The Japanese Society of Starch Science, at first, carried out many roles as a leader in the development of starch producing and provided many opportunities for communication among enterprises, universities, and public research laboratories . Glucose production by enzyme process was industrialized as a result. In recent years, the direction of study has turned to not only technical but also academical and fundamental works in the field of starch and other carbohydrate science. On the other hand, the scientific works of the enterprises greatly also developed in the production of functional oligosaccharides. This society must expect that researchers of different departments take part in the discussion of the fundamental researches.

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1. Galactanasses from Penicillium citrinum and Bacillus subtilis were purified and char-acterized. Two galactanases from P. citrinum had essentially the same properties and hydrolyzed β-1, 4-galactosidic linkages of soybean arabinogalactan (SAG) in an endo-manner. B. subtilis galactanase was an exo-enzyme which hydrolyzed SAG and oligosaccharides to produce galac-toboise from the non-reducing ends. 2. Transfer reaction catalyzed by the galactanases were studied. The P. citrinum enzyme degraded o-nitrophenyl-β-galactoside after a lag phase. A reaction mechanism which involved transglycosylation in addition to hydrolysis was proposed. The endo- and exo-galactanases showed broad acceptor specificities and transglycosylated various alcohols, phenols, sugar alcohols, and saccharides. Structural feature of the products from glycerol was different among the galactanases and Escherichia coli β-galactosidase : the galactanases transglycosylated preferentially the secondary OH-group of glycerol rather than primary OH-groups which were almost specifically transgalactosylated by the β-galactosidase. 3. Glycosides with glycerol and trimethylolpropane (TMP) moieties, which were prepared by the transfer reaction, acted as effective substrates for the lipase-catalyzed esterification.

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Arthrobacter sp. K-1 isolated from soil, produces β-fructofuranosidase. The enzyme had a molecular weight of 52, 000, and isoelectric point of 4.3. This enzyme was most active at pH 6.5-6.8 and at 55°C and stable up to 45°C at pH 6.5 for 30 min of incubation, and from pH 5.5 to 10.0 at 40t in 2 hr of incubation. The enzyme hydrolyzed sucrose, erlose, raffiinose, xylosyl-fructoside, neokestose, and stachyose, in this order, but hardly hydrolyzed 1-kestose and nystose. The enzyme catalyzed both transfructosylation and hydrolytic action, when it was incubated with sucrose alone. But in the presence of a suitable acceptor such as lactose, the enzyme catalyzed mostly transfructosylation and transferred the fructose residue preferentially to the acceptor. The enzyme had wide acceptor specificities. D-Xylose, D-galactose, L-sorbose, D- and L-fucose, D- and L-arabinose, maltose, isomaltose, cellobiose, lactose, melibiose, xylobiose, malto-triose, methyl β-glucoside, and galactoside were efficient acceptors in the transfructosylation. On the other hand, D-ribose, L-rhamnose, D-mannose, 2-deoxy-D-glucose, D-galactosamine, D-galacturonic acid, and 1-kestose were not efficient acceptors. Various primary alcohols, polyhydric alcohols including some suggar alcohols, and some glycosides acted as acceptors. The require-ment for an acceptor of the transfructosylation by the enzyme is that the saccharide must have free hydroxyl groups of the equatorial bonds at C2 and C3 on ⁴C₁ or ¹C₄ conformation. The main transfer products to aldoses and ketoses by the enzyme were non-reducing oligosaccharides, which had a fructofuranosyl residue bounded to their hemiacetal hydroxyl groups. In the case of D-galactose and L-arabinose, the enzyme produced not only non-reducing oligosaccharides, but also reducing oligosaccharides, identified as 3-O-β-D-fructofuranosyl-D-galactose and 4-O-β-D-fructofuranosyl-L-arabinose, respectively. The enzyme is useful to synthesize heterooligosaccharide containing fructose. The lactosu-crose has been produced on industrial scale by using this enzyme, and supplied in, large quantities for food applications. The lactosucrose is not digestible in the human small intestine, but it is fermented by human intestinal microorganisms, especially by Bifidobacterium. The admin-istration of the saccharide improved the intestinal bacterial flora.

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1. Cyclodextrin-producing enzyme produced by Bacillus macerans (CGTase, macerans enzyme) Crude macerans enzyme preparation was purified and the enzyme was crystallized. Elucidation of the CDs' forming path : By use of various kinds of substrates, well-known phenomenon of p-CD accumulation at final stage of the reaction was explained from time course data of CD formation and the rate of coupling. Elucidation of cyclizing reaction : Cyclizing reaction proceeds from non-reducing ends of the substrates cyclizing 6 glucose units to form α-CD and the remaining fragments. The rate of cyclization was high on substrates having more than 8 glucose units, and accelerated by the addition of helices forming reagents ; moreover, it was found that α-and β-CDs were selectively formed by the addition of 65 and 76 helices forming reagents. Branched CDs were produced by forming helical structure with surface active reagents from substrates having branches, and in the same manner, 1 or 2 hydroxyethyl substituted CDs were produced by the action of macerans enzyme on the helix formed hydroxyethyl starch.Elucidation of coupling action : Glucosyl-α-CD and 14C-labeled glucose were reacted with macerans enzyme, and the structure of the formed branched oligosaccharides was analyzed. The smallest non-radioactive branched oligosaccharide was B4(63-α-glucosylmaltotriose), and at the initial stage of the reaction, radioactive(*) Bs(mainly 66-α-glucosylmaltoheptaose)was formed to be gradually degraded to B7* (mainly 65-α-glucosylmaltohexaose), B6 (mainly 64-α-glucosylmaltopentaose) and B5 (64-α-glucosylmaltotetraose) From the results described above, the author proposed an enzyme model of active site.2. Production of CDs Methods for the production of α-CD, α-CD containing starch syrup, γ-CD were developed, and a large ringed CDs preparation method, by which δ-θ-CDs were produced, was also developed. δ-θ-CDs preparation contained a fair amount of intra branched CDs which have α-1, 6 linkage in the CDs themselves. 3. Production and the properties of branched CDs A method for the production of branched CDs which have branch(es) longer than glucosyl unit was developed, and maltosyl-CDs (G2-CDs) were effectively produced. The number of branches attached to CD ring depends on the size of CD, and 2 and 3 branches were easily attached to a-CD and p-CD, respectively. Panose was also attached to CDs to form panosyl-CDs. Branching moiety of the CDs were degraded by the action of glucoamylase to glucosyl-CDs (G1-CDs), and maltose was again attached to G1-CDs to form doubly branched G1-, G2-CDs by the action of pullulanase. Properties of branched CDs : Solubility was higher than that of original CDs, and degree of the solubility was different depending on the variety of CD, the variety of branch and the number of branches. As for the action of starch-degrading enzymes on the branches, it was found that the action of glucoamylase was considerably different depending on the variety of branched CD. Amylo-1, 6-glucosidase acted on AD and AC type of diglucosyl-α-CD((G1)2-α-CD), but not on AB type. Aspergillus oryzae α-amylase degraded G1-CD, but the rate of degradation was depressed to less than 1/10 in comparison with that of original CD, and multiply branched glucosyl-CDs were scarcely degradable. Pullulanase (branching enzyme) scarcely degrades maltosyl branches of AB type of branched CDs, but acted on the branches of AC and AD types. AD type of (G1)2-α-CD was completely resistant to the action of macerans enzyme ; AB and AC types were reacted in the presence of acceptors such as glucose and maltose. By the combination of the results described above, systems for the separation and produc-tion of AB, AC and AD types of (G1)2-α-CD were established.

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The synthesis of starch is believed to involve 3 enzymes, ADPglucose pyrophosphorylase (ADPGIc PPase, [EC 2.7.7.27] ; reaction 1), starch synthase, (EC 2.4.1.21) ; reaction 2) and branching enzyme ([EC 2.4.1.18] ; reaction 3). (1) α-glucosyl-l-P + ATP ↔ ADPGIc + PPi (2) ADPGIc + (glucosyl) _n → ADP + (glucosyl) _n+1 (3) Linear α-1, 4-glucosyl chain of α-glucan→Amylopectin (a-1, 6 and 1, 4-glucan) There are many genetic studies to indicate that all three reactions contribute to the synthesis of both starch components, amylose and amylopectin. Maize amylose extender (ae) mutants, containing more amylose and less amylopectin than normal maize have been shown to be defective in one of the branching enzyme isoforms (Plant Physiol., 67, 1141-1145 (1981)). Maize waxy (wx) mutants which contain no amylose are deficient in the starch granule bound starch synthase (Plant Physiol., 62, 383-386 (1978)) indicating that the granule bound starch synthase is involved in amylose synthesis. Starch deficient mutants of pea (Plant Physiol., 89, 1279-1284 (1989)), maize (Science, 151, 343-348 (1966)) and Arabidopsis (Plant Physiol., 86, 1131-1135 (1988)) are deficient in ADPGIc PPase activity. Thus, genetic and biochemical evidence in plants strongly indicate that ADPglucose (ADPGIc) is synthesized predominantly, if not solely, via ADPGIc PPase catalysis. The cyanobacterial, algal and higher plant enzymes are activated by 3-P-glycerate (3PGA) and inhibited by orthophosphate. An algal mutant in which the regulatory sites of the ADPGIc PPase have been affected (Planta, 185, 17-26) (1991) and control analysis (Kacsar-Burns) of the plant ADPGIc PPase, indicate that the allosteric regulation observed of the enzyme in plants reflects the in vivo regulation of plant starch synthesis. Thus, the above studies empha-size the importance of the ADPGIc PPase in starch synthesis and the need for understanding the structure-function relationships of the regulatory and catalytic domains of the ADPGIc PPase. A number of chemical modification and site-directed mutageneses studies have identified the active and substrate binding sites both in the spinach leaf and bacterial enzymes. There is considerable conservation between the bacterial and higher plant enzymes particularly at the substrate binding sites. High conservation is also observed for the 3PGA activator sites of the cyanobacterial and higher plant ADPGIc PPases. These results suggest that the two subunits of the higher plant ADPGIc PPase evolved from the bacterial enzyme subunit. Transformation of the plant systems with the bacterial enzyme gene results in an appreciable increase in starch synthesis ranging from 1.25-to 1.6-fold in potato tubers to 10-fold in tobacco calli. These results confirm that the ADPGIc PPase activity in plants is rate-limiting for starch synthesis and that the allosteric regulation is functional in the plant.

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Molecular structures of amylose and amylopectin from several sources have been analyzed in detail by newly developed methods. Amylose content has been suggested to be determined based on the iodine affinities (or blue value) of amylose, amylopectin and starch of each specimen. The value was slightly lower than the value by assuming the iodine affinities of amylose and amylopectin as 18-20 and 0, respectively. The latter value was defined as apparent amylose content. Reducing and non-reducing residues were determined by the modified Park-Johnson's colorimetric assay and the rapid Smith degradation coupled with determination of glycerol by enzymic assay. Number-average molecular weight (or d. p. n) and number-average chain length (c.1. n) were calculated from these values and total carbohydrate measured by phenol-sulfuric acid method. Molar fractions of linear and branched amylose were determined by 1) (number of branch linkage of amylose)/(number of branch linkages of β-limit dextrin) or 2) radioactivities of H3-amylose and H3-limit dextrin. Molecular weight distribution and weight-average molecular weight (or i p, w) of amylose and debranched amylopectin (chain-length distribution) were analyzed by high-performance-gel-exclusion chromatography monitoring with a low-angle laser-light-scattering photometer and a differential refractometer. The mode of chain-linking of amylopectin was analyzed by β, i, p-degradation, which supported the irregular Meyer structure. It was found that the B chains of wheat and rice amylopectins linked increased number of A chains with increased chain length. Some B chains of wheat amylopectin were (37%) found to link no A chains but only B chains. A key factor for the cause of crystalline polymorphism of starch granules was found to be the chain length of amylopectin. The temperature and various materials appear to be secondary modulators. This suggests that molecular structure is the most basic factor on the functional properties of starch, but still we know only a part. We need more effort toward the full understanding of starch.

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Researches on structural analysis, physical properties of starch and discoveries of novel starch related enzymes are summarized in this paper as an award address. 1) Physical properties and utilization of various starches Starch phosphate derivatives obtained by chemical synthesis or simple cation substitution of naturally existing phosphate of potato starch are discussed in terms of physical properties and possible utilization. Sago starch as a. neglected starch resource in tropical area and newly developed methodology during the study such as photopastegraphy, Q-amylase/pullulanase method (BAP method) are also described. 2) Fine structure of amylopectin: Fine structures of waxy maize starch were analyzed by structure of a-limit dextrin, X-ray diffraction and enzymic structural analysis of Naegeli amylodextrin, transmission electron microscopy and computer modeling. From the results of these studies, French proposed?gcluster model?hfor amylopectin molecules and Kainuma and French proposed?gdouble helix?hconformation for crystalline part of amylopectin. In 1980, Kainuma represented schematic arrangement of amylopectin molecules within a growth ring. 3) Novel starch related enzymes and oligosaccharides: During structural analysis, exo-maltohexaohydrolase (E. C. 3.2.1.98) was unexpectedly discovered as a contaminant of A. aero genes pullulanase. Since this discovery, our research efforts were concentrated to screen new starch related enzymes and enzymatic preparation of oligosaccharides. As the results, cyclodextrins and branched cyclodextrins, isomaltose, maltohexaose, erythritol -a noncaloric sweetener-were already commercially manufactured. Raw starch hydrolyzing glucoamylase of Chalara paradoxa isolated from sago palm tree in Papua New Guinea is expected for energy-saving saccharification of starch.

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Rice originated in the highland of Asia, was introduced into Japan with the movement of people and culture, and adapted all over Japan as Japonica varieties, which have a very homo geneous hereditary nature as a result of inbreeding for long, long years. Tradition says that the rice in the past was not always the same as it now is, being often red, black, and/or fragrant. The systematic breeding of rice has begun since the middle of the Meiji era, but the target has changed from time to time depending upon the socio-economic situation, to which this paper refers in detail. This paper also explains research approach on the development of rice varieties by many prefectures and the outline of a project titled?gSuper Rice Program, ?hwhich the Ministry of Agriculture, Forestry and Fisheries is carrying out . It is very difficult to look upon the rice in the future, since the international situation connected with rice is now fluctuating . However, it is certain that rice was, is and will be an important, staple food for Japanese, and at the same time, paddy field was, is and will be important to sustain a comfortable and moderate environment . And the qualities of rice will be improved in taste and diversified much more to meet the needs of consumers, food industries and medical cure.

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The application of both pressurization and enzymatic treatment to aged rice grains was investigated to improve their cooking properties. Pressurization at 100 MPa was favorable in improving the properties. For treatment of the grains with enzymes, actinase was more effective than cellulase and pectolyase. Actinase-treated grains, when cooked, gave a favorable result in terms of the stickiness/hardness ratio and brightness . Stickiness and brightness of cooked rice were developed by removing albumins and globulins from grains . In newly-harvested grains, the proteins were easily removed by extraction with water which contained rice-originating minerals. In aged grains, however, the proteins were already denatured and extracted with difficulty. Proteases hydrolyzed the denatured proteins to be extractable . As a result, starch granules existing on the surface of grains were liberated. Some proteases also hydrolyzed a starch granule-associated protein, and, consequently, made the starch highly gelatinized by heating. The protease treatment of aged grains was effective for two main reasons ; liberation of starch granules and removal of the granule-associated protein . During cooking of rice the liberated starch without the associated proteins gelatinized to give rise to stickiness . The gelatinized starch is considered to form a more isotropic film on the surface of every cooked rice grain to give brightness as well.

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In Japan, the market for convenience rice products which can be served and consumed with ittle preparation is expanding. This paper classifies these convenience rice products into 1) dry gelatinized rice, 2) expanded rice, 3) freeze-dried rice, 4) retorted rice, 5) aseptically packaged rice, 6) frozen rice and 7) the others; and discusses their typical features and manufacturing techniques

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[in Japanese]

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The texture of cooked rice appears to be greatly dependent upon the properties of starch. Here, the structures and properties of rice starch were discussed based on our recent knowledge. Non-waxy rice starches having low (Sasanishiki, Koshihikari), intermediate (unknown, IR48, IR64) and high (IR32, IR36, IR42) iodine affinity (AI) were said to have low, intermediate and high amylose contents, respectively. However, from the analyses of AI of amylose, amylopectin and starch, each starch was found to have similar amounts of amylose, because the AT of amylopectins of high AT starch were high but those of amyloses were similar. Amylopectins with higher AI contained increased amounts of long chains, which were not found in waxy rice. The mode of each chain connection was analyzed by β, i, β-degradation. The longer B chains carried increased numbers of A chains, and the structures of the chain linking within a cluster have been suggested to be the random Meyer structure. Molecular weights of amylopectins with high IA were lower than those of others, and [η] did not correlate with molecular weight. However, the amylopectins with high AT had high [η]. This implies that amylopectins with higher AI had more elongated shapes than others in an alkaline solution (1 N NaOH, 22.5°C). Amyloses from rice starches had the similar dn p. n and d. p. w. However, the ratio of linear and branched molecules and β-amylolysis limit differed by cultivars. The β-amylolysis limit of branched amylose was found to be 39%. The d.p.n of the branched amylase was 1.8-fold that of linear molecules. The side chains of branched molecules were mostly short chains with average chain-length of 20. The [η] of amylose had good correlation with the amount of branched molecules by weight. The branched molecules probably are more stable than linear molecules in aqueous solutions and have less retrogradation tendency.

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The wx locus controls amylose synthesis in the endosperm of rice . Since the amylose contents in the endosperm greatly affect the quality of rice, it is important for agricultural application to regulate the extent of amylose synthesis . In addition, the wx locus of rice is quite suitable for the molecular studies of gene regulation because the locus has some particular advantages for genetic research and it provides us various regulation mechanisms to be eluci dated at the molecular level. In this article, we review the genetic and molecular biological research, which have been carried out in our laboratory, about the regulation of gene expression at the wx locus and amylose synthesis in rice.

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Transglycosylation reactions catalyzed by the fungal p-transglycosylases as well as the bacterial fructosyltransferase are reviewed with respect to their mechanisms and specificities in the biosynthesis of corresponding oligo- and polysaccharides. β-Transglycosylases (β-TG) of Sclerotinia libertiana . and Trichoderma longibrachiatum catalyzed the intermolecular transglycosylation using cellooligosaccharides as both glycosyl donors and acceptors. High molecular weight components synthesized by the reactions were identified as linear β-1, 4-glucans having average d. p. of 14 or more. Both enzymes were defined as new enzymes with a disproportionating activity on β-1, 4-glucosidic linkage. Interaction of β-TG with native cellulose fiber was demonstrated in the enzyme of T. longibrachiatum, suggesting its functional contribution to the fungal cell wall synthesis. The manner of fructosyl transfer by Bacillus subtilis levansucrase was easily modified with the environmental conditions. The enzyme showed jS-fructosidase-like activity, catalyzing the hydrolysis of sucrose at a high temperature as well as the fructosyl transfer to the acceptor molecule when it was present at a high concentration. The enzyme lost the ability to synthesize highly branched levan of high molecular weight in the mixture with high ionic strength where it preferably synthesized low molecular weight levan of 4 or 5 branches.

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The molecular basis of cell adhesion is of fundamental importance for under standing differentiation, organization, and development in multicellular organisms, as well as numerous diseases and disorders involving abnormal cell adhesion . Out of five families of cell adhesion molecules, two (invol ving selectin-CHO and CHO-CHO interaction) have been reviewed in this lecture. U nderstanding of CHO-based adhesion events has important diagnostic and the rapeutic implications.

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Vast knowledge on the biological functions of carbohydrate, especially as recognition marker/signal, has been accumulated during the past decade and led to the establishment of a new field of science, glycobiology. Also, there have been various impacts from the technology side relating to carbohydrate. For example, application of genetic engineering to the production of glycoproteins faced the problem of proper design and regulation of glycan chains in the glycoproteins for the production of biologically active molecule, On this background, the establishment of glycotechnology, a new field of biotechnology which aims to clarify the function of carbohydrate and utilize it, has been proposed mainly by Japanese researchers. Lectins which recognize and bind specific carbohydrates have two major roles in glycotechnology. Ore is the biological role as the molecule which recognizes the signal/marker written in a specific sugar sequences. Lectins are also important as valuable tools in various technology in the field of glycotechnology. In this article, the present situation on the studies of lectins is reviewed in relation to these two aspects. Basic studies on the molecular recognition of carbohydrates by lectins are also discussed.

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A lot of carbohydrate products are now in industrial use . Modified starch must be the oldest one of the products . Starch hydrolyzates such as dextrose and maltose are useful sweet ener and high fructose corn syrup founded a big sweetener market. Sugar transferred oligosaccharides, coupling sugar and fructo-oligosaccharide have been established asnew physiological functional sugars which are good for health . Recently, many different kinds of sugars and sugar chains that exist widely in living cells have been shown to play an important role as bi ological signal substances . In this paper the usefulness of these carbohydrates is reviewed.

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Starch is a main component of cereal and vegetable foods . In addition, it is added to many kinds of foods to improve the texture and taste as an ingredient . Starch changes its physicochemical properties by gelatinization, when it is cooked. Consequently, the behaviors of pasting and the paste properties of starches, such as gelatinization temperature, and transparency, viscosity, and retrogradation tendency of the paste, etc. give important effects on cooking and the processing of foods. In this article, the properties of various kinds of starches and their relations with the functions and roles on cooking were discussed.

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Several newly developed rice types were compared with a widely used standard rice variety (nihonbare) ; the physical properties of the cooked rices were measured and evaluated by sensory analysis and visual appearance. Chemical properties and physical properties such as viscosity and gelling properties of rice starch and rice flour were measured. There was clear relationship among amylose content, high amylopectin Fr. II and eating quality. All the newly developed rices showed different characteristic properties, we were able to utilize these properties in cooked rice and processed foods by appropriate cooking method and processing.

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