Seven xyloglucans were extracted by treatment of common vegetables with aqueous 24% KOH and purified through iodine-xyloglucan complex formations. The reactivity of the extracted xyloglucan to anti-tamarind xyloglucan antibody was determined by Enzyme-Linked Immunosorbent Assay (ELISA). From the results, xyloglucans of tomato showed similar titration curves to that of tamarind xyloglucan and were more reactive to the antibody than those of green vegetables. 1H-NMR spectroscopic analyses, including 2D NMR techniques of the purified xyloglucans, showed that structural features of green vegetable xyloglucans were very similar to each other, and were different from those of tomato and tamarind seed in containing L-fucosyl residue as a component sugar.

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α- and β-anomers of methyl 6-O-(p-tolylsulfonyl) -D-glucopyranoside, -mannopyranoside, and -galactopyranoside, which have three contiguous secondary hydroxyl groups, were acetylated by lipase-catalyzed transesterification with vinly acetate. A lipase from Pseudomonas cepacia (lipase PS) revealed high activity and regioselectivity. α-Glycopyranosides were acetylated preferentially at the C-2 hydroxyl group, while corresponding β-anomers were acetylated preferentially at the C-3 hydroxyl group by the enzyme. The best selectivity was observed in the reaction of glucopyranosides.

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We evaluated an immobilized neopullulanase for the production of isomalto/branched-oligosaccharide syrup. The syrup was produced from starch by using a transglycosylation reaction of neopullulanase. The characteristics of the immobilized neopullulanase were compared with those of a native enzyme. The optimum temperature for reaction of the immobilized neopullulanase was slightly higher than that of the native enzyme. The pH range for stability of the immobilized neopullulanase was narrower than that of the native enzyme. A continuous production system of the isomalto/branched-oligosaccharide syrup was investigated by using a column of the immobilized neopullulanase. The yield of isomalto/branched-oligosaccharide was more than 60% by this system. We preliminarily evaluated the effect of the isomalto/branched-oligosaccharide syrup by using in vitro experiments. The results suggested that the syrup could be used as a growth factor for bifidobacteria and as a non-cariogenic sweetner.

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The cyclodextrin glucanotransferase (CGTase) from Brevibacterium sp. No. 9605 showed a broad acceptor specificity to various monosaccharides in the same way as Bacillus stearothermophilus CGTase. It specially produced a large amount of transfer products from D-mannose and L-rhamnose. The major component from the transfer products of D-mannose was found to be 4-O-α-D-glucosyl-D-mannose. Brevibacterium CGTase also formed a much greater amount of transfer products than Bacillus macerans and Bacillus stearothermophilus CGTases from 1, 3-dihydroxybenzene, 1, 3, 5-trihydroxyben-zene, 3-hydroxybenzyl alcohol, and (+) -catechin used as acceptors. The major component of the two transfer products of kojic acid was found to be kojic acid 7-O-a-D-glucoside.

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The effect of branching on the physicochemical and functional properties of amylose was investigat-ed by linking maltose to the essentially linear molecule with DP n 270. The amylose was prepared from potato starch by digestion of the starch paste in the presence of 1-butanol with bacterial a-amylase. The resulting precipitate of amylose 1-butanol complex was recrystallized five times with 1-butanol saturated water. The maltosyl residues were introduced into the amylose by the reverse action of Pseudomonas isoamylase. The number of branch linkages in the amylose increased from 0.40 to 2.9 after the action. The iodine affinity and blue value decreased slightly from 19.7 to 18.2 and 1.19 to 1.13, respectively. However, the j3-amylolysis limit decreased considerably from 99 to 66%, suggesting that the maltosyl residues were introduced predominantly near the reducing end, and the short side-chains had only a slight effect on the complex formation with iodine. The retrogradation tendencies of the amylose were greatly reduced by the addition of maltose.

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Two kinds of branched cycloisomaltooligosaccharides, P1 and P2, were synthesized from cycloisomaltoheptaose and maltose by the action of cyclodextrin glucanotransferase. P1 was hydrolyzed into glucose and cycloisomaltoheptaose with the same molar ratio by the action of glucoamylase. P2 was hydrolyzed into glucose and cycloisomaltoheptaose with a molar ratio of two to one. P2 was also hydrolyzed into maltose and cycloisomaltoheptaose by acid degradation. Their molecular weight were determined by mass spectra analysis to be 1296 and 1458, respectively. Carbon and proton magnetic resonance analysis suggested that they contained α-1, 4-glucosidic linkage. From these results, they were identified as 4-0-D-glucosyl-cycloisomaltoheptaose and 4-O-α-maltosyl-cycloiso-maltoheptaose. They had higher resistance to hydrolytic action by endo-dextranase than the parent cycloisomaltoheptaose.

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Presented at the 1996 award address of the Society, the results of three research studies are reported that were carried out during the period from 1983 to 1996. 1) The physicochemical properties of starch from Katakuri (Erythronium japonicum DENCE) were compared with those of potato and edible canna starches. The average particle size of Katakuri was 25.3 μm, and the amylose content of the starch was 22.8%. The average chain lengths of fractions II and III, determined by the chromatographic method, were 47.4 and 14.6, respectively, which were longer than those of the other starches. Katakuri starch granules had high digestibility, and the gelatinization temperature was about l0°C lower than that of potato and edible canna starches. The spinnability and static viscoelasticity of Katakuri starch at a low paste concentration were confirmed by the shortest retardation time and the lowest Newtonian viscosity of the three starches. 2) The molecular properties of amyloses and amylopectins from tropical starches, and the physical properties of these starches were evaluated. Edible canna, arrowroot, cassava and sago were examined, with potato and corn starches used as controls. Each starch was debranched with isoamylase and then chromatographed in a Sephadex G-75 column. The results show that potato and edible canna starches had more long-chain branches than the other four starches. It is also shown that the edible canna, sago and corn starches were rich in branches, while the potato and arrowroot starches were rich in short chains, amylose, amylopectin and the C-fraction fom each starch were determined for their weight average molecular weight (MW), radius of gyration (RG) and intrinsic viscosity [η]. The viscosity behavior was examined by using a Rotovisco CV20 viscometer. The results of the principal analysis enabled the tropical starches to be grouped according to the 15 items of physicochemical properties. The contribution ratio was 34.9% for the 1st principal component, 26.1% for the 2nd, and 25.9% for the 3rd. The six kinds of starches could thus be divided into three types. 3) The Gelatinization characteristics, sol-gel transition and elasticity of heat-moisture treated (HMT) starches were examined. Viscograms of both HMT corn and potato starches indicated considerably suppressed maximum viscosity and no breakdown. DSC curves showed two endothermic peaks in the low- and high-temperature regions for HMT corn starch, and one peak for the native potato starch. The concentration dependence of the mechanical properties of the native and HMT corn, cassava and potato starches near the sol-gel transition point was analyzed by the scaling law derived from percolation theory. The scaling law could not be applied to the native and HMT potato starches, but the critical exponents were respectively determined as 4.2 and 3.6 for the native and HMT corn starches, and as 1.8 and 2.4 for the native and cassava starches. It is suggested from these results that the intermolecular bonding of corn starch was rigid and that of cassava starch was flexible.

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A bacterial strain that remarkably produced G, from β-CD (CDase) was isolated from soil, identified as a Bacillus sphaericus, and named B. sphaericus E-244. The CDase from B. sphaericus was purified to a homogeneous state. The enzyme had far higher affinity for β-CD than other known CDases. The hydrolase produced G, from fl CD and kept it in the reaction mixture, because the CDase had the highest catalytic efficiency for β-CD among α-glucans. The gene for the CDase was cloned in Escherichia coli cells. Sequencing a portion of the plasmid revealed a unique open reading frame of 1773 nucleotides coding for a 591 amino acid polypeptide. The deduced polypeptide sequence showed about 50% homology with that of a neopullulanase. Three unknown oligosaccharides were isolated from the culture broth of a strain of B. circulars T-3040, which had been isolated from soil. They were identified as cycloisomalto heptaose, -octaose, and -nonaose, respectively, on the basis of their 1H NMR spectra, 13C-NMR spectra, IR spectra, mass spectra, reducing power, and results of enzymatic analysis using endo dextranase and exo-dextranase. They were named as cyclodextran (CI). A novel enzyme, which catalyzes the conversion of dextran to CI by intramolecular transglucosylation (cyclization reaction), was purified to a homogeneous state. The enzyme mainly catalyzed the cyclization reaction and produced three CJs at about 20% total yield. CI-8 was the main product. Coupling, disproportionation, and hydrolytic reactions were also observed. These results showed that CITase is a multi functional enzyme. The gene for CITase was cloned in E. coli cells. The deduced amino acid sequence of the mature enzyme was unique and contained 934 residues. Another strain of bacteria that mainly produced CI-7 from dextran was newly isolated from soil and named B. circulars U155. From the results on the purification of the enzyme and the cloning of the gene for the CITase, it was confirmed that this enzyme was obviously different from T-3040 CITase. The largest difference was its main product (CI-7) from dextran. The primary amino acid sequence had about 67% homology to that of strain T-3040. The inhibition mechanisms of CIs are different from those of known anticariogenic reagents. This inhibition has high specificity and is not affected by the existence of protein. CIs are expected to be successful as anticariogenic reagents, because they are colorless, not sweet, and highly soluble in water.

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Candida guilliermondii H-404 isolated from soil, produced a large amount of thermostable α-galactosidase intracellularly and extracellularly and a few amounts of other glycosidases such as β-galactosidase, α-glucosidase, and β-glucosidase, which affect the reverse reaction of α-galactosidase using lactose hydrolyzate as a substrate. The C. guilliermondii H-404 cells can be used repeatedly as the a-galactosidase preparation. α-Linked galactooligosaccharides (α-GOS A from galactose and α-GOS B from lactose hydrolyzate) were produced using the reverse reaction of the a-galactosidase from C. guilliermondii H-404. The main product of the disaccharides in both a-GOSs was the 1, 6-isomer, with 1, 3-, 1, 2-, and 1, 1-isomers as minor products. Both were available as the donor substrates in transgalactosylation of a-galactosidase in the same manner as melibiose. Both were not hydrolyzed by the digestion method in vitro, had activity to inhibit the synthesis of insoluble glucan from sucrose and had a stronger bifidus growth activity than those of other α-linked galactooligosaccharides such as melibiose, raffinose, and stachyose. α-1, 3-Galactobiose was prepared from α-GOS A by activated carbon chromatography. This oligosaccharide can inhibit the hyperacute antibody-mediated rejection that occurs when pig organs are transplanted into primates. The a-galactosidase from C. guilliermon-dii H-404 had a broader acceptor specificity than any other a-galactosidases hitherto found. Through the transgalactosylation of the a-galactosidase from C. guilliermondii H-404, raffinose was effectively synthesized from α-GOS B and sucrose, and 6-0-α-D-galactosyl-L-ascorbic acid (a-GALA) from α-GOS B and L-ascorbic acid. α-GALA retained its reducing activity and had a 7-fold higher stability than unmodified L-ascorbic acid in a neutral solution.

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The bacterium Arthrobacter sp. Q36, isolated from soil, showed an ability to produce trehalose from maltooligosaccharides. Two novel enzymes were found to be related to trehalose synthesis. One catalyzed the conversion of maltooligosaccharide into 4-O-α-maltooligosyl trehalose by intramolecular transglycosylation, showing it to be maltooligosyl trehalose synthase. The other specifically catalyzed the hydrolysis of the α-1, 4-glucosidic linkage that bound the maltooligosyl and trehalose moieties of maltooligosyl trehalose, showing it to be maltooligosyl trehalose trehalohydrolase. Through the actions of these enzymes and isoamylase on starch, the trehalose content in the reaction mixture reached around 85%. These enzymes were also found to exist in several bacteria belonging to the genus Rhizobium, the thermoacidophilic archaebacteria Sulfolobus and other bacteria. The maltooligosyl trehalose synthase gene was linked to the maltooligosyl trehalose trehalohydrolase gene by an overlap of one nucleotide in Arthrobacter sp. Q36, constituting an operon for trehalose synthesis. In Sulfolobus acidocaldarius ATCC33909, the three structure genes, maltooligosyl trehalose trehalohydro-lase, isoamylase and maltooligosyl trehalose synthase, were found to constitute an operon in that order. The deduced amino acid sequences of synthase and hydrolase from each strain have several regions common to amylolytic enzymes belonging to the amylase family.

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Although the current literature contains references to four plant debranching enzymes-isoamylase, limit dextrinase, pullulanase and R-enzyme, the last three names are alternatives for the same enzyme, and only two plant debranching enzymes are, in fact, known. Moreover, the name pullulanase includes enzymes from both plant and microbial sources, even though these differ significantly in their ability to act on glycogen. To avoid any possible confusion or ambiguity, it is suggested that the second plant debranching enzyme should be known by the original name of limit dextrinase, rather than R-enzyme, and that the name pullulanase should be confined to enzymes of bacterial origin.

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The enzymatic hydrolysis of native cellulose has been discussed from the viewpoint of synergistic action of endo- and exo-type cellulases. However, the details in the mechanism remain unclear, partly because most studies have been generally employed to use certain types of cellulosic substrates being useful in measuring enzyme activity such as carboxymethylcellulose and Avicel instead of native cellulose with high resistance for the degradation because of its high crystallinity and water insolubility. The degree of crystallinity of cellulose is one of the most important structural parameters which affects the rate of enzymatic hydrolysis. Consequently, it is important to investigate the behavior of cellulase components in crystalline and amorphous regions of native cellulose. In the present review, we investigated the adsorption and hydrolysis modes of exo- and endo-type cellulases from Irpex lacteus for cellulose with different crystallinities, and confirmed that the crystallinity of substrate greatly affects the characteristic modes of attack of both cellulases. Moreover, the changes in the size of amorphous region of cotton cellulose with the progress of enzymatic cellulolysis were observed by differential scanning calorimetry in addition to the morphological investigation of the degradation process, and the mode of action of two types of cellulases in the amorphous and crystalline regions of native cellulose was clarified.

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Everyday we eat various foods and receive taste sensations that yield nutritional information and aid in the efficient digestion of food. When animals detect a bitter or sour-tasting food, they exercise caution because foods with these tastes are often toxic or spoiled. They may even stop eating or may vomit the food. However foods having a familiar or pleasant taste may be swallowed without caution. In general, foods having sweetness, moderate saltiness, or umami taste elicited by amino acid, glutamate, inosinate or some kind of nucleotides, are palatable, but those that are sour and bitter are usually avoided. The sense of taste plays an important role in detecting food containing required nutrients, and in predicting proper digestion and metabolism for the homeostatic control of each nutrient. Taste preferences are altered to reflect physiological needs for each nutrient and to support the recovery from nutritional disorders. The central mechanism involved in both recognition for and adaptation to a deficient essential nutrient, i.e. L-lysine, has been unveiled and shows that the feeding center in the hypothalamus is a primary center nucleus to induce a neuronal plasticity responding to dietary intake of deficient nutrient intake both centrally in the brain, and peripherally, such as in the sense of taste and its concentration change. Changing taste preferences may act as an alarm, signaling protein malnutrition for saltiness or metabolic adult disease, such as hypertension for saltiness, diabetes for sweetness, etc.

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Trehalose (α, α-trehalose) is a non-reducing disaccharide consisting of glucose. This sugar has been produced conventionally by the method of extraction from the yeast cell, and was thus too expensive to be used in the food industry as a material for foods. Recently, however, a novel pathway for trehalose synthesis from glucan in bacteria was discovered, and established industrially to produce trehalose from starch using newly discovered bacterial enzymes. Currently, trehalose is being supplied on a larger scale and at a lower cost. Trehalose is very stable to heat and acid, and causes no browning reaction with amino acids or protein. The sweetness of trehalose is 45% that of sucrose, and the quality, of sweetness is higher than sucrose. Trehalose is digested and absorbed in the human small intestine to become a source of energy, and prevents retrogradation of starch and denaturation of protein. It has a wide application in foods, cosmetics and the medical field due to its many useful properties.

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