Effect of fructose and glucose supplementation on invertase mediated synthesis of oligosaccharides from sucrose

Summary The total amount of novel oligosaccharides synthesized by -D-fructofuranosidase at pH 7.5 increased three-fold using a medium composed of 1.2M sucrose, 0.5M fructose and 0.1M glucose, as compared to that with only 1.8M sucrose solution. Using 0.6M of the three sugars did not increase yield but reduced rate of sucrose hydrolysis by 72.7%. Synthesis of fructosyl/glucosyl oligosaccharides based on -fructofuranosidase mediated transglycosylation is enhanced by supplementation of sucrose solution with appropriate concentrations fructose and glucose.     

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.     

Influence of glucose,fructose and sucrose as carbon sources on kinetics and stoichiometry of lysine production by <Emphasis Type="Italic">Corynebacterium glutamicum</Emphasis>

Batch cultivations of l-lysine-producing Corynebacterium glutamicum ATCC 21253 were carried out on the different carbon sources, glucose, sucrose and fructose. The time profiles of substrate and product concentrations were evaluated to compare kinetics and stoichiometry of lysine production. The lysine yield (mol C/mol C) on glucose was 8% higher than on sucrose and 30% higher than on fructose. The highest final biomass concentration of 5.0 g/l was obtained on glucose, whereas fructose and sucrose yielded 20% less biomass. Compared to glucose, fructose resulted in significantly higher respiration rates, a higher substrate uptake rate but a lower lysine production rate during the cultivation process. This was probably due to a higher tricarboxylic cycle activity combined with a lower activity of the pentose phosphate pathway. On sucrose, specific rates and yields differed significantly from those on fructose and glucose. Transport and metabolism of sucrose, therefore, are not a simple superposition of its building blocks, glucose and fructose. Journal of Industrial Microbiology & Biotechnology (2002) 28, 338–343 DOI: 10.1038/sj/jim/7000252 Received 28 November 2001/ Accepted in revised form 06 March 2002     

Influence of Nutritional Factors on the Nature, Yield, and Composition of Exopolysaccharides Produced by Gluconacetobacter xylinus I-2281

The influence of substrate composition on the yield, nature, and composition of exopolysaccharides (EPS) produced by the food-grade strain Gluconacetobacter xylinus I-2281 was investigated during controlled cultivations on mixed substrates containing acetate and either glucose, sucrose, or fructose. Enzymatic activity analysis and acid hydrolysis revealed that two EPS, gluconacetan and levan, were produced by G. xylinus. In contrast to other acetic acid strains, no exocellulose formation has been measured. Considerable differences in metabolite yields have been observed with regard to the carbohydrate source. It was shown that glucose was inadequate for EPS production since most of this substrate (0.84 C-mol/C-mol) was oxidized into gluconic acid, 2-ketogluconic acid, and 5-ketogluconic acid. In contrast, sucrose and fructose supported a 0.35 C-mol/C-mol gluconacetan yield. In addition, growing G. xylinus on sucrose produced a 0.07 C-mol/C-mol levan yield. The composition of EPS remained unchanged during the course of the fermentations. Levan sucrase activity was found to be mainly membrane associated. In addition to levan production, an analysis of levan sucrase's activity also explained the formation of glucose oxides during fermentation on sucrose through the release of glucose. The biosynthetic pathway of gluconacetan synthesis has also been explored. Although the activity of key enzymes showed large differences to be a function of the carbon source, the ratio of their activities remained similar from one carbon source to another and corresponded to the ratio of precursor needs as deduced from the gluconacetan composition.     

Effects of six sugars on the longevity,fecundity and nutrient reserves of Microplitis mediator

Parasitoid adults can directly feed on floral nectar and honeydew containing monosaccharides and disaccharides. Oligosaccharides such as maltose, melezitose and raffinose are also found in honeydew but are rare in floral nectar. The effects of six different sugar resources on the longevity, fecundity and nutrient reserves of Microplitis mediator, a larval endoparasitoid in the cotton bollworm Helicoverpa armigera (Hübner) were determined in our laboratory. The results showed that both food and sex affected longevity of this wasp. Females and males of M. mediator fed with 1 M sucrose solution survived longer than controls fed with water (5.7- and 3.7-fold longer, respectively). When provided with sucrose, glucose or fructose solutions, the parasitoid generated 3.6- to 3.7-fold more offspring than controls, and 60–75% of these progenies were produced during the first 5 days. When separately given fructose, sucrose or glucose, this wasp accumulated fructose and total sugar at the highest level, which means a high sugar levels might lead to prolonging longevity and more offspring in M. mediator. In addition, compared with organisms fed galactose or raffinose, M. mediator fed sucrose or fructose accumulated high glycogen levels. Furthermore, in M. mediator, the lipid content declined with the advancing age. Females showed the slowest lipid metabolic rates when fed with sucrose, glucose, fructose, mannose and galactose solutions versus when fed with raffinose and control. In addition, only sucrose had a significant effect on lipid levels in males nearing the end of life.     

Sugar utilization by yeast during fermentation

Summary When glucose and fructose are fermented separately, the uptake profiles indicate that both sugars are utilized at similar rates. However, when fermentations are conducted in media containing an equal concentration of glucose and fructose, glucose is utilized at approximately twice the rate of fructose. The preferential uptake of glucose also occurred when sucrose, which was first rapidly hydrolyzed into glucose and fructose by the action of the enzyme invertase, was employed as a substrate. Similar results were observed in the fermentation of brewer's wort and wort containing 30% sucrose and 30% glucose as adjuncts. In addition, the high levels of glucose in the wort exerted severe catabolite repression on maltose utilization in theSaccharmyces uvarum (carlsbergensis) brewing strain. Kinetic analysis of glucose and fructose uptake inSaccharomyces cerevisiae revealed aK _m of 1.6 mM for glucose and 20 mM for fructose. Thus, the yeast strain has a higher affinity for glucose than fructose. Growth on glucose or fructose had no repressible effect on the uptake of either sugar. In addition, glucose inhibited fructose uptake by 60% and likewise fructose inhibited, glucose uptake by 40%. These results indicate that glucose and fructose share the same membrane transport components.     

Production of a novel transfructosylating enzyme from Bacillus macerans EG-6

A new bacterium producing a novel transfructosylating enzyme was isolated from soil and designated as Bacillus macerans EG-6. Various culture conditions for enzyme production were optimized in a flask culture. 1% (w/v) sucrose as a carbon source and a mixed nitrogen source (1% yeast extract, 1% polypeptone, and 0.5% ammonium chloride) gave the best enzyme production. Addition of phosphate and magnesium ion into the medium enhanced the enzyme yield. Optimum culture pH and temperature were 7.0 and 37?°C, respectively. Under optimal culture conditions, transfructosylating enzyme was rapidly produced in the early growth period, thereafter invertase activity was predominant as the culture proceeded. Using the culture filtrate, production of fructooligosaccharides from sucrose was preliminarily carried out. In a low sucrose concentration (200?g/l), transfructosylating activity competes with invertase activity in sucrose utilization. Subsequently, low fructooligosaccharide yield (20%) was achieved due to liberation of high amounts of glucose and fructose. The best oligosaccharide yield (43%) was achieved when 500?g/l sucrose was utilized.     

Competition of polysaccharides with sugars for the pyranose and the furanose sites in the labellar sugar receptor cell of the blowfly, Phormia regina

In the labellar sugar receptor cell of the blowfly, Phormia regina, soluble starch and dextran T500 inhibited the response to sucrose, to maltose or to glucose, but did not inhibit that to fructose. On the other hand, inulin inhibited the response to fructose, but did not inhibit that to sucrose. These results suggest that both soluble starch and dextran T500 compete with sucrose, with maltose or with glucose for the pyranose site (P site), and that inulin competes with fructose for the furanose site (F site) in a single sugar receptor cell. Each inhibition constant (K_i) was estimated to be 0.6–0.7% for soluble starch. about 4.5% for dextran T500, and about 1.3% for inulin.     

In vitro and in vivo inhibition of sucrose synthesis by sucrose

The inhibitory effects of sucrose on rates of sucrose synthesis by sucrose phosphate synthase (SPS) from the maize scutellum and on net rates of sucrose production in maize scutellum slices from added glucose or fructose were studied. Scutellum extracts were prepared by freezing and thawing scutellum slices in buffer. The extracts contained SPS and sucrose phosphate phosphatase, but were free of sucrose synthase. SPS activity was calculated from measurement of UDP formation in the presence of UDPG, fructose-6-P and sucrose. The ranges of metabolite concentrations used were those estimated to be in scutellum slices after incubation in water or fructose for periods up to 5 hr. UDPG and fructose-6-P also were added at concentrations that saturated SPS. At saturating substrate levels, sucrose inhibition of SPS was less than that when tissue levels of substrates were used. With tissue levels of substrates and sucrose concentrations up to ca 166 mM, sucrose inhibitions of sucrose synthesis in vitro by SPS were similar to those observed in vivo. However, as the sucrose concentration rose above 166 mM, SPS activity was not inhibited further, whereas there was a further sharp decline in sucrose production by the slices. It is concluded that sucrose synthesis in vivo is controlled by sucrose inhibition of SPS over a considerable range of internal sucrose concentrations.     

In Vitro Sugar Transport in Zea mays L. Kernels : II. Characteristics of Sugar Absorption and Metabolism by Isolated Developing Embryos

In vitro sugar transport into developing isolated maize embryos was studied. Embryo fresh and dry weight increased concomitantly with endogenous sucrose concentration and glucose uptake throughout development. However, endogenous glucose and fructose concentration and sucrose uptake remained constant. The uptake kinetics of radiolabeled sucrose, glucose, and fructose showed a biphasic dependence on exogenous substrate concentration. Hexose uptake was four to six times greater than sucrose uptake throughout development. Carbonylcyanide-m-chlorophenylhydrazone and dinitrophenol inhibited sucrose and glucose uptake significantly, but 3-O-methyl glucose uptake was less affected. The uptake of 1 millimolar sucrose was strongly pH dependent while glucose was not. Glucose and fructose were readily converted to sucrose and insoluble products soon after absorption into the embryo. Thus, sucrose accumulated, while glucose pools remained low. Based on the findings of this and other studies a model for sugar transport in the developing maize kernel is presented.     

A β-d-fructofuranosidase from Claviceps purpurea

Evidence suggests that sucrose is the main carbon source for growth of Claviceps spp. in the parasitic condition. The sucrose acts as substrate for an active beta-fructofuranosidase, produced by the fungus, which in the first instance converts the disaccharide into glucose and an oligofructoside. In this way, 50% of the glucose, supplied as sucrose, is made available to the parasite for assimilation. Subsequent action of the enzyme on both sucrose and the oligofructoside leads to the release of more glucose and the formation of additional oligosaccharides. The structures of the main oligosaccharides formed have been elucidated and the interactions of each compound studied. In experiments with purified enzyme in vitro the interaction of the oligosaccharides is rapid but in culture they are assimilated only slowly; in each case some free fructose is liberated. Free fructose is not assimilated in the presence of glucose and, further, inhibits growth at concentrations which might be expected to occur in the parasitic condition. A dual role has been suggested for the enzyme, with sucrose as substrate, in which glucose is made available to the growing parasite, while at the same time transfer of the fructose to form oligosaccharides prevents it from accumulating at inhibitory concentrations. Ultimately, when glucose becomes limiting, the fungus will adapt to fructose assimilation.     

Investigations into the Role of Sucrose in Potato cv. Estima Microtuber Production in vitro

Sucrose has been the carbohydrate traditionally used for potatomicrotuber production. Added to nutrient media, sucrose canact solely as a carbon source, or as an osmoticum, or both.Preliminary tests showed that the osmolarity of sucrose solutionswas increased by autoclaving, indicating some breakdown of thesugar. This was taken into consideration in experiments whichinvolved supplementing 4% sucrose media with sucrose, maltose,glucose or fructose, while keeping the osmotic potential ofthe media constant. A medium concentration of about 400 mM withonly sucrose was more suitable for microtuber production thanmedia supplemented with maltose, glucose or fructose. However,a better microtuber yield was obtained when hexoses were addedthan with unsupplemented 4% sucrose media. When glucose wassupplied at concentrations which had the same number of carbonatoms as 8% sucrose, the high osmolarity inhibited microtuberisation.Sugar movement in the tubering plantlet was followed using radio-labelledsucrose, glucose and fructose. The sucrose was translocatedand used at a faster rate than the other sugars, which tendedto remain in the roots of the plantlets. Furthermore, therewas no difference in microtuber production on media to whichthe sucrose was added before or after autoclaving, indicatingthat levels of breakdown were not severe enough to affect theprocess. Therefore, it is concluded that sucrose acts primarilyas a suitable carbon source for uptake and utilization by theplantlets, but, at 8%, it also provides a favourable osmolarityfor the development of microtubers.Copyright 1995, 1999 AcademicPress Solanum tuberosum (L.), potato, microtuber, media, sugar, sucrose, osmolarity, pH     

Reduction in mutagenicity of cigarette smoke condensate by added sugars.

The effects of adding sugars to high- and low-tar cigarettes on the mutagenicity of their smoke condensates were studied using Salmonella typhimurium TA100 and TA98 with and without metabolic activation. The sugars tested were glucose, fructose, galactose, sorbitol, sucrose and lactose. The lowest mutagenicities observed with these sugars per mg of smoke condensate assayed on TA98 with metabolic activation were 37% (high-tar cigarettes) and 22% (low-tar cigaretts) of that of smoke condensate from untreated cigarettes. Addition of sugars increased the total amounts of smoke condensates, but the mutagenicities of the total condensates were also decreased by all the sugars, the lowest values being 35% (high-tar cigarettes) and 36% (low-tar cigarettes) of that of smoke condensates from cigarettes without added sugar. On assay with TA100 with metabolic activation, decreases in both specific and total mutangenicities of condensates of high-tar cigarettes were observed with all the sugars tested except galactose and sucrose. Treatment with glucose, fructose or sorbitol decreased the specific mutagenicity of condensates of low-tar cigarettes and glucose and fructose reduced also their total mutagenicity. The effects of added sugars were more marked when assayed on TA98 than on TA100 and of the sugars tested fructose and sorbitol had the greatest effects. Addition of sugars had no effect of the mutagenicity of cigarette-smoke condensate without metabolic activation.     

Regulation of invertase levels in Avena stem segments by gibberellic Acid, sucrose, glucose, and fructose

Gibberellic acid and sucrose play significant roles in the increases in invertase and growth in Avena stem segments. About 80% of invertase is readily solubilized, whereas the rest is in the cell wall fraction. The levels of both types of invertase change in a similar manner in the response to gibberellic acid and sucrose treatment. The work described here was carried out with only the soluble enzyme. In response to a treatment, the level of invertase activity typically follows a pattern of increase followed by decrease; the increase in activity is approximately correlated with the active growth phase, whereas the decrease in activity is initiated when growth of the segments slows. A continuous supply of gibberellic acid retards the decline of enzyme activity. When gibberellic acid was pulsed to the segments treated with or without sucrose, the level of invertase activity increased at least twice as high in the presence of sucrose as in its absence, but the lag period is longer with sucrose present. Cycloheximide treatments effectively abolish the gibberellic acid-promoted growth, and the level of enzyme activity drops rapidly. Decay of invertase activity in response to cycloheximide treatment occurs regardless of gibberellic acid or sucrose treatment or both, and it is generally faster when the inhibitor is administered at the peak of enzyme induction than when given at its rising phase. Pulses with sucrose, glucose, fructose, or glucose + fructose elevate the level of invertase significantly with a lag of about 5 to 10 hours. The increase in invertase activity elicited by a sucrose pulse is about one-third that caused by a gibberellic acid pulse given at a comparable time during mid-phase of enzyme induction, and the lag before the enzyme activity increases is nearly twice as long for sucrose as for gibberellic acid. Moreover, the gibberellic acid pulse results in about three times more growth than the sucrose pulse. Our studies support the view that gibberellic acid, as well as substrate (sucrose) and end products (glucose and fructose), play a significant role in regulating invertase levels in Avena stem tissue, and that such regulation provides a mechanism for increasing the level of soluble saccharides needed for gibberellic acid-promoted growth.     

Simultaneous production of citric acid and invertase by Yarrowia lipolytica SUC+ transformants

Simultaneous production of citric acid (CA) and invertase by Yarrowia lipolytica A-101-B56-5 (SUC⁺ clone) growing from sucrose, mixture of glucose and fructose, glucose or glycerol was investigated. Among the tested substrates the highest concentration of CA was reached from glycerol (57.15 g/L) with high yield (Y_CA/S = 0.6 g/g). When sucrose was used, comparable amount of CA was secreted (45 g/L) with slightly higher yield (Y_CA/S = 0.643 g/g). In all cultures amount of isocitrate (ICA) was below 2% of total citrates. Considering invertase production, the best carbon source appeared to be sucrose (72 380 U/L). The highest yield of CA and invertase biosynthesis calculated for 1 g of biomass was obtained for cells growing from glycerol (9.9 g/g and 4325 U/g, respectively). Concentrates of extra- and intracellular invertase of the highest activity were obtained from sucrose as substrate (0.5 and 1.8 × 10⁶ U/L, respectively).     

Ethanol fermentation by an efficient strain,NRRL B-4286, of Zymomonas mobilis

We studied the effect of the initial substrate concentration over the range of 100–250 g·l⁻¹ on the fermentation kinetics in batch cultures of Zymomonas mobilis NRRL B-4286 on glucose, fructose, and sucrose, using an adapted initial inoculum. With increasing concentrations of substrate, parameters related to growth were more rapidly and strongly affected than those related to ethanol production. This strain produced 94.0 g·l⁻¹, 76.9 g·l⁻¹, and 66.5 g·l⁻¹ of ethanol at glucose, fructose, and sucrose concentrations of 200 g·l⁻¹, respectively, more than the amount produced by the efficient strain ZM4 (NRRL B-14023).     

DNA synthesis and nuclear division during formation of infection structures by bean rust uredospore germlings

Plectonema boryanum can grow in the dark with ribose, sucrose, mannitol, maltose, glucose, or fructose. Cell doubling times with 10 mM substrate are the following: 5 days with ribose, 6 days with sucrose or mannitol, 10 days with maltose, 12 days with glucose, and 13 days with fructose; with ribose plus 0.1% casamino acids it is 2.5 days. Dark-grown cells appear morphologically similar to light-grown cells. Cells grown in the dark for several years remain pigmented and resume photoautotrophic growth when placed in the light. Dim light (85 lux) increases the growth rate with ribose and with ribose plus casamino acids to nearlytwice that of the dark rate. In moderate light, growth takes place with ribose even in the presence of 1x10^-5 M DCMU.     

Mannitol production from glycerol by resting cells of Candida magnoliae

Production of mannitol from glycerol by resting cells of Candida magnoliae under aerobic condition was investigated. The resting cells were suspended in aqueous solution of glycerol in Erlenmeyer flasks and incubated on rotary shaker. The samples were analyzed by ion exclusion–HPLC equipped with refractive index and UV detector. The resting cells of C. magnoliae produced mannitol from fructose, sucrose and glycerol but not from glucose. Addition of yeast extract and/or potassium phosphate to the glycerol solution adversely affected its conversion to mannitol. The conversion of glycerol to mannitol was dependent on oxygen availability. Using resting cells, the yield of mannitol was as high as 45%. This is probably the first report of conversion of glycerol to mannitol by osmophilic yeast.     

Candida glycerinogenes不同碳源代谢的初步研究

研究了不同碳源对Candidaglycerinogenes的菌体生长、发酵液pH值及代谢产物的影响,结果发现以葡萄糖、果糖等单糖为碳源时茵体生长较快,最终生物量比以蔗糖、麦芽糖等二糖为碳源时高20％～30％;导致发酵前12h发酵液pH值明显下降的主要因素是乳酸;与葡萄糖为碳源转化为甘油相比,果糖为碳源时更易累积乙醇;以蔗糖、麦芽糖为碳源时,用于转化生成甘油的碳源明显降低,碳源主要用于茵体自身生物合成及HMP途径,以蔗糖为碳源时,用于乳酸、丙酸及柠檬酸生物合成的碳源较麦芽糖明显提高,TCA途径代谢较为活跃。     

Improvement of yield of Pleurotus eryngii var. eryngii by substrate supplementation and use of a casing overlay

Improved yield and biological efficiency (BE) of Pleurotus eryngii var. eryngii were achieved by supplementation of substrate with a commercial delayed-release nutrient and use of a casing overlay. Yield increases of 14% were achieved from cased substrates that were supplemented at time of casing with delayed-release nutrient (Remo’s). Use of a casing layer enhanced yield by 141% over non-cased substrates. When casing and substrate supplementation were combined, yield increased 179% over non-cased/non-supplemented substrates. Mushrooms harvested from cased substrates were darker in color and solids contents were lower compared to non-cased substrates. An additional break of mushrooms was harvested from non-cased “spent” substrate by fragmenting and re-supplementing the substrate prior to the application of a casing overlay. Three production methods were compared for their effect on mushroom yield: “standard”, “casing” and “casing after first break”. Casing of the substrate before first break (“casing” production method) resulted in the highest yield and biological efficiency.