US4097338A
Expired / Lapsed
US
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Fluorimetric demonstration and determination of a reduced coenzyme or derivative in an aqueous system
Assignee: Akzona Inc
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Filing: 1976-01-21
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Granted: 1978-06-27
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Expires: 1996-01-21
Abstract
Abstract The present invention relates to a rapid and sensitive method for the determination of a reduced coenzyme, particularly NADH, NADPH and derivatives thereof, in that the fluorescence of such a reduced coenzyme in an aqueous medium is measured in the presence simultaneously of an organic liquid miscible with water and a dipersion of one or more slightly soluble or insoluble substances.
Inventors
- 👤 Aarne Ilmari Konttinen
- 👤 Bastiaan Cornelis Goverde
- 👤 Peter Silvester Lambertus Janssen
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-24 01:06
CPC Classifications
C; C12; C12Q; C12Q1/00; C12Q1/008; G; G01; G01N; G01N21/00; G01N21/62; G01N21/63; G01N21/64; G01N21/6428; Y; Y10; Y10T; Y10T436/00; Y10T436/10; Y10T436/107497; Y10T436/108331; Y10T436/14; Y10T436/145555
Claims Analysis
Total Claims: 8
Claim 1
Independent Claim
144 words
A method for the determination of a reduced co-enzyme selected from the group consisting of NADH and NADPH in an aqueous system by fluorometric means, comprising the steps of preparing an aqueous sample containing said reduced co-enzyme wherein said aqueous sample contains as fluorescence intensifiers therefor (a) a water-miscible organic liquid or a mixture of such liquids, and (b) a dispersion of a water-insoluble material added thereto, said water-insoluble material selected from the group consisting of a metal carbonate, a metal sulphate, a metal phosphate, a metal silicate, a metal oxide, and a protein material obtained by precipitation of a protein in situ by the addition of another protein or a polypeptide to said aqueous sample for the precipitation of said water-insoluble material where no protein, or only enzyme protein, is present, and measuring specifically the fluorescence of said reduced co-enzyme in said sample.
Claim 2
Independent Claim
144 words
A method for the determination of a reduced co-enzyme selected from the group consisting of NADH and NADPH in an aqueous system by fluorometric means, comprising the steps of preparing an aqueous sample containing said reduced co-enzyme wherein said aqueous sample contains as fluorescence intensifiers therefor (a) a water-miscible organic liquid or a mixture of such liquids, and (b) a dispersion of a water-insoluble material added thereto, said water-insoluble material selected from the group consisting of a metal carbonate, a metal sulphate, a metal phosphate, a metal silicate, a metal oxide, and a protein material obtained by precipitation of a protein in situ by the addition of another protein or a polypeptide to said aqueous sample for the precipitation of said water-insoluble material where no protein, or only enzyme protein, is present, and measuring specifically the fluorescence of said reduced co-enzyme in said sample.
Claim 3
Dependent Claim
59 words
Method according to claim 1, in which said reduced co-enzyme is the reaction product of an enzymatic reaction and in which said water-miscible organic liquid is selected from the group consisting of methanol, ethanol, 2-propanol, ethylene glycol, propylene glycol, 2-methoxyethanol, 2-ethoxyethanol, 2(ethoxyethoxy)ethanol, glycerol, acetone, methyl-ethyl-ketone, dimethyl sulfoxide, formamide, dimethylformamide and pyridine, and a mixture of two or more thereof.
Claim 4
Dependent Claim
48 words
Method according to claim 2 in which said enzymatic reaction is preceded by at least one enzymatic or non-enzymatic reaction and in which the fluorimetrically determined quantity of reduced co-enzyme is a measure of the amount of reaction component to be determined in one of said participating reactions.
Claim 5
Dependent Claim
55 words
Method according to claim 2 in which said enzymatic reaction is preceded by a first reaction relating to creatine (phospho) kinase, a second reaction which is the ancillary reaction relating to a hexokinase, and in which the third reaction which is the indicator reaction in which the reduced co-enzyme is formed relating to glucose-6-phosphate dehydrogenase.
Claim 6
Dependent Claim
45 words
Method according to claim 2 in which said enzymatic reaction is preceded by a first reaction relating to creatine (phospho) kinase, the ancillary reaction relates to pyruvate kinase, and the indicator reaction in which the amount of reduced co-enzyme present decreases, relates to lactate dehydrogenase.
Claim 7
Dependent Claim
25 words
Method according to claim 1 in which said aqueous sample contains, as said fluorescence intensifiers, a suspension of an insoluble zinc salt in ethylene glycol.
Claim 8
Dependent Claim
34 words
Method according to claim 1 in which said water-insoluble material is selected from the group consisting of zinc carbonate, magnesium carbonate, calcium phosphate, zinc phosphate, calcuim sulphate, aluminum oxide, silica and powdered natural silicates.
References Cited (Backward Citations)
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1981-06-26 | 1981-12-26 | 1982-06-27 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1985-06-26 | 1985-12-26 | 1986-06-27 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1989-06-26 | 1989-12-26 | 1990-06-27 | $7700.00 | Window Closed / Lapsed |