Food by-products as substrates for the design of a folate bio-enriched fermented beverage: evaluation of bioavailability and effects on vitamin D receptor (VDR) and folate transporters gene expression in vitro and in vivo

• Ano de defesa: 2024
• Autor(a) principal: Cucick, Ana Clara Candelaria
• Tipo de documento: Tese
• Tipo de acesso: Acesso aberto
• Idioma: eng
• Instituição de defesa: Biblioteca Digitais de Teses e Dissertações da USP
• Palavras-chave em Português: Bactérias lácticas; Bioavailability; Biodisponibilidade; Fermentação; Fermentation; Folate; Folato; Food by-products; Lactic acid bacteria; Probiotic; Probiótico; Receptor de vitamina D (VDR); Subprodutos alimentares; Vitamin D receptor (VDR)
• Link de acesso: https://www.teses.usp.br/teses/disponiveis/112/112131/tde-19022025-160228/

Folate (vitamin B9) is an essential micronutrient that acts in cellular maintenance and metabolic processes. Rising concerns over folate deficiency, the risks of excessive synthetic folic acid intake, and the environmental impact of food waste highlight the need for sustainable solutions. One promising approach is folate bio-enrichment through fermentation with folate-producing lactic acid bacteria (LAB) and/or probiotic strains. Food by-products, such as whey and fruit residues, are rich sources of nutrients that might support LAB growth and folate production. Emerging research suggests a link between vitamin D and its receptor (VDR) in folate metabolism, yet their interplay remains insufficiently understood. The present study aimed to explore agro-industrial residues, including whey and fruit by-products, as substrates for fermentation, using selected LAB and/or probiotic folate-producing strains, to obtain a folate bio-enriched fermented beverage and to study the bioavailability of vitamin B9 produced, in vitro and in vivo. In addition, the effect of this beverage on folate transporters expression reduced folate carrier (rfc), and proton-coupled folate transporter (pcft) and in vdr expression in colon cells was evaluated in vitro and in vivo. Among 5 commercial starter and probiotic strains tested, 3 were selected based on their potential to produce folate in folate-free medium (FACM): Lacticaseibacillus (L.) rhamnosus GG, Bifidobacterium (B.) longum subsp. infantis BB-02, and Streptococcus (S.) thermophilus TH-4. When tested in co-culture, L. rhamnosus LGG consumed the folate produced by S.thermophilus, demonstrating to be a folate-efficient strain, while S. thermophilus TH-4 and B. infantis BB-02 were folate-overproducing bacteria and were selected for the next phase. The selection of cultivation conditions, such as pH, substrates and combination of strains, with grape by-product water extract demonstrating the most substantial effect on folate production, was crucial for successfully obtaining a bio-fortified fermented whey-based beverage (BFWB), containing 407 ng/mL of folate. The safety of the BFWB and of the grape by-product was demonstrated by cytotoxicity and microbiology analysis. In the next phase, the effect of the BFWB obtained on the gene expression of the vdr and folate transporters genes in colon cells was assessed, employing both in vitro and in vivo approaches. Human colon cancer cell line HCT-116 was treated with the beverage and controls, and mRNA expression levels of vdr, rfc, and pcft were measured through qPCR. For the in vivo study, a prevention model was conducted with 30 BALB/c mice fed on a folate-depleted diet for 21 days and receiving the beverage or controls to evaluate whether the BFWB, with or without added vitamin D3 (2000 IU), could prevent folate deficiency. The mice folate status was analyzed at the end of experiment, alongside mRNA expression of vdr, rfc, and pcft genes in colonic tissue. To our knowledge, it was demonstrated for the first time that a folate-deficient diet downregulated vdr expression in the colon of mice. In the cell model, the BFWB significantly upregulated the expression of the genes vdr, rfc, and pcft (p < 0.05). However, in vivo observations were more tenuous; increased vdr expression was only seen in mice receiving BFWB supplemented with vitamin D3. Regarding bioavailability, findings suggested that certain factors related to the dairy matrix of the BFWB, particularly its high ash content (2%), may have hindered folate absorption. This study underscores the potential of valorizing food by-products and safely reintegrating them into the food chain to create functional foods. This approach not only boosts nutritional value but also reduces the environmental impact associated with improper disposal