FUNCTIONAL PROPERTIES OF EXOPOLYSACCHARIDE (EPS) EXTRACT FROM L. FERMENTUM LF2 AND ITS ANALYSIS WHEN COMBINED WITH B. ANIMALIS INL1 IN YOGURT

PERALTA, G.; REINHEIMER J.A.; ALE, E.; BURNS, P.; ÁVILA, O.; CONTINI, L.; BINETTI, A.

Córdoba

Congreso; VII Congreso Internacional de Ciencia y Tecnología de Alimentos; 2018

Lactobacillus fermentum Lf2 is an autochthonous strain which was isolated as non-starter culture from a local semi-hard cheese. It produces high amounts of EPS when it grows under controlled conditions of temperature and pH, reaching ~1 g/L. In the present study, the functional roles of an EPS extract from L. fermentum Lf2 were studied by means of in vitro and in vivo assays, isolated or combined with an autochthonous probiotic strain, Bifidobacterium animalis subsp. lactis INL1, and both applied as food ingredients in yogurt. First, an in vitro analysis was done with THP-1 cell line. Macrophages derived from THP-1 were stimulated with crude or purified EPS, 60 μg/mL and 12.6 μg/mL, respectively. This last concentration was proposed taking into account that, after purification of the crude EPS extract, 21% (approximately) is recovered. A positive control (treated with lipopolysaccharide, 0.5 μg/mL), and negative control (untreated cells) were included. The cells were incubated for 4 h (37 °C, 5% CO2) for the detection of TNF-α, and 8 h for IL-6 and IL-10 determinations. The cytokine analysis was performed from the culture supernatants using the DuoSet ELISA kits (R&D Systems). Besides, an in vivo assay was done, in which BALB/c mice (7/group) received yogurt with EPS (600 mg/L, YE), bifidobacteria (5x108 CFU/mL, YB), a combination of both ingredients (YEB), or yogurt with no additives (Y, control group), for 25 days. During treatment, different bacterial groups (Lactobacillus, Bifidobacterium, B. animalis, B. bifidum, B. breve, B. catenulatum, C. leptum, C. coccoides, Staphylococcus, Enterobacteriaceae, Streptococcus, general microbial load with universal primers) and short chain fatty acids (SCFA) were determined in faeces by qPCR and HPLC, respectively (at 0, 8, 18 and 25 days). Furthermore, IgA levels were quantified in intestinal fluid, and IL-10, IFN-γ, IL-6 and TNF-α cytokines were measured in small intestine by ELISA. Histological analyses were carried out in order to evaluate the morphology of small and large intestines. The group YE presented increased concentrations of acetic, butyric and total SCFA (the sum of acetic, butyric and propionic acids) at the end of the treatment, and increasing levels of the C. coccoides cluster (SCFA producing group) over time (p< 0.05). The YEB group presented a possible symbiotic role reflected mainly in the levels of the species B. animalis at all times evaluated. These results highlight the potential application of the EPS of L. fermentum Lf2 as a functional ingredient, individually or together with B. animalis subsp. lactis INL1.