Rapid assaying method for guanase

• US 4,550,076 A
• Filed: 09/27/1982
• Issued: 10/29/1985
• Est. Priority Date: 09/25/1981
• Status: Expired due to Fees

• Current Assignee: Maruho Company Limited
• Original Assignee: Maruho Company Limited
• Inventors: Fujii, Setsuro, Muraoka, Toshiharu, Chikazawa, Nobumoto
• Primary Examiner(s): Kepplinger, Esther M.
• Application Number:
  US06/424,182
• Time in Patent Office: 1,128 Days
• Field of Search: 435/4, 435/18, 435/25, 435/26, 435/27, 435/28
• US Class Current: 435/18
• CPC Class Codes: C12Q 1/28 involving peroxidase
• CPC Class Codes: C12Q 1/34 involving hydrolase
• CPC Class Codes: G01N 2333/978 acting on carbon to nitrogen bonds other than peptide bonds (3.5)

Abstract

The guanase activity in body fluids such as blood serum can be rapidly and accurately assayed by (I) decomposing guanine with the guanase in the specimen to xanthine and ammonia at pH 7-9, preferably at pH 8, (II) decomposing the xanthine formed by former step I with xanthine oxidase to uric acid and hydrogen peroxide, (III) reacting the reactant solution of the former step II with 3-methyl-2-benzothiazolinonehydrazone, an aniline derivative such as N,N-di-lower-alkylaniline and peroxidase, and finally measuring the optical absorption of the reactant solution of the step III at 570-600 nm. The all steps can be completed within 15 minutes. Therefore, this assay is adoptable for automatic assay of guanase on usual clinically available automatic analyzers.

Claims

1. In the method of assaying guanase in the sample of the body fluid by adding guanine to said sample whereby said guanine is changed to xanthine, decomposing said xanthine to uric acid by xanthine oxidase whereby hydrogen peroxide is by-produced, reacting said hydrogen peroxide with 3-methyl-2-benzothiazolinonehydrazone and an aniline derivative in the presence of peroxidase whereby an indamine dye is produced, and finally measuring the optical absorption by said dye at 570-600 nm, the improvement for rapidly assaying the guanase comprising

(a) the reactions (I), (II), and (III) being separately and sequentially carried out;

(I) the xanthine-formation reaction,

(II) the hydrogen peroxide-formation reaction and

(III) the dye-producing reaction,

(b) the xanthine-formation-reaction being carried out within a pH in the range of 7-9, and (c) the hydrogen peroxide-formation reaction being carried out in the presence of a catalase inhibitor.

2. A rapid assaying method for guanase according to claim 1, wherein the aniline derivative is a member selected from the group consisting of N,N-di-loweralkyl anilines.

3. A rapid assaying method for guanase according to claim 1, wherein the catalase inhibitor is sodium azide.

4. A rapid assaying method for guanase according to claim 1, wherein the xanthine formation reaction is carried out within a range of pH 7-9.

5. A rapid assaying method for guanase according to any one of claim 1-4, wherein the assay is carried out by clinically acceptable automatic chemical analyser.

6. A rapid assaying method for guanase according to claim 1, wherein the xanthine formulation reaction is carried out at about pH 8.