Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.11.1.7 (peroxidase)
 65,474 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We describe the automated microassay of plasma uric acid by use of an immobilized uricase-membrane sandwich reactor. Hydrogen peroxide, formed when uric acid is oxidized, oxidatively couples two molecules of p-hydroxyphenylacetic acid in the presence of peroxidase to produce a highly fluorescent compound. The specificity of the uricase reaction is coupled with a significantly lower cost of analysis.
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PMID:Immobilized-enzyme membrane sandwich reactor used in automated micro-scale assay of plasma uric acid. 70 42

Procedures for the determination of free and total cholesterol in lipid extracts or sonicates of 10(4) cultured human skin fibroblasts are described. The method for free cholesterol employs cholesterol oxidase to generate H2O2 and peroxidase to catalyze the reaction of H2O2 with p-hydroxyphenylacetic acid to yield a stable fluorescent product. Cholesterol ester hydrolase is included for determination of total cholesterol. When samples of sonified cell suspensions are used directly, the extraction of lipids is avoided, permitting one person to carry out analyses of 30 or more subcultures in one day.
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PMID:Procedure for determination of free and total cholesterol in micro- or nanogram amounts suitable for studies with cultured cells. 73 Nov 27

A sensitive spectrofluorimetric micromethod for the determination of uric acid is presented together with its application to human and rat plasma. The method is based on the reactions or uricase and peroxidase coupled with p-hydroxyphenylacetic acid as a fluorophor. There is a very wide range of proportionality between the concentration of uric acid and the increase of fluorescence intensity. 10 ng of uric acid is still determinable. At most 25 mul of human or 50 mul of rat plasma is sufficient to obtain the accurate valve of endogenous plasma uric acid concentration. The uric acid levels of normal human and rat plasma measured by present method were within the ranges of concentrations previously reported. A comparative study of this method with a standard assay method is also presented.
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PMID:The enzymatic spectrofluorimetric determination of uric acid in microsamples of plasma by using p-hydroxyphenylacetic acid as a fluorophor. 100 Aug 45

We describe an ultramicromethod for determination of uric acid by a manual procedure. Hydrogen peroxide, formed when uric acid is oxidized in the presence of uricase, oxidatively couples two molecules of p-hydroxyphenylacetic acid in the presence of peroxidase to produce a highly fluorescent compound. The specificity of the uricase reaction is retained, and the fluorometry results in a higher sensitivity (but a lower precision) than that obtained by applying spectrophotometric methods. As little as 50 ng of uric acid can be determined, and only 10 mul of plasma is required. Values obtained for human plasma are higher than those obtained by the spectrophotometric method.
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PMID:Ultramicromethod for determination of plasma uric acid. 127 25

Hematin can substitute for horseradish peroxidase (HRP) as the catalyst in the determination of hydrogen peroxide using phenolic substrates such as p-hydroxyphenylacetate or p-cresol. Although the peroxidatic activity of hematin from bovine blood is not as great as HRP in terms of unit iron content, the activity per unit weight is substantially greater. Hematin is 500 times less expensive than HRP per unit peroxidatic activity. In hematin-catalyzed systems, reaction development and fluorescence measurement can both be conducted optimally in the same ammoniacal buffer. Hydroxyalkyl hydroperoxides are rapidly hydrolyzed to H2O2 at this pH and are also determined. On the other hand, for methyl hydroperoxide, hematin exhibits only approximately 10% of the sensitivity exhibited by HRP. Hematin is significantly more stable in solution than HRP. The use of hematin as catalyst and p-cresol as the substrate leads to a particularly inexpensive and sensitive system, permitting a limit of detection (LOD) of 7 nM H2O2 in a flow-injection configuration.
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PMID:Hematin as a peroxidase substitute in hydrogen peroxide determinations. 157 21

We have developed a simple, reliable, and sensitive method for the assay of peroxisomal fatty acyl-CoA oxidase (FAO, EC 1.13.-) in subcellular fractions. It is based on a peroxidase-linked oxidation of 4-hydroxyphenylacetic acid to a fluorescent compound [M.S. Poosch and R.K. Yamasaki (1986) Biochim. Biophys. Acta 884, 585-593]. Our method eliminates the contribution of important interfering side reactions, notably those due to the presence of reducing agents, which function as competitive substrates to 4-hydroxyphenylacetic acid. Rapidly reacting thiol groups are of particular importance, notably CoASH present endogenously (e.g. in peroxisomes and mitochondria) or formed by enzymatic hydrolysis of acyl-CoA. Alkylation of the thiol compounds by N-ethylmaleimide eliminates this disturbing side reaction, and increases the amount of fluorescent product in the coupled peroxidatic reaction. The method is suitable for routine assay of FAO activity in a wide range of tissues, notably in those with a low specific peroxisomal beta-oxidation activity and/or a high content of reducing agents. As an example of this we have included data from rat heart peroxisomal fractions. The effect of alkylation of sulfhydryl groups in the incubation mixture also applies to other oxidase reactions based on H2O2-coupled peroxidatic reactions, if the oxidase itself does not contain functional sulfhydryl groups.
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PMID:A fluorometric assay of acyl-CoA oxidase activity by a coupled peroxidatic reaction: elimination of interfering side reactions. 165 7

A selective and sensitive assay of inosine, guanosine, hypoxanthine, guanine and xanthine by high-performance liquid chromatography with immobilized enzyme reactors was developed. The separation was achieved on a Capcell Pak C18 column (15 cm x 0.46 cm I.D.) with a mobile phase of 0.1 M phosphate buffer (pH 8.0) containing 7 mM sodium 1-hexanesulphonate and 0.1 mM p-hydroxyphenylacetic acid. The fluorimetric detection of hydrogen peroxide using immobilized peroxidase and p-hydroxyphenylacetic acid was applied to the assay of these compounds, which were oxidized to yield hydrogen peroxide in the presence of immobilized enzyme (purine nucleoside phosphorylase, guanase and xanthine oxidase). Enzyme reactions occurred sufficiently without post-column addition of reagents. Enzymes that catalysed the conversion of purine compounds were co-immobilized on aminopropyl controlled-pore glass packed in stainless-steel tubing. The detection limits were 30-200 pg per injection.
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PMID:Determination of purine nucleosides and their bases by high-performance liquid chromatography using co-immobilized enzyme reactors. 211 20

Stimulated neutrophils (PMNs) produce large quantities of superoxide anion, which is the precursor for hydrogen peroxide (H2O2). We developed a new fluorimetric assay to measure the H2O2 released by zymosan A-activated PMNs utilizing the oxidation of p-hydroxyphenylacetic acid by H2O2 to its fluorescent dimer in the presence of horseradish peroxidase. Zymosan-activated PMNs isolated from nine healthy volunteers and 20 patients with acute hypoxemic respiratory failure (AHRF) released after 90 min 2.3 +/- 0.3 and 2.4 +/- 1.3 nmol H2O2/10(6) PMNs, respectively. Inhibition of the heme enzymes by 1.0 mM sodium azide (NaN3) increased the H2O2 production to 21.6 +/- 4.4 nmol H2O2/10(6) PMNs in the control group (P less than 0.001), and to 22.5 +/- 14.7 nmol H2O2/10(6) PMNs in patients with AHRF (P less than 0.001). Incubation temperature, room temperature or 37 degrees C, did not change the total amount of H2O2 produced after 90 min by zymosan-activated PMN. Addition of NaN3 improved both the sensitivity and reproducibility of the measurement of H2O2 and allowed detection of H2O2 released by PMNs with coefficients of variation of less than 5% at PMN concentrations as low as 0.1 x 10(6) cells/ml. The amount of H2O2 released by activated PMNs did not distinguish healthy controls from patients with AHRF.
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PMID:Effect of sodium azide on hydrogen peroxide production by zymosan-activated human neutrophils. 216 67

A selective and sensitive assay of substrates (hypoxanthine, xanthine and allopurinol) of xanthine oxidase by reversed-phase liquid chromatography coupled with the use of immobilized enzyme reactors is described. These compounds were oxidized by immobilized xanthine oxidase and produced hydrogen peroxide, which was determined fluorometrically using immobilized peroxidase and p-hydroxyphenylacetic acid. The detection limits of hypoxanthine, xanthine and allopurinol were approximately 50, 120 and 130 pg per injection, respectively. Immobilized xanthine oxidase inhibited by oxipurinol during the assay was reactivated by 2,6-dichlorophenolindophenol and could be used for a long period without a significant activity loss. These methods were applied to plasma and urine samples.
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PMID:Simultaneous assay of hypoxanthine, xanthine and allopurinol by high-performance liquid chromatography and activation of immobilized xanthine oxidase as an enzyme reactor. 260 92

A sensitive fluorogenic assay for acetylcholine is described. The assay is based upon the reactions: (1) hydrolysis of acetylcholine to choline and acetate, catalyzed by acetylcholinesterase. (2) Oxidation of choline to betaine and hydrogen peroxide by choline oxidase. (3) Oxidation of p-hydroxyphenylacetic acid by hydrogen peroxide to a fluorescent product, catalyzed by peroxidase. With a sensitivity comparable to chemiluminescence procedures, the assay should find application to those situations requiring the persistence of a fluorescence signal or the necessity of assaying small volumes.
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PMID:A fluorometric assay for acetylcholine with picomole sensitivity. 276 Dec 99


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