Method for immobilizing enzymes and microbial cells

• US 4,195,129 A
• Filed: 12/29/1977
• Issued: 03/25/1980
• Est. Priority Date: 11/26/1975
• Status: Expired due to Term

• Current Assignee: Kansai Paint Company Limited
• Original Assignee: Kansai Paint Company Limited
• Inventors: Iida, Takamitsu, Yamamoto, Tsutomu, Fukui, Saburo
• Primary Examiner(s): Naff, David M.
• Application Number:
  US05/865,433
• Time in Patent Office: 817 Days
• Field of Search: 195/63, 195/68, 195/DIG. 11, 195/59, 204/159.16
• US Class Current: 435/182
• CPC Class Codes: C12N 11/04 entrapped within the carrier, e.g. gel or hollow fibres
• CPC Class Codes: C12N 11/087 Acrylic polymers
• CPC Class Codes: C12N 11/089 obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds
• CPC Class Codes: C12N 11/093 Polyurethanes
• CPC Class Codes: C12N 11/096 Polyesters; Polyamides
• CPC Class Codes: C12Q 1/00 Measuring or testing processes involving enzymes, nucleic acids or microorganisms measuring or testing apparatus with condition measuring or sensing means, e.g. colony counters, C12M1/34; Compositions therefor; Processes of preparing such compositions

Abstract

A method for immobilizing enzymes or microbial cells which comprises the steps of uniformly mixing an aqueous dispersion of enzymes or microbial cells with a photo-curable resin having a number average molecular weight of 800 to 100,000, two or more photopolymerizable ethylenically unsaturated groups per molecule and hydrophilic groups, and irradiating actinic rays to the above mixture of photo-curable resin and enzymes or microbial cells. The immobilized product is advantageously used in various enzymatic reactions with stable enzyme activity.

Claims

1. A method for immobilizing enzymes or microbial cells which is characterized in the steps of:

uniformly mixing aqueous dispersion of enzymes or microbial cells with a monomer-free photo-curable resin having a number average molecular weight of 800 to 100,000, two or more photo-polymerizable ethylenically unsaturated groups per molecule and nonionic hydrophilic groups; and

irradiating actinic rays having a wave length of 2500 to 6000 A to said mixture, whereby said enzymes or microbial cells are entrapped in said resin.

2. A method for immobilizing enzymes or microbial cells as claimed in claim 1 wherein said mixture additionally contains another monomer-free photo-curable resin having a number average molecular weight of 800 to 100,000, two or more photo-polymerizable ethylenically unsaturated groups per molecule and ionic hydrophilic groups, the ratio of the resin having ionic hydrophilic groups to the resin having nonionic hydrophilic groups being in the range of 1/99 to 50/50 by weight.

3. A method for immobilizing enzymes or microbial cells as claimed in claim 2, wherein said enzymes are urease, glucose oxidase, catalase, glucoamylase, glucose isomerase, invertase, acetoacetic decarboxylase, glucose oxidase-catalase, peroxidase, lactase, D-amino acid oxidase, α

-galactosidase, aminoacylase, aspartase or penicillin amidase, and said microbial cells are the cells of Lactobacillus bulgaricus, Aerobacter aerogenes, Bacillus subtilis, Azotobacter vinelandii or Proteus vulgaris.

4. A method for immobilizing enzymes or microbial cells as claimed in claim 2, wherein said photo-curable resin having ionic hydrophilic groups is at least one member selected from the group consisting of salts of high acid value unsaturated polyesters, high acid value unsaturated epoxides, anionic unsaturated acrylic resins, cationic unsaturated acrylic resins, unsaturated polyamines and unsaturated carboxylated cellulose.

5. A method for immobilizing enzymes or microbial cells as claimed in claim 2, wherein said photo-curable resin having nonionic hydrophilic groups is at least one member selected from the group consisting of:

polyesters made from polyethylene glycol and acrylic or methacrylic acid, urethanated adduct of the product made from polyisocyanate and polyethylene glycol with 2-hydroxyethyl acrylate or methacrylate, unsaturated cellulose, unsaturated polyvinyl alcohol and unsaturated polyamide.

6. A method for immobilizing enzymes or microbial cells as claimed in claim 2, wherein said ratio is in the range of 3/97 to 20/80 by weight.

7. A method for immobilizing enzymes or microbial cells as claimed in claim 2, wherein a photosensitizer is added to said mixture of photo-curable resins.

8. A method for immobilizing enzymes or microbial cells as claimed in claim 7, wherein said photosensitizer is at least one member selected from the group consisting of α

-carbonyl alcohol, acyloin ether, α

-substituted acyloin, naphtol, hydroxyanthracene, axoamide, uranyl nitrate, ferric chloride, mercaptan, disulfide, ascorbic acid and rivoflavin.

9. A method for immobilizing enzymes or microbial cells as claimed in claim 2, wherein said number average molecular weight is 1,000 to 70,000.

10. A method for immobilizing enzymes or microbial cells as claimed in claim 4, wherein said high acid value is 40 to 200.