Probiotic microorganisms are increasingly exploited for their capacity to generate bioactive metabolites with potential health-promoting properties. In this study, we investigated the in vitro antioxidant activity of sixteen probiotic bacteria and characterized the extracellular metabolomic profiles of the spent fermentation broths (cell-free supernatants) derived from two strains showing markedly different antioxidant behaviors. Antioxidant potential was quantified using the DPPH radical scavenging assay, and total phenolic content was assessed by the Folin-Ciocalteu method. Among the tested strains, Lactiplantibacillus plantarum 8VEG3C displayed the most favorable antioxidant profile within the tested panel, with the lowest IC₅₀ value (2.47% v/v) and the highest phenolic content, whereas Lacticaseibacillus paracasei LC01 showed a substantially weaker antioxidant activity. To explore the molecular basis of these functional differences, untargeted extracellular metabolomic analysis was performed by liquid chromatography-tandem mass spectrometry (LC-MS/MS), coupled with multivariate statistical analysis and putative, database-supported annotation. The two strains generated clearly distinguishable extracellular signatures. The L. plantarum 8VEG3C supernatant was enriched in fatty acyl-related metabolites, including oleic acid, 7-oxooctadecanoic acid and γ-dodecalactone, together with aromatic amino acid-derived compounds such as indole lactate. In contrast, L. paracasei LC01 was enriched in amino acid- and small peptide-related metabolites including the putatively annotated 2-(6-hydroxyhexanoylamino)-3-phenylpropanoic acid, compatible with a more prominent involvement of the proteolytic system of lactic acid bacteria. These findings support the use of untargeted metabolomics as a functional screening platform for the rational selection of probiotic strains with postbiotic potential.

The intestinal barrier is a complex and dynamic structure that plays a critical role in maintaining gut health. Though, pathological conditions such as, dysbiosis or inflammation that could damage the intestinal barrier to varying degrees, aggravating the primary disease. Recently, fermented plant-origin foods and probiotics were used as an alternative for health remedy. For this reason, in this current study, we made the inquiries about the effect of Tunisian zgougou juice obtained upon both spontaneous (J) and driven fermentation (J(A1+A2)) with autochthonous probiotic bacteria Lactiplantibacillus plantarum (LP) species A1 and A2, the whole bacteria (A1) and (A2) and their CFS. The cell integrity and permeability was assessed based on the TEER measurement on monolayer cell culture of Caco-2 cell, then the gene expression levels of the tight junction proteins (TJP) and the gene expression level of the inflammatory cytokine TNFα were evaluated using the RT-PCR. Our experimental findings exhibited that the driven fermented had a potential protective effects on intestinal barrier function in vitro shown by the great increase the electrical barrier detection (TEER) between Caco2 cell reaching ca. 690 to 963.33 Ω/cm2, although the regulatory mechanism has not been reported yet. Otherwise LP (A1 and A2) and their CFS showed moderate results. Furthermore, J(A1+A2) promoted the mRNA relative gene expression of the different tested tight TJPs and moderated the intestinal inflammation by influencing the TNFα expression. This study revealed that probiotics and especially probiotic-based plant-origin fermented functional foods (Tunisian zgougou juice) could improve and reinforce the intestinal barrier which provide new ideas for treating intestinal injury-related diseases, and have to be a revival in worldwide popularity, mostly as a consequence of claims made about their health benefits. So that, to completely reveal the health advantages of these probiotic LAB strains for humans and this new food matrix, additional in vivo and clinical investigations are necessary, as highlighted by the limitations of in vitro models.

The intestinal barrier is a critical regulator of gut health, often compromised by dysbiosis and inflammation. Given the rising interest in fermented plant‐based foods and probiotics as adjuvants in maintaining gut health, this study explored the biological activity of zgougou juice, comparing spontaneous fermentation (J) with driven fermentation using two lactobacilli with probiotic traits (namely Lactiplantibacillus paraplantarum A1 and Lactiplantibacillus plantarum A2), as well as the effects of cell biomass and Cell‐Free Supernatants (CFS) obtained from those lactobacilli. Using a Caco‐2 cell monolayer model, we assessed barrier integrity via Transepithelial Electrical Resistance (TEER) and gene expression of Tight Junction Proteins (TJP) and the cytokine Tumor Necrosis Factor α (TNFα). Treatment of Caco‐2 monolayer with zgougou juice fermented with lactobacilli significantly reinforced the intestinal barrier, with TEER values reaching 690–963.33 Ω·cm ² . Furthermore, it upregulated TJP expression, outperforming bacterial biomass and their CFS. These findings would suggest that zgougou juice improves intestinal barrier, though further in vivo and clinical trials are necessary to claim any benefits for human health.

This study evaluated the microbiological, nutritional, sensory, and in vitro biological properties of probiotic fermented milks enriched with 5% Prickly Pear Seed (PPS) flour. Different formulations were tested: plain fermented milk (Control), control with adjunct lactobacilli with probiotic traits (Lactiplantibacillus plantarum LPAL; Lacticaseibacillus casei BGP 93; Lacticaseibacillus rhamnosus LR B; Lacticaseibacillus paracasei LC4 P1), fermented milk with PPS, and fermented milk with PPS and adjunct lactobacilli. PPS flour did not affect the starters or the adjunct lactobacilli viability. Throughout 40 days of storage, probiotic levels remained above the minimum threshold required for health benefits. PPS supplementation increased the in vitro antioxidant activity of the fermented milk formulations only when combined with some of the adjunct lactobacilli. The formulation containing PPS and L. casei BGP 93 (PPS_BGP93) exhibited superior antioxidant performance. In HCT8 cell lines, PPS and PPS_BGP93 reduced Reactive Oxygen Species levels. PPS_BGP93 also upregulated IL-10 gene expression, although this response was not consistent with that found in the presence of LPS after 40 days of storage. PPS and PPS_BGP93 displayed the highest mineral levels and total fiber (around 3%), allowing the nutrition claims to be a source of phosphorus, magnesium, and fiber. A panel test (0–9 scores) revealed that although mouthfeel was impaired by the enrichment of PPS, the PPS-enriched formulations achieved scores around 6 for overall acceptability.