CN1446925A - Single stable colorimetric reagent of enzyme in liquid state and its application - Google Patents
Single stable colorimetric reagent of enzyme in liquid state and its application Download PDFInfo
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Abstract
A high-stability liquid chromometric enzyme reagent is prepared from buffering liquid, water-soluble chromogenic agent, 4-aminoantipyrin, peroxidase, oxidase for generating hydrogen peroxide and enzyme stabilizer. Its advantages are durable stability, and high efficiency and correctness.
Description
Technical field
The present invention relates to a kind of analytical reagent and application thereof, particularly a kind of stable single liquid enzyme colorimetric reagent and application thereof.
Technical background
Use enzymic colorimetric now clinically and measure reagent, common have triglyceride level, cholesterol, glucose, uric acid, creatinine etc. are measured reagent, and its principle nearly all is that detected compound carries out colorimetric estimation by specific oxydase generation hydrogen peroxide.
Enzymic colorimetric is in the past measured reagent and is measured reagent based on powdered reagent.The weak point of powdered reagent is, need during use to redissolve, the redissolution time is long and to redissolving the water quality requirement height, the reagent bottle differences was big after different users was redissolved, and quality can not get guaranteeing, easily causes reagent waste, and its packing of dry powder single reagent can not be fitted all kinds of automatic analysers, change the special-purpose bottle of analyser after needing to redissolve again, troublesome poeration, vulnerable to pollution and process reagent waste are big.For easy to use, it was main developing into liquid reagent gradually in recent years.Because various ingredient stabilities differ, and are difficult to make stable single liquid reagent, common solution is to adopt biliquid reagent, and the enzyme and the reagent component two group reagent forms that split into that are about to different stable conditions preserve increase of service life.But because the increase of clinical detection project now makes the reagent position in storehouse of analytical instrument become crowded, because the specimen amount that detects trend increases, reagent is changed the frequency height, easily make mistakes, double reagent is measured in two steps, and spended time is long, because each mensuration project is divided two kinds of reagent easily to cause when installed reagents and obscured, cause that measurement result goes out disorderly, wastes reagent, influences work quality, some use its reagent position in storehouse of user of middle-size and small-size instrument not enough even more serious.
Summary of the invention
The objective of the invention is provides a kind of single liquid state to maintain a long-term stability for the deficiency that solves above-mentioned technology, performance is good, easy to use, on instrument programming simple, easy to identify being difficult for makes mistakes, the efficient height, the stable single liquid enzyme colorimetric reagent of suitable clinical use.
In order to achieve the above object, the single liquid enzyme colorimetric reagent that the present invention is stable, its basic comprising comprises damping fluid, developer, 4-aminoantipyrene, peroxidase, it is characterized in that it also comprises can produce the oxydase of the required hydrogen peroxide of colorimetric reaction and the enzyme stabilizers component that consumption is 0.01～500g/L with detected material effect.Described oxydase can be Phosphoric acid glycerol esters oxydase, rCO, urico-oxidase, glucose oxidase, sorbyl alcohol oxydase, sarcosine oxidase, Ketoamine oxidase, XOD, amino-acid oxidase, E.C. 1.1.99.1, pyruvic oxidase, Lactate Oxidase, fructosyl amino acid oxidase, 1,5-anhydro sorbitol oxydase.
In order further to improve the stability of single liquid enzyme colorimetric reagent, described damping fluid can be a biological buffer, the pH5.0-9.0 of biological buffer, and buffer concentration is 10～500mMol/L.Biological buffer can be Good ' S biological buffer (this English biological buffer is all trade(brand)name) pH5.0-9.0 such as ADA, ACES, BES, MES, Bis-Tris, PIPES, MOPS, MOPSO, HEPES, TES, DIPSO, TAPSO, and buffer concentration is 10～500mMol/L.Described developer can be selected water-soluble developer for use, and as TOPS, TOOS, TODB, ADPS, DBHS, TBHBA (the water-soluble developer of this English is all trade(brand)name) etc., consumption is 0.1～50mmol/L, also can be the combination of above-mentioned water-soluble developer.Water-soluble developer also can with phenol, P-hydroxybenzoic acid, 4-chlorophenol, 2 in the past, 4-two chlorophenols etc. are used in combination in right amount.Described reagent also can contain surface active agent composition, can select for use nonionogenic tenside, amphoterics, polyethers to be used singly or in combination, and consumption is 0.05～100g/L.Described reagent also can comprise the enzyme stabilizers component.
Enzyme is stablized component can select polyvalent alcohol, albumin, amino acid, proteinase inhibitor, list, disaccharides, sequestrant, antioxidant for use, can be independent use, also can be to be used in combination.Polyvalent alcohol can be glycerine, ethylene glycol, propylene glycol, N.F,USP MANNITOL, glycol ether etc., and consumption is 1～500g/L.Albumin can be bovine serum albumin, human serum albumin, ovalbumin etc., and consumption is 1～500g/L.Amino acid can be Methionin, aspartic acid, L-glutamic acid, L-Ala, leucine, methionine(Met), arginine, glycine, Threonine, tyrosine etc. and salt thereof, and consumption is 0.1～500g/L.Proteinase inhibitor has: phytic acid, glycoprotein ulinastatin, borate, 4-FPBA, proteinase inhibitor PMSF, Trypsin inhibitor,Trasylol (proteinase inhibitor Aprotinin), S-nitroso-group Triptide, Sodium cholic acid etc., consumption is 0.01～300g/L.Sequestrant has ethylenediamine tetraacetic acid (EDTA) (EDTA), iminodiethanoic acid, nitrilotriacetic acid and diethyl pentetic acid, 2-cyclohexanediaminetetraacetic acid, ethylene glycol bis (Norleucine) ether tetraacethyl, HEDTA etc. and its esters, and consumption is 0.05～100g/L.PROCLIN series sanitas, sodium azide, gentamicin, paraxin, phenols, methyl p-hydroxybenzoate, deoxidation acetate and salt, cats product etc., consumption is 0.05～200g/L.As reductive agents such as bovine serum albumin, boric acid, EDTA, tensio-active agent, gsh, mercaptoethanols stabilization is arranged all in the inventive method, for example the CO-N=base in the tensio-active agent polyvinylpyrrolidone (PVP) can both be in conjunction with phenol, form stabilized complex, boric acid also can rely on hydrogen bond to form mixture with phenolic compound, delay the oxidation of phenol, reach stable effect.An amount of metal ion can be magnesium, calcium, manganese, sodium, ammonium, potassium, cupric ion etc., and consumption is 0.01～100g/L.
A kind of stable single liquid enzyme colorimetric reagent provided by the invention, reagent can be maintained a long-term stability at the single liquid state, performance surpasses the requirement of dry powder single reagent, have easy to use, on instrument programming simple, easy to identify being difficult for makes mistakes, efficient height (can finish mensuration in common 5 minutes) meets clinical service requirements, also is applicable to the occasion of double reagent service requirements simultaneously.
In order to understand essence of the present invention better, below purposes of the present invention and effect are set forth by control experiment." national Clinical Laboratory working specification " second edition that reference examples is compiled with reference to Department of Medical Administration of Ministry of Health of the People's Republic of China.
Research through the contriver, discovery causes the labile factor of this reagents series to have several factors to cause, particularly its component of triglyceride determination reagent is complicated, therefore need one or more stabilising method effect, in most of the cases these stabilising method can be used in various Trinder reagent, will illustrate by implementing below:
Embodiment 1:
1, biological buffer substitutes traditional damping fluid, can effectively must improve reagent stability, first-selected Good ' the S of described biological buffer biological buffer, as ADA, ACES, BES, MES, Bis-Tris, PIPES, MOPS, MOPSO, HEPES, TES, DIPSO, TAPSO etc., its pH5.0-9.0 buffer concentration is that 10～200mM. present embodiment provides use biological buffer in glucose assays reagent:
Sample is a 300mg/dL glucose 37 ℃ of reactions 10 minutes, in the 500nm measurement result:
Test-results shows that Good ' S damping fluid can significantly improve stability.Embodiment 2:
| Reference examples 1 | Embodiment 1 | ||
| Phosphate buffered saline buffer pH7.0 | 100mM | MES pH of buffer 5.7 | ?50mM |
| Vapor enrichment phenol | 0.5g/L | Vapor enrichment phenol | ?0.5g/L |
| Glucose oxidase (aspergillus niger) | 10KU/L | Glucose oxidase (aspergillus niger) | ?10KU/L |
| Peroxidase | 6KU/L | Peroxidase | ?6KU/L |
| The 4-aminoantipyrene | 5mg/L | The 4-aminoantipyrene | ?5mg/L |
| Reference examples 1 | Embodiment 1 | |
| Fresh preparation is measured | 302mg/dL | ?301mg/dL |
| Reagent is put 37 ℃ and is measured after-7 days | 210mg/dL | ?302mg/dL |
Adopt the new type water-solubility developer to substitute in the past phenol or chlorophenol: as aniline between TOPS, TOOS, TODB, ADPS, HDAOS, ADOS, DHBS, TBHBA, N-ethyl-(β amino-ethyl) etc., consumption is 0.1～50mmol/L.New type water-solubility developer of the present invention also can with phenol, P-hydroxybenzoic acid, 4-chlorophenol, 2 in the past, 4-two chlorophenols etc. are used in combination in right amount.The great majority colour developing of new type water-solubility developer is the absorption peak that purple or blue look are avoided oxyphorase, can effectively reduce oxyphorase in the sample (redness) and disturb, and more can reduce the turbid interference of fat in 600nm/700nm place dual wavelength mensuration, and not have the corrodibility of phenol.
Present embodiment provides at total cholesterol and measures employing biological buffer and water-soluble developer in the reagent:
The sample cholesterol level is that 352mg/dL reacted 5 minutes at 37 ℃:
| Reference examples 2 | Embodiment 2 | ||
| Phosphate buffered saline buffer pH7.7 | 400mM | PIPES pH of buffer 7.0 | 50mM |
| Para-chlorophenol | 3.5mmol/L | TOOS | 2mmol/L |
| RCO | 500U/L | RCO | 500U/L |
| Sterol esterase | 1000U/L | Sterol esterase | 1000U/L |
| Peroxidase | 5KU/L | Peroxidase | 5KU/L |
| Sodium cholic acid | 3mmol/L | Sodium cholic acid | 3mmol/L |
| The 4-aminoantipyrene | 10mg/L | The 4-aminoantipyrene | 10mg/L |
| Triton?X-100 | 3g/L | Triton?X-100 | 3g/L |
| Reference examples 2 | Embodiment 2 | |
| Measure wavelength | 500nm | ?546nm |
| The standard pipe absorbancy | 0.280 | ?0.416 |
| Fresh preparation is measured | 354mg/dL | ?352mg/dL |
| Reagent is put 37 ℃ and is measured after-3 days | 110mg/dL | ?346mg/dL |
Test-results show the present invention with new type water-solubility developer and in the past chlorophenol mutually specific energy reduce damage greatly to enzyme activity.And new developer has higher sensitivity, can reduce the sample consumption and can reduce the interference to measuring such as serum mesobilirubin, xitix, oxyphorase, fat are turbid, improved the reliability of detected result.Embodiment 3:
Use different tensio-active agents and promote that the protein dissolving discharges detected material in the sample, and reduce the interference that the high fat of sample causes, can be that nonionogenic tenside, amphoterics, polyethers are used singly or in combination, consumption be 0.05～100g/L
Present embodiment provides with embodiment 2 and compares, and measures at total cholesterol and uses nonionogenic tenside to substitute Sodium cholic acid in the reagent separately:
The sample cholesterol level is that 352mg/dL reacted 5 minutes at 37 ℃:
| Embodiment 2 | Embodiment 3 | ||
| PIPES pH of buffer 7.0 | 50mM | PIPES pH of buffer 7.0 | 50mM |
| TOOS | 2mmol/L | TOOS | 2mmol/L |
| RCO | 500U/L | RCO | 500U/L |
| Sterol esterase | 1000U/L | Sterol esterase | 1000U/L |
| Peroxidase | 5KU/L | Peroxidase | 5KU/L |
| Sodium cholic acid | 3mmol/L | ||
| The 4-aminoantipyrene | 10mg/L | The 4-aminoantipyrene | 10mg/L |
| Triton?X-100 | 3g/L | Voranol EP 2001 HLB13.3 | 5g/L |
| Embodiment 2 | Embodiment 3 | |
| Measure wavelength | 546nm | ?546nm |
| The standard pipe absorbancy | 0.416 | ?0.418 |
| Fresh preparation is measured | 352mg/dL | ?351mg/dL |
| Reagent is put 37 ℃ and is measured after-7 days | 346mg/dL | ?353mg/dL |
Test-results shows that the embodiment of the invention 3 do not use Sodium cholic acid, use nonionogenic tenside to have high stability equally separately, its advantage: reference examples 2, embodiment 2 reagent contain Sodium cholic acid and easily disturb other clinical assays project such as bile acide, high and low density cholesterol direct method to measure, and embodiment 3 does not then have this drawback.Embodiment 4:
Present embodiment provides how to improve enzyme stability in the enzyme colorimetric reagent.
Usually enzyme is at liquid state less stable all, cause enzyme deactivation easily, protein is separated out, reagent blank rises, and polyvalent alcohol, albumin, amino acid, list, disaccharides, sequestrant, antioxidant, sanitas, proteinase inhibitor can both strengthen its stability action, can be independent use, also can be to be used in combination.
The following polyvalent alcohol of six carbon not only has stabilization and antifreeze effect (high temperature and freezing can both damage enzyme activity) is arranged: common have glycerine, ethylene glycol, propylene glycol, N.F,USP MANNITOL, a glycol ether etc., and consumption is 1～500g/L.
Protein can with the enzyme molecularity, prevent to dissociate into subunit or sex change inactivation the tendency: bovine serum albumin (having certain oxidation-resistance), human serum albumin, ovalbumin etc., consumption is 1～500g/L
Amino acid is to constitute proteinic fundamental unit, add the stability that amino acid can strengthen enzyme in enzyme liquid: Methionin, aspartic acid, L-glutamic acid, L-Ala, leucine, methionine(Met), arginine, glycine, Threonine, tyrosine etc. and salt thereof, consumption is 0.1～500g/L.
Because protease content is higher in some enzymes, the albumen, the meeting decomposing protein, because enzyme also is a protein, so required toolenzyme of proteolytic enzyme meeting decomposition reaction, cause the reagent instability, therefore add the stability that an amount of proteinase inhibitor and albumin can improve enzyme, common proteinase inhibitor has: phytic acid, glycoprotein ulinastatin, borate, 4-FPBA, proteinase inhibitor PMSF (to methanesulfonyl fluoride), Trypsin inhibitor,Trasylol (proteinase inhibitor Aprotinin), S-nitroso-group Triptide, Sodium cholic acid etc., consumption is 0.01～300g/L
These enzymic colorimetric reagent are except enzyme, also have some chemical reagent components compositions such as damping fluid, may contain a certain amount of heavy metal in these reagent, heavy metal can be facilitated the oxidation of enzyme molecule halfcystine and make enzyme deactivation, add an amount of sequestrant and can effectively prevent enzyme deactivation, and resemble EDTA class sequestrant and have certain antioxygenation: ethylenediamine tetraacetic acid (EDTA) (EDTA), iminodiethanoic acid, nitrilotriacetic acid and diethyl pentetic acid, the 2-cyclohexanediaminetetraacetic acid, ethylene glycol bis (Norleucine) ether tetraacethyl, HEDTA etc. and its esters, consumption is 0.05～100g/L.
Because it is rotten to contain appearance easy bacteria-developings such as a certain amount of protein, sugar, amino acid in the reagent, add an amount of anticorrosion and bactericidal agent stabilization is arranged: PROCLIN series sanitas, sodium azide, gentamicin, paraxin, phenols, methyl p-hydroxybenzoate, deoxidation acetate and salt, cats product etc., consumption is 0.05～200g/L
Trinder reagent needs the derivative of phenol or phenol to participate in color reaction, because phenol is easy to oxidized, cause the blank rising of reagent variable color easily, cause and measure influence, add suitable antioxidant and can obviously reduce blank: in the inventive method as bovine serum albumin, boric acid, EDTA, tensio-active agent, gsh, reductive agents such as mercaptoethanol all have stabilization, for example the CO-N=base in the tensio-active agent polyvinylpyrrolidone (PVP) can both be in conjunction with phenol, form stabilized complex, boric acid also can rely on hydrogen bond to form mixture with phenolic compound, delay the oxidation of phenol, reach stable effect.
An amount of metal ion has certain activation or stabilization to enzyme, and it can be: can be magnesium, calcium, manganese, sodium, ammonium, potassium, cupric ion etc., consumption be 0.01～100g/L.
Present embodiment is provided at and uses the technology of the present invention in the triglyceride determination reagent:
Embodiment 5:
| Reference examples 3 | Embodiment 4 | ||
| Tris-HCl pH of buffer 7.6 | 150mM | PIPES pH of buffer 7.0 | 50mM |
| Lipoproteinesterase | 3KU/L | Lipoproteinesterase | 3KU/L |
| Glycerol kinase | 2KU/L | Glycerol kinase | 2KU/L |
| The Phosphoric acid glycerol esters oxydase | 5KU/L | The Phosphoric acid glycerol esters oxydase | 5KU/L |
| Peroxidase | 5KU/L | Peroxidase | 5KU/L |
| ATP | 1mmol/L | ATP | 1mmol/L |
| Sal epsom | 17.5mmol/L | Sal epsom | 17.5 mmol/L |
| Sodium cholic acid | 3.5mmol/L | BSA | 5g/L |
| The 4-chlorophenol | 3.5mmol/L | TOOS | 2mmol/L |
| The 4-aminoantipyrene | 10mg/L | The 4-aminoantipyrene | 10mg/L |
| Triton?X-100 | 0.1g/L | The L-Asparagus cochinchinensis ammonia acid sodium | 30g/L |
| N.F,USP MANNITOL | 30g/L | ||
| Aprotinin | 50KU/L | ||
| Voranol EP 2001 HLB13.3 | 0.5g/L | ||
| Polyoxyethylene lauryl ether | 0.5g/L | ||
Present embodiment provides and further improve enzyme stability in the enzyme colorimetric reagent.
Embodiment 6:
| Embodiment 5 | |
| MOPSO pH of buffer 7.0 | ?50mM |
| Lipase (Japanese Asahi Chemical Industry) | ?500MU/L |
| Glycerol kinase | ?2KU/L |
| The Phosphoric acid glycerol esters oxydase | ?5KU/L |
| Peroxidase | ?5KU/L |
| ATP | ?1mmol/L |
| Magnesium chloride | ?10mmol/L |
| Calcium chloride | ?1mmol/L |
| BSA | ?5g/L |
| ADPS | ?2mmol/L |
| The 4-aminoantipyrene | ?10mg/L |
| P-Chlorophenol | ?0.5mmol/L |
| The L-Asparagus cochinchinensis ammonia acid sodium | ?30g/L |
| Ethylene glycol | ?50ml/L |
| FAD | ?10μmol/L |
| 4-FPBA | ?0.1g/L |
| Voranol EP 2001 HLB13.3 | ?1g/L |
| Polyoxyethylene lauryl ether | ?0.5g/L |
In order to illustrate that the present invention also can be used on the double reagent.Present embodiment can effectively be eliminated free glycerol (1000mg/dL) interference as measuring with two step method, to improve reagent stability.
Sample is the 1000mg/dL triglyceride level, measures after 5 minutes 37 ℃ of reactions:
| Embodiment 6 | |||
| Reagent 1 | Reagent 2 | ||
| PIPES pH of buffer 7.0 | ?50mM | PIPES pH of buffer 7.2 | 50mM |
| Glycerol kinase | ?1.5KU/L | Lipase (Japanese Asahi Chemical Industry) | 2000MU/L |
| The Phosphoric acid glycerol esters oxydase | ?3.75KU/L | Calcium chloride | 4mmol/L |
| Peroxidase | ?3.75KU/L | ADPS | 8mmol/L |
| ATP | ?0.75mmol/L | P-Chlorophenol | 2mmol/L |
| Magnesium chloride | ?10mmol/L | Voranol EP 2001 HLB13.3 | 1g/L |
| The 4-aminoantipyrene | ?10mg/L | Polyoxyethylene lauryl ether | 0.5g/L |
| BSA | ?5g/L | ||
| FAD | ?10μmol/L | ||
| The L-Asparagus cochinchinensis ammonia acid sodium | ?30g/L | ||
| N.F,USP MANNITOL | ?30g/L | ||
| 4-FPBA | ?0.1g/L | ||
| Gentamicin | ?50mg/L | ||
| Voranol EP 2001 HLB13.3 | ?1g/L | ||
| Polyoxyethylene lauryl ether | ?0.5g/L | ||
| Sodium azide | ?0.5g/L | ||
| Reference examples 3 | Embodiment 4 | Embodiment 5 | Embodiment 6 | |
| Measure wavelength | 500nm | 546nm | 546nm | ?546nm |
| Fresh preparation is measured | 1002mg/dL | 1003mg/dL | 1002mg/dL | ?1001mg/dL |
| Reagent is put 37 ℃ and is measured after-7 days | 162mg/dL | 1001mg/dL | 1000mg/dL | ?1003mg/dL |
Last table results of comparison, reference examples is through obvious inactivation behind the high temperature, and embodiment still keeps steady state.Embodiment 7:
Present embodiment provides the applicating example of the technology of the present invention in testing uric acid reagent:
Following examples make uric acid reagent reach long-time stable by multiple stabilization technique, reagent blank is low, because NaN3 has restraining effect to colibacillary uriKoxidase, use cats product to substitute NaN3 and can obtain high stability equally, can certainly use other anticorrosion and bactericidal agent.
Sample is that the 20mg/dL uric acid reacted measurement result 5 minutes at 37 ℃:
| Reference examples 4 | Embodiment 7 | ||
| Phosphate buffered saline buffer pH7.7 | 100mM | TAPS pH of buffer 8.0 | ?50mM |
| DHBS | 2mmol/L | topS | ?2mmol/L |
| UriKoxidase | 160U/L | UriKoxidase (recombination bacillus coli) | ?150U/L |
| Peroxidase | 1.5KU/L | Peroxidase | ?1.5KU/L |
| The 4-aminoantipyrene | 8mg/L | The 4-aminoantipyrene | ?8mg/L |
| NaN3 | 0.5g/L | EDTA.2Na | ?5mmol/L |
| Repone K | ?10mmol/L | ||
| Trisodium Citrate | ?10mmol/L | ||
| Dipotassium hydrogen phosphate | ?10mmol/L | ||
| Ethylene glycol | ?100ml/L | ||
| BSA | ?0.1％ | ||
| Polyethers | ?5g/L | ||
| Cats product | ?1g/L | ||
| Reference examples 4 | Embodiment 7 | |
| Measure wavelength | 500nm | ?546nm |
| The reagent blank absorbancy | 0.075 | ?0.008 |
| Fresh preparation is measured | 20.1mg/dL | ?20.0mg/dL |
| Reagent is put 37 ℃ and is measured after-10 days | 12.3mg/dL | ?20.1mg/dL |
| Reagent is put 4 ℃ and is measured after-18 months | 15.6mg/dL | ?20.0mg/dL |
Show from above all embodiment results, the present invention uses multiple stabilization technique to make up, Trinder reagent can be maintained a long-term stability under liquid single reagent (or double reagent) state, and the stabilization technique among the embodiment is not limited to embodiment, also can be applied in other Trinder reagent; Stabilization technique combination of the present invention is not limited among the embodiment makes up, and can multiple stabilising method make up arbitrarily as required.
Claims (9)
1, a kind of stable single liquid enzyme colorimetric reagent, its basic comprising comprises damping fluid, developer, 4-aminoantipyrene, peroxidase, it is characterized in that it also comprises can produce the oxydase of the required hydrogen peroxide of colorimetric reaction and the enzyme stabilizers component that consumption is 0.01～500g/L with detected material effect.
2, stable single liquid enzyme colorimetric reagent according to claim 1 is characterized in that described damping fluid is a biological buffer, pH of buffer 5.0-9.0, and buffer concentration is 10～500mMol/L.
3, stable single liquid enzyme colorimetric reagent according to claim 1 and 2 is characterized in that described developer is water-soluble developer, and consumption is 0.1～50mmol/L.
4, stable single liquid enzyme colorimetric reagent according to claim 1 and 2 is characterized in that described enzyme colorimetric reagent contains surface active agent composition, and consumption is 0.05～100g/L.
5, stable single liquid enzyme colorimetric reagent according to claim 3 is characterized in that described enzyme colorimetric reagent contains surface active agent composition, and consumption is 0.05～100g/L.
6, stable single liquid enzyme colorimetric reagent according to claim 1 and 2, it is characterized in that described oxydase can be Phosphoric acid glycerol esters oxydase, rCO, urico-oxidase, glucose oxidase, sorbyl alcohol oxydase, sarcosine oxidase, Ketoamine oxidase, XOD, amino-acid oxidase, E.C. 1.1.99.1, pyruvic oxidase, Lactate Oxidase, fructosyl amino acid oxidase, 1,5-anhydro sorbitol oxydase.
7, stable single liquid enzyme colorimetric reagent according to claim 3, it is characterized in that described oxydase can be Phosphoric acid glycerol esters oxydase, rCO, urico-oxidase, glucose oxidase, sorbyl alcohol oxydase, sarcosine oxidase, Ketoamine oxidase, XOD, amino-acid oxidase, E.C. 1.1.99.1, pyruvic oxidase, Lactate Oxidase, fructosyl amino acid oxidase, 1,5-anhydro sorbitol oxydase.
8, stable single liquid enzyme colorimetric reagent according to claim 4, it is characterized in that described oxydase can be Phosphoric acid glycerol esters oxydase, rCO, urico-oxidase, glucose oxidase, sorbyl alcohol oxydase, sarcosine oxidase, Ketoamine oxidase, XOD, amino-acid oxidase, E.C. 1.1.99.1, pyruvic oxidase, Lactate Oxidase, fructosyl amino acid oxidase, 1,5-anhydro sorbitol oxydase.
9, stable single liquid enzyme colorimetric reagent according to claim 5, it is characterized in that described oxydase can be Phosphoric acid glycerol esters oxydase, rCO, urico-oxidase, glucose oxidase, sorbyl alcohol oxydase, sarcosine oxidase, Ketoamine oxidase, XOD, amino-acid oxidase, E.C. 1.1.99.1, pyruvic oxidase, Lactate Oxidase, fructosyl amino acid oxidase, 1,5-anhydro sorbitol oxydase.
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