ABSTRACT
The establishment of the gut microbiota immediately after birth is a dynamic process that may impact lifelong health. At this important developmental stage in early life, human milk oligosaccharides (HMOs) serve as specific substrates to shape the gut microbiota of the nursling. The well-orchestrated transition is important as an aberrant microbial composition and bacterial-derived metabolites are associated with colicky symptoms and atopic diseases in infants. Here, we study the trophic interactions between an HMO-degrader, Bifidobacterium infantis and the butyrogenic Anaerostipes caccae using carbohydrate substrates that are relevant in the early life period including lactose and total human milk carbohydrates. Mono- and co-cultures of these bacterial species were grown at pH 6.5 in anaerobic bioreactors supplemented with lactose or total human milk carbohydrates. A. caccae was not able to grow on these substrates except when grown in co-culture with B. infantis, leading to growth and concomitant butyrate production. Two levels of cross-feeding were observed, in which A. caccae utilised the liberated monosaccharides as well as lactate and acetate produced by B. infantis. This microbial cross-feeding points towards the key ecological role of bifidobacteria in providing substrates for other important species that will colonise the infant gut. The progressive shift of the gut microbiota composition that contributes to the gradual production of butyrate could be important for host-microbial crosstalk and gut maturation.
Subject(s)
Bifidobacterium longum subspecies infantis/metabolism , Clostridiales/metabolism , Lactose/metabolism , Milk, Human/metabolism , Oligosaccharides/metabolism , Bifidobacterium longum subspecies infantis/genetics , Bifidobacterium longum subspecies infantis/growth & development , Bioreactors/microbiology , Clostridiales/genetics , Clostridiales/growth & development , Coculture Techniques , Culture Media/metabolism , HumansABSTRACT
AIM OF THE STUDY: In order to confirm the traditional use of Ligustrum vulgare L. (common privet, Oleaceae) we investigated the inhibitory activity of different extracts from leaves (LlE), flowers (LflE) and fruits (LfrE) on metallopeptidases ACE and NEP. MATERIALS AND METHODS: Powdered plant materials were first extracted with water and then with ethyl acetate and n-butanol saturated with water. The metallopeptidases activity was determined using in vitro fluorimetric assays. RESULTS: At a concentration of 100microg/ml the ethyl acetate extracts showed the highest activity. The bio-guided fractionation of the leaves extract led to the isolation of two iridoids which were identified by (1)H, (13)C and HETCOR NMR spectroscopy as oleuropein and ligstroside aglycones. Both compounds are dual ACE/NEP inhibitors with IC(50) of 20 and 25microM for ACE and IC(50) of 35 and 75microM for NEP, respectively. Secoirydoids glycosides, tyrozol and hydroxytyrozol, as well as, flavonoids present in the ethyl acetate extracts showed little or no inhibitory activity. CONCLUSIONS: Our results partially support the diuretic and hypotensive activities of common privet.
Subject(s)
Angiotensin-Converting Enzyme Inhibitors/pharmacology , Iridoids/pharmacology , Ligustrum/chemistry , Plant Extracts/pharmacology , Angiotensin-Converting Enzyme Inhibitors/administration & dosage , Angiotensin-Converting Enzyme Inhibitors/isolation & purification , Animals , Flowers , Fluorometry , Fruit , Glucosides/administration & dosage , Glucosides/isolation & purification , Glucosides/pharmacology , Inhibitory Concentration 50 , Iridoid Glucosides , Iridoids/administration & dosage , Iridoids/isolation & purification , Male , Medicine, Traditional , Neprilysin/antagonists & inhibitors , Plant Extracts/administration & dosage , Plant Leaves , Pyrans/administration & dosage , Pyrans/isolation & purification , Pyrans/pharmacology , SwineABSTRACT
Pooled human milk oligosaccharides were separated into neutral and several acidic oligosaccharide fractions by preparative anion-exchange chromatography (AEC) using AG 1-X2. The oligosaccharides were eluted stepwise using deionized water and three different concentrations of ammonium acetate buffer, pH 6.8. The elution order of the compounds was determined directly by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) analysis of the AEC effluent without any cleanup or concentration steps. Up to a concentration of 500 mM ammonium acetate, the masses of acidic oligosaccharides could be detected by screening the fractions in an automated mode. The combination of the improved chromatographic procedure, the applied MALDI matrices, and operating parameters is suitable for the detection of neutral oligosaccharides as well as acidic oligosaccharides. The method provides high sensitivity and mass accuracy, including for the high-molecular-weight monosialylated oligosaccharides up to 2751.5 Da. The applied ionic strength of the anion-exchange eluents enables a rapid and an unambiguous composition assignment by MALDI-MS for neutral, monosialylated, and disialylated oligosaccharides from human milk. The acidic fractions have to be desalted by electrodialysis and were finally analyzed by HPAEC-PAD to get a high-resolution "fingerprint" of structures present in each fraction. From these analyses, it can be concluded that the isomeric variety of monosialylated oligosaccharides occurring in human milk is higher than estimated before.
