US4016042A
Expired / Lapsed
US
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Substrate for the quantitative determination of enzymes
Assignee: DSM Nutritional Products AG
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Filing: 1975-06-30
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Granted: 1977-04-05
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Expires: 1995-06-30
Abstract
Abstract A synthetic substrate having the structure R.sup.1 -- Pro -- X -- Y -- NH -- R.sup.2, wherein R 1 is a blocking group, --NH--R 2 is a chromogenic or fluorescent group, X represents a phenylalanyl, β-cyclohexylanyl, phenylglycyl or tyrosyl group, and Y represents a protonized arginyl or lysyl group, is disclosed. This substrate is useful for the quantitative determination of proteolytic enzymes of class E.C. 3.4.4., except thrombin and thrombin-like enzymes.
Inventors
- 👤 Lars Gundro Svendsen
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-26 00:32
CPC Classifications
C; C07; C07K; C07K5/00; C07K5/04; C07K5/08; C07K5/0821; C07K5/0823; C12; C12Q; C12Q1/00; C12Q1/34; C12Q1/37; C12Q2337/00; C12Q2337/10; C12Q2337/12; Y; Y10; Y10S; Y10S530/00; Y10S530/802
Claims Analysis
Total Claims: 18
Claim 1
Independent Claim
157 words
A chromogenic or fluorescent substrate for the quantitative determination of proteolytic enzymes of class E.C. 3.4.4., which split peptide chains on the carboxyl side of arginine and of lysine, except thrombin and thrombin-like enzymes, in human and mammal body fluids and in vegetable and animal cell extracts and in glandular venoms of cold-blooded animals, which essentially consists of a compound having the following formula: R.sup.1 -- Pro -- X -- Y -- NH -- R.sup.2 I wherein R 1 represents hydrogen, or an acyl or sulfonyl group, R 2 represents an aromatic hydrocarbon group which may carry substituents, X represents a phenylalanyl, β-cyclohexylalanyl, phenylglycyl or tyrosyl group and Y represents a protonized arginyl or lysyl group, --NH--R 2 being a chromogenic or fluorescent group, and which, when subjected to the proteolytic action of the enzymes, yields a split product of formula NH 2 --R 2 the quantity of which is measurable by photometric, spectrophotometric or fluorescence-photometric methods.
Claim 2
Dependent Claim
157 words
A chromogenic or fluorescent substrate for the quantitative determination of proteolytic enzymes of class E.C. 3.4.4., which split peptide chains on the carboxyl side of arginine and of lysine, except thrombin and thrombin-like enzymes, in human and mammal body fluids and in vegetable and animal cell extracts and in glandular venoms of cold-blooded animals, which essentially consists of a compound having the following formula: R.sup.1 -- Pro -- X -- Y -- NH -- R.sup.2 I wherein R 1 represents hydrogen, or an acyl or sulfonyl group, R 2 represents an aromatic hydrocarbon group which may carry substituents, X represents a phenylalanyl, β-cyclohexylalanyl, phenylglycyl or tyrosyl group and Y represents a protonized arginyl or lysyl group, --NH--R 2 being a chromogenic or fluorescent group, and which, when subjected to the proteolytic action of the enzymes, yields a split product of formula NH 2 --R 2 the quantity of which is measurable by photometric, spectrophotometric or fluorescence-photometric methods.
Claim 3
Dependent Claim
108 words
The substrate according to claim 1 in which the acyl group R 1 has the following partial formula R 3 -- CO -- (II) wherein R 3 represents a. an aliphatic hydrocarbon radical which comprises from 1 to 17 carbon atoms and which may carry an amino group in the ω-position; b. an araliphatic hydrocarbon radical which comprises from 1 17 carbon atoms and the aryl component of which may carry an amino group; c. a cycloaliphatic hydrocarbon radical which may carry an amino group or an aminomethyl group; d. an aromatic hydrocarbon radical which may carry an alkyl, amino or aminoalkyl group; or e. a benzyloxy group.
Claim 4
Dependent Claim
19 words
The substrate according to claim 1 in which the sulfonyl group R 1 is a benzenesulfonyl or p-toluenesulfonyl group.
Claim 5
Dependent Claim
17 words
The substrate according to claim 1 in which R 2 is a p-nitrophenyl, β-naphthyl or 4-methoxy-β-naphthyl group.
Claim 6
Dependent Claim
40 words
The substrate according to claim 1 which is protonized with a mineral acid such as HCl, HBr, H 2 SO 4 or H 3 PO 4 , or an organic acid such as formic acid, oxalic acid or tartaric acid.
Claim 7
Dependent Claim
1 words
N.sup.α-3-Phenylpropionyl-Pro-Phe-Arg-pNA-hydrochloride.
Claim 8
Dependent Claim
1 words
N.sup.α-3-Phenylpropionyl-Pro-Phe-Arg-pNA-hydrochloride.
Claim 9
Dependent Claim
1 words
N.sup.α-Cyclohexylcarbonyl-Pro-Phe-Arg-pNA-hydrochloride.
Claim 10
Dependent Claim
1 words
N.sup.α-Cyclohexylcarbonyl-Pro-Phe-Arg-pNA-hydrochloride.
Claim 11
Dependent Claim
1 words
N.sup.α-Benzoyl-Pro-Phe-Arg-pNA-hydrochloride.
Claim 12
Dependent Claim
1 words
N.sup.α-Benzoyl-Pro-Phe-Arg-pNA-hydrochloride.
Claim 13
Dependent Claim
1 words
N.sup.α-Cbo-Pro-Phe-Arg-pNA-hydrochloride.
Claim 14
Dependent Claim
1 words
N.sup.α-Cbo-Pro-Phe-Arg-pNA-hydrochloride.
Claim 15
Dependent Claim
1 words
N.sup.α-Caproyl-Pro-Phe-Arg-pNA-hydrochloride.
Claim 16
Dependent Claim
1 words
N.sup.α-Caproyl-Pro-Phe-Arg-pNA-hydrochloride.
Claim 17
Dependent Claim
111 words
A process for the quantitative determination of proteolytic enzymes of class E.C. 3.4.4., which split peptide chains on the carboxyl side of arginine as well as of lysine, except thrombin and thrombin-like enzymes, in human and mammal body fluids as well as in vegetable and animal cell extracts and in glandular venoms of cold-blooded animals, which comprises reacting the said body fluids, cell extracts or glandular venoms with a substrate according to claim 1 and measuring by means of photometric, spectrophotometric or fluorescence-photometric methods the quantity of the product NH 2 --R 2 split off from the substrate as a result of the proteolytic action of the enzymes on the substrate.
Claim 18
Dependent Claim
27 words
The process according to claim 11 as applied to the quantitative determination of plasma pre-kallikrein and its activators, plasma kallikrein and its inhibitors, papain and prothrombin activators.
References Cited (Backward Citations)
US3884896A
Cited
US3886136A
Cited
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1980-04-04 | 1980-10-04 | 1981-04-05 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1984-04-04 | 1984-10-04 | 1985-04-05 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1988-04-04 | 1988-10-04 | 1989-04-05 | $7700.00 | Window Closed / Lapsed |