Presence Of Xylitol Research Articles

Influence of Additives on High Pressure Stability of β-Galactosidase from Kluyveromyces Lactis and Invertase from Saccharomyces Cerevisiae

High pressure denaturation of two yeast enzymes, Kluyveromyces lactis β-galactosidase and Saccharomyces cerevisiae invertase in aqueous solutions of salts and polyols was investigated. Polyols (more than salts) were shown to act as very effective agents against pressure denaturation for both enzymes, leading to an increase in half-lives by factors between 1–10,000. NaBr and KBr were essentially destabilizing compounds, leading to protective effects near zero in the case of β-galactosidase. The maximum stability was obtained when KCl was used as additive. The β-galactosidase half-life is increased by a factor of 60 in a 3 m KCl solution and the invertase half-life by a factor of 40. Whereas NaCl had no stabilizing effect on β-galactosidase up to 2 m, this salt was of greater efficiency for invertase whatever the concentration, leading to a protective effect of almost 30 at 3 m. The use of polyols (glycerol, erythritol, xylitol, and sorbitol) allows β-galactosidase and invertase to be stabilized against pressure deactivation. This stabilizing effect increases with the polyol concentration. For invertase in the presence of xylitol, the half-life is increased by a factor of 7 at 1 m and by a factor near 50 at 2 m. The increase in pressure resistance due to the presence of polyols was always greater with β-galactosidase than with invertase. The protective effect of a 2 m xylitol solution is above 10,000 for β-galactosidase whereas it is about 50 for invertase. After the systematic study of the influence of salts and polyols, certain hypotheses are advanced on the possible mechanisms by which additives stabilize proteins to pressure denaturation.

Growth of xylitol-resistant versus xylitol-sensitive Streptococcus mutans strains in saliva

Five Streptococcus mutans pairs (serotype c S. mutans 10449 and four clinical isolates of S. mutans: 123.1, LG1, OMFA, T10B) were used to find out if the xylitol-resistant (XR) natural mutants of the corresponding xylitol-sensitive (XS) S. mutans parental strains differ in their growth patterns in saliva. The isogenic XR natural mutants of the parental S. mutans strains were selected after sequential cultivations in the presence of xylitol and glucose. The XR/XS strains pairs were grown in individual and pooled glucose-supplemented filter-sterilized salivas (one to five sequential cultivations). The two salivas used represented subjects with good or poor support of the growth of S. mutans in vivo. Protease and peptidase activities were determined from the saliva growth media and cell suspensions. Salivary protein profiles were analyzed using SDS-PAGE and native IEF before and after the cultivations. The growth properties of the XR/XSS. mutans pairs were similar in both individual and pooled salivas. Sequential cultivation of all strains did not show any differences in growth patterns. XS strains were inhibited by the presence of xylitol (2% w/v) in pooled saliva, as shown for other glucose-supplemented media. Protease and peptidase activities of the XR/XSS. mutans pairs were low and of similar magnitude. Also, the general hydrolytic properties of most XR/XSS. mutans pairs appeared similar as judged by the small growth-induced changes in salivary protein profiles. In conclusion, saliva, the source of nutrients for salivary microorganisms in vivo, favored neither the XR nor the XS strains of S. mutans.

Evaluation of a Gluconobacter oxydans whole cell biosensor for amperometric detection of xylose

Whole cells of Gluconobacter oxydans were employed in a microbial sensor for xylose determinations using Clark-type electrodes. Bacterial cells were immobilized on chromatographic paper by simple physical adsorption and attached to the surface of the electrodes. The lower limit of xylose detection was approximately 0·5 mM and measurements were useful up to at least 20 mM xylose. Physiological buffers showed little effect on biosensor function. Responses were highly reproducible, showing a standard deviation of 6·7% over 10 consecutive measurements. Whole cell biosensors were relatively stable, retaining 60% of initial activity after 35 days of dry storage at 4°C. Xylose detection was not significantly affected by the presence of xylitol, suggesting that biosensors will be useful in monitoring conversions of these compounds. However, glucose or ethanol elicited a 10-fold higher response than xylose at equal concentrations (1 mM). Such interfering materials will need to be controlled or concurrently monitored in specific sensor applications.

Emergence of multiple xylitol-resistant (fructose PTS-) mutants from human isolates of mutans streptococci during growth on dietary sugars in the presence of xylitol.

The growth inhibition of mutans streptococci is one of the proposed mechanisms of action of xylitol, a caries-preventive natural carbohydrate sweetener. Xylitol is taken up and accumulated as non-metabolizable, toxic xylitol phosphate via a constitutive fructose PTS, and selects, during in vitro growth at the expense of glucose, for natural xylitol-resistant mutants that lack constitutive fructose PTS activity. Since long-term xylitol consumption leads to the emergence of xylitol-resistant mutans populations in humans in an oral environment containing sugars of dietary origin, we wanted to test the hypothesis that xylitol-resistant cells could be selected from mutans streptococci strains during in vitro growth on fructose, sucrose, or lactose. Three laboratory strains and three fresh mutans streptococcal isolates were repeatedly transferred in trypticase-yeast extract medium supplemented with glucose, fructose, sucrose, or lactose in the presence and absence of xylitol. Depending on the growth sugar, the presence of xylitol resulted in the selection of xylitol-resistant populations for several of the six strains tested, but not necessarily in the presence of all four sugars. All six strains rapidly became xylitol-resistant when grown on glucose in the presence of xylitol. All three fresh isolates became xylitol-resistant after 9 to 16 transfers in the presence of fructose or sucrose plus xylitol, while none of the laboratory strains became xylitol-resistant after 16 transfers in the presence of these sugars. The growth rates of 12 xylitol-resistant mutants in the presence of eight sugars suggested the existence of various types of xylitol-resistant mutants. The data partially explain the occurrence of xylitol-resistant mutans populations in long-term xylitol consumers and suggest a mechanism consistent with a selection process. Since various preliminary results suggest that xylitol-resistant natural mutants may be less virulent and less cariogenic than their parent strains, this selection process may alter, for the better, the mutans streptococci population of the plaque and play a role in the caries-preventive action of xylitol.

Effect of xylitol on the metabolism and viability of Candida shehatae

Anaerobic D-xylose fermentations were performed with C. shehatate in the presence of 0, 25, and 50 g/L of xylitol. D-Xylose was preferentially utilized over xylitol and ethanol yields (Y Etoh/S ≈ 0.26 g/g) were unaffected by xylitol. Added xylitol did inhibit conversion of xylose to xylitol at an external xylitol concentration of 50 g/L; Y Xylitol/S was reduced from 0.21 to 0.14. Cell viability declined in all of the fermentations, but was not due to the presence of xylitol. The decline in viability was attributed to oxygen deprivation, since ethanol levels only reached 10.5 g/L and the decline cell viability was the same in each fermentation, regardless of the xylitol concentration.

Initial catabolism of sorbitol in Actinomyces naeslundii and Actinomyces viscosus.

The initial steps of sorbitol catabolism were studied in 4 strains of Actinomyces naeslundii and Actinomyces viscosus that had been isolated from human dental plaque. Cell-free extracts were prepared from cells grown in the presence of either sorbitol, xylitol or glucose. The extracts from all strains grown on sorbitol had nicotinamide adenine dinucleotide-linked dehydrogenase activities for sorbitol and xylitol and reduced nicotinamide adenine dinucleotide-linked reductase activities for fructose and xylulose. Two of the strains also exhibited these activities when grown in the presence of xylitol, and all glucose-grown cells lacked them. The results indicate that sorbitol metabolism in oral actinomyces involve oxidation of sorbitol to fructose by an inducible enzyme, nicotinamide adenine dinucleotide-linked sorbitol dehydrogenase. This step is followed by the phosphorylation of fructose with guanosine triphosphate as a main phosphoryl donor. Thus, the initial catabolic pathway of sorbitol in A. naeslundii and A. viscosus is different from those described for other oral bacteria.

On the mechanism of xylitol-dependent inhibition of glycolysis in streptococcus sobrinus OMZ 176

1. 1. The mechanism of xylitol-dependent inhibition of glycolysis in Streptococcus sobrinus OMZ 176 was investigated in aerobically and anaerobically grown cells. 2. 2. Glucose-stimulated glycolysis was followed polarographically, by radio-HPLC-analyses of glycolytic intermediates, by measurement of ATP generated, and spectrophotometric monitoring of extent of NAD(P) +/NADPH-status. 3. 3. Xylitol added to suspensions of S. sobrinus inhibited O 2 uptake by approximately 20%, and led to a corresponding decrease in rate of lactate formation in aerobic and anaerobic cells. 4. 4. Xylitol also delayed the onset of the glucose-dependent rapid reduction of NAD(P) + by approximately 1 min, although the total extent of reduction was not significantly affected compared to control cells. 5. 5. The inhibitory effect of xylitol on glucose dependent ATP synthesis, however, was decreased by 70–80%. 6. 6. Hence the dramatic decrease in glucose-dependent synthesis of ATP may be the direct cause of decreased bacterial growth in the presence of xylitol. 7. 7. A mechanism explaining the observed phenomena is proposed.

Effect of polyols on the thermostability of pullulan-hydrolysing activity from Sclerotium rolfsii

The influence of various polyols on the thermostability of pullulan-hydrolysing activity fromSclerotium rolfsii was studied. The half-life of the enzyme activity at 60°C was determined to be of the order of 30 min. In the presence of xylitol and sorbitol (3 M or more) there was a significant enhancement in the thermostability of the enzyme with retention of 100% activity after incubation for 7 h at 60°C. However, ethylene glycol and glycerol were found to have no protective effect. The stabilizing efficiency was found to be dependent on the concentration of the polyhydric alcohol used and the number of OH-groups present per molecule.

Solubilization and dissolution of famotidine from solid glass dispersions of xylitol

The aqueous solubility and dissolution of famotidine from solid glass dispersions of xylitol, prepared by the fusion method, were investigated. Both famotidine and xylitol exhibited minimal degradation during the fusion process. Famotidine alone and in the presence of xylitol was found to be relatively stable in water at 37 ± 0.5°C for at least 3 days: less than 4% degradation was observed at the end of 72 h. Solubility of famotidine from solid glass dispersions and physical mixtures containing varying proportions of famotidine and xylitol at 37 ± 0.5°C was found to be higher than that of famotidine alone in water. However, the solid glass dispersions were more effective in enhancing the solubility of famotidine. A 1:40 famotidine:xylitol dispersion produced greatest solubility enhancement (31%). The solubility of famotidine from physical mixtures increased linearly with the increase in xylitol concentration, but the relationship was not linear for glass dispersions. Dissolution studies on glass dispersions with famotidine:xylitol ratios of 1:1, 1:10 and 1:20 in water at 37 ± 0.5°C revealed a marked increase in the dissolution rate of famotidine from solid glass dispersions when compared to the dissolution rate of famotidine powder alone. The increase in the dissolution rate was greatest at the lowest drug level (1:20) with 100% of the drug dissolving within one minute. Thermograms of the solid glass dispersions obtained by differential scanning calorimetry showed no evidence of chemical interaction between famotidine and xylitol. The phase diagram of the dispersion system by the capillary tube method suggested the formation of a eutectic mixture of famotidine and xylitol at a drug:carrier ratio approaching 1:40.

A New Method for the Detection and Enumeration of Xylitol-resistant (fructose-PTS-) Strains ofStreptococcus mutansfrom Pure Cultures and Human Saliva

A simple procedure for the identification and enumeration, in large bacterial populations, of Streptococcus mutans of both xylitol-resistant and xylitol-sensitive phenotypes was developed. Since xylitol-resistant cells have lost the capacity to take up xylitol from the extracellular medium, due to a defect in their fructose-PTS, there is no uptake of [ 14 C]xylitol by these cells whereas the [ 14 C]xylitol taken up by the sensitive cells is used to visualise them by autoradiography. The method was shown to be suitable for laboratory strains as well as for natural populations of S. mutans ofthe human oral cavity. The in vitro selection of the xylitol-resistant phenotype during consecutive periods of growth at the expense of glucose in the presence of xylitol was followed. Likewise, it was found that the average percentage of xylitol-resistant fresh isolates from two human subjects increased from 0 per cent to 78 per cent over a xylitol-consuming period of 17 d, while it increased from 0 per cent to 8 per cent only in the control subjects who were consuming sorbitol and mannitol instead of xylitol. The superiority of this method resides in the possibility of processing many samples simultaneously. Moreover, it can be applied successfully to the detection and enumeration of other types of PTS mutants if the radioactive xylitol is replaced with a sugar analogue of the defective PTS. A strain of S. salivarius , defective in mannose- PTS, was identified with 2-deoxy-[ 14 C]glucose. Keywords: S. mutans ; Xylitol; PTS; Detection; Enumeration; Mutant; Dental caries.

Isolation and Partial Characterization of Dermatan Sulfate Proteoglycans from Human Post-Burn Scar Tissues

Dermatan sulfate (DS) proteoglycans (PGs) were extracted from human post-burnscar (Sc) tissues with 4M guanidinium chloride and isolated from the extracts by DEAE-cellulose chromatography and by differential ethanol precipitation. The DS.PGs were further purified by Sepharose CL-6B column chromatography. The average molecular weight (Mr) of hypertrophic scar (HSc) tissue DS.PGs was 39,000 based on sedimentation equilibrium measurements. Alkaline borohydride treatment of DS.PGs liberated glycosaminoglycan (GAG) chains and the presence of xylitol indicated that these chains were attached to protein core by xylosyl residues. The average Mr of the DS.GAG chain from HSc and normal scar (NSc) samples were 23,500 and 20,000 respectively. After digestion of the HSc and NSc, DS.PGs with chondroitinase ABC in the presence of proteinase inhibitors, two peptide components with Mr values of 21,500 and 17,000 were detected by SDS-polyacrylamide gel electrophoresis using reducing conditions. Analysis of the protein core fractions derived from NSc and HSc DS.PGs by Sepharose CL-6B column chromatography showed the presence of a single NH2-terminal amino acid (aspartic acid) and also that the fractions with different KAV values had an identical NH2-terminal sequence (A1-AS). The A1-A23 sequence of NSc DS.PG (major fraction, C) : NH2Asp-Glu-Ala-O-Gly-Ile-Gly-Pro-Glu-Val-Pro-Asp-Asp-Arg-Asp-Phe-Glu-Pro-Ser-Leu-Gly-Pro-Val was the same as reported for a DS.PG isolated from human fetal membrane (HFM) tissue (Brennan et al., 1984).ELISA inhibition assay using monoclonal antibodies raised in rabbit against the NH2-terminal peptide (containing 15 amino acids) of human fetal membrane tissue were found to cross-react with HSc and NSc DS.PGs. Monoclonal antibodies to bovine skin DS.PGs protein core (Pearson et al., 1983) did not show any cross-reactivity with scar DS.PGs. These results show that the scar DS.PGs described here are different from normal bovine skin DS.PGs in the size and type of the protein core, and that in all the samples, the peptide components have the same NH2-terminal amino acid sequence.

Influence of xylitol on growth, establishment, and cariogenicity of Streptococcus mutans in dental plaque of rats.

The influence of xylitol on glucose metabolism of Streptococcus mutans C67-1S was studied in vitro and in rats in vivo. In vitro there was an inhibition by xylitol of the growth rate and acid production of S. mutans C67-1S, but in vivo no effect on the initial growth rate could be demonstrated. In vivo there was a delay in the start of growth of S. mutans which resulted in a 10-day delay of establishment. The isolated xylitol-resistant mutant of C67-1S, S. mutans C67-1S XR, showed no difference in glucose metabolism in batch cultures. S. mutans C67-1S and S. mutans C67-1S XR were both used to study caries development in SPF Osborne Mendel rats. The rats were housed under programmed feeding conditions and were fed a glucose or a glucose-xylitol diet. In the case of S. mutans C67-1S, there was a remarkable inhibition of fissure caries lesions in the presence of xylitol compared to the glucose group. But also in the S. mutans C67-1S XR groups there were fewer lesions in the presence of xylitol. These results indicated that the anticariogenic properties of xylitol do not only depend on an inhibition of bacterial metabolism, but in part on other mechanisms.

Influence of xylitol- and/or fluoride-containing toothpastes on the remineralization of surface softened enamel defects in vivo.

This paper describes the effect of xylitol and fluoride on remineralization of surface softened human enamel in vivo. Surface softened enamel blocks were mounted in the lower prosthesis of 20 full denture-wearing participants. These participants used xylitol- and/or fluoride-containing toothpastes during 4-week periods. Knoop hardness measurements were employed to estimate the degree of remineralization in each period. After 26 days of toothpaste use it was found that the enamel defects were rehardened substantially and that the hardness of sound enamel was approached. The changes of enamel hardness were not significantly affected by the presence of xylitol and/or fluoride in the toothpastes.

Further studies on the growth inhibition of some oral bacteria by xylitol.

The growth in Brain Heart Infusion medium containing glucose or sucrose of Streptococcus mutans strain OMZ 176, two strains of Strep. sanguis (ATCC 10556 and No. 39) and Actinomyces viscosus type ATCC 27044 was inhibited by xylitol but not by D-arabitol or ribitol. When fructose was substituted for glucose, xylitol had no growth-inhibiting effect. It was also shown that the bacteria produced less acid in the presence of xylitol, as measured by the pH of the cultures or by neutralization with alkali.

Extracellular hydrolase activity of the cells of the oral bacterium Streptococcus mutans isolated from man and grown on glucose or xylitol

Cells of Streptococcus mutans were maintained for 18 months in medium containing either glucose or xylitol as the only added carbohydrate. Growth in 0.25 per cent xylitol was retarded by about 60 per cent as compared with the growth in glucose medium. The growth pattern in the presence of xylitol was similar to that obtained when no carbohydrates were added. No uptake of [ 14C(U)]- xylitol was observed with cells maintained for 18 months in xylitol. Extracellular enzyme activities, tested with 10 different N- l- aminoacyl-2- naphthylamines , N- carbo-2- naphthoxy- dl- phenylalanine , casein, azocoll, congocoll and a chromophore collagenase substrate, were significantly higher in the xylitol medium. Isoelectric focusing of proteins from the extracellular medium containing xylitol revealed enzymes hydrolysing N- l- methionyl-2- naphthylamine and casein, not detected in the chromatograms of glucose-containing medium. Xylitol behaved as an inert carbohydrate with respect to the increases in the extracellular proteinase activities, and could be associated with a general increase in protein metabolism upon starvation of a fermentable carbohydrate (glucose). Similar increase in the proteinase activities occurs in whole saliva and dental plaque when xylitol is habitually consumed. Because of their non-specificity, these proteinases could attack salivary proteins and peptides in vivo.

The Effect of Dental Plaque Grown in the Presence of Xylitol or Sucrose on Bone Resorption in vitro

The Effect of Dental Plaque Grown in the Presence of Xylitol or Sucrose on Bone Resorption <i>in vitro</i>

Comparative effects of the substance-sweeteners glucose, sorbitol, sucrose, xylitol and trichlorosucrose on lowering of pH by two oral Streptococcus mutans strains in vitro

Streptococcus mutans suspensions were incubated with each of the sweeteners in suspending media of increasing complexity. The lower pH levels were generally attained by sucrose and glucose; higher pH values were obtained with sorbitol, trichlorosucrose, and xylitol, respectively. The acidogenicity of sucrose was more often decreased by the presence of trichlorosucrose than by the presence of xylitol.

The amino acid composition ofStreptococcus mutansand its culture medium supplemented with xylitol

The amino and keto acid composition of the cells of Streptococcus mutans, strain Ingbritt, maintained and grown on a Trypticase-Phytone based medium without any added carbohydrate or supplemented with xylitol or glucose, was analyzed. The results showed no remarkable differences in the portions of individual amino acids liberated by acid hydrolysis from the cellular proteins of cells grown in the above mentioned media. However, the amount of free amino acids in the water extracts of the cells grown in the glucose medium differed considerably from those obtained from cells grown in the two other media. The amounts of free amino acids of the medium at the end of the growth period were higher in the glucose containing medium than in the two other media. The content of keto acids was lower in the cells grown in the presence of xylitol or without added carbohydrate when compared to those cells grown in glucose containing medium.

Effect of xylitol on the growth of three oral strains of Candida albicans.

An earlier report claimed that the cells of Streptococcus mutans became adapted to the use of xylitol after a period of four to five months (MAKINEN, J Dent Res 51: 403-408, 1972). Later experiments showed that no adaptation to the metabolizing of this polyol was involved. It was emphasized that the cells became adapted to "tolerate" the presence of xylitol (MAKINEN, in Sugars in Nutrition, 1974). Recent experiments indicate that S mutans, in the presence of xylitol

Inhibition of ribulose diphosphate carboxylase/oxygenase by xylitol 1,5-diphosphate

Xylitol 1,5-diphosphate is a potent inhibitor of RuDP carboxylase/oxygenase activity. At an enzyme concentration of 400 μg/ml (less than 1 μM), RuDP carboxylase is inhibited 50% by 2 μM xylitol 1,5-diphosphate. The molar ratio of effector to enzyme is 2 to 1. The inhibition with respect to RuDP is mixed uncompetitive and noncompetitive. The response of enzyme to bicarbonate in the presence of xylitol 1,5-diphosphate is sigmoidal. The inhibition is time-dependent and pH-dependent, with a pK a for inhibition of about 8.6. RuDP oxygenase is inhibited at pH values below 9, but at pH 9 or above, neither RuDP oxygenase nor carboxylase is significantly inhibited. Xylitol 1,5-diphosphate does not produce a differential regulation of the RuDP carboxylase and oxygenase activities.