Enzymatic ethanol assay

• US 3,926,736 A
• Filed: 08/30/1974
• Issued: 12/16/1975
• Est. Priority Date: 08/30/1974
• Status: Expired due to Term

• Current Assignee: American Hoechst Corporation (Sanofi )
• Original Assignee: Giovanni Bucolo
• Inventors: Bucolo, Giovanni
• Primary Examiner(s): Monacell, A. Louis
• Assistant Examiner(s): Fan, C. A.
• Application Number:
  US05/502,210
• Time in Patent Office: 473 Days
• Field of Search: 195/13.5R 23/23B
• US Class Current: 435/26
• CPC Class Codes: C12Q 1/32 involving dehydrogenase

Abstract

This invention provides an improved enzymatic assay for ethanol in body fluids, whereby a single substance acts simultaneously as a buffer and as a trapping agent for one of the products of the enzymatic reaction.

Claims

1. A COMPOSITION OF MATTER AS A REAGENT IN DETERMINING THE ETHANOL CONTENT OF BODY FLUIDS CONSISTING ESSENTIALLY OF:

A. THE ENZYME ALCOHOL DEHYDROGENASE;

B. THE COENZYME NICTINAMIDE ADENINE DINUCLEOTIDE;

C. A BUFFER/ACTEALDEHYDR-TRAPPING AGENT SELECTED FROM THE GROUP CONSISTING OF 2-AMINO-2-HYDROXYMETHYL-1,3PROPANEDIOL, 2-AMINO-2-METHYL-1,3-PROPANEDIOL AND MIXTURES THEREOF;

D. AM ACID IN QUANTITY SUFFICIENT TO LOWER THE PH OF AN AQUEOUS SOLUTION OF SAID MIXTURE TO WITHIN THE RANGE OF ABOUT 8.8 TO 9.2;

SAID MIXTURE BEING FREE OF ANY OTHER ACETALDEHYDE TRAPPING AGENT.

2. The composition of claim 1 wherein said composition is present in the form of two moieties, one containing said coenzyme and the other the balance of the recited components.

3. The composition of claim 1 which includes a heavy metal sequestering agent.

4. The composition of claim 3 in which said sequestering agent is ethylene diamine tetra-acetic acid, or a sodium salt thereof.

5. An assay solution useful in the determination of the ethanol content of body fluids consisting essentially of the composition of claim 1 dissolved in sufficient water to provide a concentration of said buffer/trapping agent of at least 0.3 molar.

6. A composition in accordance with claim 1 wherein, for each 6.99 millimoles of said buffer/trapping agent, 16 - 18 mg of NAD;

and from 200 - 220 I.U. of alcohol dehydrogenase are present.

7. An assay solution useful in the determination of the ethanol content of boDy fluids consisting essentially of the composition of claim 6 dissolved in sufficient water to provide a concentration of said buffer/trapping agent of approximately at least 0.3 molar.