Separation of Oligosaccharides from Lotus Seeds via Medium-pressure Liquid Chromatography Coupled with ELSD and DAD

peer-reviewedLotus seeds were identified by the Ministry of Public Health of China as both food and medicine. One general function of lotus seeds is to improve intestinal health. However, to date, studies evaluating the relationship between bioactive compounds in lotus seeds and the physiological activity of the intestine are limited. In the present study, by using medium pressure liquid chromatography coupled with evaporative light-scattering detector and diode-array detector, five oligosaccharides were isolated and their structures were further characterized by electrospray ionization-mass spectrometry and gas chromatography-mass spectrometry. In vitro testing determined that LOS3-1 and LOS4 elicited relatively good proliferative effects on Lactobacillus delbrueckii subsp. bulgaricus. These results indicated a structure-function relationship between the physiological activity of oligosaccharides in lotus seeds and the number of probiotics applied, thus providing room for improvement of this particular feature. Intestinal probiotics may potentially become a new effective drug target for the regulation of immunity

Bioengineering approaches to simulate human colon microbiome ecosystem

Background Several diseases associated to colon microbial imbalance (dysbiosis), such as obesity, diabetes, inflammatory bowel disease, cardiovascular disease and cancer, are being reverted by modulation of gut microbiota composition through treatment with prebiotics and probiotics. Multiple in vitro models have been developed over the past three decades, with several experimental configurations, as they provide a quick, easy, and cost-effective approach to study the gut microbiome, as compared to troublesome and time-consuming in vivo studies. Scope and approach This review aims to provide an overview of the most relevant available in vitro models used to mimic the human colon microbiome dynamics, including macro-scale and microfluidic-based models. Main characteristics, functionalities, current applications and advantages or disadvantages of the models are discussed in order to provide useful information for end users (namely food and pharmaceutical researchers), when selecting the most appropriated model for assessing health claims and safety of novel functional food and drugs. Finally, the use of these colon models as a tool to study prebiotic and probiotic response in host-microbiota interaction is reviewed. Key findings and conclusions A wide range of in vitro models representing specific colon parts have been developed. However, none of these models can simultaneously cover all the key conditions found in the human colon (namely anatomical, physical, biochemical, and biological characteristics), as well as the complex microbiome-host interaction. Thus, there is a significant opportunity for further improvement of the models experimental setups towards more realistic operating systems, including mucosal surfaces, intestinal cells and tissues allowing microbiomehost crosstalk representation.SFRH/BD/139884/2018 from the Portuguese Foundation for Science and Technology (FCT). This study was supported by the FCT under the scope of the strategic funding of UIDB/04469/2020 unit and the Project ColOsH PTDC/BTM–SAL/30071/2017 (POCI–01–0145–FEDER–030071)info:eu-repo/semantics/publishedVersio

Analyse Systémique de la Modulation de la bascule respiro-fermentaire\ud chez Saccharomyces cerevisiae

L’objective de notre travail est d’étudier la transition métabolique respiro-fermentaire chez\ud Saccharomyces cerevisiae, plus spécifiquement d’évaluer l’importance du transport du carbone issu du\ud métabolisme de l’acétyl-coenzymeA vers la mitochondrie. Pour ce faire, une approche originale du\ud génie microbiologique a été mise en place basée sur l’utilisation de chémostat sur substrats mixtes.\ud Cela consiste à introduire au cours d’un chémostat oxydatif sous limitation glucose une perturbation\ud locale touchant le transport du carbone du cytosol vers la mitochondrie par l’apport d’un second\ud substrat et à analyser les conséquences sur la bascule métabolique étudiée. Il a été ainsi testé comme\ud seconds substrats, l’acide oléique, la carnitine et le succinate. La présence d’acide oléique permet de\ud retarder l’apparition de la bascule métabolique (jusqu’à 15 minutes), de rediriger le flux de carbone de\ud la voie fermentaire vers la voie oxydative (réduction de 33% de la production d’éthanol) et\ud d’augmenter en accélérostat la valeur du taux de dilution critique de 8%. Ce résultat a été retrouvé sur\ud une deuxième souche de Saccharomyces cerevisiae testée. Ce résultat marquant constitue un des rares\ud succès de réduction de l’effet Crabtree reporté dans la littérature à ce jour. Une analyse systémique a\ud été réalisée en intégrant analyse macrocinétique, métabolique et transcriptomique afin de mieux\ud comprendre l’action de l’acide oléique sur la modulation de la bascule métabolique.-------------------------------------------------------------------------------------------The aim of this work is to study the transition from respiratory to fermentative metabolism in\ud Saccharomyces cerevisiae CENPK 113-7D and more specifically to evaluate the implication of the\ud acetyl-coenzyme A derived carbon transport from cytosol to mitochondria in the onset of the\ud metabolic shift. An original approach of microbiological engineering was carried out based on the\ud utilization of chemostat with mixed substrates. The strategy consisted of introducing, during aerobic\ud glucose-limited chemostat, a local perturbation around the step to be studied by the addition of a cosubstrate\ud and in analyzing the consequences of such a perturbation on the metabolic transition. Oleic\ud acid, L-carnitine and succinate were among the tested co-substrates. Feeding the culture with oleic\ud acid led to a delay in the onset of the metabolic shift (up to 15 min), a redirection of the carbon flux\ud toward biomass production (33% decrease in the ethanol production) and increases the critical dilution\ud rate in accelerostat cultures (an 8% increase in the critical dilution rate). These results were found in a\ud second strain of Saccharomyces cerevisiae tested. This work constitutes one of the rare successful\ud reduction of the Crabtree effect reported in literature. A system biology approach, integrating\ud macrokinetic, metabolic and trancriptomic analyses, was used of ways of understanding better the\ud action of oleic acid in the onset of the metabolic shif

Impact of oleic acid as co-substrate of glucose on "short" and "long-term" Crabtree effect in Saccharomyces cerevisiae

Background: Optimization of industrial biomass directed processes requires the highest biomass yield as possible. Yet, some useful yeasts like Saccharomyces cerevisiae are subject to the Crabtree effect under glucose excess. This phenomenon can occur in large scale tank where heterogeneities in glucose concentrations exist. Therefore yeasts encounter local environments with glucose excess leading to ethanol production to the detriment of biomass formation. We previously demonstrated that oleic acid as a co-substrate in glucose-limited chemostat allowed to delay and modulate the "short-term" Crabtree effect in Saccharomyces cerevisiae. Here we further investigated the effect of oleic acid as a modulator of the Crabtree effect. Results: The impact of oleic acid as co-substrate on the Crabtree effect was investigated in terms of i) strain specificity, ii) reversibility of the potential effect with aerobic glucose-excess batches and iii) durability and maximal capacities under high ethanol stress with glucose-excess fed-batches. First, the addition of oleic acid resulted in an increase of the critical dilution rate by 8% and the specific carbon uptake rate by 18%. Furthermore, a delay was observed for the onset of ethanol production when a batch was inoculated with cells previously grown in glucose-oleate chemostat. Finally, the culture of adapted cells in a glucose-oleate fed-batch led to a redirection of the carbon flux toward biomass production, with a 73% increase in the biomass yield. Conclusions: This work demonstrated clearly that the perturbation by oleic acid as co-substrate resulted in a decrease in the "short-term" and "long-term" Crabtree effects. This impact was not strain dependent and reversible. Thus, industrial applications of this biochemical strategy could be envisaged to tackle heterogeneities issues in large scale tanks or to prepare starter yeasts for various applications

Oleic acid delays and modulates the transition from respiratory to fermentative metabolism in Saccharomyces cerevisiae after exposure to glucose excess

International audienceThis work aimed to study the transition from respiratory to fermentative metabolism in Saccharomyces cerevisiae CEN.PK 113-7D and more specifically to evaluate the implication of the acetyl-coenzymeA-derived carbon transport from cytosol to mitochondria in the onset of the metabolic shift. The strategy consisted in introducing, during aerobic glucose-limited chemostat (D=0.16 h(1)), a local perturbation around the step to be studied by the addition of cosubstrate and in analyzing the consequences of such a perturbation on the metabolic transition. Oleic acid and L-carnitine were among the tested cosubstrates because they were known to stimulate enzymes implicated in the acetyl-coenzymeA transport between the different cell compartments, such as the carnitine acetyl transferases. The metabolic transition was then comparatively quantified in sole glucose and in glucose/oleic acid chemostats in presence/absence of L-carnitine after a pulse of glucose. Feeding the culture with oleic acid (D-ole=0.0041 and 0.0073 h(1)) led to a delay in the onset of the metabolic shift (up to 15 min), a 33% decrease in the ethanol production and a redirection of the carbon flux toward biomass production. The data clearly showed a modulation of the carbon distribution among respiration and fermentation, in favor of a decrease in the "short-term" Crabtree effect by the oleic acid

Three-stage continuous culture system with a self-generated anaerobia to study the regionalized metabolism of the human gut microbiota

This work was supported by a postdoctoral scholarship support from the European Union (UE) and the Auvergne Council, to D. F.-G. (FEDER), by a PhD Scholarship support from the European Union (UE) and the Auvergne Council to W.T. (FEDER), and to a PhD Scholarship support from the French "Ministere de l'Enseignement Superieur et de la Recherche". We thank Pascal Vandekerckove from Lesaffre SA and Zsolt Popse from Global Process Concept (France) for their support. J.-F.B. thanks Celine Vidal and Claire Ardaens for their technical help.The technical and ethical difficulties in studying the gut microbiota in vivo warrant the development and improvement of in vitro systems able to simulate and control the physicochemical factors of the gut biology. Moreover, the functional regionalization of this organ implies a model simulating these differences. Here we propose an improved and alternative three-stage continuous bioreactor called 3S-ECSIM (three-stage Environmental Control System for Intestinal Microbiota) to study the human large intestine. Its main feature compared with other in vitro systems is the anaerobic atmosphere originating directly from the microbiota metabolism, leading to different gas ratios of CO2 and H-2 in each compartment Analyses of the metabolic and microbiological profiles (LC-MS and a phylogenetic microarray) show different profiles together with a maintenance of this differentiation between the three compartments, simulating respectively a proximal, a transversal and a distal colon. Moreover, the last reactor presents a high similarity with the initial fecal sample, at the microbiological diversity level. Based on our results, this in-vitro process improvement is a valuable alternative tool to dynamically study the structure and metabolism of gut microbiota, and its response to nutrients, prebiotics, probiotics, drugs or xenobiotics. (C) 2013 Elsevier B.V. All rights reserved

Colonic transit time is a driven force of the gut microbiota composition and metabolism: In vitro evidence

Jean-François Brugère thanks bachelors’students for their help, more specifically Céline Vidal, Claire Ardaens,Adeline Régnier, and Amandine Maurin, and Sylvain Denis forhis valuable help concerning in vitro systems. In memory of GeorgeT MacFarlane (died in 2015) for all of his pioneering works on gut in vitro simulations systemsThis work was supported by the European Union (UE) through the Auvergne Council (FEDER) with a PhD and a postdoctoral Scholarship support respectively to William Tottey and to David Feria-Gervasio, and by a PhD scholarship support from the French "Ministere de l'Enseignement Superieur et de la Recherche" to Nadia GaciBackground/Aims: Human gut microbiota harbors numerous metabolic properties essential for the host's health. Increased intestinal transit time affects a part of the population and is notably observed with human aging, which also corresponds to modifications of the gut microbiota. Thus we tested the metabolic and compositional changes of a human gut microbiota induced by an increased transit time simulated in vitro. Methods: The in vitro system, Environmental Control System for Intestinal Microbiota, was used to simulate the environmental conditions of 3 different anatomical parts of the human colon in a continuous process. The retention times of the chemostat conditions were established to correspond to a typical transit time of 48 hours next increased to 96 hours. The bacterial communities, short chain fatty acids and metabolite fingerprints were determined. Results: Increase of transit time resulted in a decrease of biomass and of diversity in the more distal compartments. Short chain fatty acid analyses and metabolite fingerprinting revealed increased activity corresponding to carbohydrate fermentation in the proximal compartments while protein fermentations were increased in the lower parts. Conclusions: This study provides the evidence that the increase of transit time, independently of other factors, affects the composition and metabolism of the gut microbiota. The transit time is one of the factors that explain some of the modifications seen in the gut microbiota of the elderly, as well as patients with slow transit time

Laboratory simulators of the colon microbiome

Laboratory simulators of the colon are designated to reproduce the composition and functionality of complex microbial communities. The in vitro colonic models range from batch fecal incubations, suitable for metabolic studies, to more complex continuous models, involving one or multiple connected vessels, to mimic different regions of the human colon. Laboratory simulators of the colon microbiome allow the screening of a large number of substances and are valuable in performing mechanistic studies. Moreover, multistage continuous models are relevant for microbial ecological studies, since they allow long-term experiments needed to evaluate the spatial and temporal adaptation of the colonic microbiota to dietary ingredients, drugs, and toxic or radioactive compounds, without ethical constraints. In vitro models are particularly well suited for screening prebiotics or probiotics, for special functions in the gut. The progress made in developing in vitro fermentation models, able to closely mimic the gut microbial environment, can offer remarkable insights into gut microbiota functions or microbial metabolites associated with a disease state and could be predictive for in vivo situations.Peer reviewe

Omic approaches coupled to gastrointestinal dynamic modelling to assess food bioactivity

Peer reviewe