Glutathione peroxidase assay

Se-selenite into peritoneal exudate cells of mice separation and glutathione peroxidase assay of adherent and non-adherent cells. Selenium in Biology and Medicine,... 

Isolation, Purification, Characterization, and Assay of Antioxy-genic Enzymes Isolation and characterization of superoxide dis-mutase, 105, 88 superoxide dismutase assays, 105, 93 assays of glutathione peroxidase, 105, 114 catalase in vitro, 105, 121 assays of lipoxygenase, 105, 126. 

Glutathione-peroxidase activity was measured spectrophoto-metrically at 340nm by an enzyme coupled assay procedure of Paglia and Valentine (28) as modified by Reddy, et al. (26). A molar extinction coefficient of 6.2 x 103cm 1 was used in calculations. 

Finally, if the product of the enzymatic reaction is known, the activity of the enzyme can be assayed on the basis of quantitating the product off line. Thus, for instance, glutathione peroxidase activity can be measured by quantitating the oxidized and reduced forms of glutathione by capillary electrophoresis, as demonstrated by Pascual et al. (1992). The electrophoretic separation buffer used was 100 mM tetraborate (pH 8.2), containing 100 m M SDS. 

Pascual P, Martinez-Lara E, B4rcena JA, Lopez-Barea J, Toribio F (1992) Direct assay of glutathione peroxidase using high-performance capillary electrophoresis. J Chromatogr 581 49-56. 

Early animal studies and human population surveys used whole blood as the main indicator of selenium status. Whole blood selenium can be determined after acid digestion using a fluorometric method. The more convenient carbon furnace atomic absorption spectroscopy (CFAAS) assay for plasma and/or serum selenium is now the most widely used procedure. The main components of plasma selenium are extracellular glutathione peroxidase (GSHPx 3) and selenoprotein P. 

Glutathione peroxidases (Se-dependent and -independent enzymes) activities toward cumene hydroperoxide and H2O2 as substrates were determined by the coupled enzyme method of Lawrence and Burk (1976) with minor modifications. The standard assay contained 100 mM sodium phosphate buffer (pH 7.5), 1 mM EDTA, 0.12 mM NADPH, 1 mM NaN3 (for H2O2 assay), 2 mM GSH, 0.8 mM cumene hydroperoxide or 0.4 mM H2O2, 1 unit glutathione reductase and sample. The oxidation of NADPH was followed at 340 nm e — —6.22 mM cm ). 

Eugenol and methyl eugenol are the main compounds of the essential oil of Ocimum sanctum L., which are able to decrease serum lipids. 0. sanctum leaves oil is used for the measurement of the antioxidative and antihyperlipidamic properties in rats, which were on cholesterol diet. Lipid peroxide concentration was assessed in TABRS assay, and it was found for this essential oil to decrease TBA levels in heart and liver. In heart tissue, it is also shown for the oil to exert a decreasing effect on SOD and glutathione peroxidase (GPx) without effecting CAT. On the other hand GPx, SOD, and CAT were not impacted in liver tissue (Suanarunsawat et al., 2010). 

When rats are administered with bacoside A (10 mg/kg), levels of glutathimie, vitamin C, vitamin E, and vitamin A were reduced. The activities of superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase were also assayed. Copper, iron, zinc, and selenium levels in brain and serum ceruloplasntin activity were also measured. Administration of bacoside A improved the antioxidant status and maintained the levels of trace elements [16]. 

Peroxide assays - peroxide measurement Iodine formation - tihation, ferrous oxidation xylenol orange (FOX) in vitro assay for LDL peroxidation, glutathione peroxidase (GPX) specific for fatty acid peroxides. Cyclooxygenase (COX) used to measure trace peroxides in biological fluids... 

Another endogenous HNO source relies on the oxidation of hydroxylamine (HA), or other alcohol amine, such as hydroxyurea or A -hydroxy arginine. In vivo, such a process is postulated to depend on the activity of several heme proteins, which are able to stabilize oxo ferryl species (compound I and compound II), such as peroxidases. Recently, Donzelli et al. evaluated HNO production by this mechanism (22), with a newly developed selective assay in which the reaction products, GS(0)NH, in the presence of reduced glutathione (GSH) are quantified by HPLC. Their results showed that metmyoglobin, horse radish peroxidase, and myeloperoxidase were efficient HNO producers using hydroxylamine as substrate. However, there are several remaining unresolved questions concerning the proposed mechanism (which is outlined below, Eq. (2)). 

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