Introduction: B. animalis subsp. lactis HN019 is a commercially available wellcharacterized
probiotic with documented effects on human health, such as the
ability to enhance the immune function and to balance the intestinal microbiome.
Therefore, optimizing the manufacturing process to improve sustainability,
increasing biomass yields and viability, and avoiding animal -derived nutrients
in the medium to meet vegan consumer’s needs, is currently of interest. Besides
the established use of live probiotic cells, alternative supplements indicated as
postbiotics, like non-viable cells and/or probiotics derived bioactive molecules
might be considered as potential next generation biotherapeutics. In fact,
advantages of postbiotics include fewer technological limitations, such as
easier production processes and scale-up, and even higher specificity.
Methods: In this work, medium design together with different fermentation
strategies such as batch, fed-batch and in situ product removal on lab-scale
bioreactors were combined. Medium pretreatment by ultrafiltration and protease
digestion was performed to reduce polysaccharidic contaminants and facilitate
the purification of secreted exopolysaccharides (EPS). The latter were isolated
from the fermentation broth and characterized through NMR, GC-MS and SECTDA
analyses. The expression of TLR-4, NF-kb and IL-6 in LPS challenged
differentiated CaCo-2 cells treated with EPS, live and heat-killed B. lactis cells/
broth, was evaluated in vitro by western blotting and ELISA. Zonulin was also
assessed by immunofluorescence assays.
Results and Discussion: The titer of viable B. lactis HN019was increased up to 2.9 ±
0.1 x 1010 on an animal-free semidefined medium by applying an ISPR fermentation
strategy. Medium pre-treatment and a simple downstream procedure enriched the
representativity of the EPS recovered (87%), the composition of which revealed the
presence of mannuronic acid among other sugars typically present in
polysaccharides produced by bifidobacteria. The isolated EPS, live cells and whole
heat inactivated broth were compared for the first up to date for their immunomodulatory and anti-inflammatory properties and for their ability to promote
intestinal barrier integrity. Interestingly, EPS and live cells samples demonstrated
immune-stimulating properties by downregulating the expression of TLR-4 and
NF-kb, and the ability to promote restoring the integrity of the intestinal barrier by
up-regulating the expression of zonulin, one of the tight junctions forming proteins.
Postbiotics in the form of heat killed broth only reduced NF-kb expression, whereas
they did not seem effective in the other tested conditions.

Title

Frontiers in Bioengineering and Biotechnology

Publisher

Frontiers

Publisher Country

Switzerland

Publication Type

Prtinted only

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Year

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