METHOD FOR STABILIZING LEUCO-TYPE COLORANT

• US 20080295259A1
• Filed: 02/21/2008
• Published: 12/04/2008
• Est. Priority Date: 02/22/2007
• Status: Active Grant

• Current Assignee: Asahi Kasei Pharma Corporation (Asahi Kasei Corporation)
• Original Assignee: Asahi Kasei Pharma Corporation (Asahi Kasei Corporation)
• Inventors: KAN, Akihiko, UEDA, Shigeru, MATSUOKA, Takeshi
• Granted Patent: US 8,268,017 B2
• Time in Patent Office: Days
• US Class Current: 8/401
• CPC Class Codes: C09B 67/0077 Preparations with possibly reduced vat, sulfur or indigo dyes
• CPC Class Codes: C12Q 1/28 involving peroxidase

Abstract

Provided are a method of stabilizing a leuco dye storable as a liquid for a long time, a method of reducing nonspecific color development thereof at a time of a color development reaction, and a stable liquid reagent composition using the methods. The inventors of the present invention found that coexistence of the leuco dye with a specific reducing agent resulted in suppression of self color development and its remarkably improved stability in a solution, and that when a color development reaction of the leuco dye with hydrogen peroxide was performed, coexistence of the leuco dye, in a reaction solution, with another dye which had an absorption spectrum not influencing a measurement wavelength of the leuco dye and did not react with hydrogen peroxide suppressed nonspecific color development and lowered a reagent blank value, which was applied to an analytical reagent.

Claims

1. A method of stabilizing a leuco dye solution, wherein a leuco dye coexists with at least one of reducing agents selected from the group consisting of reductive thioalcohols and reductive sulfates.

2. A method of stabilizing a leuco dye solution according to claim 1, wherein the leuco dye comprises 10-(carboxymethylaminocarbonyl)-3,7-bis(dimethylamino) phenothiazine sodium or N,N,N′

,N′

,N″

,N″

-hexa(3-sulfopropyl)-4,4′

,4″

-triaminotriphenylmethane-6sodium.

3. A method of stabilizing a leuco dye solution according to claim 1, wherein a concentration of the leuco dye in a solution is 0.01 to 2 mM.

4. A method of stabilizing a leuco dye solution according to claim 1, wherein a concentration of the reducing agent in a solution is 0.005 to 10 mM.

5. A method of stabilizing a leuco dye solution according to claim 1, wherein the reducing agent includes at least one of kinds selected from the group consisting of cysteine, cysteamine, N-acetylcysteine, thioglycerol, sodium thiosulfate, sodium sulfite, and sodium disulfite.

6. A liquid composition of a leuco dye, comprising:

at least one of reducing agents selected from the group consisting of 0.005 to 10 mM reductive thioalcohols and reductive sulfates; and

0.01 to 2 mM of 10-(carboxymethylaminocarbonyl)-3,7-bis(dimethylamino) phenothiazine sodium or N,N,N′

,N′

,N″

,N″

-hexa(3-sulfopropyl)-4,4′

,4″

-triaminotriphenylmethane-6 sodium.

7. A liquid composition according to claim 6, wherein the reducing agent is at least one of kinds selected from the group consisting of cysteine, cysteamine, N-acetylcysteine, thioglycerol, sodium thiosulfate, sodium sulfite, and sodium disulfite.

8. A method of reducing a nonspecific color development reaction of a leuco dye, wherein the leuco dye coexits with another dye in a color development reaction of hydrogen peroxide in the co-presence of peroxidase using the leuco dye, the another dye having an absorption spectrum not influencing on a measurement wavelength of a leuco dye and not reacting with hydrogen peroxide.

9. A method of reducing a nonspecific color development reaction of a leuco dye according to claim 8, wherein a maximum absorption wavelength of the another dye which has an absorption spectrum not influencing on a measurement wavelength of a leuco dye and does not react with hydrogen peroxide is in a range of 400 nm to 550 nm.

10. A method of reducing a nonspecific color development reaction of a leuco dye according to claim 8, wherein the another dye which has an absorption spectrum not influencing on a measurement wavelength of a leuco dye and does not react with hydrogen peroxide comprises at least one of kinds selected from Orange G, Orange II, Food Red No. 2, Food red No. 3, Food Red No. 40, Food Red No. 102, Food Red No. 104, Food Red No. 106, Food Yellow No. 4, and Food Yellow No. 5.

11. A method of reducing a nonspecific color development reaction of a leuco dye according to claim 8, wherein the leuco dye comprises 10-(carboxymethylaminocarbonyl)-3,7-bis(dimethylamino) phenothiazine sodium or N,N,N′

,N′

,N″

,N″

-hexa (3-sulfopropyl)-4,4′

,4″

-triaminotriphenylmethane-6 sodium.

12. A method of quantifying hydrogen peroxide, comprising:

reacting a leuco dye with hydrogen peroxide in a state where the leuco dye coexists with at least one of reducing agents selected from the group consisting of reductive thioalcohols and reductive sulfates and another dye which has an absorption spectrum not influencing on a measurement wavelength of the leuco dye and does not react with hydrogen peroxide.

13. A reagent for quantifying hydrogen peroxide, wherein the reagent is formed of a combination of a reagent containing another dye which has an absorption spectrum not influencing on a measurement wavelength of a leuco dye and does not react with hydrogen peroxide and peroxidase and another reagent containing a leuco dye and at least one of reducing agents selected from the group consisting of reductive thioalcohols and reductive sulfates.

14. A liquid and stable reagent for measuring a glycated protein using a reagent for quantifying hydrogen peroxide according to claim 13.

15. A reagent for measuring a glycated protein according to claim 14, wherein the reagent is formed of a combination of:

a reagent containing, as a leuco dye, 10-(carboxymethylaminocarbonyl)-3,7-bis (dimethylamino) phenothiazine sodium or N,N,N′

,N′

,N″

,N″

-hexa(3-sulfopropyl)-4,4′

,4″

-triaminotriphenylmethane-6 sodium, at least one of reducing agents selected from the group consisting of reductive thioalcohols and reductive sulfates, and protease which acts on a glycated protein; and

another reagent containing peroxidase, another dye which has an absorption spectrum not influencing on a measurement wavelength of a leuco dye and does not react with hydrogen peroxide, and ketoamine oxidase.

16. A reagent for measuring a glycated protein according to claim 14, wherein the glycated protein comprises hemoglobin A1c, and a total hemoglobin is quantified with the first reagent, and a hemoglobin A1 c is quantified with the second reagent, whereby a ratio of an amount of the hemoglobin A1 c to the total hemoglobin amount is determined.

17. A reagent for measuring a glycated protein according to claim 16, comprising:

a first reagent containing a protease reaction accelerator, ketoamine oxidase which acts on 1-deoxyfructosyl-L-valyl-L-histidine, peroxidase, a trinder reagent, ascorbate oxidase, and another dye which has an absorption spectrum not influencing on a measurement wavelength of leuco dye and does not react with hydrogen peroxide; and

a second reagent containing protease which can cleave 1-deoxyfructosyl-L-valyl-L-histidine from a glycated β

chain terminal of the hemoglobin A1c, 10-(carboxymethylaminocarbonyl)-3,7-bis(dimethylamino)phenothiazine sodium, cyclodextrins, and at least one of reducing agents selected from the group consisting of reductive thioalcohols and reductive sulfates.

18. A reagent for measuring a glycated protein according to claim 16, comprising:

a first reagent containing a protease reaction accelerator, protease which acts on the hemoglobin A1c to cleave 1-deoxyfructosyl-L-valyl-L-histidine from a glycated β

0 chain terminal of the hemoglobin A1 c, 10-(carboxymethylaminocarbonyl)-3,7-bis(dimethylamino) phenothiazine sodium, catalase, cyclodextrins, and a reducing agent; and

a second reagent containing ketoamine oxidase, peroxidase, and a dye which has an absorption spectrum not influencing on a measurement wavelenth of a leuco dye and does not react with hydrogen peroxide.

19. A reagent for measuring a glycated protein according to claim 17, wherein:

the protease reaction accelerator comprises an anionic surfactant selected from N-acylamino acid (salt) and N-acyl taurine (salt);

the ketoamine oxidase is derived from Neocosmospora or Curvularia;

the dye which has an absorption spectrum not influencing on a measurement wavelength of a leuco dye and does not react with hydrogen peroxide comprises Yellow No. 4, Red No. 40, or Red No. 102;

the protease which acts on the hemoglobin A1c to cleave 1-deoxyfructosyl-L-valyl-L-histidine from a glycated p chain terminal of the hemoglobin A1c is derived from Bacillus or Rhizobacter;

the cyclodextrins comprise 2-hydroxypropyl-β

-cyclodextrin;

the trinder reagent comprises ALPS or ADPS;

the reducing agent comprises sodium sulfite; and

hemocyte is hemolyzed with water to be used as a specimen.

20. A reagent for measuring a glycated protein according to claim 17, wherein the reagent measures a glycated protein in whole blood, and the first reagent further contains ketoamine oxidase.