NO753268L - - Google Patents
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- Publication number
- NO753268L NO753268L NO753268A NO753268A NO753268L NO 753268 L NO753268 L NO 753268L NO 753268 A NO753268 A NO 753268A NO 753268 A NO753268 A NO 753268A NO 753268 L NO753268 L NO 753268L
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- iodophenyl
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- KFZMGEQAYNKOFK-UHFFFAOYSA-N Isopropanol Chemical compound CC(C)O KFZMGEQAYNKOFK-UHFFFAOYSA-N 0.000 claims description 35
- 239000000243 solution Substances 0.000 claims description 30
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-Dimethylformamide Chemical compound CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 claims description 26
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 19
- 239000007864 aqueous solution Substances 0.000 claims description 13
- 108010071390 Serum Albumin Proteins 0.000 claims description 12
- 102000007562 Serum Albumin Human genes 0.000 claims description 12
- 238000000034 method Methods 0.000 claims description 12
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 claims description 10
- 229920002307 Dextran Polymers 0.000 claims description 9
- FVFWUNAWQMROIF-UHFFFAOYSA-N n-(4-iodophenyl)imino-n'-(4-nitroanilino)benzenecarboximidamide Chemical compound C1=CC([N+](=O)[O-])=CC=C1NN=C(C=1C=CC=CC=1)N=NC1=CC=C(I)C=C1 FVFWUNAWQMROIF-UHFFFAOYSA-N 0.000 claims description 9
- 229910052757 nitrogen Inorganic materials 0.000 claims description 9
- 239000012087 reference standard solution Substances 0.000 claims description 7
- 239000002904 solvent Substances 0.000 claims description 7
- 230000008961 swelling Effects 0.000 claims description 7
- 108091003079 Bovine Serum Albumin Proteins 0.000 claims description 6
- 238000010521 absorption reaction Methods 0.000 claims description 6
- 229940098773 bovine serum albumin Drugs 0.000 claims description 6
- 239000003795 chemical substances by application Substances 0.000 claims description 6
- 238000006911 enzymatic reaction Methods 0.000 claims description 5
- 238000003756 stirring Methods 0.000 claims description 5
- 239000004067 bulking agent Substances 0.000 claims description 3
- 239000012086 standard solution Substances 0.000 claims 2
- BWJJSYVJOLHQFF-AMLDTQNSSA-N N'-(4-iodoanilino)-N-(4-nitrophenyl)iminobenzenecarboximidamide Chemical compound C1=CC=C(C=C1)/C(=N/NC2=CC=C(C=C2)I)/N=NC3=CC=C(C=C3)[N+](=O)[O-] BWJJSYVJOLHQFF-AMLDTQNSSA-N 0.000 description 9
- 239000003153 chemical reaction reagent Substances 0.000 description 9
- 102000004190 Enzymes Human genes 0.000 description 8
- 108090000790 Enzymes Proteins 0.000 description 8
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 8
- 230000000694 effects Effects 0.000 description 8
- 230000002255 enzymatic effect Effects 0.000 description 8
- 239000011521 glass Substances 0.000 description 8
- 238000004458 analytical method Methods 0.000 description 6
- JORABGDXCIBAFL-UHFFFAOYSA-M iodonitrotetrazolium chloride Chemical compound [Cl-].C1=CC([N+](=O)[O-])=CC=C1N1[N+](C=2C=CC(I)=CC=2)=NC(C=2C=CC=CC=2)=N1 JORABGDXCIBAFL-UHFFFAOYSA-M 0.000 description 6
- 229930027945 nicotinamide-adenine dinucleotide Natural products 0.000 description 6
- 210000002966 serum Anatomy 0.000 description 6
- 102000003855 L-lactate dehydrogenase Human genes 0.000 description 5
- 108700023483 L-lactate dehydrogenases Proteins 0.000 description 5
- 238000004108 freeze drying Methods 0.000 description 5
- 238000002835 absorbance Methods 0.000 description 4
- 239000013074 reference sample Substances 0.000 description 4
- 239000000523 sample Substances 0.000 description 4
- 102000004420 Creatine Kinase Human genes 0.000 description 3
- 108010042126 Creatine kinase Proteins 0.000 description 3
- JVTAAEKCZFNVCJ-REOHCLBHSA-N L-lactic acid Chemical compound C[C@H](O)C(O)=O JVTAAEKCZFNVCJ-REOHCLBHSA-N 0.000 description 3
- ACFIXJIJDZMPPO-NNYOXOHSSA-N NADPH Chemical compound C1=CCC(C(=O)N)=CN1[C@H]1[C@H](O)[C@H](O)[C@@H](COP(O)(=O)OP(O)(=O)OC[C@@H]2[C@H]([C@@H](OP(O)(O)=O)[C@@H](O2)N2C3=NC=NC(N)=C3N=C2)O)O1 ACFIXJIJDZMPPO-NNYOXOHSSA-N 0.000 description 3
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- BOPGDPNILDQYTO-NNYOXOHSSA-N nicotinamide-adenine dinucleotide Chemical compound C1=CCC(C(=O)N)=CN1[C@H]1[C@H](O)[C@H](O)[C@@H](COP(O)(=O)OP(O)(=O)OC[C@@H]2[C@H]([C@@H](O)[C@@H](O2)N2C3=NC=NC(N)=C3N=C2)O)O1 BOPGDPNILDQYTO-NNYOXOHSSA-N 0.000 description 3
- 239000000758 substrate Substances 0.000 description 3
- VEPOHXYIFQMVHW-XOZOLZJESA-N 2,3-dihydroxybutanedioic acid (2S,3S)-3,4-dimethyl-2-phenylmorpholine Chemical compound OC(C(O)C(O)=O)C(O)=O.C[C@H]1[C@@H](OCCN1C)c1ccccc1 VEPOHXYIFQMVHW-XOZOLZJESA-N 0.000 description 2
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 description 2
- 101710088194 Dehydrogenase Proteins 0.000 description 2
- PCNDJXKNXGMECE-UHFFFAOYSA-N Phenazine Natural products C1=CC=CC2=NC3=CC=CC=C3N=C21 PCNDJXKNXGMECE-UHFFFAOYSA-N 0.000 description 2
- 239000000872 buffer Substances 0.000 description 2
- 239000000284 extract Substances 0.000 description 2
- 230000001575 pathological effect Effects 0.000 description 2
- 238000002360 preparation method Methods 0.000 description 2
- ALSTYHKOOCGGFT-KTKRTIGZSA-N (9Z)-octadecen-1-ol Chemical class CCCCCCCC\C=C/CCCCCCCCO ALSTYHKOOCGGFT-KTKRTIGZSA-N 0.000 description 1
- IXPNQXFRVYWDDI-UHFFFAOYSA-N 1-methyl-2,4-dioxo-1,3-diazinane-5-carboximidamide Chemical compound CN1CC(C(N)=N)C(=O)NC1=O IXPNQXFRVYWDDI-UHFFFAOYSA-N 0.000 description 1
- 102100031126 6-phosphogluconolactonase Human genes 0.000 description 1
- 108010029731 6-phosphogluconolactonase Proteins 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- 229920002134 Carboxymethyl cellulose Polymers 0.000 description 1
- FBPFZTCFMRRESA-FSIIMWSLSA-N D-Glucitol Natural products OC[C@H](O)[C@H](O)[C@@H](O)[C@H](O)CO FBPFZTCFMRRESA-FSIIMWSLSA-N 0.000 description 1
- FBPFZTCFMRRESA-KVTDHHQDSA-N D-Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 description 1
- FBPFZTCFMRRESA-JGWLITMVSA-N D-glucitol Chemical compound OC[C@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-JGWLITMVSA-N 0.000 description 1
- 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 1
- 108091006149 Electron carriers Proteins 0.000 description 1
- 108010018962 Glucosephosphate Dehydrogenase Proteins 0.000 description 1
- 229920000084 Gum arabic Polymers 0.000 description 1
- 108010044467 Isoenzymes Proteins 0.000 description 1
- 229930195725 Mannitol Natural products 0.000 description 1
- BAWFJGJZGIEFAR-NNYOXOHSSA-N NAD zwitterion Chemical compound NC(=O)C1=CC=C[N+]([C@H]2[C@@H]([C@H](O)[C@@H](COP([O-])(=O)OP(O)(=O)OC[C@@H]3[C@H]([C@@H](O)[C@@H](O3)N3C4=NC=NC(N)=C4N=C3)O)O2)O)=C1 BAWFJGJZGIEFAR-NNYOXOHSSA-N 0.000 description 1
- 241000978776 Senegalia senegal Species 0.000 description 1
- 229920002472 Starch Polymers 0.000 description 1
- 229930006000 Sucrose Natural products 0.000 description 1
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 1
- 239000007983 Tris buffer Substances 0.000 description 1
- 235000010489 acacia gum Nutrition 0.000 description 1
- 239000000205 acacia gum Substances 0.000 description 1
- 239000003708 ampul Substances 0.000 description 1
- 239000000538 analytical sample Substances 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 150000001720 carbohydrates Chemical class 0.000 description 1
- 235000014633 carbohydrates Nutrition 0.000 description 1
- 239000001768 carboxy methyl cellulose Substances 0.000 description 1
- 235000010948 carboxy methyl cellulose Nutrition 0.000 description 1
- 239000008112 carboxymethyl-cellulose Substances 0.000 description 1
- 235000010418 carrageenan Nutrition 0.000 description 1
- 239000000679 carrageenan Substances 0.000 description 1
- 229920001525 carrageenan Polymers 0.000 description 1
- 229940113118 carrageenan Drugs 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 238000007398 colorimetric assay Methods 0.000 description 1
- 230000001419 dependent effect Effects 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- IRXRGVFLQOSHOH-UHFFFAOYSA-L dipotassium;oxalate Chemical compound [K+].[K+].[O-]C(=O)C([O-])=O IRXRGVFLQOSHOH-UHFFFAOYSA-L 0.000 description 1
- 238000007824 enzymatic assay Methods 0.000 description 1
- 238000009472 formulation Methods 0.000 description 1
- 238000007429 general method Methods 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- 239000000594 mannitol Substances 0.000 description 1
- 235000010355 mannitol Nutrition 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 230000002906 microbiologic effect Effects 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 229950006238 nadide Drugs 0.000 description 1
- 239000008213 purified water Substances 0.000 description 1
- 150000003839 salts Chemical class 0.000 description 1
- 229920006395 saturated elastomer Polymers 0.000 description 1
- 235000010413 sodium alginate Nutrition 0.000 description 1
- 239000000661 sodium alginate Substances 0.000 description 1
- 229940005550 sodium alginate Drugs 0.000 description 1
- WZWGGYFEOBVNLA-UHFFFAOYSA-N sodium;dihydrate Chemical compound O.O.[Na] WZWGGYFEOBVNLA-UHFFFAOYSA-N 0.000 description 1
- 239000000600 sorbitol Substances 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 235000000346 sugar Nutrition 0.000 description 1
- 150000008163 sugars Chemical class 0.000 description 1
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
- UHVMMEOXYDMDKI-JKYCWFKZSA-L zinc;1-(5-cyanopyridin-2-yl)-3-[(1s,2s)-2-(6-fluoro-2-hydroxy-3-propanoylphenyl)cyclopropyl]urea;diacetate Chemical compound [Zn+2].CC([O-])=O.CC([O-])=O.CCC(=O)C1=CC=C(F)C([C@H]2[C@H](C2)NC(=O)NC=2N=CC(=CC=2)C#N)=C1O UHVMMEOXYDMDKI-JKYCWFKZSA-L 0.000 description 1
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N31/00—Investigating or analysing non-biological materials by the use of the chemical methods specified in the subgroup; Apparatus specially adapted for such methods
- G01N31/22—Investigating or analysing non-biological materials by the use of the chemical methods specified in the subgroup; Apparatus specially adapted for such methods using chemical indicators
-
- 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
- C12Q1/26—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving oxidoreductase
- C12Q1/32—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving oxidoreductase involving dehydrogenase
-
- 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
- C12Q1/48—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving transferase
- C12Q1/50—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving transferase involving creatine phosphokinase
-
- 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
- C12Q1/54—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving glucose or galactose
Description
"Stabil, farvet referansestandard for enzymatiske bestemmelser" "Stable, colored reference standard for enzymatic determinations"
Metoder for kvantitativ bestemmelse av visse enzymer er basert på fremstillingen av NADH (redusert nikotinamid-adenin-dinukleotid) eller NADPH (redusert nikotinamid-adenin-dinukleotid-fosfat) som et resultat av den enzymatiske omsetningen på et substrat; NADH eller NADPH reduserer igjen fargeløst 2- (p-jodfenyl) -3-(p-nitrofenyl)-5-fenyltetrazoliumklorid (kjent som INT) i nær-vær av en elektronbærer slik som fenazinmetsulfat, til 1-(p-jodfenyl) -5-(p-nitrofenyl)-3-fenylformazan (kjent som INT formazan) , som er sterkt rødfarget og kan måles spektrofotometrisk. Enzym-konsentrasjoner bestemmes ved samtidig å utføre omsetningen på en referanseprøve som inneholder'kjente mengder av enzymet og sammen-ligne spektrofotometriske avlesninger av den ukjente prøven mot referanseprøven. Methods for the quantitative determination of certain enzymes are based on the production of NADH (reduced nicotinamide adenine dinucleotide) or NADPH (reduced nicotinamide adenine dinucleotide phosphate) as a result of the enzymatic reaction on a substrate; NADH or NADPH again colorlessly reduces 2-(p-iodophenyl)-3-(p-nitrophenyl)-5-phenyltetrazolium chloride (known as INT) in the presence of an electron carrier such as phenazine metsulfate, to 1-(p-iodophenyl)- 5-(p-nitrophenyl)-3-phenyl formazan (known as INT formazan), which is strongly red colored and can be measured spectrophotometrically. Enzyme concentrations are determined by simultaneously performing the reaction on a reference sample containing known amounts of the enzyme and comparing spectrophotometric readings of the unknown sample against the reference sample.
Representative enzymatiske bestemmelser hvor den ovenfor nevnte generelle metode anvendes, omfatter de følgende: Babson, A.L. og Phillips, G.E., " A Rapid Colorimetric Assay for Serum Lactic Dehydrogenase" , Clin. Chim. Acta 12_, 210-215 (1965); Representative enzymatic determinations where the above-mentioned general method is used include the following: Babson, A.L. and Phillips, G.E., "A Rapid Colorimetric Assay for Serum Lactic Dehydrogenase", Clin. Chim. Acta 12_, 210-215 (1965);
Van Der Veen et al., " Isoenzymes of Creatine Phosphokinase in Tissue Extracts and in Normal and Pathological Sera", Clin. Chim. Acta 13., 312-316 (1966); ogNachlas, M.M. , et al., " The Determina-tion of Lactic Dehydrogenase with a Tetrasodium Salt", Anal.Biochem., 1, 317-326 (1960). Van Der Veen et al., "Isoenzymes of Creatine Phosphokinase in Tissue Extracts and in Normal and Pathological Sera", Clin. Chim. Acta 13., 312-316 (1966); and Nachlas, M.M. , et al., "The Determination of Lactic Dehydrogenase with a Tetrasodium Salt", Anal.Biochem., 1, 317-326 (1960).
Nøyaktigheten av de ovenfor nevnte metoder for bestemmelse av enzymatiske virkninger er i høy grad avhengig av fremstillingen og stabiliteten av referanseprøven som inneholder den kjente mengde av enzym som analyseprøven skal sammenlignes med. Dessuten omfatter disse enzymatiske analysemetoder dobbelt arbeid, siden analysen må utføres i hvert tilfelle mot referanseprøven, så vel som mot den ukjente prøven. Slik duplisering krever utstrakt kontroll for å være sikker på at man ikke får unøyaktige resultater. The accuracy of the above-mentioned methods for determining enzymatic effects is highly dependent on the preparation and stability of the reference sample which contains the known amount of enzyme with which the analytical sample is to be compared. Moreover, these enzymatic methods of analysis involve double work, since the analysis must be performed in each case against the reference sample as well as against the unknown sample. Such duplication requires extensive control to ensure that inaccurate results are not obtained.
Behovet for og fordelene ved en stabil, farvet referansestandard for anvendelse ved enzymatiske bestemmelser uten samtidig å utføre en kontroll er åpenbar. Slikt et produkt er imidlertid ikke kommersielt tilgjengelig. The need for and advantages of a stable, colored reference standard for use in enzymatic determinations without simultaneously performing a control is obvious. However, such a product is not commercially available.
En stabil, farvet sammenli.gningsstandard består av en vandig løsning som inneholder fra 0,001%-0,020 vekt-%, basert på vekten av hele løsningen av 1- (p-jodfenyl)-5-(p-nitrofenyl)-3-fenyl-formazan, fra 0,01-0,10 vekt-% serumalbumin, fra 2-10 vekt-% av en løsning slik som N,N<1->dimetylformamid eller dimetylsulfoksyd og fra 2-10 vekt-% isopropylalkohol. Det kan tilsettes fra 5% til 20 vekt-% av et inert svellemiddel til den vandige, fargede referansestandard-løsningen, som deretter kan lyofiliseres. Det lyofiliserte produkt rekondisjoneres lett med vann. Den vandige, fargede referansestandard-løsningen har et absorpsjonsmaksimum ved 500 nanometer og er anvendbar for bestemmelse av visse enzymvirkninger, slik som serumlaktat dehydrogenas, kreatin fosfokinas, glukose-6-fosfat dehydrogenase og lignende. A stable, colored comparison standard consists of an aqueous solution containing from 0.001%-0.020% by weight, based on the weight of the entire solution, of 1-(p-iodophenyl)-5-(p-nitrophenyl)-3-phenyl- formazan, from 0.01-0.10% by weight serum albumin, from 2-10% by weight of a solution such as N,N<1->dimethylformamide or dimethylsulfoxide and from 2-10% by weight isopropyl alcohol. From 5% to 20% by weight of an inert bulking agent can be added to the aqueous, colored reference standard solution, which can then be lyophilized. The lyophilized product is easily reconditioned with water. The aqueous, colored reference standard solution has an absorption maximum at 500 nanometers and is useful for the determination of certain enzyme activities, such as serum lactate dehydrogenase, creatine phosphokinase, glucose-6-phosphate dehydrogenase and the like.
Man har nå funnet at en farget referansestandard for anvendelse ved bestemmelse av visse enzymvirkninger som inneholder, It has now been found that a colored reference standard for use in the determination of certain enzyme activities containing,
i en vandig løsning, fra 0,001-0,020 vekt-%, basert på vekten av den totale løsningen av 1-(p-jodfenyl)-5-(p-nitrofenyl)-3-fenyl-formazan, fra 0,01-0,10 vekt-% serumalbumin, fra 2-10 vekt-% av et løsningsmiddel valgt fra gruppen som består av N,N'-dimetyl-formamid og dimetylsulfoksyd, og fra 2-10 vekt-% isopropylalkohol har et absorpsjonsmaksimum ved 500 nanometer. Tilsetningen av fra 5-20 vekt-% av et inert svellemiddel gir en løsning som kan lyofiliseres. Den lyofiliserte form av den fargede referansestandarden i følge foreliggende oppfinnelse er stabil minst i 6 måneder ved 4°C og 24°C. Den lyofiliserte formen av referansestandarden i følge foreliggende oppfinnelse kan dessuten rekondisjoneres med vann for å gi en klar løsning som kan anvendes, ruten vanskelighet, som en referansestandard ved enzymatiske bestemmelser. in an aqueous solution, from 0.001-0.020% by weight, based on the weight of the total solution of 1-(p-iodophenyl)-5-(p-nitrophenyl)-3-phenyl-formazan, from 0.01-0, 10% by weight serum albumin, from 2-10% by weight of a solvent selected from the group consisting of N,N'-dimethylformamide and dimethylsulfoxide, and from 2-10% by weight isopropyl alcohol has an absorption maximum at 500 nanometers. The addition of from 5-20% by weight of an inert swelling agent gives a solution which can be lyophilized. The lyophilized form of the colored reference standard according to the present invention is stable for at least 6 months at 4°C and 24°C. The lyophilized form of the reference standard according to the present invention can also be reconstituted with water to give a clear solution which can be used, the route difficulty, as a reference standard in enzymatic determinations.
1-(P-jodfenyl) -5-(p-nitrofenyl) -3-fenylformazan i den fargede referansestandarden i følge oppfinnelsen (kjent som INT formazan) er kommersielt tilgjengelig (National Biochemicals Corp., Cleveland, Ohio) og har følgende formel: 1-(P-iodophenyl)-5-(p-nitrophenyl)-3-phenyl formazan in the colored reference standard of the invention (known as INT formazan) is commercially available (National Biochemicals Corp., Cleveland, Ohio) and has the following formula:
Man treffer ofte på INT formazan i mikrobiologiske identifiseringsteknikker, særlig for undersøkelse av næringsmidler og patologiske prøver som dersom bakterier er til stede, reduserer det fargeløse 2-(p-jodfenyl)-3-(p-nitrofenyl)-5-fenyltetrazolium-klorid (kjent som INT) som har formelen: INT formazan is often encountered in microbiological identification techniques, particularly for the examination of foodstuffs and pathological samples which, if bacteria are present, reduce the colorless 2-(p-iodophenyl)-3-(p-nitrophenyl)-5-phenyltetrazolium chloride ( known as INT) which has the formula:
til et kraftig farget INT formazan. Visse enzymatiske omsetninger vil dessuten forårsake reduksjon av INT til INT formazan og det er ved bestemmelsen av disse enzymer at den fargede referansestandarden i følge foreliggende oppfinnelse finner anvendelse. to a heavily colored INT formazan. Certain enzymatic reactions will also cause reduction of INT to INT formazan and it is in the determination of these enzymes that the colored reference standard according to the present invention finds application.
Løsningsmidler som har vist seg å forbedre løseligsegen-skapene av INT formazan og å tillate lett rekondisjonering med vann etter lyofilisering omfatter N,N'-dimetylformamid eller di-metylsulf oksyd. Tilsetningen av isopropylalkohol letter også denne løselighetsprosessen. Det er av vesentlig betydning at disse løsningsmidler- ikke forstyrrer absorpsjonsmaksimum av den vandige,'fargede referansestandarden. Solvents that have been shown to improve the solubility properties of INT formazan and to allow easy reconditioning with water after lyophilization include N,N'-dimethylformamide or dimethylsulfoxide. The addition of isopropyl alcohol also facilitates this solubility process. It is of essential importance that these solvents do not interfere with the absorption maximum of the aqueous, colored reference standard.
Den stabile, fargede referansestandarden i følge oppfinnelsen inneholder også serumalbumin. Det er foretrukket å an-vende bovin-serumalbumin. Dette materiale antas å hjelpe til å stabilisere både den lyofiliserte og den vandige formen av referansestandarden. The stable, colored reference standard according to the invention also contains serum albumin. It is preferred to use bovine serum albumin. This material is believed to help stabilize both the lyophilized and aqueous forms of the reference standard.
Egnede inerte svellemidler for anvendelse i referansestandarden i følge foreliggende oppfinnelse omfatter naturlige og syntetiske gummier, slik som gummi arabikum, natriumalginat, ekstrakt av knuskaragen, karboksymetylcellulose, polyvinylpyrrolidinon og lignende; sukker slik som sorbitol, mannitol, sukrose og lignende;, Suitable inert swelling agents for use in the reference standard according to the present invention include natural and synthetic gums, such as gum arabic, sodium alginate, extract of crushed carrageenan, carboxymethylcellulose, polyvinylpyrrolidinone and the like; sugars such as sorbitol, mannitol, sucrose and the like;,
og karbohydrater slik som stivelse, dekstran og lignende. Det foretrukne svellemiddel er et dekstran som har en molekylvekt på ca.10 000. and carbohydrates such as starch, dextran and the like. The preferred bulking agent is a dextran which has a molecular weight of approximately 10,000.
En foretrukket farget referansestandard kan fremstilles i følge foreliggende oppfinnelse ved å fremstille en vandig løsning, egnet for lyofilisering,som inneholder 0,005-0,015 vekt-%, basert på vekten av den totale løsningen av 1-(p-jodfenyl)-5-(p-nitrofenyl)-3-fenylformazan; fra 0,02-0,075 vekt-% bovin-serumalbumin; A preferred colored reference standard can be prepared according to the present invention by preparing an aqueous solution, suitable for lyophilization, containing 0.005-0.015% by weight, based on the weight of the total solution of 1-(p-iodophenyl)-5-(p -nitrophenyl)-3-phenylformazan; from 0.02-0.075% by weight bovine serum albumin;
fra 8-14 vekt-% .dekstran som har en molekylvekt på ca. 10 000; from 8-14% by weight dextran which has a molecular weight of approx. 10,000;
2-10 vekt-% N,N'-dimetylformamid og fra 2-10 vekt-% isopropylalkohol. 2-10% by weight of N,N'-dimethylformamide and from 2-10% by weight of isopropyl alcohol.
En særlig foretrukket fargereferansestandard i følge foreliggende oppfinnelse inneholder, i en vandig løsning, 0,0123 vekt-%, basert på vekten av hele løsningen, av 1-(p-jodfenyl)-5-(p-nitrofenyl)-3-fenylformazan; 0,563 vekt-% bovin-serum-albumin; A particularly preferred color reference standard according to the present invention contains, in an aqueous solution, 0.0123% by weight, based on the weight of the entire solution, of 1-(p-iodophenyl)-5-(p-nitrophenyl)-3-phenylformazan; 0.563% by weight bovine serum albumin;
9 vekt-% dekstran som har en molekylvekt på ca. 10 000; 2,5 vekt-% N,N<1->dimetylformamid og 7,5 vekt-% isopropylalkohol. Absorpsjonen 9% by weight dextran, which has a molecular weight of approx. 10,000; 2.5% by weight N,N<1->dimethylformamide and 7.5% by weight isopropyl alcohol. The absorption
av den vandige, fargede referansestandarden av denne løsningen ved 500 nanometer er ca. 1,34. of the aqueous colored reference standard of this solution at 500 nanometers is approx. 1.34.
Den lyofiliserte formen av den fargede referansestandardenThe lyophilized form of the colored reference standard
i følge foreliggende oppfinnelse har vist seg å være stabil i minst 6 måneder ved temperaturer fra 4°C og 24°C. Den lyofiliserte stan-darden som er rekondisjonert med vann eller 0,1 N saltsyre er lett håndterbar for fortynninger som kan være nødvendig for å konstruere standardkurver for fast-punkt-kolorimetrisk enzymatiske analyser. Rekondisjonerte, fargede referansestandard-løsninger er stabile according to the present invention has been shown to be stable for at least 6 months at temperatures between 4°C and 24°C. The lyophilized standard reconstituted with water or 0.1 N hydrochloric acid is readily amenable to dilutions that may be necessary to construct standard curves for fixed-point colorimetric enzymatic assays. Reconditioned, colored reference standard solutions are stable
i 8 timer ved væreIsestemperatur, og minst 48 timer ved 4°C.for 8 hours at room temperature, and at least 48 hours at 4°C.
For å fremstille den stabile, fargede referansestandardTo produce the stable, colored reference standard
i følge foreliggende oppfinnelse må de forskjellige bestanddeler ovenfor tilsettes på en spesiell måte for å oppnå et produkt som har den nødvendige stabilitet og løselighet som tillater rekondisjonering eller lyofilisering. Slike egenskaper oppnås først ved å oppløse INT formazan fullstendig i det valgte løsningsmiddel; deretter tilsette isopropylalkohol, under god blanding; oppløs serum-albumin i vann; og langsomt tilsette løsningen av INT formazan til serum-albuminløsningen under omrøring. Ved den foretrukne ut-førelsesform i følge foreliggende oppfinnelse, når lyofilisering er ønsket, oppløses det inerte svellemiddel i vann sammen med serum - albuminet. Den resulterende løsning som oppnås, oppdeles i like according to the present invention, the various ingredients above must be added in a special way in order to obtain a product that has the necessary stability and solubility that allows reconditioning or lyophilization. Such properties are achieved first by completely dissolving INT formazan in the chosen solvent; then add isopropyl alcohol, mixing well; dissolve serum albumin in water; and slowly adding the solution of INT formazan to the serum-albumin solution while stirring. In the preferred embodiment according to the present invention, when lyophilization is desired, the inert swelling agent is dissolved in water together with the serum albumin. The resulting solution obtained is divided equally
porsjoner og underkastes de vanlige lyofiliseringsmetoder.portions and subjected to the usual lyophilization methods.
For anvendelse ved enzymatiske bestemmelser rekondisjoneres den ovenfor angitte lyofiliserte form av den fargede referansestandarden vann eller 0,1 N saltsyre. Den fargede referansestandard-løsningen tjener som en standard av kjent virkning som ukjente analyseprøver kan avleses mot for å bestemme enzymvirkningen i den ukjente prøven. Den fargede referansestandard-løsningen i følge oppfinnelsen kan fortynnes om ønsket for å konstruere kurver for fast-punkt-kolometrisk enzymatiske analyser. For use in enzymatic determinations, the above-mentioned lyophilized form of the colored reference standard is reconditioned with water or 0.1 N hydrochloric acid. The colored reference standard solution serves as a standard of known activity against which unknown assay samples can be read to determine enzyme activity in the unknown sample. The colored reference standard solution according to the invention can be diluted if desired to construct curves for fixed-point colorimetric enzymatic analyses.
Det er således skaffet til veie en farget referansestandard som har utmerket stabilitet i lyofilisert tilstand og A colored reference standard has thus been obtained which has excellent stability in the lyophilized state and
som ved rekondisjonering er lett å håndtere ved enzymatiske bestemmelser. which during reconditioning is easy to handle by enzymatic determinations.
De følgende eksempler illustrerer oppfinnelsen:The following examples illustrate the invention:
Eksempel IExample I
Fremstilling av stabil, farget referansestandard- løsning.Preparation of stable, colored reference standard solution.
A. 12,3 mg INT formazan oppløses i 2,5 ml N,N<1->dimetyl-formamid. Til denne løsning tilsettes 7,5.ml isopropanol av analysekvalitet og blandes deretter godt. A. 12.3 mg of INT formazan is dissolved in 2.5 ml of N,N<1->dimethylformamide. 7.5 ml isopropanol of analysis quality is added to this solution and then mixed well.
B. 9,0 g dekstran som har en molvekt på ca. 10 000 og 56,3 mg bovin-serumalbumin oppløses i 90 ml renset vann. C. Løsningen A tilsettes langsomt til løsningen B ved om-røring. Den kombinerte løsningen oppdeles i like deler på 4,0 ml i 10 ml ampuller. Ampullene forsynes med hette under tørr nitrogen. Hver ampulle rekondisjoneres med 10 ml av 0,1 N saltsyre og absorpsjon ved 500 nanometer er 1,340 0,010 som utgjør 540 internasjonale enheter av laktat-dehydrogenasevirkning ved 37°C; eller 420 inter nasjonale enheter av kreatinfosfokinaseyirkning ved 30°C. B. 9.0 g of dextran which has a molecular weight of approx. 10,000 and 56.3 mg of bovine serum albumin are dissolved in 90 ml of purified water. C. Solution A is slowly added to solution B with stirring. The combined solution is divided into equal parts of 4.0 ml in 10 ml ampoules. The ampoules are supplied with a cap under dry nitrogen. Each ampoule is reconditioned with 10 ml of 0.1 N hydrochloric acid and absorbance at 500 nanometers is 1.340 0.010 which is 540 international units of lactate dehydrogenase activity at 37°C; or 420 inter national units of creatine phosphokinase activity at 30°C.
Eksempel IIExample II
Kolometrisk analyse på serumlaktat- dehydrogenase.Colometric analysis of serum lactate dehydrogenase.
Reagensene for analysen fremstilles som følger: Fargereaqens: 50 mg INT oppløses i ca. 15 ml vann under lengre tids omrøring inntil man får fullstendig oppløsning. Det tilsettes og oppløses 125 mg nikotinamid-adenin-dinukleotid, etter-fulgt av tilsetningen av 12,5 mg fenazin-metsulfat. Løsningen over-føres under vasking umiddelbart til en lavaktinisk måleflaske på The reagents for the analysis are prepared as follows: Color reagent: 50 mg INT is dissolved in approx. 15 ml of water while stirring for a long time until a complete solution is obtained. 125 mg of nicotinamide adenine dinucleotide are added and dissolved, followed by the addition of 12.5 mg of phenazine metsulfate. During washing, the solution is immediately transferred to a low-actin measuring bottle
25 ml og fortynnes med vann til merket.25 ml and dilute with water to the mark.
Pufferreaqens: 1,0 g etoksylert oleylalkohol ("Lipal 10-OA," Drew ChemialCo., Boonton, N.J.) oppløses i 10 ml vann under oppvarmnihg til ca. 95°C. Løsningen fortynnes med vann til 50 ml og tilsettes 12,1 g Tris. pH justeres til 8,2 med 3 N HCl og fortynnes deretter til 100 ml. Buffer reagent: 1.0 g of ethoxylated oleyl alcohol ("Lipal 10-OA," Drew ChemialCo., Boonton, N.J.) is dissolved in 10 ml of water while heating to approx. 95°C. The solution is diluted with water to 50 ml and 12.1 g of Tris is added. The pH is adjusted to 8.2 with 3 N HCl and then diluted to 100 ml.
Substratreaqens: (0,1 M 1(+) laktat) 5,0 ml av en 20% løsning av L(+) melkesyre tilsettes til ca. 50 ml vann. pH justeres til 5,5 med IN NaOH og fortynnes til 120 ml med vann. Løsningen mettes med noen få dråper kloroform. Substrate reagent: (0.1 M 1(+) lactate) 5.0 ml of a 20% solution of L(+) lactic acid is added to approx. 50 ml of water. The pH is adjusted to 5.5 with IN NaOH and diluted to 120 ml with water. The solution is saturated with a few drops of chloroform.
Kontrollreagens: 0,2 g kaliumoksalat go 0,2 g etylen-diamintetraeddiksyre, natriumdihydrat oppløses i 100 ml vann. Fremstillingene ovenfor er beskrevet av Babson et al. Clin Chim. Acta 12i 210-215 (1965). Control reagent: 0.2 g of potassium oxalate and 0.2 g of ethylenediaminetetraacetic acid, sodium dihydrate are dissolved in 100 ml of water. The above formulations are described by Babson et al. Clin Chim. Acta 12i 210-215 (1965).
Fremgangsmåte: 0,1 ml serum og 0,2 ml pufferreagens pipetteres i hver av glassene. 0,5 ml substrat tilsettes til Procedure: 0.1 ml of serum and 0.2 ml of buffer reagent are pipetted into each of the glasses. 0.5 ml of substrate is added
ett glass og 0,5 ml av kontrollreagensen tilsettes til det andre glasset. Begge glassene blandes og varmes til 37°C. Ved nøy-aktig fastlagte intervaller tilsettes 0,2 ml fargereagens til begge glassene og innholdene blandes og returneres umiddelbart til vann-badet. Nøyaktig 5 minutter etter tilsetningen av fargereagensen tilsettes 5 ml av 0,1 N HCl til begge glassene og innholdene blandes. Forskjellen i absorpsjon mellom kontrollglasset og glasset med serum-prøven bestemt ved 500 nm i løpet av 20 minutter er 0,67. Denne absorpsjon sammenlignes med den fargede referansestandarden i eksempel 1 som har en absorpsjon på 1,340 0,010 som er ekvalent med 540 internasjonale enheter av laktat-dehydrogenase-virkning. Laktat-dehydrogenase-virkningen i serumet beregnes til å være 270 internasjonale enheter ved 37°C. one glass and 0.5 ml of the control reagent is added to the other glass. Both glasses are mixed and heated to 37°C. At precisely determined intervals, 0.2 ml of color reagent is added to both glasses and the contents are mixed and immediately returned to the water bath. Exactly 5 minutes after the addition of the color reagent, 5 ml of 0.1 N HCl is added to both glasses and the contents are mixed. The difference in absorbance between the control glass and the glass with the serum sample determined at 500 nm over 20 minutes is 0.67. This absorbance is compared to the colored reference standard of Example 1 which has an absorbance of 1.340 0.010 which is equivalent to 540 international units of lactate dehydrogenase activity. The lactate dehydrogenase activity in the serum is calculated to be 270 international units at 37°C.
Claims (12)
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| US51226174A | 1974-10-04 | 1974-10-04 |
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| Publication Number | Publication Date |
|---|---|
| NO753268L true NO753268L (en) | 1976-04-06 |
Family
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| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| NO753268A NO753268L (en) | 1974-10-04 | 1975-09-25 |
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| Country | Link |
|---|---|
| JP (1) | JPS557239B2 (en) |
| CA (1) | CA1043677A (en) |
| CH (1) | CH617268A5 (en) |
| DE (1) | DE2537499A1 (en) |
| DK (1) | DK442775A (en) |
| FR (1) | FR2287036A1 (en) |
| GB (1) | GB1468494A (en) |
| IT (1) | IT1043092B (en) |
| NO (1) | NO753268L (en) |
| SE (1) | SE408232B (en) |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| DE3048662A1 (en) * | 1980-12-23 | 1982-07-22 | Boehringer Mannheim Gmbh, 6800 Mannheim | STABILIZED PREPARATION OF TETRAZOLIUM SALTS |
| JPS59219270A (en) * | 1983-05-30 | 1984-12-10 | Wako Pure Chem Ind Ltd | Method and reagent for stabilization of tetrazolium salt with cyclodextrin |
| CN110726833B (en) * | 2018-07-17 | 2024-01-09 | 上海瀚联诊断科技有限公司 | Blood glucose quality control liquid preparation method |
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| GB1322951A (en) * | 1969-10-29 | 1973-07-11 | Warner Lambert Co | Process and agent for the determination of glucose |
-
1975
- 1975-08-22 DE DE19752537499 patent/DE2537499A1/en not_active Withdrawn
- 1975-09-05 FR FR7527274A patent/FR2287036A1/en active Granted
- 1975-09-15 GB GB3774775A patent/GB1468494A/en not_active Expired
- 1975-09-25 NO NO753268A patent/NO753268L/no unknown
- 1975-10-01 DK DK442775A patent/DK442775A/en unknown
- 1975-10-02 JP JP11829675A patent/JPS557239B2/ja not_active Expired
- 1975-10-03 SE SE7511133A patent/SE408232B/en unknown
- 1975-10-03 CH CH1285975A patent/CH617268A5/en not_active IP Right Cessation
- 1975-10-03 IT IT27920/75A patent/IT1043092B/en active
- 1975-10-03 CA CA236,982A patent/CA1043677A/en not_active Expired
Also Published As
| Publication number | Publication date |
|---|---|
| IT1043092B (en) | 1980-02-20 |
| FR2287036A1 (en) | 1976-04-30 |
| FR2287036B1 (en) | 1978-04-07 |
| CH617268A5 (en) | 1980-05-14 |
| GB1468494A (en) | 1977-03-30 |
| DE2537499A1 (en) | 1976-04-15 |
| JPS557239B2 (en) | 1980-02-23 |
| SE7511133L (en) | 1976-04-05 |
| SE408232B (en) | 1979-05-21 |
| JPS5162091A (en) | 1976-05-29 |
| DK442775A (en) | 1976-04-05 |
| CA1043677A (en) | 1978-12-05 |
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