Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.10.3.3 (ascorbate oxidase)
 778 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The chemical diversity of antioxidants makes it difficult to separate and quantify antioxidants from the vegetable matrix. Therefore, it is desirable to establish a method that can measure the total antioxidant activity level directly from vegetable extracts. The current literature clearly states that there is no "total antioxidant" as a nutritional index available for food labeling because of the lack of standard quantitation methods. Thus, this work reports the development of a simple, widely applicable antioxidant capacity index for dietary polyphenols and vitamins C and E, utilizing the copper(II)-neocuproine [Cu(II)-Nc] reagent as the chromogenic oxidizing agent. Because the copper(II) (or cupric) ion reducing ability of polyphenols is measured, the method is named by our research group "cupric reducing antioxidant capacity" abbreviated as the CUPRAC method. This method should be advantageous over the ferric reducing antioxidant power (FRAP) method because the redox chemistry of copper(II)-as opposed to that of ferric ion-involves faster kinetics. The method comprises mixing of the antioxidant solution (directly or after acid hydrolysis) with a copper(II) chloride solution, a neocuproine alcoholic solution, and an ammonium acetate aqueous buffer at pH 7 and subsequent measurement of the developed absorbance at 450 nm after 30 min. Because the color development is fast for compounds such as ascorbic acid, gallic acid, and quercetin but slow for naringin and naringenin, the latter compounds were assayed after incubation at 50 degrees C on a water bath for 20 min [after Cu(II)-Nc reagent addition] so as to force the oxidation reaction to reach completion. The flavonoid glycosides were hydrolyzed to their corresponding aglycons by refluxing in 1.2 M HCl-containing 50% MeOH so as to exert maximal reducing power toward Cu(II)-Nc. Certain compounds also needed incubation after acid hydrolysis to fully exhibit their reducing capability. The CUPRAC antioxidant capacities of synthetic mixtures of antioxidants were experimentally measured as Trolox equivalents and compared to those theoretically found by making use of the principle of additivity of absorbances assuming no chemical interaction between the mixture constituents. Because ascorbic acid is not resistant to elevated temperature incubation, it should be assayed initially by measuring the absorbance (at 450 nm) difference of original and ascorbate oxidase-added mixture solutions at the end of 1 min of Cu(II)-Nc reagent addition. Thus, the total CUPRAC antioxidant capacity of a mixture containing various antioxidants should be that finally measured after a suitable combination of hydrolysis and incubation procedures, added to the initially measured capacity due to ascorbate. The antioxidant polyphenolic compounds tested demonstrate that the highest capacities in the CUPRAC method were observed for epicatechin gallate, epigallocatechin gallate, quercetin, fisetin, epigallocatechin, catechin, and caffeic acid in this order, in accordance with theoretical expectations, because the number and position of the hydroxyl groups as well as the degree of conjugation of the whole molecule are important. The antioxidant potency of flavonoids is nearly proportional to the total number of -OH groups and is positively affected by the presence of an o-dihydroxy moiety in the B-ring. beta-Carotene, which did not react with the CUPRAC reagent in alcoholic aqueous medium, could be assayed in dichloromethane solvent. Linear calibration curves for ascorbic acid and flavonoids were redrawn in synthetic solutions containing a mixture of antioxidants, and also in real matrices such as grape and orange juices, green tea, and blackberry tea, showing an initial nonzero absorbance with the CUPRAC reagent. The parallellism of the linear calibration curves of pure compounds in a given complex matrix effectively demonstrated that there were no interferent chemical interactions among the solution constituents and that the antioxidant capacities of the tested antioxidants were additive. The CUPRAC reagent is reasonably selective, stable, easily accessible, and sensitive toward thiol-type oxidants, unlike the FRAP method. The reaction is carried out at nearly physiological pH as opposed to the unrealistic acidic pH of FRAP.
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PMID:Novel total antioxidant capacity index for dietary polyphenols and vitamins C and E, using their cupric ion reducing capability in the presence of neocuproine: CUPRAC method. 1561 84

The objective of this study was to establish a hepatic lipase (HL) assay method that can be applied to automatic clinical analyzers. Seventy-four hyperlipidemic subjects (men/women 45/29) were recruited. Lipase activity was assayed measuring the increase in absorbance at 546 nm due to quinonediimine dye production. Reaction mixture R-1 contained 50 mM Tris-HCl (pH 9.5), 0.5 mM glycerol-1,2-dioleate, 0.4% (unless otherwise noted) polyoxyethylene-nonylphenylether, 3 mM ATP, 3 mM MgCl(2), 1.5 mM CaCl(2), monoacylglycerol-specific lipase, glycerol kinase, glycerol-3-phosphate oxidase, 0.075% N,N-bis-(4-sulfobutyl)-3-methylaniline-2 Na, peroxidase, ascorbic acid oxidase. Reaction mixture R-2 contained 50 mM Tris-HCl (pH9.5), 0.15% 4-aminoantypirine. Automated assay for activity was performed with a Model 7080 Hitachi analyzer. In the lipase assay, 160 microl of R-1 was incubated at 37 degrees C with 3 microl of samples for 5 min, and 80 microl of R-2 was added. Within-run coefficient of variations was 0.9-1.0%. Calibration curve of lipase activity was linear (r = 0.999) between 0 and 320 U/l. Analytical recoveries of purified HL added to plasma were 96.6-99.8%. HL activity in postheparin plasma measured in this method had a closer correlation with HL mass by a sandwich ELISA (r = 0.888, P < 0.0001) than those in the conventional method using [(14)C-]triolein (r = 0.730, P < 0.0001). This assay method for HL activity can be applied to an automatic clinical analyzer.
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PMID:A novel method for measuring human hepatic lipase activity in postheparin plasma. 1709 Jun 61

The objective of this study was to establish a new lipoprotein lipase (LPL) and hepatic lipase (HL) activity assay method. Seventy normal volunteers were recruited. Lipase activities were assayed by measuring the increase in absorbance at 546 nm due to the quinoneine dye. Reaction mixture-1 (R-1) contained dioleoylglycerol solubilized with lauryldimethylaminobetaine, monoacylglycerol-specific lipase, glycerolkinase, glycerol-3-phosphate oxidase, peroxidase, ascorbic acid oxidase, and apolipoprotein C-II (apoC-II). R-2 contained Tris-HCl (pH 8.7) and 4-aminoantipyrine. Automated assay of lipase activities was performed with an automatic clinical analyzer. In the assay for HL + LPL activity, 160 microl R-1 was incubated at 37 degrees C with 2 microl of sample for 5 min, and 80 microl R-2 was added. HL activities were measured under the same conditions without apoC-II. HL and LPL activities were also measured by the conventional isotope method and for HL mass by ELISA. Lipase activity detected in a 1.6 M NaCl-eluted fraction from a heparin-Sepharose column was enhanced by adding purified apoC-II in a dose-dependent manner, whereas that eluted by 0.8 M NaCl was not. Postheparin plasma-LPL and HL activities measured in the present automated method had high correlations with those measured by conventional activity and mass methods. This automated assay method for LPL and HL activities is simple and reliable and can be applied to an automatic clinical analyzer.
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PMID:A novel method for measuring human lipoprotein lipase and hepatic lipase activities in postheparin plasma. 1834 10