Biotechnological production of probiotics

Lactillobacillus spp are microorganisms that dominate the vaginal flora of healthy women. Their probiotic function consists in maintaining an environment that limits the growth of pathogens, through the production of organic acids and hydrogen peroxide. Recent studies have found that some strains have an antiviral activity. Thus, it would be advantageous to develop a lactillobacillus that can be added to food stuffs or drugs having a long shelf-life, for the treatment and prevention of infections.

BioTekNet is capable of metabolically characterising and optimising the production of strains such as L. crispatus and L. plantarum and to evaluate the secretion of hydrogen peroxide, lactic acid and secondary metabolites, such as polysaccharides, responsible for bacterial adhesion to mucosal surfaces. We can also:

  • determine the ideal conditions for the conservation of the probiotic biomass in a lyophilised form;
  • determine the best growing conditions for selected strains and assess their production of organic acids from various sources of nitrogen and carbon.

One of the biggest hindrances to the production of lactillobacilli is the production of lactic acid. This inhibits their metabolism in a negative feedback loop, causing the ‘lactate effect’. It is therefore difficult to obtain high yields of this bacterium with traditional fermentation processes. Moreover, the maximum biomass that can be produced is determined not only by substrate usage but also by the reduction in pH produced by the generation of lactic acid. Even when the pH of the medium is kept optimal, the lactate effect inhibits growth. At BioTekNet, we have overcome these problems by developing a bioreactor that microfiltrates the lactic acid from the medium. This innovative bioreactor allows high yields to be reached and reduces the costs and time for the whole process. Moreover, it permits collection of the spent fermentation medium, allowing efficient extraction of the lactic acid. 

Lactillobacilli are kept and sold in a liquid, frozen, lyophilised or dried form. The processes needed to produce these forms can reduce the viability of the bacteria and, hence, negatively affect the functioning and organoleptic properties of the final product. We have therefore studied molecules, such as glycerol, saccharose, lactose, trehalose, dextrine and solutes, in order to identify cryoprotective agents compatible with lyophilisation and that might eliminate, or at least reduce, the damage caused by this process. 

We have assessed the ability of selected strains to grow on many types of carbohydrate and yield a high biomass. Our bioreactor increased the maximum cell density five-fold with respect to control conditions. Moreover, strains grow well on renewable resources, such as dairy waste and malt extract. The best cryoprotective agents ? in terms of increased viability after osmotic and thermal shock and extension of shelf-life ? were found to be trehalose, dextrin and ectoine.

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