US3947352A
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Polysaccharide matrices for use as adsorbents in affinity chromatography techniques
Assignee: Individual
•
Filing: 1974-05-31
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Granted: 1976-03-30
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Expires: 1994-05-31
Abstract
Abstract Polysaccharide matrices, e.g., agarose, for use in certain affinity chromatography procedures, are activated with sodium metaperiodate, derivatized with a symmetrical dihydrazide and thence reductively stabilized, preferably with sodium cyanoborohydride. Moreover, intermediate Schiff bases formed from the NaIO 4 -oxidized polysaccharide are selectively, reductively stabilized with said sodium cyanoborohydride.
Inventors
- 👤 Pedro Cuatrecasas
- 👤 Indu Parikh
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-23 14:14
CPC Classifications
C; C08; C08B; C08B31/00; C08B31/18; C08B31/185; B; B01; B01J; B01J20/00; B01J20/22; B01J20/281; B01J20/286; B01J20/30; B01J20/32; B01J20/3202; B01J20/3206; B01J20/3208; B01J20/3212; B01J20/3214; B01J20/3217; B01J20/3219; B01J20/3231; B01J20/3242; B01J20/3268; B01J20/3272; B01J20/3274; C08B15/00; C08B15/05; C08B15/06; C08B37/00; C08B37/0006; C08B37/0009; C08B37/0021; C08B37/0036; C08B37/0039; C08H; C08H1/00; C12; C12N; C12N11/00; C12N11/02; C12N11/10; B01J2220/00; B01J2220/50; B01J2220/54; Y; Y10; Y10S; Y10S435/00; Y10S435/814
Claims Analysis
Total Claims: 21
Claim 1
Independent Claim
76 words
In the method for the activation of a polysaccharide matrix for use as an adsorbent in affinity chromatography techniques, comprising activating the said polysaccharide by oxidizing the cis-vicinal hydroxyl groups thereof with sodium metaperiodate to generate aldehyde functions, converting said aldehydic functions into Schiff bases and thence reductively stabilizing the said resultant Schiff bases, the improvement which comprises the reductive stabilization of said intermediate Schiff bases with sodium cyanoborohydride at a pH between 6 and 6.5.
Claim 2
Dependent Claim
76 words
In the method for the activation of a polysaccharide matrix for use as an adsorbent in affinity chromatography techniques, comprising activating the said polysaccharide by oxidizing the cis-vicinal hydroxyl groups thereof with sodium metaperiodate to generate aldehyde functions, converting said aldehydic functions into Schiff bases and thence reductively stabilizing the said resultant Schiff bases, the improvement which comprises the reductive stabilization of said intermediate Schiff bases with sodium cyanoborohydride at a pH between 6 and 6.5.
Claim 3
Dependent Claim
36 words
In the method as defined by claim 1, the improvement further comprising the preferential, selective reductive stabilization of the said intermediate Schiff bases in the absence of a reduction of the aldehyde functions of the polysaccharide.
Claim 4
Dependent Claim
13 words
In the method as defined by claim 2 wherein the polysaccharide is agarose.
Claim 5
Independent Claim
78 words
The method for the preparation of a hydrazide-polysaccharide derivative for use as an adsorbent in affinity chromatography techniques, which comprises activating a polysaccharide matrix by oxidizing the cis-vicinal hydroxyl groups thereof with sodium metaperiodate to generate aldehyde functions, reacting said periodate-oxidized polysaccharide with a bifunctional, symmetrical dihydrizide to convert the same to the corresponding hydrazido/hydrazone derivative, and thence reductively stabilizing said hydrazido/hydrazone derivative with sodium cyanoborohydride at a pH between 6 and 6.5 to produce an unsemetric hydrazide.
Claim 6
Independent Claim
78 words
The method for the preparation of a hydrazide-polysaccharide derivative for use as an adsorbent in affinity chromatography techniques, which comprises activating a polysaccharide matrix by oxidizing the cis-vicinal hydroxyl groups thereof with sodium metaperiodate to generate aldehyde functions, reacting said periodate-oxidized polysaccharide with a bifunctional, symmetrical dihydrizide to convert the same to the corresponding hydrazido/hydrazone derivative, and thence reductively stabilizing said hydrazido/hydrazone derivative with sodium cyanoborohydride at a pH between 6 and 6.5 to produce an unsemetric hydrazide.
Claim 7
Dependent Claim
15 words
The method as defined by claim 4, wherein the bifunctional, symmetrical dihydrazide is succinic dihydrazide.
Claim 8
Dependent Claim
16 words
The method as defined by claim 5, wherein the resultant unsymmetric hydrazide bears unreacted hydrazido moieties.
Claim 9
Dependent Claim
51 words
In a method for the affinity chromatographic technique of coupling proteins or ligands, bearing carbonyl or amino moieties, to a polysaccharide matrix, the improvement which comprises derivatizing the matrix as defined in claim 6, coupling a said protein or ligand to the polyhydrazide-matrix and thence reductively stabilizing said protein- or ligand-hydrazido-polysaccharide.
Claim 10
Dependent Claim
20 words
The method as defined by claim 7, wherein the reductive stabilization of the coupled complex is effected with sodium cyanoborohydride.
Claim 11
Dependent Claim
24 words
The method as defined by claim 7, wherein a spacer molecule is interposed between the derivatized matrix and the protein or ligand coupled thereto.
Claim 12
Dependent Claim
23 words
The method as defined by claim 7, wherein the polysaccharide is selected from the group consisting of cellulose, starch, cross-linked dextran and agarose.
Claim 13
Dependent Claim
12 words
The method as defined by claim 10, wherein the polysaccharide is agarose.
Claim 14
Dependent Claim
31 words
The method as defined by claim 7, wherein the molecule coupled to the polysaccharide matrix is selected from the group consisting of glycolipid, ganglioside, glycoprotein, glycopeptide, oligosaccharide, brain ganglioside and thyroglobin.
Claim 15
Dependent Claim
10 words
The product of the method as defined by claim 1.
Claim 16
Dependent Claim
10 words
The product of the method as defined by claim 4.
Claim 17
Dependent Claim
10 words
The product of the method as defined by claim 7.
Claim 18
Dependent Claim
10 words
The product of the method as defined by claim 9.
Claim 19
Dependent Claim
21 words
The method as defined by claim 7, wherein the protein or ligand to be coupled is preliminarily oxidized with sodium metaperiodate.
Claim 20
Dependent Claim
29 words
In an affinity chromatography column comprising an insoluble, solid polysaccharide adsorbent, the improvement which comprises, as the said adsorbent or matrix thereof, the product as defined by claim 14.
Claim 21
Dependent Claim
27 words
In the method as defined by claim 2, the improvement where in the polysaccharide is selected from the group consisting of cellulose, starch, cross-linked dextran and agarose.
References Cited (Backward Citations)
US3850798A
Cited
US3850798B1
Cited
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1979-03-30 | 1979-09-29 | 1980-03-30 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1983-03-30 | 1983-09-29 | 1984-03-30 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1987-03-30 | 1987-09-29 | 1988-03-30 | $7700.00 | Window Closed / Lapsed |