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US4473494A
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Preparation of stroma-free, non-heme protein-free hemoglobin
Assignee: United States Department of the Army
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Filing: 1983-05-04
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Granted: 1984-09-25
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Expires: 2003-05-04
Abstract
Abstract A method for the production of stroma-free, non-heme protein-free hemoglobin by use of zinc ion to promote precipitation of a zinc ion-bound insoluble hemoglobin complex, followed by membrane ultrafiltration of the zinc-hemoglobin complex from the filtrate fluid medium, preferably with a saline dialyzate.
Inventors
- 👤 Ross W. Tye
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-29 11:41
CPC Classifications
C; C07; C07K; C07K14/00; C07K14/795; C07K14/805; A; A61; A61K; A61K38/00; Y; Y10; Y10S; Y10S530/00; Y10S530/827; Y10S530/829; Y10S530/83
Claims Analysis
Total Claims: 12
Claim 1
Independent Claim
74 words
A method for the production of stroma-free, non-heme protein-free hemoglobin comprising promoting precipitation of a zinc ion-bound insoluble hemoglobin complex by allowing a solution of stroma-free hemoglobin to complex with an at least tenfold molar excess of soluble zinc salt, then ultrafiltering said zinc ion-bound hemoglobin complex precipitate from the filtrate fluid containing the non-heme protein by use of a membrane having a pore size in the range of about 0.22μ to about 0.5μ.
Claim 2
Independent Claim
74 words
A method for the production of stroma-free, non-heme protein-free hemoglobin comprising promoting precipitation of a zinc ion-bound insoluble hemoglobin complex by allowing a solution of stroma-free hemoglobin to complex with an at least tenfold molar excess of soluble zinc salt, then ultrafiltering said zinc ion-bound hemoglobin complex precipitate from the filtrate fluid containing the non-heme protein by use of a membrane having a pore size in the range of about 0.22μ to about 0.5μ.
Claim 3
Dependent Claim
17 words
The method according to claim 1 wherein the stroma-free hemoglobin is produced by crystallization, centrifugation, or ultrafiltration.
Claim 4
Dependent Claim
21 words
The method according to claim 1 wherein the stroma-free hemoglobin is derived from a human, or is bovine, ovine or porcine.
Claim 5
Dependent Claim
40 words
The method according to claim 1 wherein the solution of stroma-free hemoglobin is diluted to less than 0.5 g% prior to precipitation by zinc ion and said zinc ion-bound hemoglobin complex precipitate is dialyzed with normal saline by said ultrafiltration.
Claim 6
Dependent Claim
25 words
The method according to claim 1 wherein the solution of stroma-free hemoglobin is adjusted to a salinity in the range of 50 to 250 mM.
Claim 7
Dependent Claim
20 words
The method according to claim 1 wherein the stroma-free hemoglobin has a pH in its range of 7 to 9.
Claim 8
Dependent Claim
12 words
The method according to claim 6 wherein the pH is about 7.5.
Claim 9
Dependent Claim
39 words
The method for the production of stroma-free, non-heme protein-free hemoglobin according to claim 1 further comprising treating the ultrafiltered zinc-ion bound hemoglobin complex precipitate with a solubilizing solution quantity of EDTA, and refiltering the released hemoglobin through said membrane.
Claim 10
Independent Claim
133 words
A method for the production of stroma-free, non-heme protein-free hemoglobin comprising promoting precipitation of a zinc ion-bound insoluble hemoglobin complex by allowing less than 0.52% solution of ultrafiltered stroma-free hemoglobin derived from a human to complex with an at least tenfold molar excess of soluble zinc salt, said stroma-free hemoglobin solution having a salinity in the range of 50 to 250 mM and a pH in the range of 7 to 9, ultrafiltering with a saline dialyzate said zinc-ion bound hemoglobin complex precipitate from the filtrate fluid containing the non-heme protein by use of a membrane having a pore size in the range of about 0.22μ to about 0.5μ, treating the ultrafiltered zinc-ion bound hemoglobin complex precipitate with a solubilizing solution quantity of ethylenediaminetetraacetic acid, and refiltering the released hemoglobin through said membrane.
Claim 11
Independent Claim
133 words
A method for the production of stroma-free, non-heme protein-free hemoglobin comprising promoting precipitation of a zinc ion-bound insoluble hemoglobin complex by allowing less than 0.52% solution of ultrafiltered stroma-free hemoglobin derived from a human to complex with an at least tenfold molar excess of soluble zinc salt, said stroma-free hemoglobin solution having a salinity in the range of 50 to 250 mM and a pH in the range of 7 to 9, ultrafiltering with a saline dialyzate said zinc-ion bound hemoglobin complex precipitate from the filtrate fluid containing the non-heme protein by use of a membrane having a pore size in the range of about 0.22μ to about 0.5μ, treating the ultrafiltered zinc-ion bound hemoglobin complex precipitate with a solubilizing solution quantity of ethylenediaminetetraacetic acid, and refiltering the released hemoglobin through said membrane.
Claim 12
Dependent Claim
17 words
A method according to claim 9 wherein the pH of the strome-free hemoglobin solution is about 7.5.
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) | 1987-09-25 | 1988-03-26 | 1988-09-25 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1991-09-25 | 1992-03-26 | 1992-09-25 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1995-09-25 | 1996-03-26 | 1996-09-25 | $7700.00 | Window Closed / Lapsed |