Method and test composition for determination of hydrogen peroxide

• US 4,384,042 A
• Filed: 07/29/1981
• Issued: 05/17/1983
• Est. Priority Date: 07/30/1980
• Status: Expired due to Term

• Current Assignee: Kyowa Hakko Kogyo Company Limited (Kirin Holdings Company Limited)
• Original Assignee: Kyowa Hakko Kogyo Company Limited (Kirin Holdings Company Limited)
• Inventors: Shimizu, Yoshiaki, Miike, Akira, Tatano, Toshio, Watanabe, Katsuyuki
• Primary Examiner(s): Kepplinger, Esther M.
• Application Number:
  US06/288,123
• Time in Patent Office: 657 Days
• Field of Search: 435/10, 435/11, 435/14, 435/15, 435/18, 435/19, 435/25, 435/28, 435/805, 435/810
• US Class Current: 435/25
• CPC Class Codes: C12Q 1/28 involving peroxidase
• CPC Class Codes: C12Q 2326/00 Chromogens for determinations of oxidoreductase enzymes
• CPC Class Codes: G01N 33/52 Use of compounds or compositions for colorimetric, spectrophotometric or fluorometric investigation, e.g. use of reagent paper and including single- and multilayer analytical elements immunological elements G01N33/54386; involving labelled immunochemicals G01N33/58; for haemoglobin or occult blood G01N33/72
• CPC Class Codes: Y10S 435/805 Test papers
• CPC Class Codes: Y10S 435/81 Packaged device or kit

Abstract

Disclosed is a method for the colorimetric determination of hydrogen peroxide in a sample by reacting a particular chromogen with the hydrogen peroxide in the presence of peroxidase and measuring the absorbancy of the reaction solution in the visible ray region. Also disclosed is a test composition for carrying out the determination.

Claims

1. A method for the determination of hydrogen peroxide in a sample which comprises reacting the hydrogen peroxide with a chromogen represented by the formulae ##STR12## wherein Z represents hydroxyl, amino or substituted amino, Y represents oxygen atom or sulfur atom, R₁ represents hydrogen, alkyl, alkenyl, aryl, amino or mono-substituted amino, R₂ represents hydrogen, hydroxyl, alkyl, alkenyl, aryl, amino, alkyl-amino or alkoxy, R₃, R₄, R₅ and R₆ represent hydrogen, alkyl, alkenyl, acyl, aryl, halogen, sulfo, nitro, carboxyl, hydroxyl, hydroxyalkyl or alkoxy, R₃ and R₄ or R₅ and R₆ may form alkenylene, X represents --S--, --O--, ##STR13## R₇ and R₈ represent hydrogen, alkyl, alkenyl or aryl, in the presence of peroxidase and measuring the absorbancy of the reaction solution in the visible ray region.

2. A method according to claim 1, wherein Z is a dimethylamino group.

3. A method according to claim 1, wherein R₂ is a dimethylamino group.

4. A method according to claim 1, wherein both Z and R₂ are dimethylamino group, R₁ is a substituted amino group and R₃, R₄, R₅ and R₆ are hydrogen.

5. A method according to claim 1, wherein said hydrogen peroxide is a product formed by enzymatic reaction.

6. A method according to claim 5, wherein said enzymatic reaction is the oxidation of a substrate using oxidase.

7. A method according to claim 6, wherein said oxidase is selected from the group consisting of uricase, cholesterol oxidase, xanthin oxidase, choline oxidase, pyruvate oxidase, glycerin-3-phosphate oxidase, acyl Co A oxidase, glycerol oxidase, glucose oxidase and galactose oxidase.

8. A method according to claim 5, wherein said hydrogen peroxide-producing reaction and the reaction of hydrogen peroxide with said chromogen are conducted simultaneously.

9. A method according to claim 1, wherein said reaction is carried out in a buffer solution.

10. A test composition for the guantitative determination of hydrogen peroxide in a sample which comprises a sufficient amount of peroxidase and a chromogen defined in claim 1 to react with said hydrogen peroxide and form a pigment proportional to the amount of hydrogen peroxide in said sample.

11. A test composition according to claim 10 wherein said composition further a contains buffer reagent.

12. A test composition according to claim 10, wherein said composition further contains a member selected from the group consisting of a surfactant, antiseptics and ascorbate oxidase.

13. A test composition according to claim 10, which additionally comprises an enzymatic hydrogen peroxide-producing system in an amount sufficient to produce hydrogen peroxide proportional to the amount of a substrate in said sample.

14. A test composition according to claim 13, wherein said enzymatic hydrogen peroxide-producing system includes an oxidase for said substrate.