US4268628A
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Method for the determination of α-amylase
Assignee: Roche Diagnostics GmbH
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Filing: 1979-03-19
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Granted: 1981-05-19
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Expires: 1999-03-19
Abstract
Abstract Novel method for the determination of α-amylase which method comprises contacting a sample suspected of containing α-amylase with a starch derivative carrying a substituent capable of dyestuff-forming coupling; separating from the resulting starch-containing phase the low molecular weight soluble fission products formed by splitting of said starch derivative by α-amylase; coupling the said couplable substituent with another dyestuff forming component; and measuring the dyestuff so formed as a measure of the initial α-amylase content.
Inventors
- 👤 Sigmar Klose
- 👤 Hans-Georg Batz
- 👤 Michael Stoltz
- 👤 Alexander Hagen
- 👤 Gunter Weimann
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-11-08 15:34
CPC Classifications
C; C08; C08B; C08B31/00; C12; C12Q; C12Q1/00; C12Q1/34; C12Q1/40; C12Q2334/00; Y; Y10; Y10S; Y10S435/00; Y10S435/805
Claims Analysis
Total Claims: 20
Claim 1
Independent Claim
213 words
Method for the determination of alpha-amylase which method comprises contacting at a pH of from 5 to 9, a sample suspected of containing alpha-amylase with a starch derivative carrying couplable substituents capable of dyestuff-forming coupling, to split said starch derivative by said alpha-amylase, forming a starch-containing phase and low molecular weight soluble fission products containing said couplable substituents; separating from the resulting said starch-containing phase the low molecular weight soluble fission products, coupling said couplable substituents with a dyestuff-forming component, and measuring the dyestuff so formed as a measure of the initial alpha-amylase content; wherein said starch derivative is substituted with a compound of the formula ##STR7## wherein R is a hydrogen atom or an alkyl radical containing up to 4 carbon atoms or an --(CH 2 ) n --X radical, X is a function capable of forming covalent bond with a hydroxyl group of the starch, and n is 1, 2, 3 or 4 or with a compound of the formula ##STR8## wherein R 1 is a hydrogen atom or a protective group which is easily split off, and X and n have the same meanings as in formula (I) or wherein the starch derivative is of the formula ##STR9## wherein X 1 -N- or ##STR10## and S is a starch residue.
Claim 2
Dependent Claim
213 words
Method for the determination of alpha-amylase which method comprises contacting at a pH of from 5 to 9, a sample suspected of containing alpha-amylase with a starch derivative carrying couplable substituents capable of dyestuff-forming coupling, to split said starch derivative by said alpha-amylase, forming a starch-containing phase and low molecular weight soluble fission products containing said couplable substituents; separating from the resulting said starch-containing phase the low molecular weight soluble fission products, coupling said couplable substituents with a dyestuff-forming component, and measuring the dyestuff so formed as a measure of the initial alpha-amylase content; wherein said starch derivative is substituted with a compound of the formula ##STR7## wherein R is a hydrogen atom or an alkyl radical containing up to 4 carbon atoms or an --(CH 2 ) n --X radical, X is a function capable of forming covalent bond with a hydroxyl group of the starch, and n is 1, 2, 3 or 4 or with a compound of the formula ##STR8## wherein R 1 is a hydrogen atom or a protective group which is easily split off, and X and n have the same meanings as in formula (I) or wherein the starch derivative is of the formula ##STR9## wherein X 1 -N- or ##STR10## and S is a starch residue.
Claim 3
Dependent Claim
34 words
Method as claimed in claim 1 wherein said starch derivative contains said couplable substituents capable of dyestuff-forming coupling in an amount of one substituent per 6 to 30 glucose units in said starch derivative.
Claim 4
Dependent Claim
34 words
Method as claimed in claim 2 wherein said starch derivative contains said couplable substituents capable of dyestuff-forming coupling in an amount of one substituent per 15 to 25 glucose units in said starch derivative.
Claim 5
Dependent Claim
66 words
Method as claimed in claim 1 wherein said starch derivative is substituted with a compound of the formula ##STR11## wherein R is a hydrogen atom or an alkyl radical containing up to 4 carbon atoms or a --X--(CH 2 ) n radical, X is a function capable of forming covalent bond with a hydroxyl group of the starch and n is 1, 2, 3, or 4.
Claim 6
Dependent Claim
35 words
Method as claimed in claim 4 wherein X is an oxirane group, a halogen atom, an acid chloride group, an isocyanate group, an isothiocyanate group, a tosylate group, a dichlorotriazine group or a mesylate group.
Claim 7
Dependent Claim
16 words
Method as claimed in claim 4 wherein a diazonium salt is used as the dyestuff-forming component.
Claim 8
Dependent Claim
24 words
Method as claimed in claim 4 wherein a compound capable of oxidative coupling with a phenol or aniline is used as the dyestuff-forming component.
Claim 9
Dependent Claim
19 words
Method as claimed in claim 7 wherein said dyestuff forming component is selected from N-alkylpyridone-(4)-hydrazone derivative, aminodimethyl-aniline and 1-phenyl-2,3-dimethyl-4-amino-3-pyrazolin-5-one.
Claim 10
Dependent Claim
13 words
Method as claimed in claim 8 wherein the N-alkylpyridone-(4)-hydrazone derivative used is N-methylbenzthiazolone-hydrazone.
Claim 11
Dependent Claim
35 words
Method as claimed in claim 7 wherein the oxidative coupling is carried out with ferric chloride, potassium ferricyanide, copper sulphate, silver nitrate, sodium hypochlorite, lead dioxide, ceric sulphate, hydrogen peroxide or a hydrogen peroxide-forming reaction.
Claim 12
Dependent Claim
61 words
Method as claimed in claim 1 wherein the starch employed has been reacted with a compound of the formula ##STR12## wherein R 1 is a hydrogen atom or a protective group which is easily split off and X is a function capable of forming covalent bond with a hydroxyl group of the starch and n is 1, 2, 3, or 4.
Claim 13
Dependent Claim
16 words
Method as claimed in claim 11 wherein the protective group R 1 is an acyl radical.
Claim 14
Dependent Claim
43 words
Method as claimed in claim 1 wherein said starch derivative is one in which every 10th to 50th glucose unit has been oxidized is reacted with an aniline derivative to give a Schiff base or with a phenyl-hydrazine derivative to give a hydrazone.
Claim 15
Dependent Claim
20 words
Method as claimed in claim 13, wherein the resulting Schiff base or hydrazone is reduced to give the corresponding amine.
Claim 16
Dependent Claim
21 words
Method as claimed in claim 1 wherein the low molecular weight, soluble fission product is separated by dialysis, filtration or centrifugation.
Claim 17
Dependent Claim
60 words
Method as claimed in claim 15 wherein in the case of a soluble starch derivative, the low molecular weight, soluble fission products are separated off by dialysis, a proportion of the fission products being dialysed from a first continuous stream of liquid into a second stream of liquid which is passed along on the other side of the dialysis membrane.
Claim 18
Dependent Claim
15 words
Method as claimed in claim 16 wherein a dialysis membrane with high permeability is used.
Claim 19
Dependent Claim
17 words
Method as claimed in claim 1 wherein said starch derivative is provided absorbed into an absorbent carrier.
Claim 20
Dependent Claim
18 words
Method as claimed in claim 18 wherein said carrier is in the form of test paper or rodlet.
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USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1984-05-18 | 1984-11-17 | 1985-05-19 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1988-05-18 | 1988-11-17 | 1989-05-19 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1992-05-18 | 1992-11-17 | 1993-05-19 | $7700.00 | Window Closed / Lapsed |