Release Of Fructose Research Articles

The catalytic domains of Streptococcus mutans glucosyltransferases: a structural analysis.

Streptococcus mutans, found in the human oral cavity, is a significant contributor to the pathogenesis of dental caries. This bacterium expresses three genetically distinct types of glucosyltransferases named GtfB (GTF-I), GtfC (GTF-SI) and GtfD (GTF-S) that play critical roles in the development of dental plaque. The catalytic domains of GtfB, GtfC and GtfD contain conserved active-site residues for the overall enzymatic activity that relate to hydrolytic glycosidic cleavage of sucrose to glucose and fructose, release of fructose and generation of a glycosyl-enzyme intermediate in the reducing end. In a subsequent transglycosylation step, the glucosyl moiety is transferred to the nonreducing end of an acceptor to form a growing glucan polymer chain made up of glucose molecules. It has been proposed that both sucrose breakdown and glucan synthesis occur in the same active site of the catalytic domain, although the active site does not appear to be large enough to accommodate both functions. These three enzymes belong to glycoside hydrolase family 70 (GH70), which shows homology to glycoside hydrolase family 13 (GH13). GtfC synthesizes both soluble and insoluble glucans (α-1,3 and α-1,6 glycosidic linkages), while GtfB and GtfD synthesize only insoluble or soluble glucans, respectively. Here, crystal structures of the catalytic domains of GtfB and GtfD are reported. These structures are compared with previously determined structures of the catalytic domain of GtfC. With this work, apo structures and inhibitor-complex structures with acarbose are now available for the catalytic domains of GtfC and GtfB. The structure of GtfC with maltose allows further identification and comparison of active-site residues. A model of sucrose binding to GtfB is also included. The new structure of the catalytic domain of GtfD affords a structural comparison of the three S. mutans glycosyltransferases. Unfortunately, the catalytic domain of GtfD is not complete since crystallization resulted in the structure of a truncated protein lacking approximately 200 N-terminal residues of domain IV.

How is Trehalulose Formed by Australian Stingless Bees? - An Intermolecular Displacement of Nectar Sucrose.

Trehalulose, a rare sucrose isomer, is a dominant sugar in stingless bee honey, with traces of the trisaccharide erlose. Incubating sucrose solutions with macerated stingless bee parts (head, thorax, and abdomen) from Tetragonula carbonaria, we observed that sucrose isomerization occurs predominantly in the head incubations, with trehalulose constituting 76.2-80.0% of total detected sugar. By contrast, sucrose hydrolysis occurred in stingless bee abdomen incubations, with glucose and fructose observed as 48.6-51.7% and 48.3-49.7%, respectively, of total detected sugar. Incubating glucose/fructose (1:1) solutions with any bee part did not result in trehalulose formation. In addition, by tracing the 13C isotope-labeled monosaccharide moieties throughout the isomerization from sucrose to trehalulose and erlose, for the first time, the mechanism was established as an enzymatic double displacement reaction. Sucrose acts as a glucose donor giving a β-d-glucosyl enzyme intermediate with fructose release as demonstrated by mixed isotope products. Glucosylation of fructose (inter- or intramolecularly) with isomerization forms trehalulose (favorable), while glucosylation of sucrose forms erlose (less favorable).

Magnetically assisted commercially attractive chemo-enzymatic route for the production of 5-hydroxymethylfurfural from inulin

Biomass has recently been extensively exploited as an attractive alternative to the scarce fossil fuels and has been widely used for the production of fine chemicals. The present work defines an integrated approach for high yield of 5-hydroxymethylfurfural (HMF) from inulin via a two-step process. Initially, insolubilization of inulinase from Aspergillus niger on chitosan-coated magnetic nanoparticles (cMNP) was achieved, and under optimal conditions, hydrolysis of 5% inulin by immobilized inulinase (2.5 U/mg) led to fructose release of 35.8 g/L at pH 6.0 and temperature 60 °C in 3 h. Subsequently, the fructose was dehydrated to HMF in the presence of TiO2-magnetic silica spheres (TiO2-MSS). Upon optimizing the process parameters, HMF yield of 96.58% was achieved with 15% fructose in 15 min at 140 °C with a TiO2-MSS load of 10% (w/w). Concerning the operational stability of the immobilized biocatalyst/catalyst, the immobilized inulinase was recycled up to 10 cycles of inulin hydrolysis with 9.2 g/L of fructose released at the 10th cycle. TiO2-MSS also showed high operational stability and were recycled up to 10 cycles of HMF production with 5.4 g/L of HMF produced at 10th cycle. The mass balance for inulin to HMF was performed which showed the efficiency of the process.

Identification of a Novel Oligosaccharide in Maple Syrup as a Potential Alternative Saccharide for Diabetes Mellitus Patients

The incidence of diabetes mellitus (DM) is increasing rapidly and is associated with changes in dietary habits. Although restrictions in the use of sweeteners may prevent the development of DM, this might reduce the quality of life of patients with DM. Therefore, there has been a great deal of research into alternative sweeteners. In the search for such sweeteners, we analyzed the carbohydrate content of maple syrup and identified a novel oligosaccharide composed of fructose and glucose, linked at the C-4 of glucose and the C-6 of fructose. This oligosaccharide inhibited the release of fructose from sucrose by invertase (IC50: 1.17 mmol/L) and the decomposition of maltose by α-(1-4) glucosidase (IC50: 1.72 mmol/L). In addition, when orally administered together with sucrose to rats with DM, the subsequent plasma glucose concentrations were significantly lower than if the rats had been administered sucrose alone, without having any effect on the insulin concentration. These findings suggest that this novel oligosaccharide might represent a useful alternative sweetener for inclusion in the diet of patients with DM and may also have therapeutic benefits.

Promotion catalytic role of ethanol on Brønsted acid for the sequential dehydration-etherification of fructose to 5-ethoxymethylfurfural

The function of ethanol and Brønsted acid for the dehydration-etherification of fructose to 5-ethoxymethylfurfural (EMF) has been theoretically investigated at G4 level in ethanol solution. Initially, fructose prefers to dehydration other than etherification with ethanol in the presence of Brønsted acid. The proton H+ should be solvated on ethanol other than fructose, resulting in [C2H5OH2]+ as the catalytically active species. Both protonation and [C2H5OH2]+ exhibit good catalytic performance, but ethanol does not. Furthermore, [C2H5OH2]+ displays better catalytic performance than protonation, which reflects the promotion catalytic role of C2H5OH on the proton H+. The turnover frequency analysis shows that the SN2 nucleophilic substitution for the etherification of 5-hydroxymethylfurfural (HMF) to EMF is the rate-controlling step in the whole reaction catalyzed by [C2H5OH2]+. The catalytic performance of [C2H5OH2]+ stems from the positive charge of OH2 group, which assists both the H2O release of fructose to HMF and the etherification of HMF to EMF. For the dehydration of fructose, the catalytic superiority of [C2H5OH2]+ to protonation comes from the shift of the intramolecular H-shift catalyzed by protonation to the intermolecular H-shift catalyzed by [C2H5OH2]+, which lowers the activation energy barrier for the H2O release. The present study is useful for understanding the roles of ethanol and Brønsted acid in acid-catalyzed dehydration-etherification of carbohydrates to biofuel in ethanol solution.

Process options for conversion of Agave tequilana leaves into bioethanol

This paper reports on mild acid pretreatment options for the conversion of Agave tequilana leaves into composite sugars for ethanol fermentation. The effect of five different pretreatment conditions (time, temperature and acid concentrations) were assessed in terms of cellulose digestibility, hemicellulose solubilisation and lignin content in leaves of 1.25 years old A. tequilana plants from Rockhampton and 2.5 year plants from Kalamia. Dilute acid pretreatment and enzyme saccharification of A. tequilana leaf bagasse significantly improved total glucose recovery. A recovery of 273mg/g (70% theoretical) was attained when the bagasse was pretreated with 2.0% H2SO4 for 60min at 121°C and saccharified with 6% (w/w) CTec 2. Saccharomyces cerevisiae efficiently fermented crude A. tequilana bagasse and juice hydrolysates within 13h and 7h respectively, yielding up to 38.6g/L and 12.4g/L. This corresponds to glucose to ethanol conversion rate of 68 and 61% for A. tequilana leaf bagasse and juice, respectively. With further developments, including fermentation of C5 sugars and inulinase saccharification of juices (release of fructose), this process could deliver greater yields, reinforcing its potential as a biofuel feedstock.

Rapid and efficient bioconversion of chicory inulin to fructose by immobilized thermostable inulinase from Aspergillus tubingensis CR16

Abstract Background Fructose, a monosaccharide, has gained wide applications in food, pharmaceutical and medical industries because of its favourable properties and health benefits. Biocatalytic production of fructose from inulin employing inulinase is the most promising alternative for fructose production. For commercial production, use of immobilized inulinase is advantageous as it offers reutilization of enzyme and increase in stability. In order to meet the demand of concentrated fructose syrup, inulin hydrolysis at high substrate loading is essential. Results Inulinase was immobilized on chitosan particles and employed for fructose production by inulin hydrolysis. Fourier transform infrared spectroscopy (FTIR) analysis confirmed linkage of inulinase with chitosan particles. Immobilized biocatalyst displayed significant increase in thermostability at 60 and 65 °C. Statistical model was proposed with an objective of optimizing enzymatic inulin hydrolytic process. At high substrate loading (17.5 % inulin), using 9.9 U/g immobilized inulinase at 60 °C in 12 h, maximum sugar yield was 171.1 ± 0.3 mg/ml and productivity was 14.25 g/l/h. Immobilized enzyme was reused for ten cycles. Raw inulin from chicory and asparagus was extracted and supplied in 17.5 % for enzymatic hydrolysis as a replacement of pure inulin. More than 70 % chicory inulin and 85 % asparagus inulin were hydrolyzed under optimized parameters at 60 °C. Results of high performance liquid chromatography confirmed the release of fructose after inulin hydrolysis. Conclusions The present findings prove potentiality of immobilized thermostable inulinase from Aspergillus tubingensis CR16 for efficient production of fructose syrup. Successful immobilization of inulinase on chitosan increased its stability and provided the benefit of enzyme reutilization. Box-Behnken design gave a significant model for inulin hydrolysis. Extraction of raw inulin from chicory and asparagus and their enzymatic hydrolysis using immobilized inulinase suggested that it can be a remarkable cost-effective process for large-scale fructose production.

Effect of inulin on the growth and metabolism of a probiotic strain of Lactobacillus rhamnosus in co-culture with Streptococcus thermophilus

Metabolic studies are very important to improve quality of functional dairy products. For this purpose, the behaviors of pure cultures of Streptococcus thermophilus (St) and Lactobacillus rhamnosus (Lr) as well a co-culture of them (St–Lr) were investigated during skim milk fermentation, and the inulin effect as prebiotic was assessed. Lr was able to metabolize 6 g/100 g more galactose than St and St–Lr. Final lactic acid production by Lr was higher (9.8 g/L) compared to St (9.1 g/L) and St–Lr (9.1 g/L). Acetic acid concentration varied from 0.8 g/L (St–Lr) to 1.5 g/L (Lr) and that of ethanol from only 0.2 g/L (St–Lr) to 0.4 g/L (Lr), which suggests the occurrence in Lr of a NADH oxidase activity and citrate co-metabolization via pyruvate, both dissipating a part of the reducing power. Diacetyl and acetoin accumulated at the highest levels (18.4 and 0.8 mg/L, respectively) with St–Lr, which suggests possible synergistic interactions between these microorganisms as well as the Lr capability of co-metabolizing citrate in the presence of lactose. Inulin stimulated both biomass growth and levels of all end-products, as the likely result of fructose release from its partial hydrolysis and subsequent metabolization as an additional carbon and energy source.

β-Fructofuranosidase and sucrose phosphorylase of rumen bacterium Pseudobutyrivibrio ruminis strain 3

The subject of this study was the fructan and sucrose degrading enzymes of bacterium Pseudobutyrivibrio ruminis strain 3. It was stated that cell extract from bacteria growing on inulin contained β-fructofuranosidase (EC 3.2.1.80 and/or EC 3.2.1.26) and sucrose phosphorylase (EC 2.4.1.7), while the bacteria maintained on sucrose showed only phosphorylase. Partially purified β-fructofuranosidase digested inulooligosaccharides and sucrose to fructose or fructose and glucose, respectively, but was unable to degrade the long chain polymers of commercial inulin and Timothy grass fructan. Digestion rate of inulooligosaccharides fit Michaelis-Menten kinetics with V(max) 5.64 μM/mg/min and K(m) 1.274%, respectively, while that of sucrose was linear. Partially purified sucrose phosphorylase digested only sucrose. The digestion products were fructose, glucose-1P and free glucose. The reaction was in agreement with Michaelis-Menten kinetics. The V(max) were 0.599 and 0.584 μM/mg/min, while K(m) were 0.190 and 0.202% for fructose release and glucose-1P formation, respectively, when bacteria grew on inulin. The V(max) were, however, 1.37 and 1.023 μM/mg/min, while K(m) were 0.264 and 0.156%, if bacteria were grown on sucrose. The free glucose was hardly detectable for the enzyme originated from inulin grown bacteria, but glucose levels ranged from 0.05 to 0.25 μM/mg/min, when cell extract from bacteria grown on sucrose was used. Release of free glucose was observed when no inorganic phosphate was present in reaction mixture.

Phosphorolytic cleavage of sucrose by sucrose-grown ruminal bacterium Pseudobutyrivibrio ruminis strain k3

Rumen bacterium Pseudobutyrivibrio ruminis strain k3 utilized over 90% sucrose added to the growth medium as a sole carbon source. Zymographic studies of the bacterial cell extract revealed the presence of a single enzyme involved in sucrose digestion. Thin layer chromatography showed fructose and glucose-1-phosphate (Glc1P) as end products of the digestion of sucrose by identified enzyme. The activity of the enzyme depended on the presence of inorganic phosphate and was the highest at the concentration of phosphate 56 mmol/L. The enzyme was identified as the sucrose phosphorylase (EC 2.4.1.7) of molar mass approximately 54 kDa and maximum activity at pH 6.0 and 45 degrees C. The calculated Michaelis constant (Km) for Glc1P formation and release of fructose by partially purified enzyme were 4.4 and 8.56 mmol/L while the maximum velocities of the reaction (Vlim) were 1.19 and 0.64 micromol/L per mg protein per min, respectively.

Sucrose phosphorylase of the rumen bacteriumPseudobutyrivibrio ruminisstrain A

To verify the taxonomic affiliation of bacterium Butyrivibrio fibrisolvens strain A from our collection and to characterize its enzyme(s) responsible for digestion of sucrose. Comparison of the 16S rRNA gene of the bacterium with GenBank showed over 99% sequence identity to the species Pseudobutyrivibrio ruminis. Molecular filtration, native electrophoresis on polyacrylamide gel, zymography and thin layer chromatography were used to identify and characterize the relevant enzyme. An intracellular sucrose phosphorylase with an approximate molecular mass of 52 kDa exhibiting maximum activity at pH 6.0 and temperature 45 degrees C was identified. The enzyme was of inducible character and catalysed the reversible conversion of sucrose to fructose and glucose-1-P. The reaction required inorganic phosphate. The K(m) for glucose-1-P formation and fructose release were 3.88 x 10(-3) and 5.56 x 10(-3) mol l(-1) sucrose, respectively - while the V(max) of the reactions were -0.579 and 0.9 mumol mg protein(-1) min(-1). The enzyme also released free glucose from glucose phosphate. Pseudobutyrivibrio ruminis strain A utilized sucrose by phosphorolytic cleavage. Bacterium P. ruminis strain A probably participates in the transfer of energy from dietetary sucrose to the host animal.

Electron microscopic detection and activity of glucosyltransferase B, C, and D in the in situ formed pellicle

Objective Glucosyltransferases (GTFs) represent a virulence factor of mutans streptococci. The aim of the present in situ study was to investigate the distribution of different GTF-isoforms in the pellicle. Design Bovine enamel slabs were fixed on buccal and palatal sites of individual splints worn by five subjects for 30 and 120 min to allow pellicle formation. Pellicle specimens were processed for transmission electron microscopy (TEM) and field emission in-lens scanning electron microscopy (FEI-SEM). Gold-immunolabelling was used for detection of GTF-isoforms B, C and D. Furthermore, glucosyltransferase activity of 3-, 30- and 120-min pellicles was tested via determination of fructose release. Results All isoforms of the enzyme were found to be randomly distributed within all layers of the pellicle. In cross-sections (TEM), GTF D was the most abundant isoform. More labelled molecules were detected on buccal sites compared with palatal surfaces, the number of molecules detected increased with time. The amount of GTF B, C and D found on the pellicle surface by FEI-SEM showed no correlation with pellicle formation time or localisation in the oral cavity. Overall, GTF D was detected more frequently on the surface than GTF B and C. All pellicles tested showed GTF-activity. Conclusion The study shows for the first time the presence of the GTF-isoforms B, C and D within all layers of the in situ formed pellicle. This emphasises the impact of streptococcal products on the composition of the pellicle and illustrates a mechanism used by bacteria to colonize dental surfaces.

Development of a rapid, quantitative glucosyltransferase assay based on a screen‐printed fructose enzyme electrode and application to optimization studies on gtfD expression in recombinant Escherichia coli

A biosensor for fructose determination was used as basis of an assay for the determination of glucosyltransferase (GTF) activities and applied to monitoring recombinant enzyme production. GTFs catalyze the synthesis of glucans from sucrose leading to the release of fructose. Specific fructose determinations in the microM concentration range were achieved with a fructose electrode based on fructose dehydrogenase, which was immobilized on a screen-printed platinum electrode. This electrode was used as basis of the new assay for GTF activity determinations. Depending on the amount of enzyme, the assay was completed within 15-30 min compared to 1-2 h for the traditional photometric assay. From the amount of fructose released in a given reaction time, GTF activities were determined down to approx. 20 U/L. Even unpurified samples from a recombinant GTF-S production process could be analyzed without any problems, and a good correlation was obtained to data obtained from the photometric assay. Analysis of samples from cultures of various rGTF-S-producing recombinant E. coli strains grown on different media with SDS-PAGE and with the new assay identified the same strain and culture medium as optimum for recombinant GTF-S production.

Kinetic study of the acid hydrolysis of various oligofructose samples.

The kinetic of acid hydrolysis of five commercially available oligofructose samples used as food ingredients has been investigated as a function of the dry matter concentration, reaction pH, and temperature. The initial fructose release rate is found to be roughly proportional to the inverse of the average polymerization degree in number. A pseudo first order kinetic is found with respect to the fructosyl chain end concentration and to the proton concentration. Arrhenius plot is found to reasonably fit the data in a relatively wide temperature range (7-130 degrees C). The results allow the estimation of the fructose release rate in many foodstuff processing conditions.

Seasonal variation of fructan‐β‐fructosidase (FEH) activity and characterization of a β‐(2‐1)‐linkage specific FEH from tubers of Jerusalem artichoke (Helianthus tuberosus)

SUMMARYThe fructan‐β‐fructosidase activity (1‐FEH; EC 3.2.1.80) that degrades inulin in tubers of Helianthus tuberosus L. appears to be developmentally regulated; it was low in growing tubers but increased during dormancy and sprouting. In spite of relatively high 1‐FEH activity in vitro, fructose concentration was very low in developing and dormant tubers and increased markedly only during sprouting. A fructan‐β‐fructosidase from such sprouting tubers was purified 41‐fold to a single protein band on one‐dimensional sodium dodecylsulphate‐polyacrylamide gels. The purification procedure included ammonium sulphate precipitation, lectin‐affinity chromatography on concanavalin A, anion‐exchange and cation‐exchange chromatography. The enzyme had an apparent molecular mass of 75000 measured by size‐exclusion chromatography, and 79000 measured by one‐dimensional sodium dodecylsulphate‐polyacrylamide gel electrophoresis. It exhibited a high substrate specificity, hydrolysing terminal β‐(2‐1)‐fructosyl‐fructose‐linkages in linear and branched fructan oligomers; β‐(2‐6)‐linkages were hardly hydrolysed. Hydrolysis of inulin oligomers followed normal saturation kinetics: Km values for 1,1‐kestotetraose and 1,1,1‐kestopentaose were 8‐3 mM and 12 mM, respectively. Fructosyl residues were hydrolysed from inulin oligomers by a multi‐chain mechanism. The fructan‐β‐fructosidase showed optimal enzyme activity at pH 5–2, and it retained its full activity after pre‐incubation for 1 h at up to 40°C. The release of fructose from 5 mm 1,1‐kestotetraose was reduced by 25 % when 1‐FEH was assayed in the presence of 10 mM sucrose. It is proposed that the inhibition of 1‐FEH activity by sucrose is a mechanism for controlling fructan degradation in planta.

Release of fructose and hexose phosphates from perivascular cells induced by low density lipoprotein and acceleration of protein glycation in vitro

We investigated whether low density lipoprotein (LDL) under oxidative stress might induce the release of fructose, glucose-6-phosphate and fructose-6-phosphate from perivascular cells, and also whether these substances might accelerate the formation of advanced glycation end products (AGE) from proteins in vitro. When vascular smooth muscle cells were incubated with LDL in Ham's F10 at 37 °C for 48 h, release of all these substances was increased dose-dependently by oxidized LDL. Fructose release was increased in a dose-dependent manner by glucose. Indomethacin (20 μM) significantly ( P < 0.01) suppressed the release of fructose (25.4 ± 15.7% of control) and hexose phosphates (29.4 ± 4.0) with the inhibition of release of lactate dehydrogenase (35.5 ± 4.9) as well as probucol, whereas an aldose reductase inhibitor, epalrestat, significantly ( P < 0.001) inhibited only the fructose release (0.9 ± 0.8). Release of fructose and hexose phosphates from vascular endothelial cells was also induced by oxidized LDL. AGE immunoreactivities and AGE-related fluorescence formed from proteins and glucose were significantly increased ( P < 0.001) in the presence of small amounts of the cellular glucose metabolites (6.6%) with glucose (93.4%). These data suggest that release of potent AGE initiators, fructose and hexose phosphates, from perivascular cells induced by oxidized LDL may be an important phenomenon for vascular complications.

Relationship between Structure and Antigenicity of O1 Vibrio cholerae Lipopolysaccharides

The relationship between the release of fructose from O1 Vibrio cholerae lipopolysaccharides (LPS) by dilute acetic acid hydrolysis and the decrease in their antigenicity was examined. Decrease in the antigenicity of LPS was not parallel with the release of fructose, and occurred very much later than the latter. Periodate oxidation of LPS resulted in the total elimination of the fructose and glucose, and two-thirds of the heptose constituents, but no difference in the antigenicity of LPS was observed before and after oxidation. These findings indicate that the fructose present in O1 V. cholerae LPS is not substantially involved in their specific antigenicity. In the O1 V. cholerae LPS, the fructose is in the branch structure, most probably in the core region.

Intraischemic metabolic effects of different disaccharides on protected canine kidneys

The addition of the disaccharides maltose (10, 20, 30 mM) and sucrose (30, 60 mM) to Bretschneider's organ protective HTK solution was evaluated to improve renal protection by an enhanced glycolytic energy supply. Canine kidneys were perfused for 8 min with either HTK solution or HTK solution containing additional disaccharides. After nephrectomy the kidneys were incubated at 25 degrees C and metabolic parameters were determined at regular intervals. Maltose and sucrose are slowly cleaved during renal ischemia but maltose distinctly faster than sucrose. Maltose increases intraischemic ATP supply. However, 30 mM maltose was no better than 10 mM. 60 mM sucrose was about as effective for glycolysis as 10 mM maltose. However, possibly due to fructose release there was an accelerated decrease of adenine nucleotides with sucrose. Although fructose enters glycolysis it seems to have negative side-effects. Hence, probably neither sucrose nor fructose are appropriate for renal substrate supply during ischemia.

Gibberellin-induced hydrolysis of endosperm cell walls in gibberellin-deficient tomato seeds prior to radicle protrusion

The weakening of the mechanical restraint of the endosperm layer in tomato (Lycopersicon esculentum Mill.) seeds, a prerequisite for germination, has been studied with the use of seeds of the gibberellin (GA)-deficientgib-1 mutant. Incubation ofgib-1 endosperms, including part of the testa, in 10 μM GA4+7, resulted within 12 h in the release of fructose, glucose, galactose and mannose into the incubation medium. Only small amounts of sugars diffused out of thegib-1 endosperms during incubation in water. Chemical hydrolysis of endosperm cell walls ofgib-1 seeds showed that they are mainly composed of mannose, and smaller quantities of glucose and galactose. Treatment with GA4+7 induced in the endosperms the production of endo-β-mannanase activity that was not detectable during incubation in water, and also increased the activities of mannohydrolase and α-galactosidase as compared with the water controls. No cellulase activity was found. It is concluded that in tomato seeds the weakening of endosperms prior to radicle protrusion is mediated by a GA-induced enzymatic degradation of the mannan-rich cell walls.

Studies of the Effects of an Elevated Glucose Concentration on the Ultrastructure and Composite Metabolism of the Intact Rabbit Aortic Intima-Media Preparation

To assess whether the ultrastructure and composite metabolism of the arterial wall are subject to acute modification by exposure to an elevated glucose concentration, paired samples of the intact rabbit aortic intima-media preparation were incubated with 5 mM or 20 mM glucose for 1 h under conditions in which the ultrastructural and metabolic stability of samples incubated with 5 mM glucose has previously been demonstrated. Incubation with 20 mM glucose did not alter the qualitative electron-microscopic appearance of the tissue, and no significant changes were found on a quantitative morphometric evaluation of the aortic endothelium. A detectable, composite-free intracellular glucose concentration was not demonstrable in samples incubated with 5 mM or 20 mM glucose by a comparison of the ratio of glucose space:inulin space. Samples incubated with 5 mM glucose had significant sorbitol concentrations and unexpectedly high fructose concentrations but did not release fructose into the medium at detectable rates. Incubation with 20 mM glucose resulted in an increase in tissue sorbitol without an increase in tissue fructose or a detectable rate of fructose release. Tissue water content and myoinositol concentration were unaltered by incubation with 20 mM glucose, as were the rates of O2 uptake and lactate production whether the gas phase was 5% CO2:95% O2 or 5% CO2:95% air. The ultrastructure' of intact aortic intima-media is not acutely altered by exposure to an elevated glucose concentration. Ambient glucose concentration is not the primary determinant of composite-free intracellular glucose concentration in arterial wall, and an increase in medium glucose concentration does not result in a significant increase in glucose utilization via the polyol pathway or induce the concomitants of increased polyol pathway activity demonstrable in a conventionally prepared aortic intima-media preparation.