US3862885A - Determination of uric acid in blood with uricase - Google Patents
Determination of uric acid in blood with uricase Download PDFInfo
- Publication number
- US3862885A US3862885A US200582A US20058271A US3862885A US 3862885 A US3862885 A US 3862885A US 200582 A US200582 A US 200582A US 20058271 A US20058271 A US 20058271A US 3862885 A US3862885 A US 3862885A
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- US
- United States
- Prior art keywords
- hydrogen peroxide
- recited
- blood
- chromogen
- uric acid
- 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 - Lifetime
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- 210000004369 blood Anatomy 0.000 title claims abstract description 36
- 239000008280 blood Substances 0.000 title claims abstract description 36
- LEHOTFFKMJEONL-UHFFFAOYSA-N Uric Acid Chemical compound N1C(=O)NC(=O)C2=C1NC(=O)N2 LEHOTFFKMJEONL-UHFFFAOYSA-N 0.000 title abstract description 47
- TVWHNULVHGKJHS-UHFFFAOYSA-N Uric acid Natural products N1C(=O)NC(=O)C2NC(=O)NC21 TVWHNULVHGKJHS-UHFFFAOYSA-N 0.000 title abstract description 47
- 229940116269 uric acid Drugs 0.000 title abstract description 47
- 108010092464 Urate Oxidase Proteins 0.000 title abstract description 35
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 claims abstract description 122
- 238000000034 method Methods 0.000 claims abstract description 66
- 229940123748 Catalase inhibitor Drugs 0.000 claims abstract description 9
- 239000000872 buffer Substances 0.000 claims abstract description 9
- 230000003647 oxidation Effects 0.000 claims abstract description 7
- 238000007254 oxidation reaction Methods 0.000 claims abstract description 7
- PXIPVTKHYLBLMZ-UHFFFAOYSA-N Sodium azide Chemical group [Na+].[N-]=[N+]=[N-] PXIPVTKHYLBLMZ-UHFFFAOYSA-N 0.000 claims description 18
- 102000003992 Peroxidases Human genes 0.000 claims description 8
- 108040007629 peroxidase activity proteins Proteins 0.000 claims description 8
- 239000007853 buffer solution Substances 0.000 claims description 5
- 239000002270 dispersing agent Substances 0.000 claims description 5
- CDAWCLOXVUBKRW-UHFFFAOYSA-N 2-aminophenol Chemical compound NC1=CC=CC=C1O CDAWCLOXVUBKRW-UHFFFAOYSA-N 0.000 claims description 4
- 229920000036 polyvinylpyrrolidone Polymers 0.000 claims description 4
- 239000001267 polyvinylpyrrolidone Substances 0.000 claims description 4
- 235000013855 polyvinylpyrrolidone Nutrition 0.000 claims description 4
- NUIURNJTPRWVAP-UHFFFAOYSA-N 3,3'-Dimethylbenzidine Chemical compound C1=C(N)C(C)=CC(C=2C=C(C)C(N)=CC=2)=C1 NUIURNJTPRWVAP-UHFFFAOYSA-N 0.000 claims description 3
- QNGVNLMMEQUVQK-UHFFFAOYSA-N 4-n,4-n-diethylbenzene-1,4-diamine Chemical compound CCN(CC)C1=CC=C(N)C=C1 QNGVNLMMEQUVQK-UHFFFAOYSA-N 0.000 claims description 3
- SNCJAJRILVFXAE-UHFFFAOYSA-N 9h-fluorene-2,7-diamine Chemical compound NC1=CC=C2C3=CC=C(N)C=C3CC2=C1 SNCJAJRILVFXAE-UHFFFAOYSA-N 0.000 claims description 3
- JPVYNHNXODAKFH-UHFFFAOYSA-N Cu2+ Chemical compound [Cu+2] JPVYNHNXODAKFH-UHFFFAOYSA-N 0.000 claims description 3
- 239000002202 Polyethylene glycol Substances 0.000 claims description 3
- 239000008351 acetate buffer Substances 0.000 claims description 3
- 239000007979 citrate buffer Substances 0.000 claims description 3
- 229910001431 copper ion Inorganic materials 0.000 claims description 3
- -1 o-dianididine Chemical compound 0.000 claims description 3
- 239000008363 phosphate buffer Substances 0.000 claims description 3
- 229920001223 polyethylene glycol Polymers 0.000 claims description 3
- NNFCIKHAZHQZJG-UHFFFAOYSA-N potassium cyanide Chemical compound [K+].N#[C-] NNFCIKHAZHQZJG-UHFFFAOYSA-N 0.000 claims description 3
- BZORFPDSXLZWJF-UHFFFAOYSA-N N,N-dimethyl-1,4-phenylenediamine Chemical compound CN(C)C1=CC=C(N)C=C1 BZORFPDSXLZWJF-UHFFFAOYSA-N 0.000 claims description 2
- 239000004372 Polyvinyl alcohol Substances 0.000 claims description 2
- KXZJHVJKXJLBKO-UHFFFAOYSA-N chembl1408157 Chemical compound N=1C2=CC=CC=C2C(C(=O)O)=CC=1C1=CC=C(O)C=C1 KXZJHVJKXJLBKO-UHFFFAOYSA-N 0.000 claims description 2
- 229920002451 polyvinyl alcohol Polymers 0.000 claims description 2
- 239000003054 catalyst Substances 0.000 claims 2
- 125000000129 anionic group Chemical group 0.000 abstract description 10
- 239000004094 surface-active agent Substances 0.000 abstract description 10
- 239000000126 substance Substances 0.000 abstract description 9
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 abstract description 5
- 229910052760 oxygen Inorganic materials 0.000 abstract description 5
- 239000001301 oxygen Substances 0.000 abstract description 5
- 210000002966 serum Anatomy 0.000 description 16
- 238000012360 testing method Methods 0.000 description 13
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 description 12
- 239000000243 solution Substances 0.000 description 10
- 238000006243 chemical reaction Methods 0.000 description 8
- 102000016938 Catalase Human genes 0.000 description 7
- 108010053835 Catalase Proteins 0.000 description 7
- 230000000694 effects Effects 0.000 description 7
- ZZZCUOFIHGPKAK-UHFFFAOYSA-N D-erythro-ascorbic acid Natural products OCC1OC(=O)C(O)=C1O ZZZCUOFIHGPKAK-UHFFFAOYSA-N 0.000 description 6
- 102000004190 Enzymes Human genes 0.000 description 6
- 108090000790 Enzymes Proteins 0.000 description 6
- 229930003268 Vitamin C Natural products 0.000 description 6
- 229940088598 enzyme Drugs 0.000 description 6
- 239000000203 mixture Substances 0.000 description 6
- 235000019154 vitamin C Nutrition 0.000 description 6
- 239000011718 vitamin C Substances 0.000 description 6
- 239000003795 chemical substances by application Substances 0.000 description 5
- 238000010521 absorption reaction Methods 0.000 description 4
- POJWUDADGALRAB-UHFFFAOYSA-N allantoin Chemical compound NC(=O)NC1NC(=O)NC1=O POJWUDADGALRAB-UHFFFAOYSA-N 0.000 description 4
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-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 4
- 239000003638 chemical reducing agent Substances 0.000 description 4
- 238000000354 decomposition reaction Methods 0.000 description 4
- 230000002255 enzymatic effect Effects 0.000 description 4
- 238000005562 fading Methods 0.000 description 4
- 238000005259 measurement Methods 0.000 description 4
- WDGFFVCWBZVLCE-UHFFFAOYSA-N purpurogallin Chemical compound C1=CC=C(O)C(=O)C2=C1C=C(O)C(O)=C2O WDGFFVCWBZVLCE-UHFFFAOYSA-N 0.000 description 4
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 3
- 239000008103 glucose Substances 0.000 description 3
- 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 2
- POJWUDADGALRAB-PVQJCKRUSA-N Allantoin Natural products NC(=O)N[C@@H]1NC(=O)NC1=O POJWUDADGALRAB-PVQJCKRUSA-N 0.000 description 2
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 2
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 2
- 108010015776 Glucose oxidase Proteins 0.000 description 2
- 239000004366 Glucose oxidase Substances 0.000 description 2
- 240000004808 Saccharomyces cerevisiae Species 0.000 description 2
- 230000002378 acidificating effect Effects 0.000 description 2
- 229960000458 allantoin Drugs 0.000 description 2
- 229910000365 copper sulfate Inorganic materials 0.000 description 2
- ARUVKPQLZAKDPS-UHFFFAOYSA-L copper(II) sulfate Chemical compound [Cu+2].[O-][S+2]([O-])([O-])[O-] ARUVKPQLZAKDPS-UHFFFAOYSA-L 0.000 description 2
- XUJNEKJLAYXESH-UHFFFAOYSA-N cysteine Natural products SCC(N)C(O)=O XUJNEKJLAYXESH-UHFFFAOYSA-N 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- IINNWAYUJNWZRM-UHFFFAOYSA-L erythrosin B Chemical compound [Na+].[Na+].[O-]C(=O)C1=CC=CC=C1C1=C2C=C(I)C(=O)C(I)=C2OC2=C(I)C([O-])=C(I)C=C21 IINNWAYUJNWZRM-UHFFFAOYSA-L 0.000 description 2
- 238000009472 formulation Methods 0.000 description 2
- 229940116332 glucose oxidase Drugs 0.000 description 2
- 235000019420 glucose oxidase Nutrition 0.000 description 2
- RWSXRVCMGQZWBV-WDSKDSINSA-N glutathione Chemical compound OC(=O)[C@@H](N)CCC(=O)N[C@@H](CS)C(=O)NCC(O)=O RWSXRVCMGQZWBV-WDSKDSINSA-N 0.000 description 2
- 235000003969 glutathione Nutrition 0.000 description 2
- 229960003180 glutathione Drugs 0.000 description 2
- 150000002500 ions Chemical class 0.000 description 2
- 239000008055 phosphate buffer solution Substances 0.000 description 2
- 239000000049 pigment Substances 0.000 description 2
- 235000018102 proteins Nutrition 0.000 description 2
- 102000004169 proteins and genes Human genes 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- 239000001509 sodium citrate Substances 0.000 description 2
- 239000012064 sodium phosphate buffer Substances 0.000 description 2
- HFQQZARZPUDIFP-UHFFFAOYSA-M sodium;2-dodecylbenzenesulfonate Chemical compound [Na+].CCCCCCCCCCCCC1=CC=CC=C1S([O-])(=O)=O HFQQZARZPUDIFP-UHFFFAOYSA-M 0.000 description 2
- 239000003381 stabilizer Substances 0.000 description 2
- WQNHWIYLCRZRLR-UHFFFAOYSA-N 2-(3-hydroxy-2,5-dioxooxolan-3-yl)acetic acid Chemical compound OC(=O)CC1(O)CC(=O)OC1=O WQNHWIYLCRZRLR-UHFFFAOYSA-N 0.000 description 1
- YRSOHTBCCGDLSN-UHFFFAOYSA-N 4-(4-amino-3-methylphenyl)-2-methylaniline;hydron;chloride Chemical compound Cl.C1=C(N)C(C)=CC(C=2C=C(C)C(N)=CC=2)=C1 YRSOHTBCCGDLSN-UHFFFAOYSA-N 0.000 description 1
- BTBUEUYNUDRHOZ-UHFFFAOYSA-N Borate Chemical compound [O-]B([O-])[O-] BTBUEUYNUDRHOZ-UHFFFAOYSA-N 0.000 description 1
- RYGMFSIKBFXOCR-UHFFFAOYSA-N Copper Chemical compound [Cu] RYGMFSIKBFXOCR-UHFFFAOYSA-N 0.000 description 1
- XUJNEKJLAYXESH-REOHCLBHSA-N L-Cysteine Chemical compound SC[C@H](N)C(O)=O XUJNEKJLAYXESH-REOHCLBHSA-N 0.000 description 1
- 229910019142 PO4 Inorganic materials 0.000 description 1
- ULUAUXLGCMPNKK-UHFFFAOYSA-N Sulfobutanedioic acid Chemical class OC(=O)CC(C(O)=O)S(O)(=O)=O ULUAUXLGCMPNKK-UHFFFAOYSA-N 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- JPIYZTWMUGTEHX-UHFFFAOYSA-N auramine O free base Chemical compound C1=CC(N(C)C)=CC=C1C(=N)C1=CC=C(N(C)C)C=C1 JPIYZTWMUGTEHX-UHFFFAOYSA-N 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 238000003759 clinical diagnosis Methods 0.000 description 1
- 239000003086 colorant Substances 0.000 description 1
- 229910052802 copper Inorganic materials 0.000 description 1
- 239000010949 copper Substances 0.000 description 1
- 235000018417 cysteine Nutrition 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000002405 diagnostic procedure Methods 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 229960001484 edetic acid Drugs 0.000 description 1
- 229940011411 erythrosine Drugs 0.000 description 1
- 235000012732 erythrosine Nutrition 0.000 description 1
- 239000004174 erythrosine Substances 0.000 description 1
- 230000002349 favourable effect Effects 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 239000003112 inhibitor Substances 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- OOYGSFOGFJDDHP-KMCOLRRFSA-N kanamycin A sulfate Chemical group OS(O)(=O)=O.O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CN)O[C@@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O2)O)[C@H](N)C[C@@H]1N OOYGSFOGFJDDHP-KMCOLRRFSA-N 0.000 description 1
- 239000011259 mixed solution Substances 0.000 description 1
- 150000002978 peroxides Chemical class 0.000 description 1
- 235000021317 phosphate Nutrition 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- 238000001556 precipitation Methods 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- HNJBEVLQSNELDL-UHFFFAOYSA-N pyrrolidin-2-one Chemical compound O=C1CCCN1 HNJBEVLQSNELDL-UHFFFAOYSA-N 0.000 description 1
- 238000004445 quantitative analysis Methods 0.000 description 1
- 230000029219 regulation of pH Effects 0.000 description 1
- 150000003839 salts Chemical class 0.000 description 1
- 239000000523 sample Substances 0.000 description 1
- 239000012488 sample solution Substances 0.000 description 1
- NLJMYIDDQXHKNR-UHFFFAOYSA-K sodium citrate Chemical compound O.O.[Na+].[Na+].[Na+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O NLJMYIDDQXHKNR-UHFFFAOYSA-K 0.000 description 1
- 150000003467 sulfuric acid derivatives Chemical class 0.000 description 1
- 229920002554 vinyl polymer Polymers 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/62—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving uric acid
Definitions
- ABSTRACT A process for quantitively determining uric acid in blood with uricase characterized by making microbeoriginated uricase act on uric acid in blood in the presence of a catalase inhibitor and a buffer effective to maintain the pH at 5.5 to 7.0 and measuring the quantity of the generated hydrogen peroxide under a condition of a pH of 4.0 to 7.0 in the presence of an anionic surface active agent, a chromogen developing a color with hydrogen peroxide and a substance having a function of separating free oxygen from hydrogen peroxide and catalysing the oxidation of the chromogen.
- This invention relates to a process for the quantitative determination of uric acid in blood by using uricase.
- This invention comprises making uricase act on uric acid at a pH range of 5.5 to 7.0 so the blood may be converted into allantoin and at the same time, generate hydrogen peroxide thus bringing free oxygen decomposed from hydrogen peroxide catalyzed by the peroxidase into contact with a chromogen whereby a color is developed.
- the method of enzymatic determination of a substance in which the substance is converted into an intermediate substance by reacting the substance with an enzyme and then the quantity of the intermediate substance is measured by using another enzyme is already known.
- glucose oxidase is used for the oxidation of glucose as an auxiliary enzyme and hydrogen peroxide produced by the oxidation of glucose is quantitatively determined by its color development by the oxidation of a chromogen using oxygen liberated by the acid of peroxidase.
- a process for the enzymatic determination of uric acid in blood using animal-originated uricase is already known in Japanese Patent Publication No. 4440/1966 and U.S. Patent Spec. No.
- This process is a method of the quantitative determination of hydrogen peroxide generated by making animal uricase act on uric acid under a condition of a pH of 8.5 to 10.
- the generated hydrogen peroxide is so unstable and particularly in blood, is so readily influenced by catalase or reducing substance in the blood serum that it is impossible to make a high precision measurement of uric acid in the blood.
- the present inventors developed an accurate, quick and easy process for the quantitative determination of uric acid in blood by applying the principle of such enzymatic analyzing process.
- the present invention is as to a process which is characterized by the measurement of hydrogen peroxide generated by making uricase act on uric acid in blood under a condition of a pH of 5.5 to 7.0, and particularly perferable to make microbe-produced uricase act on uric acid under a condition of a pH of 5.7 to 6.5.
- a pH range of below 5.5 the uricase action on uric acid will remarkably decrease.
- a bad influence of reducing substance such as catalase and vitamin C etc. in blood are observed.
- microbe uricase particularly preferable yeast-originated uricase
- the optimum pH of the microbe uricase reaction shifts to the relatively acidic side as compared with the animal-originated uricase, therefore the decomposition of hydrogen peroxide by catalase in blood or autodegradation of hydrogen peroxide seems to be surpressed and hydrogen peroxide can be quantitatively caught by the hydrogen peroxide detecting system.
- Microbe-produced uricase obtained from yeast is such that its reaction pH is deviated to the acidic side about 1 more than that of animal originated uricase. Additionally it has sufficient activity even at a pH of 5 or 6 at which animal-originated uricase does not act and under this condition. Therefore the quantitative determination of uric acid in blood which was not possible by a known enzymatic process is now made possible. Surprisingly, a significant improvement in accuracy was observed by the process of the'present invention compared to that of known process, particularly in the following point; when uricase is made to act on uric acid near the pH of 8.0 to 9.0 hydrogen peroxide produced thereby will be so quickly decomposed by the catalase present in the blood serum, or so readily autodegradated by hydroxy.
- reducing substance such as vitamin C. glutathion, cysteine and SH protein
- the quantitatively determined value of uric acid is lower than its true value and, thus, it will not be able to be utilized pratically.
- the measured value was 55 compared with the value, 100 obtained by the uricametric UV process, and it evidently shows us how large the loss of hydrogen peroxide is and also shows that the practical use of the known process is thought to be impossible, whereas. in the process of the present invention, the measured value is 97 and the practical use of the process is possible.
- the effects of reducing substance such as vitamin C, glutathion, cystein and SH protein, and catalase which consume hydrogen peroxide are strong, whereas, in preserved blood serum, its influence is little.
- the present inventors have clarified how important the effect of pH regulation is for eliminating these influences.
- uric acid in blood It is absolutely necessary to know the true value of uric acid in blood.
- the process of the present invention which is useful, quick and easy for the quantitative measurement of uric acid in blood with uricase in a clinical diagnosis, it becomes possible to practice a diagnostic testing method.
- any buffer which works at a pH range of 5.5 to 7.0 can be used.
- a phosphate buffer. citrate buffer and acetate buffer are used in the present invention.
- the generated hydrogen peroxide can be measured by any method. Howover, usually it is measured by developing a color of a chromogen with hydrogen peroxide in the presence of a substance, for example peroxidase, which catalyzed the reaction of chromogen with free oxygen produced by decomposition of hydrogen peroxide and then colorimetrically determining it.
- a substance for example peroxidase
- peroxidase which catalyzed the reaction of chromogen with free oxygen produced by decomposition of hydrogen peroxide and then colorimetrically determining it.
- chromogen one can list o-tolidine, 2,7- diaminofluorene, dimethyl-p-phenylenediamine, diethyl-p-phenylenediamine, o-dianisidine and oaminophenol. It is preferable to develop a color by contacting hydrogen peroxide with said chromogen in a buffer solution ofa pH of 4 to 7, most preferably a pH of 4
- catalase As a hydrogen peroxide decomposing enzyme, catalase, is contained in the blood, it is preferable to add a catalase inhibitor in order to prevent the decomposition of the produced hydrogen peroxide.
- catalase inhibitor for example, sodium azide, potassium cyanide and sodium cyanide can be enumerated. Sodium azide among them is particularly preferable.
- the concentration of said added anionic'surface active agent it will be preferable to make the concentration of said added anionic'surface active agent more than 0.001
- the anionic surface active agent is added to a chromogen-containing reagent solution for preparing testing paper for detecting hydrogen peroxide, it will be preferable to add it so that is concentration may be more than 0.01
- an anionic surface active agent is added to a chromogen as mentioned above, the chromogen oxidated by the produced hydrogen peroxide will become insoluble, cohere and precipitate.
- a dispersing agent as a polyethylene glycol. polyvinyl alcohol or polyvinyl pyrrolidone.
- Such dispersing agent has an effect as of a color increasing agent simultaneously with an effect as of a pigment dispersing agent.
- the chromogen color can be made easy to judge with the naked eye by adding some pigment such as erythrosin or auramine, etc. so that the tone may be varied.
- the process of the present invention can be worked in the presence of not only ethylenediaminetetracetate as a stabilizer for uricase but also any other suitable stabilizers.
- the process of the present invention can be applied to blood in general and is particularly useful to the quantitative determination of uric acid in blood serum.
- a proper amount of copper ions for example, a small amount of copper sulfate into the blood srum in the process of the present invention. No bad influence of the coexistence of copper ions on the process of the present invention has been recognized.
- the process of the present invention practically comprises making a hydrogen peroxide generating system containing uricase, a catalase inhibitor and a buffer effective to maintain the pH at 5.5 to 7.0, act on uric acid, and then detecting the hydrogen peroxide generated with a hydrogen peroxide detecting system containing a chromogen, a substance having a function of catalyzing the oxidation of the chromogen and an anionic surface active agent.
- a hydrogen peroxide detecting system containing a chromogen, a substance having a function of catalyzing the oxidation of the chromogen and an anionic surface active agent.
- the generation and detection of hydrogen peroxide can be carried out simultaneously in one system in the present invention.
- one unit of the uricase activity is defined to be the quantity of the enzyme to catalyze uric acid so as to be converted to allantoin at a rate of one micromol per minute at a temperature of 25C.
- one unit of the peroxidase activity is defined to be the quantity of the enzyme having a capacity of forming 1 mg. of purpurogallin per 20 seconds at a pH of 6.0 at 20C.
- the reaction system was colorimetrically quantitatively determined with a standard uric acid sample solution (of 10.0 mg./dl. of uric acid) having reacted to develop a color under the same condition and obtained a favorable result in which the quantity of uric acid in blood was 10.2 mg./dl.
- EXAMPLE 2 Filter paper was impregnated with a solution prepared to be of a total amount of 100 ml by adding Peroxidase 190 purpurogallin units/mg.) 0. O-tolidine hydrochloride Citric anhydride 1. Sodium citrate 6 Polyvinyl pyrrolidone 2. Sodium laurylbenzene sulfonate 0. Erythrosine 0 Alcohol 30 to distilled water and was dried at the room temperature to prepare hydrogen peroxide testing paper.
- a process for quantitatively determining uric acid in blood with a yeast originated uricase which comprises reacting the uricase with the uric acid in blood at a pH of 5.7-6.5 to generate hydrogen peroxide and measuring the generated hydrogen peroxide.
- the buffer is a phosphate buffer, a citrate buffer or an acetate buffer.
- chromogen is o-tolidine. 2,7-diaminofluorene. dimethyl-pphenylenediamine, diethyl-p-phenylenediamine, odianididine, or o-aminophenol.
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- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Life Sciences & Earth Sciences (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Microbiology (AREA)
- Biochemistry (AREA)
- Physics & Mathematics (AREA)
- Molecular Biology (AREA)
- Biotechnology (AREA)
- Biophysics (AREA)
- Analytical Chemistry (AREA)
- Immunology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Investigating Or Analysing Biological Materials (AREA)
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP45103966A JPS5033795B1 (en, 2012) | 1970-11-25 | 1970-11-25 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| US3862885A true US3862885A (en) | 1975-01-28 |
Family
ID=14368090
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| US200582A Expired - Lifetime US3862885A (en) | 1970-11-25 | 1971-11-19 | Determination of uric acid in blood with uricase |
Country Status (2)
| Country | Link |
|---|---|
| US (1) | US3862885A (en, 2012) |
| JP (1) | JPS5033795B1 (en, 2012) |
Cited By (12)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US3979262A (en) * | 1974-02-12 | 1976-09-07 | Hoffmann-La Roche Inc. | Compositions and methods for the determination of oxidizing agents |
| DE2558536A1 (de) * | 1975-12-24 | 1977-07-07 | Boehringer Mannheim Gmbh | Verfahren zur kinetischen substratbestimmung und reagens zu seiner durchfuehrung |
| DE2625834A1 (de) * | 1976-06-09 | 1977-12-15 | Boehringer Mannheim Gmbh | Verfahren zur bestimmung von substraten oder enzymaktivitaeten |
| FR2372426A1 (en, 2012) * | 1976-11-25 | 1978-06-23 | Merck Patent Gmbh | |
| DE2855433A1 (de) * | 1978-02-13 | 1979-08-16 | Miles Lab | Testmittel und anzeiger zum nachweis von harnsaeure |
| EP0012446A1 (en) * | 1978-12-14 | 1980-06-25 | Kyowa Hakko Kogyo Co., Ltd | Acidic uricase, its production and its use for the determination of uric acid |
| US4810633A (en) * | 1984-06-04 | 1989-03-07 | Miles Inc. | Enzymatic ethanol test |
| US4837395A (en) * | 1985-05-10 | 1989-06-06 | Syntex (U.S.A.) Inc. | Single step heterogeneous assay |
| WO1993015218A1 (en) * | 1992-01-31 | 1993-08-05 | Actimed Laboratories, Inc. | Inhibition of catalase activity in biological fluids |
| US5266472A (en) * | 1991-05-17 | 1993-11-30 | Instrumentation Laboratory S.R.L. | Stabilization of the enzyme urate oxidase in liquid form |
| ITCS20080020A1 (it) * | 2008-11-20 | 2010-05-21 | Garofalo Alessandro | Metodo e kit per la determinazione della concentrazione di perossidi in un liquido organico |
| CN106615888A (zh) * | 2017-03-16 | 2017-05-10 | 青岛大学 | 一种影响嘌呤代谢的酵母果糖组合物 |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS54124198U (en, 2012) * | 1978-09-07 | 1979-08-30 |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US3335069A (en) * | 1964-12-14 | 1967-08-08 | Miles Lab | Composition and method for determining uric acid |
| US3367842A (en) * | 1965-02-17 | 1968-02-06 | Miles Lab | Test composition and device for the detection of galactose in fluids |
| US3475276A (en) * | 1965-07-28 | 1969-10-28 | Ono Pharmaceutical Co | Method of producing uricase from yeast |
| US3677903A (en) * | 1969-05-19 | 1972-07-18 | Donald L Bittner | Determination of uricase activity |
-
1970
- 1970-11-25 JP JP45103966A patent/JPS5033795B1/ja active Pending
-
1971
- 1971-11-19 US US200582A patent/US3862885A/en not_active Expired - Lifetime
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US3335069A (en) * | 1964-12-14 | 1967-08-08 | Miles Lab | Composition and method for determining uric acid |
| US3367842A (en) * | 1965-02-17 | 1968-02-06 | Miles Lab | Test composition and device for the detection of galactose in fluids |
| US3475276A (en) * | 1965-07-28 | 1969-10-28 | Ono Pharmaceutical Co | Method of producing uricase from yeast |
| US3677903A (en) * | 1969-05-19 | 1972-07-18 | Donald L Bittner | Determination of uricase activity |
Cited By (16)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US3979262A (en) * | 1974-02-12 | 1976-09-07 | Hoffmann-La Roche Inc. | Compositions and methods for the determination of oxidizing agents |
| DE2558536A1 (de) * | 1975-12-24 | 1977-07-07 | Boehringer Mannheim Gmbh | Verfahren zur kinetischen substratbestimmung und reagens zu seiner durchfuehrung |
| DE2625834A1 (de) * | 1976-06-09 | 1977-12-15 | Boehringer Mannheim Gmbh | Verfahren zur bestimmung von substraten oder enzymaktivitaeten |
| FR2372426A1 (en, 2012) * | 1976-11-25 | 1978-06-23 | Merck Patent Gmbh | |
| DE2855433A1 (de) * | 1978-02-13 | 1979-08-16 | Miles Lab | Testmittel und anzeiger zum nachweis von harnsaeure |
| EP0012446A1 (en) * | 1978-12-14 | 1980-06-25 | Kyowa Hakko Kogyo Co., Ltd | Acidic uricase, its production and its use for the determination of uric acid |
| US4317878A (en) * | 1978-12-14 | 1982-03-02 | Kyowa Hakko Kogyo Co., Ltd. | Test composition containing acidic uricase used for quantitative determination of uric acid in sample |
| US4810633A (en) * | 1984-06-04 | 1989-03-07 | Miles Inc. | Enzymatic ethanol test |
| US4837395A (en) * | 1985-05-10 | 1989-06-06 | Syntex (U.S.A.) Inc. | Single step heterogeneous assay |
| US5089383A (en) * | 1985-05-10 | 1992-02-18 | Syntex (U.S.A.) Inc. | Heterogeneous assay having delayed signal production |
| US5266472A (en) * | 1991-05-17 | 1993-11-30 | Instrumentation Laboratory S.R.L. | Stabilization of the enzyme urate oxidase in liquid form |
| WO1993015218A1 (en) * | 1992-01-31 | 1993-08-05 | Actimed Laboratories, Inc. | Inhibition of catalase activity in biological fluids |
| US5610025A (en) * | 1992-01-31 | 1997-03-11 | Actimed Laboratories, Inc. | Inhibition of interfering endogenous enzyme activity in assays of biological fluids |
| ITCS20080020A1 (it) * | 2008-11-20 | 2010-05-21 | Garofalo Alessandro | Metodo e kit per la determinazione della concentrazione di perossidi in un liquido organico |
| CN106615888A (zh) * | 2017-03-16 | 2017-05-10 | 青岛大学 | 一种影响嘌呤代谢的酵母果糖组合物 |
| CN106615888B (zh) * | 2017-03-16 | 2020-02-11 | 青岛大学 | 一种影响嘌呤代谢的酵母果糖组合物 |
Also Published As
| Publication number | Publication date |
|---|---|
| JPS5033795B1 (en, 2012) | 1975-11-04 |
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