Koen Molly

Development of a 5-step multi-chamber reactor as a simulation of the human intestinal microbial ecosystem.

A five-stage reactor was developed to simulate the gastro-intestinal microbial ecosystem of humans. The small intestine was simulated by a two-step “fill and draw” system, the large intestine by a three-step reactor. A representative supply medium was developed to support a microbial community resembling that of the human gastro-intestinal tract. The entire system was validated by monitoring fermentation fluxes and products, i.e. indicator bacterial groups, volatile fatty acids, enzymatic activities and headspace gases. The simulator was operated with varying concentrations and combinations of arabinogalactan, xylan, pectin, dextrins and starch. The resulting patterns of microbial diversity and activity were analyzed and compared with data for in-vivo gastro-intestinal microbial communities as described in the literature and found to be reprensentative.

The importance of meat enzymes in ripening and flavour generation in dry fermented sausages. First results of a European project

The importance of bacterial and meat enzymes in lipolysis and proteolysis was evaluated during dry sausage ripening. The data suggest that lipolysis is to a great extent brought about by muscle and fat tissue. Polyunsaturated fatty acids are liberated from the polar lipid fraction and their specific liberation is higher than for monounsaturated and saturated fatty acids. Lipolysis seemed to be more pronounced in sausages with pork than with beef. Initial protein degradation seemed to be originating from cathepsin D like muscle enzymes. In a later stage, bacterial enzymes become more important in further degradation of the protein fragments formed.

POLYMERS FOR COLON SPECIFIC DRUG DELIVERY

Azo-linked polymeric prodrugs of 5-aminosalicylic acid (5-ASA) were prepared and evaluated in simulated human intestinal microbial ecosystem. Release of 5-ASA was demonstrated. Polyamides containing azo groups in the backbone were prepared and tested in vitro in a reductive buffer or in the bioreactor medium. It was demonstrated that for the hydrophobic polymer reduction stops at the hydrazine stage whereas for a hydrophilic analogue reduction with formation of amines occurred.

Lipolysis in a Belgian sausage: Relative importance of endogenous and bacterial enzymes

The importance of bacterial and meat enzymes in lipolysis and carbonyl formation was evaluated during dry sausage ripening. Sausages were prepared with and without addition of an antibiotic-antimicotic mixture. In some experiments, an extra inoculum of Micrococcaceae was added and in two experiments, glucose was omitted. Total viable bacterial counts after 21 days were lowered by at least 2 log units in the presence of antibiotics. Free fatty acid productions after 3 and 21 days, in the presence of antibiotics were not significantly lower than observed in the control sausages. Total carbonyl compounds (benzidine reaction compounds) were significantly lowered by the presence of antibiotics compared to the control sausages except when glucose was omitted from the recipe. The data suggest that lipolysis is almost exclusively brought about by muscle and fat tissue. Polyunsaturated fatty acids are liberated from the polar lipid fraction and their specific liberation is higher than for monounsaturated and saturated fatty acids. Carbonyl production from lipids seems to be independent of bacterial activity.

the influence of lentinus edodes (shiitake mushroom) preparations on bacteriological and morphological aspects of the small intestine in piglets

Among substances intended to replace growth promoting antibiotics in pig nutrition, non-digestible oligosaccharides or polysaccharides could be potential alternative compounds. Therefore, the influence of β-1,3-1,6 glucans on bacteriological, biochemical and morphological aspects of the small intestine in weaned piglets was investigated. As sources of β-glucans, Lentinan (extract of Lentinus edodes mycelium) or dried L. edodes mycelium were added to the diet. Four homogenous groups of 5 newly weaned piglets (4 weeks of age) received one of four diets: control diet (C), C supplemented with Avilamycin (50 mg/kg, positive control), C supplemented with 0.1% of Lentinan and C supplemented with 5% of dried L. edodes mycelium powder. A first group of 10 piglets was euthanized after 11 days and the remaining 10 on day 12 of the experiment. The gastrointestinal tract was divided in segments and samples taken from digesta (stomach, proximal and distal jejunum, caecum), mucosal scrapings (jejunum) and ring shaped tissue samples (1 cm) of proximal and distal jejunum. Bacterial counts were made with digesta and mucosal samples, and short-chain fatty acids (SCFA), lactic acid and ammonia concentrations were determined. Tissue samples of both jejunal sites were embedded in paraffin wax for morphometrical (villus length, crypt depth) and histological observations (numbers of intraepithelial lymphocytes (IEL), goblet cells, apoptotic enterocytes on villi, mitotic cells in crypts). Only the diet containing 5% of dried L. edodes consistently resulted in lower viable counts (ca. 1 – 2 log10 CFU) of total bacteria, E. coli, streptococci and lactic acid bacteria, and luminal and mucosal effects agreed very well. With this diet, acetate and butyrate concentrations in the distal jejunum were doubled, which is favourable in view of the trophic effect on enterocytes and colonocytes. Villus length (V) was increased with both diets containing β-glucans while crypt depth (C) was not altered, but V/C was higher. IEL counts were decreased by both diets although bacterial numbers, which is only one parameter of bacterial load, were only diminished with the L. edodes feed. The three supplemented feeds lowered the number of apoptotic enterocytes on the villi, but these numbers were very low (control diet : 44 cells per 100 villi), making clear interpretation difficult. The mitotic index was slightly lower with the L. edodes feed, although not statistically significant. Decreased viable counts observed with the latter diet is a favourable effect as it is accepted that a lower bacterial load causes lower turnover rates of the intestinal epithelial cells, while there is also less competition for specific substrates. A higher V/C ratio, a smaller number of IEL in the epithelium and a lower apoptotic index also indicate slower turnover rate of the mucosa when Lentinan and L. edodes diets were fed. The inconsistent effects observed with Lentinan were probably due to the low amount added to the diet. It should be taken into account that the influence of L. edodes mycelium powder was more likely due to the presence of antibacterial compounds (eg. lenthionine, lentinamycin, terpenoids, polyphenols), rather than to an immunostimulating action of β-glucans with increased release of IgA onto the mucosa surface.

Incorporation of galactomannans in the diet of newly weaned piglets: Effect on bacteriological and some morphological characteristics of the small intestine

In search of substances replacing antibiotics as growth promoters for farm animals, non-digestible oligosaccharides (NDO) or non-starch polysaccharides (NSP) have been proposed as possible alternatives. In this context, the influence of galactomannans on bacteriological and morphological aspects of the gastrointestinal tract in weanling pigs was investigated. Four groups of five newly weaned piglets received one of the following diets: control feed (C), C supplemented with guar gum (1%), C supplemented with locust bean gum (1%) and C supplemented with 10% of carob tree seeds meal as source of locust bean gum. The animals were euthanized after 11 – 12 days and digesta were sampled in stomach, jejunum (proximal and distal) and caecum, while mucosal scrapings and ring shaped tissue samples were taken of proximal and distal jejunum. On these samples bacteriological, biochemical and morphological determinations were carried out. Total count of bacteria in digesta and mucosal scrapings was not influenced by the different diets, with the exception of the proximal jejunum where a small decrease (0.5 log10 CFU) was noted with the guar gum and carob tree seeds diet. The number of E. coli increased by feeding both gums and carob tree seeds. With the latter diet, higher counts of streptococci were observed. In agreement with the lower concentration of lactic acid in jejunal contents, guar gum decreased the number of lactobacilli. Locust bean gum decreased the molar proportion of acetate in caecal contents while butyrate and valerate were augmented. Feeding the carob tree seeds resulted in shorter villi and a lower villus height/crypt depth ratio in the jejunum mucosa, which was an indication for a faster renewal rate of the epithelium. Both locust bean gum feeds significantly lowered the mitotic index in the crypts of the small intestine. Only with the carob tree seeds diet, viscosity of jejunal contents was increased. In conclusion, the effects of the addition of 1% of pure guar gum or locust bean gum were inconsistent and not very outspoken, whereas 10% of carob tree seeds meal in the diet resulted in influences on intestinal characteristics at the bacteriological and morphological level.