A critical review on in vitro and ex vivo models of the intestinal epithelium of humans and monogastric animals

Recently, the bioactive potential of several functional ingredients and biomolecules has been evaluated regarding human and animal nutrition. The digestive process from food intake to absorption and metabolism are important events that induce changes in ingredients, which affect their bioactivity. Consequently, there is a need to assess the bioavailability and bioaccessibility of these compounds. The methodology for the simulation of the human gastrointestinal tract has been standardized (INFOGEST protocol), while a gastrointestinal protocol for other animals (e.g., ruminants or broilers) has yet to be established. However, INFOGEST allows us only to predict bioaccessibility, leaving a gap regarding a methodology able to assess bioavailability by mimicking intestinal permeability and absorption. Several approaches—including in vitro, ex vivo, in situ and in vivo methods—can be found in the literature, aiming to tackle transepithelial routes, but leading to different results concerning the bioefficiency of the compounds studied. Therefore, this review aims to assess the current state-of-the-art regarding monogastric intestinal dynamics, absorption, and permeability events. Moreover, it compiled methodologies for simulating intestinal absorption in several biological systems, while reasoning their advantages, disadvantages, applications in ingredient development and the existing gaps.info:eu-repo/semantics/publishedVersio

Do all prebiotics have the same impact on my gut microbiota? a case study using an in vitro gastrointestinal model to assess functional oligosaccharides added to milk

info:eu-repo/semantics/publishedVersio

Strategies to assess the impact of sustainable functional food ingredients on gut microbiota

Nowadays, it is evident that food ingredients have different roles and distinct health benefits to the consumer. Over the past years, the interest in functional foods, especially those targeting gut health, has grown significantly. The use of industrial byproducts as a source of new functional and sustainable ingredients as a response to such demands has raised interest. However, the properties of these ingredients can be affected once incorporated into different food matrices. Therefore, when searching for the least costly and most suitable, beneficial, and sustainable formulations, it is necessary to understand how such ingredients perform when supplemented in different food matrices and how they impact the host's health. As proposed in this manuscript, the ingredients' properties can be first evaluated using in vitro gastrointestinal tract (GIT) simulation models prior to validation through human clinical trials. In vitro models are powerful tools that mimic the physicochemical and physiological conditions of the GIT, enabling prediction of the potentials of functional ingredients per se and when incorporated into a food matrix. Understanding how newly developed ingredients from undervalued agro-industrial sources behave as supplements supports the development of new and more sustainable functional foods while scientifically backing up health-benefits claims.info:eu-repo/semantics/publishedVersio

Sugarcane straw polyphenols as potential food and nutraceutical ingredient

The sugarcane processing industry generates a large amount of straw, which has a negative environmental impact, and high costs are associated with their elimination, wasting their potential bioactive value attributed to their richness in polyphenols. In this study, an ethanolic extract produced from sugarcane straw was screened for its phenolic compounds content, and the potential use of this extract in the development of a food ingredient was further evaluated. Fifty different secondary metabolites belonging to the hydroxybenzoic acids, hydroxycinnamic acids, and flavonoids were identified by liquid chromatography–electrospray ionization–ultrahigh-resolution—quadrupole time of flight–mass spectrometry (LC-ESI-UHR-QqTOF-MS). The predominant phenolic compounds found were 4-hydroxybenzaldehyde, chlorogenic acid, and 5-O-feruloylquinic acid. The obtained extracts showed strong potential as food preservatives by exhibiting (a) antioxidant activity using both 2.2′-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) diammonium salt radical cation (ABTS) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) methods; and (b) antimicrobial capacity, with a minimum inhibitory concentration of 50 mg/mL for Staphylococcus aureus, 74% inhibition for Bacillus cereus, and 44% for Salmonella enterica; and (c) the capacity to inhibit a food browning enzyme, tyrosinase (28–73% for 1–8 mg/ mL). Moreover, the extracts showed antidiabetic potential by inhibiting the enzymes α-glucosidase (15–38% for 1.25–5.00 mg/mL) and dipeptidyl peptidase-IV (DPP-IV) (62–114% for 0.31–5.00 mg/mL). The extract (0.625 mg/mL) also exhibited the capacity to reduce proinflammatory mediators (i.e., interleukins 6 and 8, and tumor necrosis factor alpha) when Caco-2 cells were stimulated with interleukin 1 beta. Thus, sugarcane straw extract, which is rich in phenolic compounds, showed high potential to be used in the development of food-preservative ingredients owing to its antioxidant and antimicrobial potential, and to be explored as a food supplement in diabetes prevention and as coadjuvant to reduce intestinal inflammation by reducing proinflammatory mediators.info:eu-repo/semantics/publishedVersio

Can supplemented skim milk (skm) boost your gut health?

The incorporation of functional ingredients, such as prebiotics and probiotics in food matrices, became a common practice in the human diet to improve the nutritional value of the food product itself. Worldwide, skim milk (SKM) is one of the most consumed food matrices, comprising all the essential nutrients desired for a balanced diet. Thus, the modulation of the human gut microbiota by SKM supplemented with different well-known functional ingredients was evaluated. Four well-studied prebiotics, fructo-oligosaccharides (FOS), galacto-oligosaccharides (GOS), mannanoligosaccharides (MOS) and inulin, and one probiotic product, UL-250® (Saccharomyces boulardii) were added at 1% (w/v) to SKM and subjected to a gastrointestinal in vitro model. The impact of each combination on gut microbiota profile and their fermentation metabolites (i.e., short-chain fatty acids–SCFA) was assessed by quantitative polymerase chain reaction (qPCR) and high-performance liquid chromatography (HPLC), respectively. The addition of FOS to SKM had promising results, showing prebiotic potential by promoting the growth of Lactobacillus, Bifidobacterium, and Clostridium cluster IV. Moreover, the increment of SCFA levels and the decrease of total ammonia nitrogen were observed throughout colonic fermentation. Overall, these results demonstrate that the combination SKM + FOS was the most beneficial to the host’s health by positively modulating the gut microbiota.info:eu-repo/semantics/publishedVersio

Preservation of human gut microbiota inoculums for in vitro fermentations studies

The use of fecal inoculums for in vitro fermentation models requires a viable gut microbiota, capable of fermenting the unabsorbed nutrients. Fresh samples from human donors are used; however, the availability of fresh fecal inoculum and its inherent variability is often a problem. This study aimed to optimize a method of preserving pooled human fecal samples for in vitro fermentation studies. Different conditions and times of storage at -20 degrees C were tested. In vitro fermentation experiments were carried out for both fresh and frozen inoculums, and the metabolic profile compared. In comparison with the fresh, the inoculum frozen in a PBS and 30% glycerol solution, had a significantly lower (p < 0.05) bacterial count (<1 log CFU/mL). However, no significant differences (p < 0.05) were found between the metabolic profiles after 48 h. Hence, a PBS and 30% glycerol solution can be used to maintain the gut microbiota viability during storage at -20 degrees C for at least 3 months, without interfering with the normal course of colonic fermentation.info:eu-repo/semantics/publishedVersio

Applied microbiology on poultry industry: from nutrition to the gut microbiota

Poultry products (i.e., meat and eggs) are one of the major protein sources for the human diet. The animal’s diet is one of the key elements that the poultry industry has been focused on, to improve the animal’s performance, maintaining their healthy growth and, ultimately, high quality end products. The incorporation of functional ingredients in feed formulations, aiming to provide extra benefits and/or prevent diseases, has been considered efficient in maintaining the animal’s productivity and simultaneously ensure its well-being. Nutrient’s bioavailability varies throughout digestion and absorption within the poultry’s gastrointestinal tract (GIT). A reliable in vitro model, as the one developed and used in this study, capable of mimicking all digestion, absorptive and cecal fermentation processes, is a useful tool to study the potential benefits of feed supplemented with functional and/or bioactive ingredients. The developed in vitro gastrointestinal model simulates the chemical, enzymatic, and mechanical conditions prevailing in the chicken´s GIT, from beak to cecum. Fresh broiler’s cecal samples were used as inoculum for batch cecal fermentation and the impact of different feed formulations, on bacteria modulation, organic acids, and total ammonia nitrogen production, were assessed. Overall, this approach enables to evaluate, as close to reality as possible, the potential of target additives, providing a trustworthy tool for the development of functional feeds.info:eu-repo/semantics/acceptedVersio

Human and animal in vitro gastrointestinal models: importance and applications

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Importance of gastrointestinal in vitro models for the poultry industry and feed formulations

The animal’s diet is a crucial factor, as poultry feed formulations influences greatly their development, well-being and final products quality, i.e., meat and eggs. Therefore, the search for feed additives that provide concomitantly better performances, low-cost usage, guarantying the animal well-being and products safety, became a priority to the poultry industry. Although in an early stage, research has been focused on developing the optimal cost-efficient feed formulations, taking into account the chicken’s physiology and function of the gastrointestinal tract and intestinal microbiota. This review discusses a number of concepts and novel approaches towards the optimization of poultry’s feed formulations, by critically encompassing the animal’s growth and performance. Additionally, it highlights the in vitro gastrointestinal models capabilities as a potential solution to test highly nutritive, well-balanced and efficient feed formulations within a circular economy framework.info:eu-repo/semantics/publishedVersio

Development of a Chicken Gastrointestinal Tract (GIT) Simulation Model: Impact of Cecal Inoculum Storage Preservation Conditions

A chicken gastrointestinal tract (GIT) simulation model was developed to help predict the potential effects of feed additives supplementation on chicken’ microbiota. The chemical and enzymatic conditions for oral, gastric, intestinal, and cecum fermentation phases were designed to closely resemble the chicken GIT conditions. For cecum fermentation, the inoculum was obtained from the cecal contents of 18 38-day broiler chickens. The impact of inoculum preservation on bacteria viability was assessed by comparing two methods of preservation with fresh inoculum: (1) 5% dimethyl sulfoxide (DMSO) at −80 °C and (2) 30% glycerol at −20 °C. The fermentation with fresh and frozen (DMSO method) inoculums was performed and compared using standard chicken feed (SCF) and SCF with 1% fructooligosaccharides (FOS), and inoculum control (IC) condition without feed matrix was used as a baseline. Inoculum’s viability was assessed throughout 90 days of storage by culture media platting, while bacterial growth and metabolites production during fermentation was evaluated by quantitative polymerase chain reaction (qPCR), high-performance liquid chromatography (HPLC), and total ammonia nitrogen quantification. The DMSO method was shown to be the most suitable for cecal inoculum storage. Higher growth of beneficial cecal bacteria for fresh inoculum was observed in SCF while for frozen inoculum, was the SCF + FOS condition. Also, frozen inoculum had lower activity of butyrate producers and proteolytic bacteria, showing different fermentation profiles. The GIT model developed showed to be useful to test the effect of feed additives supplementation