Abstract: D-ketohexose 3-epimerase, a novel epimerase, is obtained by cultivating bacteria of the genus Pseudomonas including Pseudomonas cichorii ST-24 (FERM BP-2736). D-Ketohexose 3-epimerase epimerizes D-ketohexose, D-ketopentose and L-ketopentose at their C-3 positions to form their corresponding epimeric counterparts in a high yield at a high conversion rate. Interconversion reaction using D-Ketohexose 3-epimerase yields mixture of intact ketose and its epimeric counterpart which can impart an appropriate sweetness, gloss and improve taste quality when used in foods, beverages, feeds, pet foods, dentifrice, cachou, sublingual agents and internal medicines.

Abstract: A method of providing and sustaining a difference in pH of a first reaction of an immobilized enzyme and a second reaction in a bulk liquid surrounding the immobilized enzyme is carried out with a bilayer pellet containing coimmobilized enzymes. The pellet can contain an enzyme that produces a desired product immobilized in an inner core and urease immobilized in an outer layer. The bulk liquid contains urea and a substrate for the enzyme in the core, and has an acidic pH. The urease reacts with urea diffusing into the outer layer from the bulk liquid to produce ammonia. The ammonia consumes hydrogen ions diffusing into the inner core from the acidic bulk liquid. This provides the enzyme in the inner core with a pH higher than the acidic pH of the bulk liquid suitable for the enzyme to react with the substrate as it diffuses into the inner core.

Abstract: A method for selecting amino acid residues is disclosed which upon replacement will give rise to an enzyme with an altered substrate specificity. New mutant glucose isomerases with an altered substrate specificity are provided according to this method. These altered properties are useful in starch degradation and in other sugar conversion reactions.

Abstract: The present invention relates to a general method of enzymatic synthesis of isotopically labeled carbohydrates, sugars and nucleosides. Labeled citric acid cycle intermediates, amino acids and ribose mononucleotides may be rapidly and conveniently synthesized from labeled pyruvate, lactate or L-alanine. The method employs a novel nicotinamide dinucleotide regeneration system which permits use of low NADH levels. The method may be manipulated to allow labeling at a variety of carbon/hydrogen sites.

Abstract: Physically strong particles containing an immobilized enzyme are prepared for use in a fixed bed-column. The particles are prepared by adding a homopolymer of 1-amino ethylene or a copolymer of 1-amino ethylene and N-vinyl formamide to an aqueous medium containing an enzyme, adding a cross-linking agent to cross-link the enzyme and polymer and cause flocculation, and dewatering, sub-dividing and drying the resultant flocculant. Preferably, the enzyme is glucose isomerase and the cross-linking agent is glutaraldehyde, polyazetidine or diisocyanate.

Abstract: This invention is in the field of glucose isomerization enzymes. More specifically, the invention is directed to a novel xylose isomerase, a process for the preparation of this enzyme, the use of this enzyme in glucose isomerization processes, and glucose isomerization processes.The enzyme is preferrably derived from Thermotoga maritima or Thermotoga neapolitana. The enzyme has a temperature optimum above 90.degree. C., pH optimum in the range of from 6 to 7 and a residual activity at 90.degree. C. of more than 40% after 30 minutes and/or residual activity at 98.degree. C. of more than 20% after 30 minutes. The enzyme can also be in immobilized form.

Abstract: A bilayered immobilized enzyme pellet and a process to manufacture this pellet are provided for use in a process involving the simultaneous isomerization of xylose to xylulose and fermentation of xylulose to ethanol. The bilayered pellet is able to maintain the environment where the isomerization reaction occurs within its optimum pH of 7.0 to 8.0 while the fermentation reaction occurs within its optimum pH range of 4.0 to 5.0. This process allows both xylose and glucose sugars to be effectively used as a feedstock for ethanol production by isomerizing the xylose to xylulose and then making the xylulose immediately available for the fermentation process. Because the xylose has been converted to its ketose isomer, xylulose, yeasts which can ferment glucose and xylulose can be used in this process.

Abstract: A mannose isomerase having excellent properties for industrial use, such as high thermal stability and resistance to high substrate concentrations, can be produced by culturing a strain of Pseudomonas (sp. AM-9582), and extracting it from the cells of AM-9582. Mannose can be effectively produced from fructose of high concentrations using the enzyme.

Abstract: Amylodextrin compositions are produced from starch hydrolysates having a dextrose equivalent (DE) less than 10 or from milled substrates such as cereals, oilseeds, and vegetable fibers. The compositions give aqueous gels with unexpectedly increased strengths and improved fat-sparing characteristics, including more desirable flavor and texture qualities. The amylodextrin compositions are segregated from hydrolyzed aqueous cereal flour or starch hydrolysate solutions, for example, by precipitation with the proper amount of a water-miscible organic solvent, such as ethanol, acetone, or 2-propanol.

Abstract: This invention provides a method for the maintenance of continuous activity of an immobilized enzyme reaction by initially underloading an adsorbent with a desired enzyme, monitoring the activity of the enzyme and periodically adding fresh enzyme to maintain a constant, continuous level of activity until the maximum carrying capacity of the support is reached. The method is preferably carried out when continuously isomerizing glucose to fructose with glucose isomerase adsorbed to a weakly basic anion exchange resin.

Abstract: This invention allows high-purity maltose to be manufactured both simply and economically by sequentially going through the steps of liquefaction of starch, saccharification of the resulting liquefied substance by combining with general-purpose enzymes and further saccharification with an enzyme which hydrolyzes oligosaccharides of trisaccharide or more, and also allows the economical and favorable manufacturing of maltitol, the reduced product of the above maltose, by going through an additional reduction step.

Abstract: A mannose isomerase having excellent properties for industrial use, such as high thermal stability and resistance to high substrate concentrations, can be produced by culturing a strain of Pseudomonas (sp. AM-9582), and extracting it from the cells of AM-9582. Mannose can be effectively produced from fructose of high concentrations using the enzyme.

Abstract: Process for manufacturing xylitol and xylitol-rich products, characterized in that it consists of:enzymatically isomerizing at M.sub.1 D-xylulose syrup into syrup containing D-xylose and D-xylulose then, without extracting the xylose,catalytically hydrogenating this syrup at M.sub.2, resulting thus in a xylitol-rich syrup, said xylitol-rich syrup being either dehydrated, or subjected to chromatographic processing or to a treatment of extraction by crystallization at M.sub.3.

Abstract: A process for producing fructose-1,6-diphosphate comprising the steps of: (a) enzymatically converting adenosine 5'-diphosphate to adenosine 5'-triphosphate using an acetate kinase-containing microorganism or an extract of the microorganism and phosphate donor; and (b) enzymatically converting a substrate capable of being converted to glucose or fructose to fructose-1,6-diphosphate using the adenosine 5'-triphosphate resulting from step (a) and the acetate kinase-containing microorganism or the extract of the microorganism.

Abstract: A method for preparing cellobiose from sucrose in a high yield at low cost without difficulty by using three enzymes of sucrose phosphorylase, glucose isomerase and cellobiose phosphorylase in the presence of orthophosphate. The additional method of the invention comprises 4 steps: to treat sucrose with sucrose phosphorylase in the presence of orthophosphate to produce fructose and glucose-1-phosphate; to treat the fructose obtained in the preceding step with glucose isomerase to produce glucose; to treat the glucose obtained in the preceding step and the glucose-1-phosphate obtained in the first step with cellobiose phosphorylase to produce cellobiose and orthophosphate; and to recover at least a part of cellobiose from the reaction mixture in the preceding step and to recycle at least a part of the remaining reaction mixture containing orthophosphate to the first step.

Abstract: A process for obtaining N-acetylneuraminic acid from N-acetylglucosamine is disclosed. The process is carried out in a reactor which contains both N-acylglucosamine-2-epimerase (E.C. 5.1.3.8) which isomerizes GlcNAc into ManNAc, and N-acetylneuraminic acid pyruvate lyase (E.C. 4.1.3.3) which catalyzes the reaction of the resulting ManNAc with pyruvic acid to give Neu5Ac. GlcNAc and Pyr are fed into the reactor and Neu5Ac is obtained in the outflow. The process is preferably carried out continuously and in particular in an enzyme membrane reactor at pH 7.5 and 25.degree. C., especially using residence times of 0.2 to 10 h, and with an excess of GlcNAc in comparison with Pyr which is subsequently added if necessary. Epimerase and lyase are preferably present in the reactor in a ratio of activities which is equivalent to the reciprocal value of the quotient of the conversion rates.

Abstract: A method of preparing a mixture of galactooligosaccharides having the following formula:Gal-(Gal)n-Glcwhere Gal represents a galactose residue, Glc represents a glucose residue, and n represents an integer between 1 and 4, and monosaccharides by having microorganisms containing .beta.-galactosidase or .beta.-galactosidase act on lactose, the method comprising:having glucose isomerase coexist in a liquid to be reacted or adding glucose isomerase after completion of the reaction, whereby a portion of glucose prepared by the .beta.-galactosidase processing is converted into fructose. Then, sweetener is prepared by separating monosaccharides from the mixture.

Abstract: A process for converting AMP into ATP which comprises (a) using an enzyme which converts AMP into ADP and has been produced from microorganisms having an optimum growth temperature of 50.degree. C. to 85.degree. C. and an enzyme which converts ADP into ATP and has been produced from microorganisms having an optimum growth temperature of 50.degree. C. to 85.degree. C. is disclosed. In addition, there is disclosed a process for producing a physiologically active substance by a multienzyme process which comprises forming ATP from AMP by the step (a), (b) synthesizing a physiologically active substance with the resulting ATP, converting AMP resulting from the reaction in step (b) into ATP by the reaction in step (a), and repeatedly utilizing the converted ATP for synthesis of the physiologically active substance in step (b). By using the process it is possible to stably and efficiently carry out conversion of AMP into ATP over a long period of time.

Abstract: The subject invention concerns a process for stabilizing intact or ruptured microbial cells having glucose isomerase associated therewith. Specifically exemplified is a process for stabilizing glucose isomerase producing cells of a microorganism belonging to the genus Ampullariella. In the invention process the whole or ruptured microbial cells are contacted with a partially carboxyalkylated- or partially phosphonoalkylated-cationic polyelectrolyte, for example, a partially carboxymethylated polyethyleneimine to flocculate and stabilize the cells. The flocculated cells are further stabilized by encapsulation with a partially carboxyalkylated- or partially phosphonoalkylated-cationic polyelectrolyte. The encapsulation can be done prior to or after the flocculated cells are crosslinked. The net effect is manifested by a dramatic increase in the half-life of the glucose isomerase.

Abstract: Polysulfonium salts that can react with nucleophilic groups and covalently cross-link are used to immobilize enzymes or enzyme-containing cellular material. Some of the polysulfonium salts can both flocculate and covalently cross-link. Replacement of the cross-linker, glutaraldehyde, with the polysulfonium salt results in greater retention of enzyme activity during immobilization. Immobilization is carried out by forming a mixture of an enzyme or enzyme-containing cellular material and the polysulfonium salt and subjecting the mixture to conditions such that sulfonium ions react with nucleophilic groups contained by the enzyme or cellular material to form a covalently cross-linked and water insoluble product. The enzyme or cellular material may be flocculated with a flocculating agent prior to cross-linking with the polysulfonium salt. The polysulfonium salt can be a polymer containing sulfonium groups.

Abstract: A process is disclosed for the production of substantially pure fructose from sucrose-containing substrates. The process comprises converting the sucrose to levan and glucose, purifying the levan by membrane technology, hydrolyzing the levan to form fructose monomers, and recovering the fructose.

Abstract: Constructions and methods for mutagenesis of nucleic acids involve chemical mutagenesis of a cassette comprising a structural gene linked to a non-functional restorable fragment of a marker gene. Mutants are detected by screening for the presence of the reconstituted marker among the ligation products of the cassette to a vector containing the non-functional restorable remainder of the marker gene. Xylose isomerase mutants, characterized by a change from glu (GAG) to lys (AAG) at amino acid position 262 in the xylA protein of E. coli were obtained by partial marker cassette mutagenesis. These mutants enzymes exhibited twice the rate of isomerization of glucose to fructose exhibited by the wild type.

Abstract: A large DNA cloning system is disclosed which is based on yeast artificial chromosomes. Cloning vectors are disclosed which allow the cloning of large segments of greater than 50 kb of exogenous DNA. The cloning vector comprises DNA sequences of an autonomous replication sequence, a centromere, a selectable yeast marker, two sequences that seed telomere function in vivo, and a cloning site within an interruptible yeast gene for insertion of the exogenous DNA segments.

Abstract: Rapid identification of different species of microorganism selected from fungi and yeast like algae is accomplished by culturing the microorganism for several hours under normal conditions on a non-inhibitory mycological medium which stimulates the microorganism to make characteristic enzymes by which the microorganism can be identified, distributing the culture (in suspension) onto several supports containing different substrates which are capable of reacting with the enzymes so produced by the different species of microorganisms; and rapidly incubating the admixture to produce a distinctly colored or colorable reaction product.

Abstract: A process for producing fructose sweetened cereal products by enzymatically converting a portion of the cellulose fraction in a cereal comprising cereal; fiber to fructose using cellulase and glucose isomerase is claimed. The process may be carried out at moisture contents exceeding 25% (w/w) and moisture contents between about 40% to 80% are preferred.

Abstract: The present invention relates to a continuous process for the enzymatic preparation of isomaltulose. A periplasmatic sucrose-mutase is produced by fermentation of microorganisms which form sucrose-mutase. The cell-free crude enzyme extract is prepared by digestion of the cells and by cross-flow microfiltration. In a single-stage, simultaneous purification and immobilization of the sucrose-mutase from the cell-free crude extract conditioned by diafiltration is obtained by selective bonding to an anionizable carrier matrix. Direct conversion of sucrose into isomaltulose is produced by the sucrose-mutase bonded to the anionizable carrier matrix, preferably in cartridge or cartouche form.

Abstract: Breakfast cereals are sweetened by treating cereal grains or at least one cereal grain fraction such as bran, with enzymes comprising glucoamylase and glucose isomerase to produce fructose while retaining cereal particle discreteness or integrity. Enzymatic treatment with alpha-amylase may be initiated prior to, during, or after cooking. The enzymatically treated, cooked cereal grains are formed into breakfast cereal shapes and the enzymes are inactivated to provide a shelf-stable cereal product. The cereal products exhibit a sweet, pleasing complex-honey-like taste and aroma. Producing fructose provides a greater level of sweetness for a given amount of starch conversion into low molecular weight reducing sugars such as mono- and di-saccharides. In achieving a given level of sweetness, more starch or high molecular weight dextrins may be retained for their matrix forming ability or for improved machineability of the enzymatically treated cereal grains into breakfast cereal shapes.

Abstract: A low cost process for producing concentrated fructose is described which involves selective, preferential biological utilization of glucose in mixtures of glucose and fructose, in order to thereby yield concentrated fructose. Broadly speaking, the method involves contacting a mixture containing respective amounts of glucose and fructose with a microorganism which preferentially utilizes glucose as compared with fructose, incubating the mixture until the relative concentration of fructose is substantially in excess of that of glucose, and recovering concentrated fructose. In preferred forms, the microorganism is Pullularia pullulans, and the starting material may be a mixture of gluctose and fructose, sucrose, a carbohydrate such as inulin or starch; in the latter instances, the P. pullulans, by virtue of excretion of invertase, acts to degrade the starting material to give the desired glucose-fructose mixture.

Abstract: An enzyme producing microorganism or an enzyme produced thereby is immobilized by contacting the microorganism or enzyme with polyethylenimine and adding chitosan and glutaraldehyde to form a reaction product. In a preferred embodiment, the reaction product is extruded onto the revolving plate of a spheronizing device to form spherical enzyme aggregates having enhanced physical and biocatalytic properties.

Abstract: Enzyme-containing microbial cells are contacted with a macro-porous non-ionic resin under reduced pressure below atmospheric to cause the cells to become incorporated into pores of the resin. The resin is then contacted with a multi-functional, amine reactive material to immobilize the cells within the pores. Prior to contacting with the resin, cells may be disrupted to produce a system containing cell fragments, intact cells and solubilized enzyme. A preferred enzyme is glucose isomerase.

Abstract: A composition comprising immobilized cells obtained by applying a dispersion of cells and curable prepolymer material selected from the group consisting of polyazetidine prepolymers, carboxymethyl cellulose, polyurethane hydrogel prepolymers and polymethylene isocyanates. as a coating to a solid inert carrier and curing the prepolymer on the carrier at a temperature below the temperature at which enzyme activity of the cells is significantly reduced. The composition may be used to produce various materials such as L-aspartic acid, L-alanine, 6-Aminopenicillanic acid, high fructose corn syrup, prednisolone or phenylalanine.

Abstract: A process is provided for the preparation of magnetic particles to which a wide variety of molecules may be coupled. The magnetic particles can be dispersed in aqueous media without rapid settling and conveniently reclaimed from media with a magnetic field. Preferred particles do not become magnetic after application of a magnetic field and can be redispersed and reused. The magnetic particles are useful in biological systems involving separations.

Abstract: Xylose isomerase, from strains of the Streptomyces murinus cluster, a method for production of such xylose isomerase, immobilized xylose isomerase and a method for isomerization of glucose to fructose therewith.

Abstract: Immobilized enzymes covalently bound to an inorganic carrier by an amino group through a bifunctional spacer have improved properties when the carrier is formed of amorphous, approximately spherical silica particles obtained from synthetic calcium silicate. The silica particles have an average particle size of from 15 to 80 .mu.m, an apparent particle volume of from 1.3 to 3 cm.sup.3 /g, and a specific surface area of from 250 to 800 m.sup.2 /g.

Abstract: Extracellular enzymes are stabilized with a carboxyalkylated or phosphonoalkylated polymer having a molecular weight of at least 500 Daltons. Exemplified is the stabilization of the enzyme glucose isomerase used in a process to convert D-glucose to D-fructose. In a preferred embodiment of the invention the feedstream containing the substrate is contacted initially with the stabilizer and then with the enzyme. In this system the stabilizer and enzyme are maintained in separate reactors. This separation, advantageously, results in a higher half-life for the enzyme.

Abstract: Disclosed is a process for the preparation of a solution containing glucose and fructose (isoglucose) by the conversion of a glucose-containing solution on a catalyst having glucose isomerase activity and produced on the basis of a SiO.sub.2 carrier. The productivity of the catalyst may be significantly increased by the addition of SiO.sub.2 to the glucose solution, and the catalyst according to the present invention is not damaged by temporary process shutdowns.

Abstract: Microbial cells are immobilized with a curable polyaziridine or polyfunctional aziridine prepolymer to obtain an insoluble, crosslinked polymer containing the cells. The microbial cells immobilized may be cells having L-aspartase or L-phenylalanine transaminase activity for the production of L-aspartic acid or L-phenylalanine. The polymer containing the cells may be formed as a coating on a solid inert carrier.

Abstract: Process for preparing fructose by treating starch with alpha-amylase, contacting the resulting liquefied starch with glucoamylase to hydrolyzed said starch to glucose, and isomerizing at least part of the resulting glucose to fructose by contacting said glucose with glucose isomerase. The three enzymes are obtained from organisms of the Basidiomycetes class of fungi.

Abstract: An immobilized glucose isomerase having increased productivity and stability is prepared by mixing a smectite filler and 50-100 mesh granular activated carbon with flocculated cells of an organism of the genus Actinoplanes and forming the resulting mixture into discrete particles.

Abstract: Enzymatically active protein-enzyme complex membranes are prepared by treating a swollen protein membrane with an aqueous solution of a compatible active enzyme. These membranes are used to effect enzymatic reactions such as hydrolyzing starch, sucrose, urea or cellulose, lysis of cells or isomerizing D-glucose.

Abstract: An improved process is provided for conversion of crude starch-containing materials such as broken-polished rice and corn grits to starch hydrolyzates and high fructose syrups. The process employs short steeping and milling steps before enzymatic liquefaction and saccharification of the starch. The process causes a minimum solubilization of protein giving readily purified products.

Abstract: A process for obtaining glucose from thinned starch by partially hydrolyzing the latter to give from 50% to 92% glucose followed by separation of the hydrolysis product to afford a glucose-enriched product with recycling of the glucose-depleted stream affords benefits unattainable by conventional commercial processes. Substantial reductions in process time and reversion products and a substantial increase in productivity are among some of the benefits.

Abstract: The invention relates to a method for producing a glucose isomerizing enzyme which is functional at temperatures over 90.degree., utilizing a strain microorganisms having the identifying characteristics of Arthrobacter sp. ATCC 21748.

Abstract: The limitation of an immobilized amyloglucosidase in hydrolyzing thinned starch to afford not more than about 93% glucose with isomaltose levels above about 1.5% can be overcome in a process for converting thinned starch to fructose using four closely coupled reactor stages. The first stage is a saccharification reactor using amyloglucosidase which converts thinned starch to a product containing from 50% to 85% glucose. This product is used in a first stage isomerization reactor, the effluent from which is sent to another saccharification reactor using immobilized amyloglucosidase where hydrolysis is continued until no more than about 6% disaccharides and higher oligosaccharides are present. Where this effluent is used as a feedstock for further conversion of glucose to fructose, it is operationally equivalent to a feedstock containing at least 94% glucose but with isomaltose levels under about 1.5%.

Abstract: Glucose is enzymatically isomerized to fructose at a temperature of from about 90.degree. C. to about 140.degree. C. by contact with chemically stabilized glucose isomerase.

Abstract: A metal ion that inhibits enzyme deactivation is bonded to a carrier and inhibits enzyme deactivation when contacted with a substrate for the enzyme. A carrier-bound metal ion such as ions of iron is particularly suitable for inhibiting deactivation of glucose isomerase when isomerizing glucose to fructose. Glucose isomerase life is remarkably prolonged by contacting carrier-bound iron ions with a glucose substrate solution prior to isomerizing with glucose isomerase.

Abstract: A mutant Streptomyces thermoviolaceus, NRRL 15615, elaborates a thermostable glucose isomerase in high yield, constitutively. The GI produced shows a negligible loss in activity when heated at 90.degree. C. in high fructose corn syrup for 5 minutes relative to the same treatment at 80.degree. C. The mutant microorganism produces about 1500 units of glucose isomerase per gram of dry weight cells in the absence of xylose.

Abstract: A process for producing carbohydrates from vegetal juice comprises extracting a vegetal juice from raw vegetal material, filtering the vegetal juice to remove material suspended therein, subjecting the resultant vegetal juice to an enzymatic reaction which consists of four stages in the following sequence. The vegetal juice in a first stage is admixed with a mixture of 3.2.1.4-.beta.-1,4-glucan glucanhydrolase and 3.2.1.15-poly-.alpha.-1,4-galacturonic glucanhydrolase, then the vegetal juice is admixed with 3.2.1.20-.alpha.-D-glucoside glucohydrolase. In the second stage the vegetal juice is admixed with 3.2.1.20-.alpha.-D-glucoside glucohydrolase; in the third state the vegetal juice is contacted with 3.2.1.21-.beta.-D-glucoside glucohydrolase, 5.3.1.4-L-arabinose-ketol-isomerase and 5.3.1.5-D-xylose-ketol-isomerase; and in the fourth stage the vegetal juice is admixed with 3.2.1.1-.alpha.-1,4-glucan-4-glucanhydrolase.

Abstract: When isomerizing part of the glucose content of a glucose-containing solution to fructose, the productivity of glucose isomerase immobilized on an SiO.sub.2 carrier is increased considerably by pre-contacting the glucose-containing solution with particles of SiO.sub.2 or aluminum silicate. An apparatus is used having a reaction vessel containing the immobilized glucose isomerase and a pre-column containing the particles of SiO.sub.2 or aluminum silicate.

Abstract: A mutant Streptomyces coelicolor, NRRL 15398, produces glucose isomerase constitutively at a level at least as great as the parent does inductively in common growth media. The isomerase can be effectively used to isomerize glucose to fructose in a continuous process using immobilized enzyme.