Development of chitosan-coated agar-gelatin particles for probiotic delivery and targeted release in the gastrointestinal tract

This study reports the development of a novel and simple formulation for probiotic delivery using chitosan-coated agar-gelatin gel particles. This methodology involves the production of agar-gelatin particles by thermally treating a mixture of agar and gelatin solutions at high temperatures (121°C) and subsequently coating with chitosan. The particles were able to protect the probiotic strain Lactobacillus plantarum NCIMB 8826 during incubation for 2 hours in simulated gastric fluid (pH 2) as no statistically significant loss (P > 0.05) in cell concentration was observed, and also resist dissolution in simulated intestinal fluid (pH 7.2). Interestingly, this protection is related to the fact that the intense thermal treatment affected the physicochemical properties of agars, and resulted in the formation of a strong and tight polymer network, as indicated by the X-ray diffraction (XRD) analysis. Using an in vitro faecal batch fermentation model simulating the conditions of the distal part of the large intestine (pH 6.7-6.9), it was demonstrated by quantitative real time PCR that the majority of L. plantarum cells were released from the agar-gelatin particles within 30 to 48 hours. Overall, this work led to the development of a novel methodology for the production of probiotic containing particles which is simpler compared to current encapsulation technologies, and has a lot of potential to be used for the controlled release of probiotics and potentially other solid bioactives in the large intestine

Impact of 2′-fucosyllactose on gut microbiota composition in adults with chronic gastrointestinal conditions: batch culture fermentation model and pilot clinical trial findings

Intestinal dysbiosis has been described in patients with certain gastrointestinal conditions including irritable bowel syndrome (IBS) and ulcerative colitis. 2′-fucosyllactose (2′-FL), a prebiotic human milk oligosaccharide, is considered bifidogenic and butyrogenic. To assess prebiotic effects of 2′-FL, alone or in combination with probiotic strains (potential synbiotics), in vitro experiments were conducted on stool from healthy, IBS, and ulcerative colitis adult donors. In anaerobic batch culture fermenters, Bifidobacterium and Eubacterium rectale-Clostridium coccoides counts, and short-chain fatty acids (SCFAs) including butyrate increased during fermentation with 2′-FL and some of the 2′-FL/probiotic combinations. In a subsequent open-label pilot trial, the effect of a 2′-FL-containing nutritional formula was evaluated in twelve adults with IBS or ulcerative colitis. Gastrointestinal Quality of Life Index (GIQLI) total and gastrointestinal symptoms domain scores, stool counts of Bifidobacterium and Faecalibacterium prausnitzii, and stool SCFAs including butyrate, increased after six weeks of intervention. Consistent with documented effects of 2′-FL, the batch culture fermentation experiments demonstrated bifidogenic and butyrogenic effects of 2′-FL during fermentation with human stool samples. Consumption of the 2′-FL-containing nutritional formula by adults with IBS or ulcerative colitis was associated with improvements in intra- and extra-intestinal symptoms, and bifidogenic and butyrogenic effects

Impact of short-chain galactooligosaccharides on the gut microbiome of lactose-intolerant individuals

Approximately 75% of the global human population are lactose malabsorbers. In a previous clinical trial, it was shown that feeding a high-purity galactooligosaccharide (>95% GOS) could improve symptoms of lactose-intolerant subjects, attaining lactose tolerance in a majority of subjects. To investigate the mechanism, we examined the microbiome of human subjects before and after GOS feeding. The results show a significant shift in the microbiome of responsive individuals, including lactose-fermenting microbes in their stools. The high-purity prebiotic GOS resulted in adaptive shifts in the microbiome and correlated with improvement in clinical symptoms

Raw and sous-vide-cooked red cardoon stalks (Cynara cardunculus L. var. altilis DC): (poly)phenol bioaccessibility, anti-inflammatory activity in the gastrointestinal tract, and prebiotic activity

The in vitro anti-inflammatory and prebiotic activity and the content and profile of bioaccessible (poly)phenols and catabolites of raw and sous-vide-cooked red cardoon (Cynara cardunculus L. var. altilis DC) were investigated during gastrointestinal (GI) digestion. Raw cardoon after in vitro GI digestion had 0.7% bioaccessible (poly)phenols, which protected against lipopolysaccharide-induced inflammation by counteracting IL-8, IL-6, TNF-α, and IL-10 secretions in differentiated Caco-2 cells. Contrarily, GI-digested sous vide cardoon showed higher (poly)phenol bioaccessibility (59.8%) and exerted proinflammatory effects in Caco-2 cells. (Poly)phenols were highly metabolized during the first 8 h of in vitro fermentation, and nine catabolites were produced during 48 h of fermentation. Colonic-fermented raw and sous-vide-cooked cardoon did not show anti-inflammatory activity in HT-29 cells but presented potential prebiotic activity, comparable to the commercial prebiotic FOS, by stimulating health-promoting bacteria such as Bifidobacterium spp. and Lactobacillus/Enterococcus spp. and by increasing the production of total SCFAs, especially acetate

Malnutrition and gut microbiota in children

Malnutrition continues to threaten the lives of millions across the world, with children being hardest hit. Although inadequate access to food and infectious disease are the primary causes of childhood malnutrition, the gut microbiota may also contribute. This review considers the evidence on the role of diet in modifying the gut microbiota, and how the microbiota impacts childhood malnutrition. It is widely understood that the gut microbiota of children is influenced by diet, which, in turn, can impact child nutritional status. Additionally, diarrhoea, a major contributor to malnutrition, is induced by pathogenic elements of the gut microbiota. Diarrhoea leads to malabsorption of essential nutrients and reduced energy availability resulting in weight loss, which can lead to malnutrition. Alterations in gut microbiota of severe acute malnourished (SAM) children include increased Proteobacteria and decreased Bacteroides levels. Additionally, the gut microbiota of SAM children exhibits lower relative diversity compared with healthy children. Thus, the data indicate a link between gut microbiota and malnutrition in children, suggesting that treatment of childhood malnutrition should include measures that support a healthy gut microbiota. This could be of particular relevance in sub-Saharan Africa and Asia where prevalence of malnutrition remains a major threat to the lives of millions

Milk- and solid-feeding practices and daycare attendance are associated with differences in bacterial diversity, predominant communities, and metabolic and immune function of the infant gut microbiome

The development of the infant intestinal microbiome in response to dietary and other exposures may shape long-term metabolic and immune function. We examined differences in the community structure and function of the intestinal microbiome between four feeding groups, exclusively breastfed infants before introduction of solid foods (EBF), non-exclusively breastfed infants before introduction of solid foods (non-EBF), EBF infants after introduction of solid foods (EBF+S), and non-EBF infants after introduction of solid foods (non-EBF+S), and tested whether out-of-home daycare attendance was associated with differences in relative abundance of gut bacteria. Bacterial 16S rRNA amplicon sequencing was performed on 49 stool samples collected longitudinally from a cohort of 9 infants (5 male, 4 female). PICRUSt metabolic inference analysis was used to identify metabolic impacts of feeding practices on the infant gut microbiome. Sequencing data identified significant differences across groups defined by feeding and daycare attendance. Non-EBF and daycare-attending infants had higher diversity and species richness than EBF and non-daycare attending infants. The gut microbiome of EBF infants showed increased proportions of Bifidobacterium and lower abundance of Bacteroidetes and Clostridiales than non-EBF infants. PICRUSt analysis indicated that introduction of solid foods had a marginal impact on the microbiome of EBF infants (24 enzymes overrepresented in EBF+S infants). In contrast, over 200 bacterial gene categories were overrepresented in non-EBF+S compared to non-EBF infants including several bacterial methyl-accepting chemotaxis proteins (MCP) involved in signal transduction. The identified differences between EBF and non-EBF infants suggest that breast milk may provide the gut microbiome with a greater plasticity (despite having a lower phylogenetic diversity) that eases the transition into solid foods

Determination of the prebiotic activity of wheat arabinogalactan peptide (AGP) using batch culture fermentation

Purpose To test the prebiotic activity of wheat arabinogalactan-peptide (AGP), which is a soluble dietary fibre composed of arabinogalactan polysaccharide linked to a 15-residue peptide, which accounts for up to 0.4% of the dry weight of wheat flour. Methods The prebiotic activity of AGP prepared from white wheat flour was tested using in-vitro fermentation by colonic bacteria in automated pH controlled anaerobic stirred batch cultures and compared to fructooligosaccharide (FOS) and wheat flour arabinoxylan (AX). Bacterial populations were measured using fluorescence in-situ hybridisation (flow-FISH) and short-chain fatty acid (SCFA) concentrations were measured using HPLC. Results Fermentation of AGP resulted in a significant bifidogenic activity and increased concentrations of SCFAs, mainly acetate after 24 h of fermentation. Conclusions These results were comparable to those obtained with AX and confirm the prebiotic potential of AGP. Furthermore, fermentation of a mixture of AGP and AX was faster compared to the single substrates and more similar to FOS, indicating that combinations of fermentable carbohydrates with different structures are potentially more effective as prebiotics than single substrates

Selección in vitro de microorganismos con potencial probiótico

200 p.La Comunidad de Castilla y León aporta el 63% de la producción nacional de leche de oveja, destinada casi en su práctica totalidad a la fabricación de queso en sus múltiples variedades. Sin embargo, y a pesar del valor nutricional inherente a la leche de oveja, son mínimas las iniciativas que se han planteado hasta ahora para elaborar alimentos probióticos partiendo de esta fuente como materia prima. De este modo, en esta Tesis Doctoral se ha iniciado la selección y caracterización de bacterias ácido-lácticas que presenten un buen potencial probiótico para su posterior incorporación en productos lácteos incrementando, por lo tanto, el valor añadido de estos productos. Para ello, este estudio se ha llevado a cabo utilizando tanto bacterias aisladas de productos lácteos elaborados con leche de oveja, de vaca o de una mezcla de ambas como bacterias procedentes de de la Colección Americana de Cultivos Tipo (ATCC). Como cepa control se ha empleado la conocida Lactobacillus rhamnosus GG (ATCC 53103) con reconocido potencial probiótico. A lo largo de este trabajo se ha se han seleccionado seis cepas de BAL (incluida la cepa control Lb. rhamnosus GG) que han cumplido los aspectos de bioseguridad para su consumo aquí estudiados, además de, presentar los criterios in vitro más reconocidos de los microorganismos probióticos. Las seis cepas de BAL han sido capaces de sobrevivir frente a las condiciones extremas del tracto gastrointestinal (bajo pH y presencia de enzimas digestivas), se han adherido a la superficie de las células intestinales Caco-2, lo que podría permitir su implantación en el tracto intestinal, han mostrado actividad antimicrobiana frente a microorganismos patógenos oportunistas frecuentemente encontrados en alimentos y han sido capaces de estimular la producción de determinadas citoquinas (IL-6 e IL-8) lo que podría ser útil en el uso de estas bacterias para la estimulación del sistema inmune en individuos sanos. Además se ha observado la capacidad de utilizar determinados compuestos prebióticos como fuente de carbono. Desde el punto de vista tecnológico, se ha comprobado que las seis cepas bacterianas son capaces de crecer tanto en leche de oveja como de vaca consiguiéndose durante el proceso de fermentación llevado a cabo por estas cepas una producción óptima de ácido láctico. Además de la producción de este compuesto, se ha observado la presencia de enzimas con actividad aminopeptidásica, propiedades que influyen positivamente en las características organolépticas de los productos lácteos que puedan ser fermentados con estos microorganismos. Además todas las cepas de Lactobacillus seleccionadas, han mostrado actividad ß-galactosidásica por lo que también podrían ser utilizadas en la elaboración de productos lácteos que eviten o alivien los síntomas de la intolerancia a la lactosa en personas con deficiencia en esta enzima. Por último, destacar que todas las cepas seleccionadas muestran una buena viabilidad cuando se almacenan en fuentes lácteas a bajas temperaturas (4ºC), característica que asegura y garantiza su efectividad y funcionalidad probiótica en los propios productos alimenticios que con su presencia pudieran comercializarse. De este modo, los resultados y conocimientos que se derivan de este estudio pueden ser interesantes a la hora de predecir la funcionalidad de estas cepas in vivo antes de emprender los ensayos clínicos pertinentes, necesarios, para confirmar finalmente una cepa bacteriana como probiótic

Impact of short-chain galactooligosaccharides on the gut microbiome of lactose-intolerant individuals

Directed modulation of the colonic bacteria to metabolize lactose effectively is a potentially useful approach to improve lactose digestion and tolerance. A randomized, double-blind, multisite placebo-controlled trial conducted in human subjects demonstrated that administration of a highly purified (>95%) short-chain galactooligosaccharide (GOS), designated “RP-G28,” significantly improved clinical outcomes for lactose digestion and tolerance. In these individuals, stool samples were collected pretreatment (day 0), after GOS treatment (day 36), and 30 d after GOS feeding stopped and consumption of dairy products was encouraged (day 66). In this study, changes in the fecal microbiome were investigated using 16S rRNA amplicon pyrosequencing and high-throughput quantitative PCR. At day 36, bifidobacterial populations were increased in 27 of 30 of GOS subjects (90%), demonstrating a bifidogenic response in vivo. Relative abundance of lactose-fermenting Bifidobacterium, Faecalibacterium, and Lactobacillus were significantly increased in response to GOS. When dairy was introduced into the diet, lactose-fermenting Roseburia species increased from day 36 to day 66. The results indicated a definitive change in the fecal microbiome of lactose-intolerant individuals, increasing the abundance of lactose-metabolizing bacteria that were responsive to dietary adaptation to GOS. This change correlated with clinical outcomes of improved lactose tolerance