Growth and exopolysaccharide production during free and immobilized cell chemostat culture of Lactobacillus rhamnosus RW-9595M

Abstract

Aims: Biomass and exopolysaccharide (EPS) production were studied during chemostat cultures in whey permeate medium with Lactobacillus rhamnosus RW-9595M-free cells and cells immobilized on solid porous supports (ImmobaSil).

Methods and results: A continuous culture with free cells was conducted for 9 days at dilution rates (D) between 0.3 and 0.8 h(-1) in yeast extract (YE)/mineral supplemented whey permeate. Maximum EPS production (1808 mg l(-1)) and volumetric productivity (542.6 mg l(-1) h(-1)) were obtained for a low D of 0.3 h(-1). A continuous fermentation in a two-stage bioreactor system, composed of a first stage with immobilized cells and a second stage inoculated with free cells produced in the first reactor, was carried out for 32 days. The influence of YE concentration, temperature and dilution rate, and their interactions on biomass, EPS and lactic acid production was investigated. A statistically significant model was found only for lactic acid production. Marked cell morphological and physiological changes led to the formation of very large cell-containing aggregates and a low mean soluble EPS production (138 mg l(-1)). Aggregate volumetric productivity of the two-stage system varied between 5.7 and 49.5 g l(-1) h(-1) for different fermentation conditions and times. Aggregates contained a very high biomass concentration, estimated at 74% of aggregate dry weight by nitrogen analysis and 4.3 x 10(12) CFU g(-1) by a DNA extraction method and a high nonsoluble polysaccharide content (14.2%). At age 24 days, insoluble EPS concentration and volumetric productivity were 1250 mg l(-1) and 2240 mg l(-1) h(-1) respectively. The physiological changes were shown to be reversible when cells were incubated during three successive batch cultures.

Conclusions: EPS production and volumetric productivity during continuous free-cell chemostat cultures with L. rhamnosus RW-9595M are among the highest values reported for lactobacilli in literature. Immobilization and continuous culture resulted in low soluble EPS production and large morphological and physiological changes of L. rhamnosus RW-9595M, with formation of macroscopical aggregates mainly composed of biomass and nonsoluble EPS.

Significance and impact of the study: This is the first study on continuous EPS production by immobilized LAB. Immobilization and culture time-induced cell aggregation and could be used to produce new synbiotic products with very high viable cell and EPS concentrations.