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US4224034A
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Assay of iron and iron binding protein reagents and methods
Assignee: INDIANA DEVELOPMENT FINANCE AUTHORITY
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Filing: 1978-09-07
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Granted: 1980-09-23
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Expires: 1998-09-07
Abstract
Abstract The discovery of 9-(2-pyridyl)-acenaphtho[1,2-e]-as-triazine and its sulfonated form, having particular advantages when used as both a chelator and indicator of ferrous ions in organic or aqueous mediums; a process for the synthesis of those compounds in which hydrazine is used as both a reactant and as a solvent in production of intermediate compounds, eliminating the need of ethanol and the disadvantageous ether-extraction step; the use of N,N-dimethylformamide as a reaction solvent in the production of triazines; the use of the new triazine compounds to measure iron, transferrin, and UIBC/TIBC in biological systems; the use of dimethylsulfoxide and certain processes to eliminate the interferences commonly encountered in these assays; and the use of the triazine compounds to measure both total and/or reduced iron in a variety of matrices.
Inventors
- 👤 Jerry W. Denney
- 👤 Mark C. Outcalt
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-11-02 15:48
CPC Classifications
G; G01; G01N; G01N33/00; G01N33/48; G01N33/50; G01N33/84; G01N33/90; Y; Y10; Y10S; Y10S436/00; Y10S436/91
Claims Analysis
Total Claims: 28
Claim 1
Independent Claim
30 words
A process for detecting the presence of ferrous ions, which includes the step of adding, to the specimen being assayed, 9-(2-pyridyl)-acenaphtho[1,2-e]-as-triazine as both a chelator and indicator of ferrous ion.
Claim 2
Dependent Claim
30 words
A process for detecting the presence of ferrous ions, which includes the step of adding, to the specimen being assayed, 9-(2-pyridyl)-acenaphtho[1,2-e]-as-triazine as both a chelator and indicator of ferrous ion.
Claim 3
Dependent Claim
24 words
In the quantitative determination of ferrous ion in biological samples, the step of adding 9-(2-pyridyl)-acenaphtho[1,2-e]-as-triazine, as both a chelator and indicator of ferrous ion.
Claim 4
Dependent Claim
24 words
In the quantitative determination of ferrous ion in biological samples, the step of adding 9-(2-pyridyl)-acenaphtho[1,2-e]-as-triazine, as both a chelator and indicator of ferrous ion.
Claim 5
Dependent Claim
32 words
In the process set forth in either of claims 1 or 2, the step of adding, to the specimen being assayed, sulfonated 9-(2-pyridyl)-acenaphtho[1,2-e]-as-triazine as both a chelator and indicator of ferrous ion.
Claim 6
Dependent Claim
26 words
The process as set forth in claim 3, in which the concentration of sulfonated 9-(2-pyridyl)-acenaphtho[1,2-e]-as-triazine in the final reaction mixture is at least about 0.02% (W/V).
Claim 7
Dependent Claim
35 words
The process as set forth in claim 3, in which the final concentration of sulfonated 9-(2-pyridyl)-acenaphtho[1,2-e]-as-triazine in the final reaction mixture is at least three times the molar quantity of iron introduced by the sample.
Claim 8
Dependent Claim
26 words
The process as set forth in claim 3, in which dimethylsulfoxide is added to the reaction mixture to increase the liberation of iron bound to transferrin.
Claim 9
Dependent Claim
24 words
The process as set forth in claim 6, in which the concentration of dimethylsulfoxide in the final reaction mixture is at least 7.0% (V/V).
Claim 10
Dependent Claim
34 words
A process for the quantitative determination of unsaturated transferrin in biological samples, which includes the step of adding, to the specimen being assayed, sulfonated 9-(2-pyridyl)-acenaphtho[1,2-e]-as-triazine as a chelator and indicator of ferrous ion content.
Claim 11
Dependent Claim
34 words
A process for the quantitative determination of unsaturated transferrin in biological samples, which includes the step of adding, to the specimen being assayed, sulfonated 9-(2-pyridyl)-acenaphtho[1,2-e]-as-triazine as a chelator and indicator of ferrous ion content.
Claim 12
Dependent Claim
22 words
The process as set forth in claim 8, in which the final concentration of sulfonated 9-(2-pyridyl)-acenaphtho[1,2-e]-as-triazine is at least about 0.02% (W/V).
Claim 13
Dependent Claim
35 words
The process as set forth in claim 8, in which the final concentration of sulfonated 9-(2-pyridyl)-acenaphtho[1,2-e]-as-triazine in the final reaction mixture is at least three times the molar quantity of iron introduced by the sample.
Claim 14
Dependent Claim
34 words
A process for the quantitative determination of total transferrin in biological samples, which includes the step of adding, to the specimen being assayed, sulfonated 9-(2-pyridyl)-acenaphtho[1,2-e]-as-triazine as a chelator and indicator of ferrous ion content.
Claim 15
Dependent Claim
34 words
A process for the quantitative determination of total transferrin in biological samples, which includes the step of adding, to the specimen being assayed, sulfonated 9-(2-pyridyl)-acenaphtho[1,2-e]-as-triazine as a chelator and indicator of ferrous ion content.
Claim 16
Dependent Claim
37 words
A process for the quantitative or qualitative determination of ferrous ions in an organic or aqueous matrix, which includes the step of adding, to the specimen being assayed, 9-(2-pyridyl)-acenaphtho[1,2-e]-as-triazine as a chelator and indicator of said ions.
Claim 17
Dependent Claim
37 words
A process for the quantitative or qualitative determination of ferrous ions in an organic or aqueous matrix, which includes the step of adding, to the specimen being assayed, 9-(2-pyridyl)-acenaphtho[1,2-e]-as-triazine as a chelator and indicator of said ions.
Claim 18
Dependent Claim
30 words
The process as set forth in claim 12, which includes the step of adding, to the specimen being assayed, sulfonated 9-(2-pyridyl)-acenaphtho[1,2-e]-as-triazine as a chelator and indicator of ferrous ion content.
Claim 19
Dependent Claim
33 words
In a process for the direct serum quantitation of ferrous ion present in serum, the step of adding, to the specimen being assayed, dimethylsulfoxide to increase the liberation of iron bound to transferrin.
Claim 20
Dependent Claim
33 words
In a process for the direct serum quantitation of ferrous ion present in serum, the step of adding, to the specimen being assayed, dimethylsulfoxide to increase the liberation of iron bound to transferrin.
Claim 21
Dependent Claim
45 words
In the process of quantitatively determining the amount of iron in a serum sample, the procedural step of maintaining the incubation of the sample in the buffer system employed therewith for an extended period sufficient to minimize interferences encountered in the use of hemolyzed samples.
Claim 22
Dependent Claim
45 words
In the process of quantitatively determining the amount of iron in a serum sample, the procedural step of maintaining the incubation of the sample in the buffer system employed therewith for an extended period sufficient to minimize interferences encountered in the use of hemolyzed samples.
Claim 23
Dependent Claim
19 words
The process as set forth in claim 15, in which the extended period is at least about 5 minutes.
Claim 24
Dependent Claim
59 words
In a process for the determination of the oxidative state of iron ion present in a solution and/or the determination of the ratio of the oxidized to reduced form thereof, the step of adding, to the specimen being assayed, 9-(2-pyridyl)-acenaphtho[1,2-e]-as-triazine or its sulfonated form as an indicator of the oxidation state of the iron ions present in the solution.
Claim 25
Dependent Claim
59 words
In a process for the determination of the oxidative state of iron ion present in a solution and/or the determination of the ratio of the oxidized to reduced form thereof, the step of adding, to the specimen being assayed, 9-(2-pyridyl)-acenaphtho[1,2-e]-as-triazine or its sulfonated form as an indicator of the oxidation state of the iron ions present in the solution.
Claim 26
Dependent Claim
184 words
A process according to either of claims 1 or 2, including the method for automatically correcting for variations of iron in the reagents or apparatus used in a determination of unsaturated iron binding capacity of a serum specimen, including the steps of: (a) Saturating a serum specimen containing an unknown amount of transferrin with a reagent containing ferrous ions in an excess amount relative to the absolute binding ability of the transferrin present in the serum specimen; (b) Determining the amount of iron not bound by transferrin in Step "a" by reaction with a reagent containing an iron indicator compound; (c) Substituting a sample of iron free water for the specimen in Step "a" above, and determining the total iron added by the reagents by use of the same reagents used in Steps "a" and "b" above; and (d) subtracting the amount of iron remaining in solution after reaction with a serum sample, as obtained in Step "b," from the amount of iron added, as measured in Step "c" above, giving a direct measure of saturated iron binding capacity of the original serum specimen.
Claim 27
Dependent Claim
27 words
The process as set forth in either of claims 14 or 15, including the step of adding 9-(2-pyridyl)-acenaphtho[1,2-e]-as-triazine as both a chelator and indicator of ferrous ion.
Claim 28
Dependent Claim
28 words
The process as set forth in either of claims 14 or 15, including the step of adding sulfonated 9-(2-pyridyl)-acenaphtho[1,2-e]-as-triazine as both a chelator and indicator of ferrous ion.
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) | 1983-09-23 | 1984-03-24 | 1984-09-23 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1987-09-23 | 1988-03-24 | 1988-09-23 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1991-09-23 | 1992-03-24 | 1992-09-23 | $7700.00 | Window Closed / Lapsed |