Rozenn Ravallec

Probiotic potential of Enterococcus faecalis strains isolated from meconium

107 bacterial isolates with Gram positive staining and negative catalase activity, presumably assumed as lactic acid bacteria, were isolated from samples of meconium of 6 donors at Roubaix hospital, in the north of France. All these bacterial isolates were identified by MALDI-TOF mass spectrometry as Enterococcus faecalis. However, only six isolates among which E. faecalis 14, E. faecalis 28, E. faecalis 90, E. faecalis 97, and E. faecalis 101 (obtained from donor 3), and E. faecalis 93 (obtained from donor 5) were active against some Gram-negative bacteria and Gram-positive bacteria , through production of lactic acid, and bacteriocin like inhibitory substances. The identification of these isolates was confirmed by 16rDNA sequencing and their genetic relatedness was established by REP-PCR and pulsed field gel electrophoresis methods. Importantly, the aforementioned antagonistic isolates were sensitive to various classes of antibiotics tested, exhibited high scores of coaggregation and hydrophobicity, and were not hemolytic. Taken together, these properties render these strains as potential candidates for probiotic applications.

In vitro evidence for gut hormone stimulation release and dipeptidyl-peptidase IV inhibitory activity of protein hydrolysate obtained from cuttlefish (Sepia officinalis) viscera

Abstract Two cuttlefish ( Sepia officinalis ) viscera protein hydrolysates were obtained with different enzymes extracted from cuttlefish and smooth hound ( Mustellus mustellus ). Their ability to stimulate the secretion of cholecystokinin (CCK) and glucagon-like peptide 1 (GLP-1), using the enteroendocrine STC-1 cell line, and to inhibit the DPP-IV activity during a simulated gastrointestinal digestion was assayed. The physico-chemical parameters of hydrolysates and their effects on intestinal cell viability were also determined. The hydrolysate obtained with cuttlefish enzymes (CVPH1) appeared to be the most promising for all assessed bioactivities. Thus CVPH1 was able to stimulate CCK and active GLP-1 releasing activities of enteroendocrine cells without any cytotoxicity and to inhibit DPP-IV activity. Moreover, these actions were enhanced after gastrointestinal digestion and CVPH1 was also able to inhibit the intestinal DPP-IV activity of Caco-2 cells. These very promising findings highlight, via two different mechanisms, the positive effect of CVPH1 on GLP-1 actions.

An improvement of surfactin production by B. subtilis BBG131 using design of experiments in microbioreactors and continuous process in bubbleless membrane bioreactor

Culture medium elements were analysed by a screening DoE to identify their influence in surfactin specific production by a surfactin constitutive overproducing Bacillus subtilis strain. Statistics pointed the major enhancement caused by high glutamic acid concentrations, as well as a minor positive influence of tryptophan and glucose. Successively, a central composite design was performed in microplate bioreactors using a BioLector®, in which variations of these impressive parameters, glucose, glutamic acid and tryptophan concentrations were selected for optimization of product-biomass yield (YP/X). Results were exploited in combination with a RSM. In absolute terms, experiments attained an YP/X 3.28-fold higher than those obtained in Landy medium, a usual culture medium used for lipopeptide production by B. subtilis. Therefore, two medium compositions for enhancing biomass and surfactin specific production were proposed and tested in continuous regime in a bubbleless membrane bioreactor. An YP/X increase of 2.26-fold was observed in bioreactor scale.

Simulated GI digestion of dietary protein: Release of new bioactive peptides involved in gut hormone secretion

Dietary proteins have been reported to induce a strong feeling of satiety that has been partially explained by gut hormone level increase. Up to date, various protein hydrolysates have demonstrated in vitro and in vivo their potential to stimulate gut hormone secretion related to food intake decrease and their mechanisms of action have just started to be resolved. In this context, this study aimed at identifying new peptide sequences involved in gut hormone secretion released by protein in vitro gastrointestinal digestion. Targeted gut hormones were Cholecystokinin (CCK) and Glucagon-Like Peptide 1 (GLP-1). The activity of DPP-IV was also considered as it strongly modulates GLP-1 action. In a previous study, simulated digestion of dietary protein has generated hydrolysates with enhancing effect on CCK and GLP-1 secretion in STC-1 cells as well as DPP-IV inhibitory properties. Successive purification steps were performed to isolate peptide fractions involved in these bioactivities whose sequence was determined by LC-MS-MS. Three peptide sequences ANVST, TKAVEH and KAAVT were pointed out for their stimulating effects on GLP-1 secretion. The sequence VAAA was isolated for its DPP-IV inhibitory properties. Two peptide groups were strongly involved in CCK release sharing a certain occurrence of aromatic amino acid residues.

Food peptidomics of in vitro gastrointestinal digestions of partially purified bovine hemoglobin: low‐resolution versus high‐resolution LC‐MS/MS analyses

Consumers and governments have become aware how the daily diet may affect the human health. All proteins from both plant and animal origins are potential sources of a wide range of bioactive peptides and the large majority of those display health‐promoting effects. In the meat production food chain, the slaughterhouse blood is an inevitable co‐product and, today, the blood proteins remain underexploited despite their bioactive potentiality. Through a comparative food peptidomics approach we illustrate the impact of resolving power, accuracy, sensitivity, and acquisition speed of low‐resolution (LR)‐ and high‐resolution (HR)‐LC‐ESI‐MS/MS on the obtained peptide mappings and discuss the limitations of MS‐based peptidomics. From in vitro gastrointestinal digestions of partially purified bovine hemoglobin, we have established the peptide maps of each hemoglobin chain. LR technique (normal bore C18 LC‐LR‐ESI‐MS/MS) allows us to identify without ambiguity 75 unique peptides while the HR approach (nano bore C18 LC‐HR‐ESI‐MS/MS) unambiguously identify more than 950 unique peptides (post‐translational modifications included). Herein, the food peptidomics approach using the most performant separation methods and mass spectrometers with high‐resolution capabilities appears as a promising source of information to assess the health potentiality of proteins.

Protein Digestion-Derived Peptides and the Peripheral Regulation of Food Intake

The gut plays a central role in energy homeostasis. Food intake regulation strongly relies on the gut–brain axis, and numerous studies have pointed out the significant role played by gut hormones released from enteroendocrine cells. It is well known that digestive products of dietary protein possess a high satiating effect compared to carbohydrates and fat. Nevertheless, the processes occurring in the gut during protein digestion involved in the short-term regulation of food intake are still not totally unraveled. This review provides a concise overview of the current data concerning the implication of food-derived peptides in the peripheral regulation of food intake with a focus on the gut hormones cholecystokinin and glucagon-like peptide 1 regulation and the relationship with some aspects of glucose homeostasis.

Using Caco-2 cells as novel identification tool for food-derived DPP-IV inhibitors

Dietary proteins have recently been investigated as a new source of DPP-IV inhibitory peptides with limited side effects and promising applications. Numerous studies have highlighted and identified peptide sequences able to inhibit DPP-IV activity in vitro, mostly from milk proteins. However, the correlation to in vivo studies remains scarce because standard in vitro assays with purified enzyme do not accurately simulate key factors impacting peptide bioactivity such as intestinal and brush border enzymes or cellular permeability. Therefore, a DPP-IV activity inhibition assay is here proposed using non differentiated confluent Caco-2 cells to rapidly assess food-derived peptide inhibitory potential in approaching intestinal conditions. DPP-IV gene expression was first checked and specific DPP-IV substrate was used to implement the assay. Using a specific DPP-IV inhibitor confirmed that non differentiated Caco-2 cells express measurable DPPIV activity. This in situ assay was then applied to digests which already demonstrated a DPP-IV inhibitory potential with a standard assay using purified enzyme. Bovine hemoglobin and cuttlefish hydrolysate digests from simulated gastrointestinal digestion exerted a dose response inhibition on DPP-IV activity but displayed different inhibitory potentials.

Flavonoids stimulate cholecystokinin peptide secretion from the enteroendocrine STC-1 cells

Animal experiments showed that flavonoids might have the potential for an anti-obesity effect by reducing weight and food intake. However, the exact mechanisms that could be involved in these proposed effects are still under investigation. The complex process of food intake is partially regulated by gastrointestinal hormones. Cholecystokinin (CCK) is the best known gastrointestinal hormone to induce satiety signal that plays a key role in food intake regulation. It is released from the endocrine cells (I cell) in response to the ingestion of nutrients into the small intestine. In this study, we investigated the possible effects of flavonoids (quercetin, kaempferol, apigenin, rutin and baicalein) on stimulation of CCK release in vitro using enteroendocrine STC-1 cells. In comparison with the control, quercetin, kaempferol and apigenin resulted in a significant increase in CCK secretion with quercetin showing the highest activity. On the other hand, no significant effect was seen by rutin and baicalein. To our knowledge, this is the first report to study the stimulation of CCK peptide hormone secretion from STC-1 cells by quercetin and kaempferol, rutin, apigenin and baicalein. Based on the cell-based results in this work, it can be suggested that the reported activity of flavonoids against food intake and weight could be mediated by stimulation of CCK signal which in turn is responsible for food intake reduction, but future animal and human studies are needed to confirm this conclusion at organism level.

Chicory Roots for Prebiotics and Appetite Regulation: A Pilot Study in Mice

The objectives of this work are to address the prebiotic effects of chicory ( Cichorium intybus) together with its possible role in appetite control. We compared nine chicory genotypes in order to determine if variations in the content of metabolites in the roasted roots would lead to modifications in release of satiety hormones and in composition of gut microbiota. To this aim, a 5-week dietary-intervention study was achieved using mice fed with distinct chicory-based preparations. A 16S rRNA gene-based metagenetic analysis of fecal microbiota was performed. In vitro gastrointestinal digestions were performed in order to study the effect of chicory intestinal digests on gut hormone regulation in enteroendocrine cells. Firmicutes/Bacteroidetes ratio and gut bacterial groups, such as Alloprevotella, Blautia, Alistipes, and Oscillibacter, were found to be modulated by chicory. On the other hand, CCK and GLP-1 satiety hormones were demonstrated to be significantly increased by chicory in vitro.