title: Developing SyrinOX total antioxidant capacity assay for measuring antioxidants in humans.
abstract: Accurate monitoring of the antioxidant status or of oxidative stress in patients is still a big challenge in clinical laboratories. This study investigates the possibility of applying a newly developed total antioxidant capacity assay method based on laccase or peroxidase oxidized syringaldazine [Tetramethoxy azobismethylene quinone (TMAMQ)] which is referred to here as SyrinOX, as a diagnostic tool for monitoring both oxidative stress and antioxidant status in patients. Attempts to adapt the Randox total antioxidant procedure [simultaneous incubation of the radical generating system (metmyoglobin and H(2) O(2) ) and antioxidant sample] for SyrinOX were abandoned after it was discovered that the H(2) O(2) reacted with enzymatically generated TMAMQ and ABTS radicals at a rate of 6.4 × 10(-2) /μM/s and 5.7 × 10(-3) /μM/s respectively. Thus this study for the first time demonstrates the negative effects of H(2) O(2) in the Randox system. This leads to erroneous results because the total antioxidant values obtained are the sum of radicals reduced by antioxidants plus those reacting with the radical generating system. Therefore they should be avoided not only for this particular method but also when using other similar methods. Consequently, SyrinOX is best applied using a three-step approach involving, production of TMAMQ, recovery and purification (free from enzyme and other impurities) and then using TMAMQ for measuring the total antioxidant capacity of samples. Using this approach, the reaction conditions for application of SyrinOX when measuring the total antioxidant capacity of plasma sample were determined to be 50% (v/v) ethanol/50 mM sodium succinate buffer pH 5.5, between 20 and 25 °C for at least 1 h.
authors.affiliation: Institute of Environmental Biotechnology, Graz University of Technology, Graz, Austria.
authors.forename: Endry N
authors.initials: EN
authors.lastname: Prasetyo
authors.forename: Otto
authors.initials: O
authors.lastname: Knes
authors.forename: Gibson S
authors.initials: GS
authors.lastname: Nyanhongo
authors.forename: Georg M
authors.initials: GM
authors.lastname: Guebitz
mesh_terms: D000975:Antioxidants / Q000032:analysis* / Q000494:pharmacology; D052160:Benzothiazoles; D001681:Biological Assay / Q000379:methods* / Q000592:standards; D015415:Biomarkers / Q000097:blood; D002021:Buffers; D002138:Calibration; D002839:Chromans / Q000494:pharmacology; D000431:Ethanol / Q000737:chemistry; D006801:Humans; D006835:Hydrazones / Q000737:chemistry*; D006861:Hydrogen Peroxide / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D007202:Indicators and Reagents / Q000737:chemistry*; D007700:Kinetics; D008786:Metmyoglobin / Q000737:chemistry; D010084:Oxidation-Reduction; D018384:Oxidative Stress / Q000187:drug effects; D011237:Predictive Value of Tests; D012015:Reference Standards; D015203:Reproducibility of Results; D012997:Solvents / Q000737:chemistry; D013057:Spectrum Analysis; D019802:Succinic Acid / Q000737:chemistry; D013451:Sulfonic Acids / Q000737:chemistry; D013696:Temperature; D013844:Thiazoles / Q000737:chemistry
chemical_list: D000975:Antioxidants; D052160:Benzothiazoles; D015415:Biomarkers; D002021:Buffers; D002839:Chromans; D006835:Hydrazones; D007202:Indicators and Reagents; D012997:Solvents; D013451:Sulfonic Acids; D013844:Thiazoles; D008786:Metmyoglobin; C009370:syringaldazine; C002502:2,2'-azino-di-(3-ethylbenzothiazoline)-6-sulfonic acid; D000431:Ethanol; D019802:Succinic Acid; D006861:Hydrogen Peroxide; C010643:6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid