DE1773839C3 - Method and diagnostic agent for the determination of hydroperoxides or peroxidatically active substances - Google Patents
Method and diagnostic agent for the determination of hydroperoxides or peroxidatically active substancesInfo
- Publication number
- DE1773839C3 DE1773839C3 DE19681773839 DE1773839A DE1773839C3 DE 1773839 C3 DE1773839 C3 DE 1773839C3 DE 19681773839 DE19681773839 DE 19681773839 DE 1773839 A DE1773839 A DE 1773839A DE 1773839 C3 DE1773839 C3 DE 1773839C3
- Authority
- DE
- Germany
- Prior art keywords
- substances
- peroxidatically
- hydroperoxides
- determination
- peroxidase
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired
Links
- 239000000126 substance Substances 0.000 title claims description 24
- 150000002432 hydroperoxides Chemical class 0.000 title claims description 7
- 239000000032 diagnostic agent Substances 0.000 title description 2
- MHAJPDPJQMAIIY-UHFFFAOYSA-N hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 claims description 18
- 229940072417 Peroxidase Drugs 0.000 claims description 12
- 102000003992 Peroxidases Human genes 0.000 claims description 12
- 108090000437 Peroxidases Proteins 0.000 claims description 12
- 239000003153 chemical reaction reagent Substances 0.000 claims description 6
- 238000006243 chemical reaction Methods 0.000 claims description 4
- 238000002845 discoloration Methods 0.000 claims description 3
- 230000003287 optical Effects 0.000 claims 1
- WQZGKKKJIJFFOK-GASJEMHNSA-N D-Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 14
- 239000008103 glucose Substances 0.000 description 14
- 239000000243 solution Substances 0.000 description 13
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 8
- 239000008363 phosphate buffer Substances 0.000 description 7
- 238000001514 detection method Methods 0.000 description 6
- 108010015776 EC 1.1.3.4 Proteins 0.000 description 5
- 229940116332 GLUCOSE OXIDASE Drugs 0.000 description 5
- 239000004366 Glucose oxidase Substances 0.000 description 5
- 210000002700 Urine Anatomy 0.000 description 5
- 235000019420 glucose oxidase Nutrition 0.000 description 5
- -1 mercapto, amino Chemical group 0.000 description 5
- 125000000217 alkyl group Chemical group 0.000 description 4
- OAKJQQAXSVQMHS-UHFFFAOYSA-N hydrazine Chemical compound NN OAKJQQAXSVQMHS-UHFFFAOYSA-N 0.000 description 4
- 210000004369 Blood Anatomy 0.000 description 3
- 125000003277 amino group Chemical group 0.000 description 3
- 239000008280 blood Substances 0.000 description 3
- 239000007979 citrate buffer Substances 0.000 description 3
- 239000000203 mixture Substances 0.000 description 3
- 125000004430 oxygen atoms Chemical group O* 0.000 description 3
- 125000003396 thiol group Chemical group [H]S* 0.000 description 3
- GZCGUPFRVQAUEE-KCDKBNATSA-N D-(+)-Galactose Natural products OC[C@@H](O)[C@H](O)[C@H](O)[C@@H](O)C=O GZCGUPFRVQAUEE-KCDKBNATSA-N 0.000 description 2
- 229940088598 Enzyme Drugs 0.000 description 2
- 102000004190 Enzymes Human genes 0.000 description 2
- 108090000790 Enzymes Proteins 0.000 description 2
- RGHNJXZEOKUKBD-SQOUGZDYSA-N Gluconic acid Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@@H](O)C(O)=O RGHNJXZEOKUKBD-SQOUGZDYSA-N 0.000 description 2
- LHGVFZTZFXWLCP-UHFFFAOYSA-N Guaiacol Chemical compound COC1=CC=CC=C1O LHGVFZTZFXWLCP-UHFFFAOYSA-N 0.000 description 2
- 102000001554 Hemoglobins Human genes 0.000 description 2
- 108010054147 Hemoglobins Proteins 0.000 description 2
- FDGQSTZJBFJUBT-UHFFFAOYSA-N Hypoxanthine Chemical compound O=C1NC=NC2=C1NC=N2 FDGQSTZJBFJUBT-UHFFFAOYSA-N 0.000 description 2
- LEHOTFFKMJEONL-UHFFFAOYSA-N Trioxopurine Chemical compound N1C(=O)NC(=O)C2=C1NC(=O)N2 LEHOTFFKMJEONL-UHFFFAOYSA-N 0.000 description 2
- 229940116269 Uric Acid Drugs 0.000 description 2
- LRFVTYWOQMYALW-UHFFFAOYSA-N Xanthine Chemical compound O=C1NC(=O)NC2=C1NC=N2 LRFVTYWOQMYALW-UHFFFAOYSA-N 0.000 description 2
- 230000001476 alcoholic Effects 0.000 description 2
- 125000003545 alkoxy group Chemical group 0.000 description 2
- 150000001875 compounds Chemical class 0.000 description 2
- 230000000875 corresponding Effects 0.000 description 2
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 2
- DHMQDGOQFOQNFH-UHFFFAOYSA-N glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 description 2
- NTYJJOPFIAHURM-UHFFFAOYSA-N histamine Chemical compound NCCC1=CN=CN1 NTYJJOPFIAHURM-UHFFFAOYSA-N 0.000 description 2
- 229910052739 hydrogen Inorganic materials 0.000 description 2
- 239000001257 hydrogen Substances 0.000 description 2
- 125000004435 hydrogen atoms Chemical class [H]* 0.000 description 2
- 239000007788 liquid Substances 0.000 description 2
- OKKJLVBELUTLKV-UHFFFAOYSA-N methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 2
- 125000004573 morpholin-4-yl group Chemical group N1(CCOCC1)* 0.000 description 2
- 150000002978 peroxides Chemical class 0.000 description 2
- 239000000080 wetting agent Substances 0.000 description 2
- JRBJSXQPQWSCCF-UHFFFAOYSA-N 3,3'-Dimethoxybenzidine Chemical compound C1=C(N)C(OC)=CC(C=2C=C(OC)C(N)=CC=2)=C1 JRBJSXQPQWSCCF-UHFFFAOYSA-N 0.000 description 1
- 102000016777 Aldehyde oxidases Human genes 0.000 description 1
- 108091022173 Aldehyde oxidases Proteins 0.000 description 1
- XJKJWTWGDGIQRH-BFIDDRIFSA-N Alginic acid Chemical compound O1[C@@H](C(O)=O)[C@@H](OC)[C@H](O)[C@H](O)[C@@H]1O[C@@H]1[C@@H](C(O)=O)O[C@@H](C)[C@@H](O)[C@H]1O XJKJWTWGDGIQRH-BFIDDRIFSA-N 0.000 description 1
- 102000016893 Amine Oxidase (Copper-Containing) Human genes 0.000 description 1
- 108010028700 Amine Oxidase (Copper-Containing) Proteins 0.000 description 1
- HFACYLZERDEVSX-UHFFFAOYSA-N Benzidine Chemical compound C1=CC(N)=CC=C1C1=CC=C(N)C=C1 HFACYLZERDEVSX-UHFFFAOYSA-N 0.000 description 1
- IGXWBGJHJZYPQS-SSDOTTSWSA-N D-Luciferin Chemical compound OC(=O)[C@H]1CSC(C=2SC3=CC=C(O)C=C3N=2)=N1 IGXWBGJHJZYPQS-SSDOTTSWSA-N 0.000 description 1
- 150000008574 D-amino acids Chemical class 0.000 description 1
- 108010015133 EC 1.1.3.9 Proteins 0.000 description 1
- 108010093894 EC 1.17.3.2 Proteins 0.000 description 1
- 108010004237 EC 1.4.3.19 Proteins 0.000 description 1
- 108010003989 EC 1.4.3.3 Proteins 0.000 description 1
- 102000004674 EC 1.4.3.3 Human genes 0.000 description 1
- 108010062431 EC 1.4.3.4 Proteins 0.000 description 1
- 102000010909 EC 1.4.3.4 Human genes 0.000 description 1
- 239000004471 Glycine Substances 0.000 description 1
- 229960001867 Guaiacol Drugs 0.000 description 1
- 108010008292 L-Amino Acid Oxidase Proteins 0.000 description 1
- 102000007070 L-Amino Acid Oxidase Human genes 0.000 description 1
- 150000008575 L-amino acids Chemical class 0.000 description 1
- 210000004185 Liver Anatomy 0.000 description 1
- 239000005089 Luciferase Substances 0.000 description 1
- 108060001084 Luciferase family Proteins 0.000 description 1
- RHCSKNNOAZULRK-UHFFFAOYSA-N Mescaline Chemical compound COC1=CC(CCN)=CC(OC)=C1OC RHCSKNNOAZULRK-UHFFFAOYSA-N 0.000 description 1
- KKXWZLQZFLNLPC-UHFFFAOYSA-N N'-(2-hydroxy-3-methoxybenzoyl)pyridine-2-carbohydrazide Chemical compound COC1=CC=CC(C(=O)NNC(=O)C=2N=CC=CC=2)=C1O KKXWZLQZFLNLPC-UHFFFAOYSA-N 0.000 description 1
- KFKOWVIALAGDNQ-UHFFFAOYSA-N N'-(2-hydroxy-3-methoxybenzoyl)pyridine-3-carbohydrazide Chemical compound COC1=CC=CC(C(=O)NNC(=O)C=2C=NC=CC=2)=C1O KFKOWVIALAGDNQ-UHFFFAOYSA-N 0.000 description 1
- 108020005203 Oxidases Proteins 0.000 description 1
- 241000282320 Panthera leo Species 0.000 description 1
- AUNGANRZJHBGPY-SCRDCRAPSA-N Riboflavin Chemical compound OC[C@@H](O)[C@@H](O)[C@@H](O)CN1C=2C=C(C)C(C)=CC=2N=C2C1=NC(=O)NC2=O AUNGANRZJHBGPY-SCRDCRAPSA-N 0.000 description 1
- 210000002966 Serum Anatomy 0.000 description 1
- NUIURNJTPRWVAP-UHFFFAOYSA-N Tolidine Chemical compound C1=C(N)C(C)=CC(C=2C=C(C)C(N)=CC=2)=C1 NUIURNJTPRWVAP-UHFFFAOYSA-N 0.000 description 1
- 108010092464 Urate Oxidase Proteins 0.000 description 1
- 240000000358 Viola adunca Species 0.000 description 1
- 235000005811 Viola adunca Nutrition 0.000 description 1
- 235000013487 Viola odorata Nutrition 0.000 description 1
- 235000002254 Viola papilionacea Nutrition 0.000 description 1
- 102100002634 XDH Human genes 0.000 description 1
- UCTWMZQNUQWSLP-UHFFFAOYSA-N adrenaline Chemical compound CNCC(O)C1=CC=C(O)C(O)=C1 UCTWMZQNUQWSLP-UHFFFAOYSA-N 0.000 description 1
- 150000001299 aldehydes Chemical class 0.000 description 1
- 229960001126 alginic acid Drugs 0.000 description 1
- 235000010443 alginic acid Nutrition 0.000 description 1
- 229920000615 alginic acid Polymers 0.000 description 1
- 239000000783 alginic acid Substances 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- 238000004164 analytical calibration Methods 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 238000011088 calibration curve Methods 0.000 description 1
- 239000003054 catalyst Substances 0.000 description 1
- 230000024881 catalytic activity Effects 0.000 description 1
- 150000004985 diamines Chemical class 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000006185 dispersion Substances 0.000 description 1
- 239000011888 foil Substances 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 239000000174 gluconic acid Substances 0.000 description 1
- 229950006191 gluconic acid Drugs 0.000 description 1
- 235000012208 gluconic acid Nutrition 0.000 description 1
- 229960001340 histamine Drugs 0.000 description 1
- 125000001841 imino group Chemical group [H]N=* 0.000 description 1
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 description 1
- 235000014655 lactic acid Nutrition 0.000 description 1
- 239000004310 lactic acid Substances 0.000 description 1
- 238000000034 method Methods 0.000 description 1
- 125000000449 nitro group Chemical group [O-][N+](*)=O 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- MYMOFIZGZYHOMD-UHFFFAOYSA-N oxygen Chemical compound O=O MYMOFIZGZYHOMD-UHFFFAOYSA-N 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N oxygen atom Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- XBDQKXXYIPTUBI-UHFFFAOYSA-M propionate Chemical compound CCC([O-])=O XBDQKXXYIPTUBI-UHFFFAOYSA-M 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 229910052717 sulfur Inorganic materials 0.000 description 1
- 125000004434 sulfur atoms Chemical group 0.000 description 1
- 229920002554 vinyl polymer Polymers 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
- 229940075420 xanthine Drugs 0.000 description 1
- WQZGKKKJIJFFOK-PHYPRBDBSA-N α-D-galactose Chemical compound OC[C@H]1O[C@H](O)[C@H](O)[C@@H](O)[C@H]1O WQZGKKKJIJFFOK-PHYPRBDBSA-N 0.000 description 1
- WQZGKKKJIJFFOK-VFUOTHLCSA-N β-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 1
Description
H1COH 1 CO
=N —N=C-R (1) H3CO OH= N -N = CR (1) H 3 CO OH
= N-N = C-= N-N = C-
in welcher Y eine Hydroxyl-, Mercapto- oder Aminogruppe und Z eine Hydroxylgruppe bedeutet, wobei Z zusammen mit Y auch ein Sauerstoffatom darstellen kann, und R Wasserstoff, eine niedere Alkyl-, Mercapto-, Amino-, Pyridino- und Morpholinogruppe, eine gegebenenfalls arylierte Pyrazangruppe oder einen gegebenenfalls mit einer oder mehreren Alkyl-, Hydroxy-, Alkoxy- und Nitrogruppen substituierten Phenylrest bedeutet, verwendet.in which Y is a hydroxyl, mercapto or amino group and Z is a hydroxyl group, where Z together with Y can also represent an oxygen atom, and R can represent hydrogen, a lower alkyl, mercapto, amino, pyridino and morpholino groups, one optionally arylated Pyrazane group or optionally with one or more alkyl, hydroxy, alkoxy and Nitro-substituted phenyl radical is used.
2. Diagnostisches Mittel zur Bestimmung von Hydroperoxiden bzw. Substanzen, die unter Freisetzung von Hydroperoxid reagieren, bestehend aus Peroxidase oder einer peroxidatisch wirksamen Substanz und einem Chromogen, dadurch gekennzeichnet, daß als Chromogen Substanzen der Formel I enthalten sind.2. Diagnostic means for the determination of hydroperoxides or substances that are released of hydroperoxide react, consisting of peroxidase or a peroxidatically active Substance and a chromogen, characterized in that substances are used as the chromogen of formula I are included.
3. Diagnostisches Mittel zur Bestimmung von Hämoglobin oder anderen peroxidatisch wirksamen Substanzen, bestehend aus Hydroperoxid oder Hydroperoxid bildenden Substanzen und einem Chromogen, dadurch gekennzeichnet, daß als Chromogen Substanzen der Formel I enthalten sind.3. Diagnostic agent for the determination of hemoglobin or other peroxidatically active Substances consisting of hydroperoxide or hydroperoxide-forming substances and a chromogen, characterized in that it contains substances of the formula I as chromogen are.
in welcher Y eine Hydroxyl-, Mercapto- oder Aminogruppe und Z eine Hydroxylgruppe bedeutet, wobei Z zusammen mit Y auch ein Sauerstoffatom darstellen kann, und R Wasserstoff, eine niedere Alkyl-, Mercapto-, Amino-, Pyridino- und Morpholinogruppe, eine gegebenenfalls arylierte Pyrazangruppe oder einen gegebenenfalls mit einer oder mehreren Alkyl-, Hydroxy-, Alkoxy- und Nitrogruppen substituierten Phenylrest bedeutet.in which Y is a hydroxyl, mercapto or amino group and Z is a hydroxyl group, where Z together with Y can also represent an oxygen atom, and R can represent hydrogen, a lower alkyl, Mercapto, amino, pyridino and morpholino groups, an optionally arylated pyrazane group or one optionally substituted by one or more alkyl, hydroxy, alkoxy and nitro groups Means phenyl radical.
Die Erfindung besteht nunmehr darin, daß man bei einem Verfahren der eingangs genannten Art
Chromogene der Formel I verwendet.
Für den Fall, daß Y eine Hydroxyl-, Mercapto- oder Aminogruppe und Z eine Hydroxylgruppe bedeutet,
können die Verbindungen 1 selbstverständlich auch in der tautomeren Form II vorliegenThe invention now consists in using chromogens of the formula I in a process of the type mentioned at the outset.
In the event that Y is a hydroxyl, mercapto or amino group and Z is a hydroxyl group, the compounds 1 can of course also be in the tautomeric form II
f V-C-NH-NH-C —R (II)f V-C-NH-NH-C —R (II)
H1CO OHH 1 CO OH
Die Erfindung bezieht sich auf ein Verfahren zur Bestimmung von Hydroperoxiden bzw. von Substanzen, die unter Freisetzung von Hydroperoxid reagieren, sowie von Peroxidase bzw. peroxidatisch wirksamen Substanzen mit Hilfe eines Chromogens unter Verwendung der Reaktion der Hydroperoxide mit Peroxidase bzw. den peroxidatisch wirksamen Substanzen durch Auswertung der Verfärbung.The invention relates to a method for determining hydroperoxides or substances which react to release hydroperoxide, as well as peroxidase or peroxidatically active substances with the help of a chromogen using the reaction of hydroperoxides with peroxidase or the peroxidatically active substances by evaluating the discoloration.
Es sind eine Reihe von Verbindungen bekannt, die von Wasserstoffperoxid und Peroxidase als Katalysator zu einem Farbstoff oxidiert werden (vgl. CH-PS 339755, GB-PS 1035911 und US-PS 3290228). Substanzen, die am häufigsten verwendet werden. sind z. B. Benzidin, o-Dianisidin, o-Tolidin, Guajakol usw. Diese Verbindungen sind aber teilweise nicht sehr stabil und können nach den neuesten Erkenntnissen gesundheitsschädlich sein, so daß ihre Handhabung nicht ungefährlich erscheint. Der Erfindung liegt die Aufgabe zugrunde, Substanzen zu finden, die als Chromogene für ein Verfahren der obengenannten Art geeignet und zugleich stabil und unschädlich sind.A number of compounds are known that use hydrogen peroxide and peroxidase as a catalyst are oxidized to a dye (cf. CH-PS 339755, GB-PS 1035911 and US-PS 3290228). Substances that are most commonly used. are z. B. benzidine, o-dianisidine, o-tolidine, guaiacol etc. However, some of these connections are not very stable and can be based on the latest findings be harmful to health, so that their handling does not appear to be harmless. The invention The object is to find substances that can be used as chromogens for a method of the above Kind and at the same time stable and harmless.
in welcher Y' ein Sauerstoff- oder Schwefelatom oder eine lminogruppe und Z' ein Sauerstoffatom darstellt und R die oben angegebene Bedeutung hat.in which Y 'is an oxygen or sulfur atom or an imino group and Z 'represents an oxygen atom and R has the meaning given above.
Die Bestimmung von Hydroperoxiden nach dem erfindungsgemäßen Verfahren ist vorzugsweise für gekoppelte und ungekoppelte Enzymreaktionen geeignet, wie z. B. Bestimmung von Glucose, Galactose, Aminosäuren, Harnsäure, Peroxiden, Hämoglobin, Peroxidase oder anderen peroxidatisch wirksamen Substanzen sowie Enzymaktivitäten. Die routinemäßige Bestimmung von derartigen Substraten ist· wegen ihrer elementaren Wichtigkeit aus der klinischen Chemie und aus der Lebensmittelchemie nicht mehr wegzudenken. Bei der Bestimmung von Glucose beispielsweise wird diese von Glucoseoxidase zu Gluconsäure oxidiert, wobei der Luftsauerstoff zu Wasserstoffperoxid reduziert wird. Das Wasserstoffperoxid oxidiert dann mittels Peroxidase oder einer peroxidatisch wirksamen Substanz den erfindungsgemäß verwendeten Indikator zum entsprechenden Farbstoff. Weitere Beispiele fur derartige analytisch brauchbaren Systeme, die unter Freisetzung von Wasserstoffperoxid reagieren, sind L-Aminosäureoxidase + L-Aminosäuren, D-Aminosäureoxidase + D-Aminosäuren, Uricase + Harnsäure, Xanthinoxidase + Hypoxanthin bzw. Xanthin, Glycinoxidase + Glycin, Monoaminooxidase + Monoamin (wie Adrenalin, Mescalin usw.), Diaminoxidase + Diamine (wie Histamin). LuciferaseThe determination of hydroperoxides by the method according to the invention is preferably for coupled and uncoupled enzyme reactions suitable, such as. B. Determination of glucose, galactose, Amino acids, uric acid, peroxides, hemoglobin, peroxidase or other peroxidatically active Substances as well as enzyme activities. The routine determination of such substrates is because of their elementary importance from clinical chemistry and from food chemistry indispensable. When determining glucose, for example, it is determined by glucose oxidase oxidized to gluconic acid, the oxygen in the air being reduced to hydrogen peroxide. The hydrogen peroxide then oxidized by means of peroxidase or a peroxidatically active substance according to the invention used indicator for the corresponding dye. More examples of such analytically useful systems that are under release reacting from hydrogen peroxide are L-amino acid oxidase + L-amino acids, D-amino acid oxidase + D-amino acids, uricase + uric acid, xanthine oxidase + hypoxanthine or xanthine, Glycine oxidase + glycine, monoamine oxidase + monoamine (such as adrenaline, mescaline, etc.), diamine oxidase + Diamines (like histamine). Luciferase
+ Luciferin, D-Asparaginsäureoxidase + iJ-Asparaginsäure, Leber - Aldehydoxidase + Aldehyd, Galactoseoxidase + Galactose, Edsons Flavinenzym + Milchsäure.+ Luciferin, D-aspartic acid oxidase + iJ-aspartic acid, Liver - aldehyde oxidase + aldehyde, galactose oxidase + galactose, Edson's flavin enzyme + Lactic acid.
Die Auswertung der Verfärbung erfolgt beispielsweise optisch oder bei Verwendung von Testpapierstreifen oder Testfilmen durch Vergleich der Farbstärke mit Blindproben bekannter Zusammensetzungen bzw. Farbtafeln.The discoloration is evaluated, for example optically or when using test paper strips or test films by comparing the color strength with blanks of known compositions or color tables.
Die Erfindung ist in den folgenden Beispielen näher erläutert.The invention is explained in more detail in the following examples.
Beispiel 1 Nachweis von Glucose in Lösungenexample 1 Detection of glucose in solutions
Je 1 mMol der in Tabelle I aufgefÜhrtsn Indikatoren der Formel I wird in 1 ml Methanol gelöst und mit 1 ml einer Lösung von 143 mg Glucoseoxidase und 7,5 mg Peroxidase in 100 ml 0,1 molarem Phosphatpuffer (pH 7,0) versetzt. Bei Zugabe von 1 ml einer Glucoselösung (etwa 10 bis 1000 mg%) erhält man die in der letzten Spalte der nachfolgenden Tabelle angegebenen Farbreaktionen, deren Intensität von der Glucosekonzentration abhängt.1 mmol of each of the indicators of the formula I listed in Table I is dissolved in 1 ml of methanol and treated with 1 ml of a solution of 143 mg of glucose oxidase and 7.5 mg of peroxidase in 100 ml of 0.1 molar phosphate buffer (pH 7.0). When 1 ml of a glucose solution (about 10 to 1000 mg%) is added, the color reactions given in the last column of the table below are obtained, the intensity of which depends on the glucose concentration.
Dieselben Ergebnisse erhält man, wenn man den Phosphatpuffer durch einen Citratpuffer vom pH 7 ersetzt.The same results are obtained if the phosphate buffer is replaced by a citrate buffer of pH 7 replaced.
Vcr- Z bindung Nr.Vcr- Z binding No.
—OH -OH-OH OH
—OH -OH-OH OH
1111th
13 —OH -OH —N H N13 —OH — OH —N H N
NO2 NO 2
— N H/ ---
- NH
Substanzen ISubstances I.
15 —OH -OH15 -OH -OH
Ver- Z bindung Nr.Ver Z binding No.
Farbecolor
desof
ReakprT" 35 16 —OH -OH
dukls Reak prT "35 16 -OH -OH
dukls
HO CH3 HO CH 3
1 —OH -OH1 -OH -OH
2 —OH -OH2 -OH -OH
3 —OH -OH3 -OH -OH
4 —OH -OH4 -OH -OH
5 -OH -OH5 -OH -OH
OCH.,OCH.,
4040
rosapink
1717th
1818th
rotviolett red-violet
rotviolett red-violet
5050
fleischfarben flesh colored
rotRed
5555
violettRed
violet
\HO
\
violettblue
violet
0—0—
—o— —ο—-O- —Ο—
—ο— —ο——Ο— —Ο—
—ο— —ο——Ο— —Ο—
— Ο— -SH- Ο— -SH
OCH3 OCH 3
Farbecolor
desof
Reak-React
lions-lions
V-ro-V-ro-
duktsdukts
rotviolett red-violet
rotviolett red-violet
orangerot Orange red
rotRed
rotorange Red orange
rotviolett red-violet
rotviolett red-violet
rotviolett red-violet
rosapink
rosapink
rosapink
rosa rosapink pink
rotRed
rotorange Red orange
rosapink
Beispiel 2
Reagenzpapier zum Nachweis von Glucose im HarnExample 2
Reagent paper for the detection of glucose in urine
7,5 mg Peroxidase und ;43 mg Glucoseoxidase werden in 100 ml eines 0,1 molaren Citrat- oder Phosphatpuflers vom pH 5.6 gelöst. Mit dieser Lösung imprägniert man Filterpapier (23 i 6 der Fa. Schleicher & Schiill). Danach werden 0,1 g N-(2-Hydroxy-3-methoxybenzoyI)-N'-(pyridin-3-carbonyl)- hydrazin (Substanz 4 der Tabelle I) in 100 ml Alkohol gelöst. Mit dieser Lösung wird das Filterpapier noch einmal imprägniert. Beim Eintauchen in glucosehaltigen Harn zeigt das weiße Papier ab etwa 50 mg% Glucose eine fleischfarbenrote Färbung, die je nach7.5 mg peroxidase and; 43 mg glucose oxidase are dissolved in 100 ml of a 0.1 molar citrate or phosphate buffer with a pH of 5.6. With this solution filter paper (23 i 6 from Schleicher & Schiill) is impregnated. Then 0.1 g of N- (2-hydroxy-3-methoxybenzoyI) -N '- (pyridine-3-carbonyl) - hydrazine (substance 4 of Table I) dissolved in 100 ml of alcohol. With this solution, the filter paper will still be once impregnated. When immersed in urine containing glucose, the white paper shows from about 50 mg% Glucose has a flesh-red color, depending on the
to Intensität den Glucosegehalt angibt.to intensity indicates the glucose content.
Beispiel 3
Reagenzfilm zum Nachweis von GlucoseExample 3
Reagent film for the detection of glucose
45 g »Propiofan« (wäßrige Dispersion von PoIyvinylpropionat der Fa. BASF), 35 g einer Lösung von 37 g »Algipon« (Alginsäure der Fa. Henkel, Düsseldorf) in 2 1 eines 0,5molaren Phosphatpuflers vom pH 5,7, 1 g »Texapon P« (Netzmittel der Fa. Deliydag, Düsseldorf), 10 ml Wasser, 0,075 g Peroxidase und 0,15 g Glucoseoxidase werden zu einem homogenen Brei verrührt. Zu dieser Mischung gibt man 6 ml einer 5%igen methanolischen Lösung von N - (2 - Hydroxy - 3 - methoxybenzoyl) - N' - (pyridin-3-carbonyI)-hydrazin. Mit der so hergestellten Mischung, in der 0,3% Indikator enthalten sind, werden Folien in einer Schichtdicke von 300 μ beschichtet und getrocknet.45 g of "Propiofan" (aqueous dispersion of polyvinyl propionate from BASF), 35 g of a solution of 37 g of "Algipon" (alginic acid from Henkel, Düsseldorf) in 2 liters of a 0.5 molar phosphate buffer with a pH of 5.7, 1 g of "Texapon P" (wetting agent from Deliydag, Düsseldorf), 10 ml of water, 0.075 g of peroxidase and 0.15 g of glucose oxidase become one mixed into a homogeneous pulp. 6 ml of a 5% strength methanolic solution are added to this mixture N - (2 - Hydroxy - 3 - methoxybenzoyl) - N '- (pyridine-3-carbonyI) hydrazine. With the mixture produced in this way, which contains 0.3% indicator, are Foils coated in a layer thickness of 300 μ and dried.
Mit wäßrigen Lösungen verschiedener Glucosekonzentrationen (z. B. Blut, Serum, Harn) erhält man auf dem farblosen Film abgestufte fleischfarbenrote Färbungen.With aqueous solutions of different glucose concentrations (e.g. blood, serum, urine), graded flesh-red color is obtained on the colorless film Colorations.
In analoger Weise erhält man bei Verwendung von 2,5-Di-(2-hydroxy-3-methoxyphenyl)-oxadiazol-1,3,4 (Substanz 18 der Tabelle) als Indikator rosa Färbungen. In an analogous manner, using 2,5-di- (2-hydroxy-3-methoxyphenyl) -oxadiazole-1,3,4 is obtained (Substance 18 of the table) as an indicator pink colorations.
Beispiel 4
Reagenzpapier zum Nachweis von Peroxiden in FlüssigkeitenExample 4
Reagent paper for the detection of peroxides in liquids
7,5 mg Peroxidase werden in 100 ml eines 0,1 molaren 3-carbonyl)-hydrazin in 100 ml Alkohol gelöst und Citrat- oder Phosphatpuffers vom pH 5,6 gelöst. das Filterpapier nochmals imprägniert. Beim Ein-Mit dieser Lösung wird ein Filterpapier (2316 der 35 tauchen in eine Lösung von Wasserstoffperoxid wird Fa. Schleicher & Schüll) imprägniert. Danach wird das Papier noch bei einer Konzentration von 0.2 ;/ml 1 g N-(2-Hydroxy-3-methoxybenzoyl)-N'-(pyridin- deutlich fleischfarben verfärbt.7.5 mg of peroxidase are dissolved in 100 ml of a 0.1 molar 3-carbonyl) hydrazine in 100 ml of alcohol and Citrate or phosphate buffer of pH 5.6 dissolved. the filter paper is impregnated again. At one-with this solution becomes a filter paper (2316 of 35 will be immersed in a solution of hydrogen peroxide Schleicher & Schüll) impregnated. Then the paper is still at a concentration of 0.2; / ml 1 g of N- (2-hydroxy-3-methoxybenzoyl) -N '- (pyridine- clearly discolored flesh-colored.
Beispiel 5
Reagenzpapier zum Nachweis von Blut im HarnExample 5
Reagent paper for the detection of blood in urine
Ein Filterpapier (2316 der Fa. Schleicher & Schiill) wird mit einem 0,1 molaren Phosphatpuffer vom pH 7,0, in welchem 1% »Texapon P« (Netzmittel der Fa. Dshydag, Düsseldorf) gelöst ist, imprägniert. Danach wird das imprägnierte Papier mit einer alkoholischen Lösung von N-(2-Hydroxy-3-methoxybenzoyl)-N'-(pyridin-3-carbonyl)-hydrazin getränkt und getrocknet. Bringt man auf das so hergestellte Papier einen Tropfen eines bluthaltigen Urins und einen Tropfen 3%igen Wasserstoffperoxids, so färbt sich das Testpapier noch bei einer Verdünnung von 1:500 000 fleischfarben.A filter paper (2316 from Schleicher & Schiill) with a 0.1 molar phosphate buffer of pH 7.0, in which 1% »Texapon P« (wetting agent from Dshydag, Düsseldorf) is dissolved, impregnated. The impregnated paper is then treated with a alcoholic solution of N- (2-hydroxy-3-methoxybenzoyl) -N '- (pyridine-3-carbonyl) hydrazine soaked and dried. If you put a drop of a blood-containing paper on the paper produced in this way Urine and a drop of 3% hydrogen peroxide, the test paper will still be colored with one Dilution of 1: 500,000 flesh-colored.
Quantitativer Nachweis von Glucose in Flüssigkeiten Jeweils 0,02 ml verschiedener Glucoselösungen mit 55 sung von 600 mg Glucoseoxidase und 100 mg PerQuantitative detection of glucose in liquids 0.02 ml each of different glucose solutions with 55 solution of 600 mg glucose oxidase and 100 mg per
Konzentrationen von 0 bis 300 mg% werden zu 2 ml einer alkoholischen 0,00143molaren Indikatorlösung von N - (2 - Hydroxy - 3 - methoxybenzoyl)-N'-(pyridin-2-carbonyl)-hydrazin (Substanz 3 der Tabelle) pipettiert. Hierzu gibt man 2 ml einer Löoxidase in 100 ml eines 0,5molaren Phosphatpuffers vom pH 5,7, schüttelt um, mißt bei einer Wellenlänge von 492 nm und berechnet mit Hilfe einer Eichkurve die entsprechenden Glucosewerte.Concentrations from 0 to 300 mg% become 2 ml of an alcoholic 0.00143 molar indicator solution of N - (2 - hydroxy - 3 - methoxybenzoyl) -N '- (pyridine-2-carbonyl) hydrazine (substance 3 of Table) pipetted. To this end, 2 ml of a Löoxidase in 100 ml of a 0.5 molar phosphate buffer are added pH 5.7, shake, measure at a wavelength of 492 nm and calculate with the aid of a Calibration curve shows the corresponding glucose values.
Claims (1)
Priority Applications (18)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| DE1598133A DE1598133B2 (en) | 1966-09-01 | 1966-09-01 | Method and diagnostic agent for the determination of hydroperoxides or peroxidatically active substances |
| US653329A US3558435A (en) | 1966-09-01 | 1967-08-04 | Diagnostic agents for use in the determination of hydroperoxides and of peroxidate-active substances and methods for manufacturing and using the same |
| CH1204467A CH500488A (en) | 1966-09-01 | 1967-08-28 | Process and diagnostic agent for the determination of hydroperoxides or peroxidatically active substances |
| GB39489/67A GB1131560A (en) | 1966-09-01 | 1967-08-29 | Process and diagnostic agent for the determination of hydroperoxides |
| FI672342A FI48782C (en) | 1966-09-01 | 1967-08-30 | Diagnostic substance for use in the analytical determination of hydrogen peroxide or peroxidatively active substances. |
| AT796767A AT278708B (en) | 1966-09-01 | 1967-08-30 | Method and diagnostic means for the determination of hydroperoxides or substances which react to release hydroperoxide and peroxidase or peroxidatically active substances |
| NL676711965A NL154003B (en) | 1966-09-01 | 1967-08-31 | PROCEDURE FOR THE DETERMINATION OF HYDROPEROXIDES, PEROXYDASE OR PEROXYDATIC ACTIVE SUBSTANCES, PROCEDURE FOR PREPARING A DIAGNOSTIC AGENT FOR THIS AND MOLDED ARTICLES THEREFORE. |
| FR119529A FR1535921A (en) | 1966-09-01 | 1967-08-31 | Method and diagnostic agent for the determination of hydroperoxides or substances exhibiting peroxidase activity |
| DE19681773839 DE1773839C3 (en) | 1968-07-15 | Method and diagnostic agent for the determination of hydroperoxides or peroxidatically active substances | |
| DE19681773899 DE1773899A1 (en) | 1966-09-01 | 1968-07-24 | Rotary piston counter |
| US834178A US3668076A (en) | 1966-09-01 | 1969-06-17 | Diagnostic agent |
| FR6923688A FR2014605A1 (en) | 1966-09-01 | 1969-07-11 | |
| GB34991/69A GB1215778A (en) | 1966-09-01 | 1969-07-11 | Process and diagnostic agent for the determination of hydrogen peroxide |
| FI692091A FI49882C (en) | 1966-09-01 | 1969-07-14 | Diagnostic agent for use in the analytical determination of hydrogen peroxide or peroxidatically active substances. |
| NL6910781.A NL163022C (en) | 1966-09-01 | 1969-07-14 | METHOD FOR DETERMINING HYDROPEROXIDES, PEROXYDASE OR PEROXYDATIC ACTIVE SUBSTANCES, METHOD FOR PREPARING A DIAGNOSTIC AGENT FOR SUCH AND ARTICLES THEREFORE OBTAINED. |
| AT675569A AT288603B (en) | 1966-09-01 | 1969-07-14 | Process and diagnostic agent for the determination of hydroperoxides or peroxidatically active substances |
| CH1079369A CH552215A (en) | 1966-09-01 | 1969-07-14 | DIAGNOSTIC MEANS FOR THE DETECTION OR QUANTITATIVE DETERMINATION OF HYDROPEROXYDES OR OF SUBSTANCES THAT REACT RELEASE OF HYDROPEROXYDE, OR OF PEROXYDASE OR PEROXYDATIC SUBSTANCES AND THEIR USES. |
| JP44056044A JPS5016678B1 (en) | 1966-09-01 | 1969-07-15 |
Applications Claiming Priority (3)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| DEB0088720 | 1966-09-01 | ||
| DE19681773839 DE1773839C3 (en) | 1968-07-15 | Method and diagnostic agent for the determination of hydroperoxides or peroxidatically active substances | |
| DE19681773899 DE1773899A1 (en) | 1966-09-01 | 1968-07-24 | Rotary piston counter |
Publications (3)
| Publication Number | Publication Date |
|---|---|
| DE1773839A1 DE1773839A1 (en) | 1971-11-25 |
| DE1773839B2 DE1773839B2 (en) | 1976-05-06 |
| DE1773839C3 true DE1773839C3 (en) | 1976-12-16 |
Family
ID=
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