Toyo Jozo Kabushiki Kaisha

Asahi Kasei Kogyo Kabushiki Kaisha

Toyo Jozo Company, Ltd.

Hidehiko Ishikawa

Hideo Misaki

Kazuo Matsuura

Shigeyuki Imamura

Masaki Takada

Naoki Muto

Shigeru Ikuta

Eiichi Yoshino

Fumitaka Inoue

Eiji Matsumura

Eiji Muramatsu

An assay method for a component in a specimen containing any one of ATP, deamide-NAD and an amide donor which comprises performing a main reaction which comprises incubating the specimen with NAD synthetase in the presence of ATP, deamide-NAD, an amide donor and Mg.sup.++ to generate NAD; performing a coenzyme cycling reaction by combining the oxidation-reduction reaction system with coenzyme NAD and the oxidation-reduction reaction system with coenzyme reduced NAD, and measuring a consumed or generated component in the cycling reaction. The NAD synthetase can be produced by culturing the microorganism Bacillus licheniformis B-0844 FERM P-6809, in a culture medium, and isolating the thus-produced NAD synthetase therefrom.

Assay method using NAD synthetase

An assay method for a component in a specimen containing any one of ATP, deamide-NAD and an amide donor which comprises performing a main reaction which comprises incubating the specimen with NAD synthetase in the presence of ATP, deamide-NAD, an amide donor and Mg.sup.++ to generate NAD; performing a coenzyme-cycling reeaction by combining the oxidation-reduction reaction system with coenzyme NAD and the oxidation-reduction reaction system with coenzyme reduced NAD, and measuring a consumed or generated component in the cycling reaction. The NAD synthetase can be produced by culturing the microorganism Bacillus licheniformis B-0844 FERM P-6809, in a culture medium, and isolating the thus-produced NAD synthetase therefrom.

Assay method using nad synthetase and a process for production of the

An enzyme which acts on a reducing terminal of a monosaccharide or oligosaccharide without requiring NAD or NADP and which catalyzes the reaction ##STR1## wherein R is a saccharide chain residue or hydrogen, A is a hydrogen acceptor other than NAD or NADP, AH or AHn is a reduced form acceptor and n is 1 or 2. This maltose dehydrogenase is produced by culturing a microorganism belonging to genus Staphylococcus, specifically, sp. B-0875 FERM BP-385, and isolating the thus-produced maltose dehydrogenase from the culture medium. An assay method for the determination of saccharide or the activity of a saccharide liberating enzyme, comprises reacting this enzyme with a substrate in the presence of a hydrogen acceptor, and measuring the amount of a detectable change.

Novel maltose dehydrogenase, process for its production, and analytical

L-glutamic acid oxidase having the following biochemical properties:

L-glutamic acid oxidase (H.sub.2 O.sub.2 -generating), its production

An enzyme having enoyl-CoA hydratase activity, 3-hydroxyacyl-CoA dehydrogenase activity and 3-ketoacyl-CoA thiolase activity, all in the same enzyme, is produced by culturing the microorganism strain Pseudomonas fragi B-0771 FERM-P No. 5701, and isolating the enzyme thus produced from the culture medium. Such an enzyme is useful in an assay method for a fatty acid component in a sample, which fatty acid is originally present in the sample or is liberated from a fatty acid ester in the sample, comprising:

Assay method for lipid component, assay composition, and process for

An assay method for amylase activity in a biological specimen such as serum, saliva or urine. The enzyme amylase in the specimen is used to decompose a substrate which is a glucose polymer having a modified reducing terminal glucose residue or a cyclic glucose polymer. A component of the decomposed substrate is measured as an indication of amylase activity in the specimen. The residue may be amylose, amylopectin, starch, starch hydrolyzate, an etherified reducing terminal, an esterified reducing terminal, gluconolactone or a gluconic acid residue or its derivative. Decomposed substrate assay may be effected by contacting the same with maltose dehydrogenase and NAD or NADP, whereupon the assay is performed by measuring the amount of reduced NAD or reduced NADP, by reacting the same with reduced-form hydrogen transport colorimetric reaction reagent. This reagent may be a tetrazolium salt and diaphorase, or tetrazolium salt and phenazinemethosulfate. To remove pre-existing glucose and maltose present in the specimen, the specimen may be pretreated with alpha-glucosidase or kinase in the presence of Mg.sup.++ and ATP, the kinase being for example hexokinase. The preferred maltose dehydrogenase is produced by culturing Bacillus megaterium B-0779 FERM-P No. 5662.

Assay method for amylase activity and method of producing maltose

L-amino acid oxidase is obtained by culturing a microorganism which can produce L-amino acid oxidase and belongs to the genus Colletotrichum, e.g. Colletotrichum sp. M5073, deposition number FERMP 5441, in a culturing medium and recovering the oxidase from cells of the microorganism.

Process for producing L-amino acid oxidase

A process for the production of acyl-Coenzyme A oxidase, comprises culturing an acyl-Coenzyme A-oxidase-producing microorganism belonging to genus Macrophomina, genus Cladosporium, genus Aspergillus, genus Monascus, genus Saccharomyces or genus Arthrobacter in a nutrient medium, and isolating the thus-formed acyl-CoA oxidase therefrom. The preferred species of microorganism are Macrophomina phaseoli ATCC 14383, Cladosporium resinae IFO 6367, Aspergillus candidus M-4815 FERM-P No. 5226, Monascus sp. M-4800 FERM-P No. 5225, Saccharomyces cerevisiae Y 0036 FERM-P No. 5174, and Arthrobacter sp. B-720 FERM-P No. 5224, respectively.

Process for the production of acyl-Coenzyme A oxidase

Ascorbate oxidase is produced by extraction from plants of the genus Sechium, particularly the species thereof which is Sechium edule Sw. The crushed plant tissues are extracted with an aqueous alkaline solvent, preferably at about pH 11. The extract is then subjected to centrifugation, concentration under vacuum, salting out with ammonium sulfate and solvent fractionation with acetone, to prepare crude ascorbate oxidase, which is then further purified by dialysis, ion exchange chromatography, adsorption chromatography and gel filtration. The ascorbate oxidase thus obtained has an optimum pH of about 7, a km value of 0.3 mM, an isoelectric point of around pH 6.3 and a molecular weight of about 100,000.

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Shizuoka, Japan

Hidehiko Ishikawa