In vivo total antioxidant capacity: comparison of two different analytical methods

Abstract

Several methods to assess the total antioxidant capacity (TAC) are available. However, the final value of measured TAC in the sample depends on the procedure used in every specific assay. This makes crucial the comparison of different analytical methods. The aim of our study was to evaluate analytical characteristics and laboratory reliability of two different assays: the ferric-reducing ability (FRAP) assay and a new spectrophotometric test (OXY-adsorbent test, Diacron, Italy). Unselected outpatients referred to the Institute of Clinical Physiology were studied (n = 187, 58 females, 129 males, mean age: 65 +/- 13 years). All blood samples were maintained on ice, centrifuged within 15 minutes after blood collection and then stored at -80 degrees C until performance of assay procedures. OXY assay: The lower limit of sensitivity was 6 micromol HClO/ml. The assay was found to be linear up to 440 micromol HClO/ml (r = -0.99, p < 0.001). Absorbance was linear over a wide concentration range with solutions containing uric acid in purified form (0-1000 micromol/l, r = -0.996, p < 0.001), serum (r = -0.99, p < 0.01) or plasma serially diluted (r = -0.99, p < 0.01). Mean value in plasma samples accounted for 366.2 +/- 7.2 micromol HClO/ml. Mean OXY value in females (353.4 +/- 13.2 micromol HClO/ml) was not different from that detected in males (372 +/- 8.6 micromol HClO/ml). A significant difference was observed between subjects without and with hypertension in serum OXY levels (344.8 +/- 9.9 and 383.2 +/- 10 micromol HClO/ml, p < 0.01, respectively). FRAP assay: The lower limit of sensitivity was 15 micromol/l. Linearity was observed up to 1000 micromol/l (r = 0.998, p < 0.001). Absorbance was linear over a wide concentration range with solutions containing uric acid in purified form (0-1000 micromol/l, r = 0.997, p < 0.001), serum (r = 0.99, p < 0.01) or plasma serially diluted (r = 0.99, p < 0.01). FRAP mean value in plasma samples, evaluated in 102 patients, accounted for 514.1 +/- 19.1 micromol/l. Mean FRAP in females (469 +/- 22.5 micromol/l) was not different from that detected in males (535 +/- 25.6 micromol/l). FRAP vs. OXY: A significant direct relationship was observed when comparing FRAP with OXY levels in the whole population (r = 0.22, p < 0.05). Neither of the methods are expensive and they are speedy and simple to perform. Values are reproducible and linearly correlated to the concentration of antioxidants present in the samples. For this reason, these methods may be considered practicable indicators of total antioxidant capacity, for routinely potential use in every laboratory and useful in all the studies concerning the evaluation of oxidative stress.

Elevated Levels of Oxidative Stress as a Prognostic Predictor of Major Adverse Cardiovascular Events in Patients with Coronary Artery Disease

“…In particular, we and others have demonstrated that isoprostanes, markers of lipid peroxidation and reduced antioxidant capacity, are related to an increased risk for cardiovascular disease and correlated with the number of cardiovascular risk factors 5,6) . Other evidence indicates that thiobarbituric acid reactive substance (TBARS, indicators of lipid peroxidation) levels are correlated with endothelial dysfunction and coronary artery disease 11,12) .…”

“…Other evidence indicates that thiobarbituric acid reactive substance (TBARS, indicators of lipid peroxidation) levels are correlated with endothelial dysfunction and coronary artery disease 11,12) . Specifically, our previous data indicated associations between d-ROMs, OXY-adsorbent and Oxidative-INDEX and the presence and severity of CAD, and the relationship between these biomarkers of oxidative stress and the extent and number of risk factors, including the presence of diabetes, dyslipidemia, hypertension, and smoking habit, and with inflammatory biomarkers 5,6,[8][9][10][11] . Interestingly, a significant inverse correlation between d-ROMs and OXY-adsorbent tests (r =−0.3, p ≤ 0.01) was also observed in our CAD patient cohort.…”

“…This assay is based on the endogenous antioxidant capacity to oppose the oxidant action of added hypochlorous acid (HClO). Specifically, the effectiveness of the anti-oxidant barrier against free radicals is estimated by simulating a massive attack by HClO, added in excess to the system 5) . After 10 min, residual HClO undergoes mean±SD) with angiographically proven CAD, admitted to the Coronary Care Unit of the Clinical Physiology Institute-CNR in Pisa.…”

“…For this purpose, reactive oxygen metabolites and the total antioxidant status were assessed by commercially available assays (d-ROMs and OXY Adsorbent Test, respectively; Diacron, Grosseto, GR, Italy) 4,5) . The global oxidative stress index (Oxidative-INDEX), including both the oxidative and antioxidant couterparts, and therefore potentially more complete, was calculated 6) .…”

Oxidative stress and its association with coronary artery disease and different atherogenic risk factors

“…The shift in the antioxidant/prooxidant equilibrium in favour of oxidative stress represents a crucial determinant in aging and other chronic and degenerative diseases [8][9][10]. Therefore, additional information may be revealed by the evaluation of the overall antioxidant capacity together with index of free radical generation [11][12].…”