlled batch cultures with Lb. rhamnosus RW-9595M in SWP. Means and standard deviations for three fermentations.
Fig. 3.1 Viable cell counts (●), lactose (■) and lactic acid (◆) concentrations, and EPS production (▲) during pH-controlled batch cultures with Lb. rhamnosus RW-9595M. Means and standard deviations for two fermentations.
Fig. 3.2 Free-cell counts (●), immobilized cell concentration measured by DNA extraction method (■) and polysaccharide concentration in the broth (vertical bars) during four repeated batch cultures with Lb. rhamnosus RW-9595M immobilized on ImmobaSil^{^®} silicone rubber solid supports.
Fig. 4.1 Viable cell counts (vertical bars), EPS concentration (●), and EPS volumetric (■) and specific productivities (▲) during free-cell chemostat culture with Lb. rhamnosus RW9595M for different dilution rates. Means and standard deviations for two replications of D.
Fig. 4.2 Optical micrograph (10X) of a semi-fine slice (methylene blue staining) of the ImmobaSil^{^®} support colonized with Lb. rhamnosus RW-9595M after 7 day chemostat culture.
Fig. 4.3 Photomicrograph of Lb. rhamnosus RW-9595M: a) in the FCB effluent after 14 days of chemostat culture (63X) b) in the fermented medium at the end of a free-cell batch culture (100X).
Fig. 4.4 Response surface of lactic acid concentration in the first immobilized-cell bioreactor (ICB) for T = 34°C, covariates stop = 0.5 and age = 12 days.
Fig. 4.5 Response surface of lactic acid concentration in the second free-cell bioreactor (FCB) for T = 34°C, covariates stop = 0.5 and age = 12 days.
Fig. 5.1 Scanning electron microscopy of colonized ImmobaSil^{^®} supports: a) pore at the surface (magnification 1600 X, b) and c) inner pores (magnification 3600 X and 12000 X, respectively).
Fig. 5.2 Optical microscopy observations of semi-fine slices of aggregates stained with methylene blue: a) high cell density (20 X), b) and c) formation of channels (63 X).
Fig. 5.3 Scanning electron microscopy of a cut through the aggregate: a) at a magnification of 11000 X, b) at a magnification of 7800 X with crystal.
Fig. 5.4 Change of aggregate dry weight in the fermented broth in the first ICB (solid bars) and the second FCB (dashed bars) bioreactors as a measure of aggregate accumulation during the 32-day continuous culture. Age 1 day corresponds to the start of the experimental design after an 8-day initial stabilization period. Fermentation conditions were changed every day according to the central composite rotatable design.
Fig. 5.5 Change of aggregate dry weight in effluent samples (solid bars) and volumetric productivity of the two-stage continuous culture (dashed bars) during the 32-day experiment. Age 1 day corresponds to the start of the experimental design after an 8-day initial stabilization period. Fermentation conditions were changed every day according to the central composite rotatable design.

Liste des tableaux

Tab. 1.1 Production d'EPS par les LAB.
Tab. 1.2 Avantages et désavantages de la technologie des cellules immobilisées.
Tab. 1.3 Immobilisation par inclusion dans des supports solides poreux.
Tab. 2.1 EPS concentration and recovery rate (R) determined by the UF method for 0.1 M NaCl and 5% WP solutions spiked with EPS.
Tab. 4.1 Real and coded values of the central composite experimental design for the continuous immobilized cell fermentation.


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