Study of developmental changes on hexoses metabolism in rat cerebral cortex

We have studied the developmental changes of glucose, mannose, fructose and galactose metabolism in rat cerebral cortex. As the animals aged, glucose, mannose and fructose oxidation to CO₂ increased, whereas galactose oxidation decreased. Lipid synthesis from glucose and fructose also increased with age, that from mannose decreased and galactose did not change. Cytochalasin B, a potent non-competitive inhibitor of sodium-independent glucose transport, significantly impaired glucose, mannose and galactose metabolism, but had no effect on fructose metabolism. Both galactose or fructose did not change, whereas mannose declined the glucose metabolism. Glucose decreased fructose, galactose and mannose metabolism. Our results show that besides glucose, the metabolism of mannose, galactose and fructose present developmental changes from fetal to adult age, and reinforce the literature data indicating that mannose and galactose are transported by glucose carriers, while fructose is not.     

Compared roles of glucose, galactose and fructose as glycogen precursors during the acute response to insulin in cultured rat foetal hepatocytes

1. The efficiency of the contribution of hexoses to basal- and stimulated-glycogenesis, when studied in cultured 18 day-old rat foetal hepatocytes in the presence of glucose, was as follows: galactose greater than glucose greater than fructose. 2. Glucose deprivation had opposite effects on the contributions of [14C]galactose (decreased) and [14C]fructose (increased) to glycogenesis, which occurred independently of insulin and were reversed by glucose concentrations as low as 30-100 microM. 3. The stimulation of glycogenesis by insulin measured with [14C]glucose (3.2-fold) was superior to that obtained with either [14C]galactose or [14C]fructose (2.7-fold in both cases), which revealed a specific beneficial effect of insulin on glucose contribution.     

Characterization and regulation of galactose transport in Neurospora crassa

Two galactose uptake systems were found in the mycelia of Neurospora crassa. In glucose-grown mycelia, galactose was transported by a low-affinity (Km = 400 mM) constitutive system which was distinct from the previously described glucose transport system I (R. P. Schneider and W. R. Wiley, J. Bacteriol. 106:479--486, 1971). In carbon-starved mycelia or mycelia incubated with galactose, a second galactose transport activity appeared which required energy, had a high affinity for galactose (Km = 0.7 mM), and was shown to be the same as glucose transport system II. System II also transported mannose, 2-deoxyglucose, xylose, and talose and is therefore a general monosaccharide transport system. System II was derepressed by carbon starvation, completely repressed by glucose, mannose, and 2-deoxyglucose, and partially repressed by fructose and xylose. Incubation with galactose yielded twice as much activity as starvation. This extra induction by galactose required protein synthesis, and represented an increase in activity of system II rather than the induction of another transport system. Glucose, mannose, and 2-deoxyglucose caused rapid degradation of preexisting system II; fructose and xylose caused a slower degradation of activity.     

Kinetic characteristics of the two glucose transport systems in Neurospora crassa

Glucose is transported across the cell membrane of Neurospora crassa by two physiologically and kinetically distinct transport systems. System II is repressed by growth of the cells in 0.1 m glucose. System I is synthesized constitutively. The apparent K(m) for glucose uptake by system I and system II are 25 and 0.04 mm, respectively. Both uptake systems are temperature dependent, and are inhibited by NaN(3) and 2,4-dinitrophenol. Glucose uptake by system II was not inhibited by fructose, galactose, or lactose. However, glucose was shown to be a noncompetitive inhibitor of fructose and galactose uptake. The transport rate of [(14)C]3-0-methyl-d-glucose (3-0-MG) was higher in cells preloaded with unlabeled 3-0-MG than in control cells. The rate of entry of labeled 3-0-MG was only slightly inhibited by the presence of NaN(3) in the medium. Further, NaN(3) caused a rapid efflux of accumulated [(14)C]3-0-MG. These data imply that the energetic step in the transport process prevents efflux.     

A useful model to study the effect of high sugar concentrations upon growth and enzymic activities of toad embryos and larvae

The aim of this study was to develop an oviparous model suitable for studying the differential effects and mechanisms by which a high concentration of extracellular glucose and other sugars produce diabetes complications, particularly body growth retardation during development. Hence, we studied the experimental conditions necessary to obtain measurable effects of high sugar concentrations (5-mM glucose, mannitol, fructose and galactose) upon body growth and development of Bufo arenarum embryos and larvae, and upon the activity of aspartate aminotransferase (AST), gamma-glutamyltransferase (GGT), and alkaline phosphatase (APP). Unfed animals kept in glucose showed lower body weight than controls at all stages, a condition only observed at stage 26 for animals kept in galactose and fructose. All animals reached the same stage of development regardless of the solution in which they were kept. Glucose and fructose significantly decreased the activity of all enzymes tested, while galactose only affected GGT activity. The model provides the first experimental evidence for the deleterious effect exerted in vivo by different sugars upon developing embryos and larvaes of Bufo arenarum. The results prove that this model might help to elucidate the effects and the pathogenic mechanisms of hyperglycemia upon growth and development of embryos exposed to environments with high sugar concentrations. It might also become a useful tool for testing the effectiveness of drugs designed to prevent the deleterious effect of such exposure.     

Sugars and Organic Acids of Vitis vinifera

Glucose, fructose, galactose, sucrose, maltose, melibiose, raffinose, and stachyose were identified in the leaves, bark, roots, and berries of Vitis vinifera L. var. Thompson Seedless. In addition to these sugars, verbascose and manninotriose were found in the leaves and bark.

Malic, tartaric, citric, isocitric, ascorbic, cis-aconitic, oxalic, glycolic, glyoxylic, succinic, lactic, glutaric, fumaric, pyrrolidone carboxylic, α-ketoglutaric, pyruvic, oxaloacetic, galacturonic, glucuronic, shikimic, quinic, chlorogenic, and caffeic acids were identified in the leaves, bark, roots, and berries.

Glucose, fructose, sucrose, malate, tartrate, and citrate were determined quantitatively in the leaf, petiole, xylem, bark, tendril, bud, puduncle pedicel, berry, lateral roots, and main roots at 4 separate physiological stages of growth. In addition, changes in the concentrations of fructose, glucose, malate, and tartrate in leaves were measured during a 36-day period starting from budburst.

     

Plasmodium gallinaceum: development in Aedes aegypti maintained on various carbohydrate diets

Fructose, galactose, glucose, maltose, melibiose, and trehalose were evaluated for their nutritional and survival values for Aedes aegypti. The development of the exogenous stages of Plasmodium gallinaceum was evaluated by counting and averaging the number of oocysts developing on the midguts of Aedes aegypti.The nutritional and survival value for A. aegypti was greatest on glucose, sucrose, and fructose and lowest on galactose and melibiose. Mosquitoes maintained on fructose produced the greatest numbers of oocysts. Only two other sugars, galactose and melibiose produced more oocysts than their respective controls. Glucose and maltose, both of which had high nutritive and survival value for A. aegypti were less efficient than the control (sucrose) for the development of P. gallinaceum.     

Differential sensitivities to glucose and galactose repression of gluconeogenic and respiratory enzymes from Saccharomyces cerevisiae

The synthesis of isocitrate lyase was induced by the presence of ethanol in the chemostat reaching a specific activity of 200 mU·mg^-1 at this induced state. In glucoselimited, derepressed cells, 20 mU·mg^-1 were detected and under repressed conditions isocitrate lyase activity was not detected.The sensitivity of gluconeogenic enzymes: cytoplasmic malate dehydrogenase; fructose 1,6-bisphosphatase and isocitrate lyase as well as the mitochondrial enzymes NADH dehydrogenase and succinate cytochrome c oxidase to glucose and galactose repression were studied in chemostat cultures. Our results show that galactose was less effective as a repressor than glucose. Malate dehydrogenase was completely inactivated by glucose, whereas galactose only produced a 78% decrease of specific activity. Fructose 1,6-bisphosphatase and isocitrate lyase were completely inactivated by both sugars but at different rate. Glucose produced an 85% decrease of specific activity of the mitochondrial enzymes whereas galactose only decrease an 67%.     

Glucose Inhibits ACC Oxidase Activity and Ethylene Biosynthesis in Ripening Tomato Fruit

Experiments were carried out to evaluate the effect of glucose on ripening and ethylene biosynthesis in tomato fruit (Lycopersicon esculentum Mill.). Fruit at the light-red stage were vacuum infiltrated with glucose solutions post-harvest and changes in 1-aminocyclopropane-1-carboxylic acid (ACC) synthase, ACC, ACC oxidase, and ethylene production monitored over time. ACC oxidase activity was also measured in pericarp discs from the same fruits that were treated either with glucose, fructose, mannose, or galactose. While control fruit displayed a typical peak of ethylene production, fruit treated with glucose did not. Glucose appeared to exert its effect on ethylene biosynthesis by suppressing ACC oxidase activity. Fructose, mannose, and galactose did not inhibit ACC oxidase activity in tomato pericarp discs. Glucose treatment inhibited ripening-associated colour development in whole fruit. The extent of inhibition of colour development was dependent upon the concentration of glucose. These results indicate that glucose may play an important role in ethylene-associated regulation of fruit ripening.     

Nutrient requirements in germination of conidiospores of Aspergillus niger V. Tieghen

Germination of conidiospores of A. niger was stimulated by the addition of some carbohydrates and nitrogenous compounds to the growth medium. Glucose at 0.5 and 1.0 mg/ml concentrations was the best sugar for germination. Sucrose and fructose were the second best. Glutamic acid and valine were the most effective nitrogenous compounds for the spore germination of the fungus. Glucose, sucrose, and glutamic acid initiated germination within the first 6 h of incubation. When cellobiose, galactose and valine were used germination started after 9 h. With leucine, cysteine and arginine, however, not only the percentage of germination reduced, but also the latent period increased, significantly. A period of 15 h was needed before germination had been started.     

Extracellular melibiose and fructose are intermediates in raffinose catabolism during fermentation to ethanol by engineered enteric bacteria.

Contrary to general concepts of bacterial saccharide metabolism, melibiose (25 to 32 g/liter) and fructose (5 to 14 g/liter) accumulated as extracellular intermediates during the catabolism of raffinose (O-alpha-D-galactopyranosyl-1, 6-alpha-D-glucopyranosyl-beta-D-fructofuranoside) (90 g/liter) by ethanologenic recombinants of Escherichia coli B, Klebsiella oxytoca M5A1, and Erwinia chrysanthemi EC16. Both hydrolysis products (melibiose and fructose) were subsequently transported and further metabolized by all three organisms. Raffinose catabolism was initiated by beta-fructosidase; melibiose was subsequently hydrolyzed to galactose and glucose by alpha-galactosidase. Glucose and fructose were completely metabolized by all three organisms, but galactose accumulated in the fermentation broth with EC16(pLOI555) and P2. MM2 (a raffinose-positive E. coli mutant) was the most effective biocatalyst for ethanol production (38 g/liter) from raffinose. All organisms rapidly fermented sucrose (90 g/liter) to ethanol (48 g/liter) at more than 90% of the theoretical yield. During sucrose catabolism, both hydrolysis products (glucose and fructose) were metabolized concurrently by EC16(pLOI555) and P2 without sugar leakage. However, fructose accumulated extracellularly (27 to 28 g/liter) at early stages of fermentation with KO11 and MM2. Sequential utilization of glucose and fructose correlated with a diauxie in base utilization (pH maintenance). The mechanism of sugar escape remains unknown but may involve downhill leakage via permease which transports precursor saccharides or novel sugar export proteins. If sugar escape occurs in nature with wild organisms, it could facilitate the development of complex bacterial communities which are based on the sequence of saccharide catabolism and the hierarchy of sugar utilization.     

Inhibition of glycogenolysis by 2,5-anhydro-D-mannitol in isolated rat hepatocytes

2,5-Anhydro-D-mannitol, an analog of D-fructofuranose, inhibited basal and glucagon-stimulated glycogenolysis and glucose production in hepatocytes isolated from fed rats. Glucose formation from galactose was unaffected by the inhibitor. 2,5-Anhydro-D-mannitol-1-phosphate inhibits phosphorylase alpha with a Ki value of 2.4 mM. This same phosphorylated metabolite accumulates to the extent of 9.2 mumol/g wet wt in treated hepatocytes suggesting that phosphorolysis is the locus of the inhibition of glucose production from glycogen. Our results suggest that 2,5-anhydro-D-mannitol can be used to produce a model of hereditary fructose intolerance and that it merits further study as a hypoglycemic agent.     

The Growth of Acer pseudoplatanus Cells in a Synthetic Liquid Medium: Response to the Carbohydrate, Nitrogenous and Growth Hormone Constituents

A synthetic culture medium which supports a high level of growth of a scrially propagated cell suspension culture of Acer pseudoplatanus is described. The sucrose of this medium can be effectively replaced by glucose or fructose or a mixture of glucose and fructose or galactose or maltose or soluble starch. When the carbohydrate is glucose or fructose no other sugars appear in the culture medium in significant amounts. Glucose is absorbed in greater quantity than fructose from an equimolar mixture of these sugars. When sucrose is supplied both glucose and fructose appear in the medium. Glucose appears in maltose medium, and maltose and glucose in soluble starch medium. Under the standard conditions of culture, media containing 2 % sucrose or 2 % glucose become depleted of sugar before the 25th day of incubation. Enhanced yield of the cultures can be obtained by raising the initial sucrose concentration to 6 %. – A supply of nitrate is essential for maximum yield and healthy growth. Growth, in the presence of nitrate, is significantly enhanced by a supply of urea. Addition of casein hydrolysate or of a mixture of amino acids enhances growth in the presence of nitrate and urea and particularly when nitrate is omitted. – When kinetin is omitted or incorporated at the standard level (0.25 mg/I), 2,4-dichlorophenoxyacetic acid (2,4-D) at 1.0 mg/l is essential for continuation of growth at a high level. It cannot be replaced by indol-3yl-acetic acid (IAA). 1-naphthaleneacetic acid (NAA) at 10 mg/l permits of a low level of growth with abnormal aggregation. When the level of kinetin is raised to 10 mg/l a high level of growth occurs in the absence of added auxin but the cultures become brown and tend to show increasing aggregation on subculture.     

Multisite control of the Crabtree effect in ascites hepatoma cells.

AS-30D hepatoma cells, a highly oxidative and fast-growing tumor line, showed glucose-induced and fructose-induced inhibition of oxidative phosphorylation (the Crabtree effect) of 54% and 34%, respectively. To advance the understanding of the underlying mechanism of this process, the effect of 5 mM glucose or 10 mM fructose on the intracellular concentration of several metabolites was determined. The addition of glucose or fructose lowered intracellular Pi (40%), and ATP (53%) concentrations, and decreased cytosolic pH (from 7.2 to 6.8). Glucose and fructose increased the content of AMP (30%), glucose 6-phosphate, fructose 6-phosphate and fructose 1,6-bisphosphate (15, 13 and 50 times, respectively). The cytosolic concentrations of Ca2+ and Mg2+ were not modified. The addition of galactose or glycerol did not modify the concentrations of the metabolites. Mitochondria isolated from AS-30D cells, incubated in media with low Pi (0.6 mM) at pH 6.8, exhibited a 40% inhibition of oxidative phosphorylation. The data suggest that the Crabtree effect is the result of several small metabolic changes promoted by addition of exogenous glucose or fructose.     

Production of d-Mannitol and Glycerol by Yeasts

D-Mannitol has not so far been known as a major product of sugar metabolism by yeasts. Three yeast strains, a newly isolated yeast from soy-sauce mash, Torulopsis versatilis, and T. anomala, were found to be good mannitol producers. Under optimal conditions, the isolate produced mannitol at good yield of 30% of the sugar consumed. Glucose, fructose, mannose, galactose, maltose, glycerol, and xylitol were suitable substrates for mannitol formation. High concentrations of yeast extract, Casamino Acids, NaCl, and KCl in media affected significantly the mannitol yield, whereas high levels of inorganic phosphate did not show any detrimental effect.     

Microbial fluxes of free monosaccharides and total carbohydrates in freshwater determined by PAD-HPLC

Abstract A new sensitive pulsed amperometric detection (PAD) method for measurements of mono- and disaccharides in nM concentrations was used in combination with high performance liquid chromatography (HPLC) to study fluxes of dissolved free and combined carbohydrates (DFCHO and DCCHO) in lake water. In a diel study concentrations of individual free saccharides typically were 5–50 nM, while total DFCHO concentrations ranged from 67 to 224 nM. No diel trends in concentration changes were obvious. At in situ light-dark conditions, dominant DFCHO were galactose, glucose, fructose and mannose/xylose. In addition to these saccharides, an increased abundance of melibiose and arabinose was measured in a parallel dark incubation. In a 118 h laboratory incubation of 1.0 μm filtered lake water, concentrations of DFCHO decreased from 194 nM (at 12 h) to a minimum of 54 nM (at 73 h). Dominant DFCHO were glucose, fructose and cellobiose. During the incubation DCCHO varied from 1.27 to 2.20 μM. Glucose, galactose and cellobiose made up 40, 30 and 10 mol-%, respectively, of the DCCHO. Fructose was degraded during hydrolysis of the DCCHO. A decline of DCCHO at 55 h was reflected in a simultaneous increase of DFCHO, but otherwise no similarities between the two saccharide pools were found. Incrased DCCHO concentrations and a high assimilation of glucose and fructose that was not reflected in a decline of their concentrations, both indicate that carbohydrates were produced during the experiment. Polysaccharides were probably excreted by the bacteria. Net assimilation of glucose and fructose sustained 14–19% (diel study) and 32% (long-term study) of the net bacterial carbon requirement.     

Analysis of various sugars by means of immobilized enzyme coupled flow injection analysis

Glucose, maltose, sucrose, lactose, xylose, sorbose, galactose, fructose and gluconolactone were analyzed by means of immobilized pyranose oxidase as well as by the combination of immobilized glucose oxidase with immobilized glycoamylase, invertase, mutarotase, maltase (α-glucosidase) and glucose isomerase by flow injection analysis (FIA). For the simultaneous analysis of glucose and other sugars three different flow-injection configurations were applied and compared. The average error of prediction of the analyses were better than 3% in model media and better than 6% in yeast extract containing media.     

Characterization of Thermoanaerobacter Glucose Isomerase in Relation to Saccharidase Synthesis and Development of Single-Step Processes for Sweetener Production

Regulation of glucose isomerase synthesis was studied in Thermoanaerobacter strain B6A, which fermented a wide variety of carbohydrates including glucose, xylose, lactose, starch, and xylan. Glucogenic amylase activities and β-galactosidase were produced constitutively, whereas the synthesis of glucose isomerase was induced by either xylose or xylan. Production of these saccharidase activities was not significantly repressed by the presence of glucose or 2-deoxyglucose in the growth media. Glucose isomerase production was optimized by controlling the culture pH at 5.5 during xylose fermentation. The apparent temperature and pH optima for these cell-bound saccharidase activities were as follows: glucose isomerase, 80°C, pH 7.0 to 7.5; glucogenic amylase, 70°C, pH 5.0 to 5.5; and β-galactosidase, 60°C, pH 6.0 to 6.5 Glucose isomerase, glucogenic amylase, and β-galactosidase were produced in xylose-grown cells that were active and stable at 60 to 70°C and pH 6.0 to 6.5. Under single-step process conditions, these saccharidase activities in whole cells or cell extracts converted starch or lactose directly into fructose mixtures. A total of 96% of initial liquefied starch was converted into a 49:51 mixture of glucose and fructose, whereas 85% of initial lactose was converted into a 40:31:29 mixture of galactose, glucose, and fructose.     

Effect of glucose on ATP dephosphorylation in rat spermatids

Round spermatids were isolated from rat testes and the effects of different energy-yielding substrates on the cellular ATP content were estimated. The ATP content was constant and high (6-8 nmol/10(6) cells) during metabolism of exogenous lactate. During incubation for 30 min in the absence of exogenous lactate, there was a remarkably slow decline of the ATP content, indicating ATP production from other substrates. It was shown that this could reflect beta-oxidation of fatty acids, but not the mobilization of an endogenous pool of acetylcarnitine. Glucose metabolism in the absence of exogenous lactate resulted in a rapid decline of the ATP content. This effect of glucose was correlated with a high fructose 1,6-biphosphate content (6-7 nmol/10(6) cells) and could be prevented by the addition of lactate. It is suggested that metabolism of glucose (and also mannose and fructose, but not galactose) in the absence of exogenous lactate can result in ATP dephosphorylation.     

The Effect of Various Mono–and Disaccharides on the Growth of Sphagnum nemoreum Thalli in Sterile Cultures

Sphagnum nemoreum Scop. thalli were grown under sterile conditions in order to study their ability to use certain carbohydrates for their growth, with special reference to the sugars occurring free in Sphagnum peat. It was found that Sphagnum nemoreum thallus could not grow at ail in vitro unless some organic carbon source war present. Growth was dependent on both the quality and the quantity of the sugar. Sucrose (1 per cent) proved to be the best carbon source and light intensity did not have any marked influence on growth under these conditions. Glucose and fructose were also growth promoting. Glucose is the main free sugar in the peat. Mannose (0.25 per cent) was almost as good a carbon source as sucrose in the same concentration. It is known that mannose accumulates in the Sphagnum peat during humification. and so does rhanmose, which was utilized to some extent. These two sugars are inhibitory to many mono–and dicotyledons. Ribose, galactose. arabinose and xylose were found to be toxic to Sphagnum.