The oral cavity in healthy subjects has a well-balanced microbiota that consists of more than 700 species. However, a disturbance of this balance, with an increase of harmful microbes and a decrease of beneficial microbes, causes oral disorders such as periodontal disease or dental caries. Nowadays, probiotics are expected to confer oral health benefits by modulating the oral microbiota. This study screened new probiotic candidates with potential oral health benefits and no harmful effects on the oral cavity. We screened 14 lactobacillus strains and 36 streptococcus strains out of 896 oral isolates derived from healthy subjects. These bacteria did not produce volatile sulfur compounds or water-insoluble glucan, had higher antibacterial activity against periodontal bacteria, and had higher adherence activity to oral epithelial cells or salivary-coated hydroxyapatite in vitro. We then evaluated the risk of primary cariogenicity and infective endocarditis of the selected oral isolates. As a result, Lactobacillus crispatus YIT 12319, Lactobacillus fermentum YIT 12320, Lactobacillus gasseri YIT 12321, and Streptococcus mitis YIT 12322 were selected because they showed no cariogenic potential in an artificial mouth system and a lower risk of experimental infective endocarditis in a rat model. These candidates are expected as new probiotics with potential oral health benefits and no adverse effects on general health.
Lactate-utilizing butyrate-producers were isolated from human feces and identified based on the sequences of 16S rRNA gene. Anaerostipes caccae strain L2, one of the seven human fecal isolates, was administered to rats with galacto-oligosaccharides (GOS) as bifidogenic carbohydrates for stimulating lactate formation in the hindgut. Ingestion of GOS alone increased concentrations of cecal lactate and butyrate compared with control rats (P<0.05). Additional administration of strain L2 on GOS tended to enhance the promoting effect of GOS on cecal butyrate formation (P=0.06) and lowered the mean value of cecal lactate concentration (P=0.32). Consequently, cecal and fecal butyrate concentrations in rats administered with both strain L2 and GOS were significantly higher than those in the control rats (P<0.01 and P<0.05, respectively). Significant changes were observed in the other fermentation acids, such as succinate, acetate, and propionate, depending on the ingestion of strain L2. Administered strain L2 was retrieved from the cecal content of a rat based on randomly amplified polymorphic DNA analysis. The results suggest that synbiotic ingestion of lactate-utilizing butyrate-producers and GOS alters the microbial fermentation and promotes the formation of beneficial fermentation acids, including butyrate, in the gut.
An enzymatic hydrolysate of wheat gluten was further digested in vitro with porcine pepsin and pancreatin to obtain an indigestible peptide. Indigestible pyroglutamyl peptide was isolated from the digest by strong cation-exchange, size-exclusion, and reversed-phase chromatographies. The pyroglutamyl peptide was digested with pyroglutamate aminopeptidase, and the digest was reacted with phenyl isothiocyanate. The resultant phenylthiocarbamyl (PTC) peptides were purified by reversed-phase HPLC by using binary gradient elution with ammonium acetate buffer, pH 6.0, and acetonitrile. The PTC peptides were analyzed with an automatic peptide sequencer on the basis of the Edman degradation method with a modified program. Some pyroglutamyl peptides were also analyzed by fast-atom bombardment ionization mass spectrometry without the pyroglutamate amino peptidase digestion. Consequently, pyroGlu-Asn-Pro-Gln, pyroGlu-Gln-Gln-Pro-Gln, pyroGlu-Gln-Pro-Gln, pyroGlu-Gln-Pro-Gly-Gln-Gly-Gln, pyroGlu-Gln, pyroGlu-Gln-Pro, pyroGlu-Ile-Pro-Gln, pyroGlu-Ile-Pro, pyroGlu-Gln-Pro-Leu, pyroGlu-Gln-Phe-Pro-Gln, pyroGlu-Ser-Phe-Pro-Gln, pyroGlu-Phe-Pro-Gln, and pyroGlu-Gln-Pro-Pro-Phe-Ser were identified.
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