Development of Whey Protein Concentrate-Pectin-Alginate Based Delivery System to Improve Survival of B. longum BL-05 in Simulated Gastrointestinal Conditions.

Bifidobacterium longum BL-05 encapsulated beads were developed by using whey protein concentrate (WPC) and pectin (PE) as encapsulating material through extrusion/ionic gelation technique with the objective to improve survival of probiotics in harsh gastrointestinal conditions. B. longum BL-05 was grown in MRS (de man rogosa and sharpe) broth, centrifuged and mixed with polymeric gel solution. Bead formulations E4 (2.5% WPC + 1.5% PE) and E5 (2% PE) showed the highest value for encapsulation efficiency, size, and textural properties (hardness, cohesiveness, springiness) due to increasing PE concentration. The survivability and viability of free and encapsulated B. longum BL-05 was assessed through their resistance to simulated gastric juice (SGJ), tolerance to bile salt, release profile in simulated intestinal fluid (SIF), and storage stability during 28 days at 4 °C. The microencapsulation provided protection to B. longum BL-05 and encapsulated cells were exhibited significant (p < 0.05) resistance to SGJ and SIF as compared to free cells. Bead formulations E3 (5.0% WPC + 1.0% PE) and E4 (2.5% WPC + 1.5% PE) exhibited more resistance to SGJ (at pH 2 for 2 h) and at 2% bile salt solution but comparatively slow release as compared to other bead formulations. Free cells lost their viability when stored at 4 °C after 28 days but microencapsulated cells demonstrated promising results during storage and viable cell count was > 107 CFU/g. This study revealed that extrusion using WPC and PE as encapsulating material could be considered as one of the novel technologies for protection and effective delivery of probiotics.

Characterization of a Sea Buckthorn Extract and Its Effect on Free and Encapsulated Lactobacillus casei.

Probiotics are bacteria that can provide health benefits to consumers and are suitable to be added to a variety of foods. In this research, viability of immobilized Lactobacillus casei in alginate with or without sea buckthorn lipid extract were studied during heat treatment and with an in vitro gastrointestinal model. The characterization of the lipid extract was also done using the UV-Vis spectrometry (UV-Vis), high-performance liquid chromatography photodiode array detection method (HPLC-PDA), gas chromatography coupled with mass spectrometry (GS-MS) and Cryo scanning electron microscopy (Cryo-SEM). During heat treatment, the entrapped probiotic cells proved high viability (>6 CFU log/g), even at temperatures above 50 °C. The rich in monounsaturated fatty acids sea buckthorn fraction improved the in vitro digestion passage regarding the probiotic viability. The survival of the probiotic cells was 15% higher after 2 h in the acidic medium of the simulated gastric fluid in the sample where L. casei was encapsulated with the sea buckthorn extract compared with the samples where no extract was added. Thus, this approach may be effective for the future development of probiotic-supplemented foods as foods with health welfare for the consumers.

In vitro gastric survival of commercially available probiotic strains and oral dosage forms.

Although the intestinal microbial community is still incompletely understood, there is strong evidence of the benefits of using probiotics to address some medical states or conditions. As a result, the probiotics oral supplements market has exploded during the last few years. However, while their sensitivity to gastric juices, acidic pH and bile is well known, most of these oral forms would not guarantee any survival of the strains in such conditions. In this work, we have studied the resistance to simulated gastric juices of several commercially available probiotics products. These included sixteen strains and ten oral forms such as enteric/non-enteric capsules/tablets and microencapsulated strains. Results demonstrated that all tested strains showed high sensitivity to acidic conditions and suggested that most of these microorganisms would not show any viability when immersed in the stomach at fasting. Most probiotics oral forms did not provide any protection to strains, unless these forms presented strong enteric protection. Consequently, the efficacy of non-enteric products to fully provide to the patient the benefits related to the consumption of probiotics supplement would be strongly questionable. This study underlines the chasm between the current opinion about probiotics protection needs and the products proposed by many companies in the dietary supplements area.

Investigations on the possible impact of a glyphosate-containing herbicide on ruminal metabolism and bacteria in vitro by means of the 'Rumen Simulation Technique'.

AIMS: This study was performed in a well-established in vitro model to investigate whether the application of a glyphosate-containing herbicide might affect the bacterial communities and some biochemical parameters in a cow's rumen. METHODS AND RESULTS: The test item was applied in two concentrations (high and low) for 5 days. In a second trial, fermentation vessels were inoculated with Clostridium sporogenes before the high dose was applied. Effluents were analysed by biochemical, microbiological and genetic methods. A marginal increase in short-chain fatty acid production and a reduction in NH3 -N were observed. There were minor and rather equivocal changes in the composition of ruminal bacteria but no indications of a shift towards a more frequent abundance of pathogenic Clostridia species. Clostridium sporogenes counts declined consistently. CONCLUSIONS: No adverse effects of the herbicide on ruminal metabolism or composition of the bacterial communities could be detected. In particular, there was no evidence of a suspected stimulation of Clostridia growth. SIGNIFICANCE AND IMPACT OF THE STUDY: Antibiotic activity of glyphosate resulting in microbial imbalances has been postulated. In this exploratory study, however, intraruminal application of concentrations reflecting potential exposure of dairy cows or beef cattle did not exhibit significant effects on bacterial communities in a complex in vitro system. The low number of replicates (n = 3/dose) may leave some uncertainty.

Effects of essential oils from medicinal plants acclimated to Benin on in vitro ruminal fermentation of Andropogon gayanus grass.

BACKGROUND: Plants from West Africa commonly used in both human and veterinary medicine contain various secondary metabolites. However, their potential in mitigating ruminal methane production has not been explored. This study examined the effects of seven essential oils (EOs) from plants acclimated to Benin at four dosages (100, 200, 300 and 400 mg L(-1)), on in vitro rumen microbial fermentation and methane production using Andropogon gayanus grass as a substrate. RESULTS: Compared to control, Laurus nobilis (300-400 mg L(-1) ), Citrus aurantifolia (300-400 mg L(-1)) and Ocimum gratissimum (200-400 mg L(-1)) decreased (P < 0.05) methane production (mL g(-1) DM) by 8.1-11.8%, 11.9-17.8% and 7.9-30.6%, respectively. Relative to the control, reductions in methane (mL g(-1) DM) of 11.4%, 13.5% and 14.2% were only observed at 400 mg L(-1) for Eucalyptus citriodora, Ocimum basilicum and Cymbopogon citratus, respectively. These EOs lowered methane without reducing concentrations of total volatile fatty acids or causing a shift from acetate to propionate production. All EOs (except M. piperita) reduced (P < 0.05) apparent dry matter (DM) disappearance of A. gayanus. CONCLUSIONS: The current study demonstrated that EOs from plants grown in Benin inhibited in vitro methane production mainly through a reduction in apparent DM digestibility.

In hydrolyzed cow's milk Helicobacter pylori becomes nonculturable and the growth of Salmonella typhi and Escherichia coli is inhibited.

The colony forming unit (CFU) of H. pylori is reduced rapidly in lipase hydrolyzed cow's milk and a similar reduction was found in a physiological saline solution when it was supplemented with soluble C4 to C10 fatty acids of milk fat composition. Slight CFU decreases were observed for E. coli and S. typhi in hydrolyzed milk buffered to pH 3, while the counts in milk and physiological saline solution at pH 3 stayed almost unchanged for 24 h. E. coli proliferated in glucose-peptone medium, better at pH 4.7 than at pH 3. On the other hand, supplementation of the medium with soluble fatty acids of milk composition completely inhibited growth for 32 h. Supplementation of the medium with fatty acids reduced the growth of S. typhi to approximately 1/20 at pH 4.7. Therefore, milk hydrolyzed by gastric lipase may damage H. pylori, producing a nonculturable state. With E. coli and S. typhi, hydrolyzed milk does not induce inactivation to a nonculturable state but inhibits their proliferation potently. The latter is considered to be a state prior to VBNC (viable but nonculturable). However, the antibiotic effect will disappear when the fatty acids are absorbed by the intestine.

Mutagenicity of arbutin in mammalian cells after activation by human intestinal bacteria.

Arbutin (hydroquinone-beta-D-glucopyranoside) is present in various food plants. Its aglycone, hydroquinone, is mutagenic and carcinogenic. We investigated whether hydroquinone may be released under conditions encountered in the human gastrointestinal tract. Arbutin was stable in artificial gastric juice. Fecal slurries from nine human subjects completely converted arbutin (2 mM) into hydroquinone. Four of nine representative human intestinal species investigated, namely Eubacterium ramulus, Enterococcus casseliflavus, Bacteroides distasonis, and Bifidobacterium adolescentis, deglycosylated arbutin at rates of 21.08, 16.62, 8.43 and 3.59 nmol x min(-1) x (mg protein)(-1), respectively. In contrast, homogenates from small intestinal mucosa and cytosolic fractions from colon mucosa deglycosylated arbutin at substantially lower rates: 0.50 and 0.09 nmol x min(-1) x (mg protein)(-1), respectively. Arbutin, unlike hydroquinone, did not induce gene mutations in Chinese hamster V79 cells in the absence of an activating system. However, in the presence of cytosolic fractions from E. ramulus or B. distasonis, arbutin was strongly mutagenic. Cytosolic fraction from Escherichia coli, showing no arbutin glycosidase activity, was not able to activate arbutin in this model system. The release of the proximate mutagen hydroquinone from arbutin by intestinal bacteria in the immediate vicinity of the colon mucosa may pose a potential risk.

Protection of bifidobacteria encapsulated in polysaccharide-protein gel beads against gastric juice and bile.

Bifidobacterium cells were encapsulated in a mixed gel composed of alginate, pectin, and whey proteins. Two kinds of capsules were obtained: gel beads without membranes and gel beads with two membranes formed by the transacylation reaction. In vitro studies were carried out to determine the effects of simulated gastric pH and bile salts on the survival of free and encapsulated Bifidobacterium bifidum. The protective effects of gel beads without membranes and gel beads coated with two membranes formed by the transacylation reaction were evaluated. After 1 h in an acidic solution (pH 2.5), the free-cell counts decreased by 4.75 log units, compared with a <1-log decrease for entrapped cells. The free cells did not survive after 2 h of incubation at pH 2.5, while immobilized-cell counts decreased by about 2 log units. After incubation (1 or 3 h) in 2 and 4% bile salt solutions, the bifidobacterium mortality level for membrane-free gel beads (4 to 7 log units) was higher than that for free cells (2 to 3 log units). However, counts of bifidobacteria immobilized in membrane-coated gel beads decreased by <2 log units. Cell encapsulation in membrane-coated protein-polysaccharide gel beads could be used to increase the survival of healthy probiotic bacteria during their transit through the gastrointestinal tract.

Helicobacter pylori may survive ampicillin treatment in the remnant stomach.

Helicobacter pylori (H. pylori) is a Gram-negative curved rod-like or spiral bacterium that chronically infects the human gastric mucosa, and is a major risk factor for gastritis, gastric and duodenal ulcer and adenocarcinoma of the stomach. After partial gastrectomy, some patients may have persistent H. pylori infection for five years or more. In this study, we detected three bacteria, i.e., Klebsiella pneumoniae, Enterobacter aerogenes, and Escherichia coli, in the gastric juice of patients with a remnant stomach. Some of these bacteria produced beta-lactamase. These findings are potentially important since such bacteria could provide H. pylori with the chance to acquire drug resistance and to transfer drug resistance genes. This could be one reason why H. pylori is difficult to eradicate in the remnant stomach.

[Poisoning with spotted and red mushrooms--pathogenesis, symptoms, treatment]. / Zatrucia muchomorami plamistym i czerwonym--patogeneza, objawy, leczenie.

Amanita pantherina and Amanita muscaria are commonly occurring mushrooms in Polish forests. They contain ibotenic acid and muscimol: the substances reacting with neurotransmitter receptors in central nervous system. The ingestion of these mushrooms produces a distinctive syndrome consisting of alternating phases of drowsiness and agitation with hallucinations, and sometimes with convulsions. The diagnosis of Amanita pantherina or Amanita muscaria poisoning is established by means of mycologic investigation of gastric lavage. The treatment is only symptomatic, and the prognosis is usually good.

Effect of ascorbic acid on the intragastric environment in patients at increased risk of developing gastric cancer.

Ascorbic acid has been shown to decrease nitrosation in vivo, and epidemiological data suggest that the consumption of foods rich in this vitamin is associated with a reduced risk for gastric cancer. In order to study this suggestion further, fasting gastric juice samples were obtained from 62 high-risk patients (seven with atrophic gastritis, ten with pernicious anaemia, ten with partial gastrectomy, 21 with vagotomy and drainage and 14 with highly selective vagotomy), before, during four weeks' treatment with 1 g ascorbic acid four times daily, and four weeks after treatment. Samples were analysed for pH, total and nitrate-reducing bacterial counts, nitrite and N-nitroso compounds. Treatment with ascorbic acid lowered the median pH only in the vagotomized patients (p less than 0.001) but resulted in a reduction in median nitrate-reducing bacterial counts and in nitrite and N-nitroso compound concentrations in all groups, except for an increase in the nitrate-reducing bacterial count in atrophic gastritis patients and in nitrite in those with pernicious anaemia. These data suggest that treatment with a high dose of ascorbic acid reduces the intragastric formation of nitrite and N-nitroso compounds.

A clinicopathologic study of enterocyte-adherent Escherichia coli: a cause of protracted diarrhea in infants.

Fifteen infants (age 3-28 wk) suffered from severe diarrhea with acute dehydration and poor growth. Persistent watery stools and suboptimal nutrition necessitated central venous alimentation with prolonged hospitalization. Repeated stool and small intestinal fluid cultures yielded the classical enteropathogenic Escherichia coli serotype 0119:B14. In all patients, biopsy of the jejunum or rectal mucosa, or both, showed moderate to severe damage, irregular atrophy of surface epithelium, and subnuclear vacuolization of crypt epithelium. Ultrastructural studies revealed bacteria adherent to mucosal cells with flattening of microvilli, loss of the cellular terminal web, and cupping of the plasma membrane around individual bacteria. Heavily colonized cells had marked intracellular damage. Assays for heat-labile, heat-stable, and vero cell toxins were negative for these Escherichia coli isolates. Oral neomycin and nutritional support resulted in clearing of Escherichia coli 0119:B14 from stool and small bowel with improvement in histologic characteristics. Damage to enterocytes and villi by adherent nontoxigenic Escherichia coli 0119:B14 results in protracted diarrhea in infants.

Effect of methane inhibitors on the metabolism of rumen microbes in vitro.

In incubations in vitro with rumen fluid, the effect of two methane inhibitors, linseed oil hydrolysate (LOH) and chloral hydrate (CH) on the efficiency of microbiol growth was investigated. Total and net microbial growth were determined from 32PO43- and NH3--N incorporation respectively and expressed as g N incorporated per kg organic matter fermented (gN/kgOMf). In a first series on incubations, it was found that LOH had no influence on overall microbial growth efficiency, while with CH, a small but significant decrease of total and net growth efficiency was measured. Further experiments showed that this was not due to accumulation of hydrogen gas in the CH incubations. Microscopic examination showed a toxic effect of LOH on protozoa, but with CH, no such effect was observed. This observation, together with earlier work where a considerable increase in microbial growth efficiency was found in vitro after defaunation of the rumen suggested the following hypothesis: both inhibitors lowered bacterial growth. In the case of LOH, this effect is marked by the defaunating action of LOH, the latter resulting in an increased growth efficiency of the bacterial fraction. This hypothesis was confirmed by incubations with washed cell suspensions (WCS) of mixed rumen bacteria, where growth efficiency was indeed decreased by both inhibitors. The possible mechanism explaining this phenomenon was discussed.