Αμαλία Δ. Καραγκούνη, Ομότιμη Καθηγήτρια, Τομέας Βοτανικής, Τμήμα Βιολογίας, Σχολή Θετικών Επιστημών, Εθνικό και Καποδιστριακό Πανεπιστήμιο Αθηνών
The Gut Microbiota (GM) is the most complex microbial community in the human body, associated with various activities that promote health. The β- (1 → 3, 1 → 6) -D-glucans, abundant in the fungal cell wall, are polysaccharides with differences in their chemical structure and bioactivity. The mechanisms through which beta-glucan-rich edible fungi regulate the gut microbial community have not been fully elucidated yet. However, increasing experimental evidence supports their potential prebiotic activity and hence, their health-promoting properties, anti-cancer and immunomodulatory activities included.
The analysis of the metabolic pattern (profile) produced as a result of the fermentation of dietary polysaccharides - nutrients by the intestinal microbiota, that is, the systematic identification of low molecular weight metabolites produced, provides extremely useful data as to the role of GM not only in normal physiology but also in its deregulation and pathogenesis. Metabolomic studies targeting specific metabolic pathways are currently a cornerstone of biological research and require the integration of high-performance analytical, state-of-the-art techniques such as NMR and Mass Spectroscopy (LC-MS, GC-MS, CE-TOF-MS, UPLC-TOF-MS).
In this thesis, we initially investigated the cytotoxic (MTT assay) and geno-protective (Comet assay) effect of the in vitro fermentation products of Pleurotus ostreatus 1123 and Ganoderma lucidum by fecal microbiota of healthy volunteers in Caco-2 (epithelial adenocarcinoma colon) and U-937 (monocyte-macrophage) human cell lines. Subsequently, all metabolites resulting from the in vitro fermentation of Pleurotus ostreatus 1123 and Ganoderma lucidum were analysed by NMR and a similar analysis followed for Pleurotus ostreatus LGM22, Pleurotus eryngii Zheng (216), Cyclocybe cylindracea CC2 and Cyclocybe cylindracea 505 in vitro fermentation products, all cultivated on Wheat Straw. The Chenomx NMR Suite 6.0 main metabolites database and bibliographic data were used to identify the spectrum resonance peaks. Using the Mnova program (Mestrelab Research) the 1H NMR spectra were corrected for phase and baseline and normalized based on the TSP resonance peak at 0 ppm. Subsequently, Chemometry was applied to extract chemical information from the NMR spectra. The main chemometric tools used, after responding to a large number of variables, were PCA and the regression of a few least squares by separating the systematic but non-correlated Y-variable (orthogonal) variable matrix X (OPLS). Chemometric analysis of the data was performed with the statistical software program SIMCA-P 14.0 (UMETRICS, Umea, Sweden).
The results of this work showed that the products of in vitro fermentation of edible mushrooms by the fecal microbiota of healthy volunteers exert a significant anti-genotoxic effect while they are characterized by distinct differences in the metabolic profile composition. Short-chain fatty acids (SCFAs), linked to the metabolism of probiotic bacteria, as well as a plethora of metabolites, specific for the fermentation process and its effectiveness, are produced.
beta-glucans, SCFA, Intestinal Microbiota, Comet assay, NMR-based Metabolomics