Effects of Free Radicals on the Fluidity of Myocardial Membranes

Free radicals, including superoxide anions (O₂^?_?), hydroxyl radical (HO'), and hypohalite radical (OCl'), as well as oxidants such as hydrogen peroxide (H₂O₂) and hypochlorous acid (HOCl), have been indicated in the pathogenesis of myocardial ischemic and reperfusion injury. In this report, we compared the integrity of the myocardial membrane when exposed to these free radicals/oxidants. Isolated rat heart membrane preparations were exposed to chemically generated free radicals with or without their respective scavengers. Membrane fluidity was monitored by fluorescence polarization using the diphenylhexatriene probe, as well as by electron spin resonance (ESR) spectroscopy using 2,2,6,6-tetramethyl piperidine-n-oxyl as the spin labeling agent. HO', H₂O₂, and OCl' + HOCl increased the fluorescence polarization (FP) and microvis-cosity significantly by 1.7-fold, 1.8-fold, and 1.7-fold, respectively, as compared to an only 1.2– fold increase in FP by O₂^?_? O₂^?_? did not alter the fatty acid profiles of the membrane phospholipids. However, HO' and H₂O₂ reduced the arachidonic acid contents in phosphatidylcholine (PC), phosphatidylethanolamine (PE), and phosphatidylinositol (PI). These radicals also stimulated the lipid peroxidation by several-fold, while that by O₂^?_? was only insignificant. These results suggest that HO' and H₂O₂ decreased the membrane fluidity and induced lipid peroxidation by releasing the arachidonic acid from PC, PE. and PI.     

Reduced free radical generation during reperfusion of hypothermically arrested hearts

Several studies indicate the presence of hydroxyl radical (OH·) as well as its involvement in the myocardial reperfusion injury. A transition metal-like iron is necessary for the conversion of superoxide anion (O₂ ^–) to a highly reactive and cytotoxic hydroxyl radical (OH·). In the present study, we have examined the generation of OH· and free iron in reperfused hearts following either normothermic (37°C) or hypothermic ischemia (5°C). Employing the Langendorff technique, isolated rat hearts were subjected to global ischemia for 30 min at 37°C or 5°C and were then reperfused for 15 min at 37°C. The results of the study suggest that both the OH· generation in myocardium and free iron release into perfusate were significantly lower in hearts made ischemic at 5°C as compared to 37°C. Release of myoglobin and lactic acid dehydrogenase into perfusate also followed a similar pattern. Furthermore, in in vitro studies, chemically generated O₂ ^– at 5°C caused a significantly lower rate of oxidation of oxymyoglobin as well as generation of OH° and free iron as compared to 37°C. These results suggest that (1) reperfusion of hypothermic ischemic heart is associated with a reduction in the generation of OH· and cellular damage compared to that of normothermic ischemic heart, and (2) myoglobin, an intracellular protein, is a source of free iron and plays a role in the reperfusion injury mediated by free radicals.Abbreviations OH· hydroxyl radical - O₂ ^– superoxide anion - ODFR oxygen-derived free radicals - KHB Krebs-Henseleit buffer - LDH lactate hydrogenase - SOD superoxide dismutase     

Modulation of fatty acid-binding capacity of heart fatty acid-binding protein by oxygen — derived free radicals

Summary In this study, we examined the effects of exposure of heart fatty acid-binding protein (h-FABP) to chemically generated O₂ ^– or OH · with respect to its oleate binding and to its electrophoretic properties. Purified rat h-FABP at 40 M scavenged as much as 30% O₂ ^– and 85% OH ·. On the other hand, when 2 nmol (4 M) FABP were exposed to free radicals, the maximum oleate binding capacity as measured by Scatchard analysis was reduced only by 14% and 27% for O₂ ^– and OH ·, respectively. The electrophoretic pattern of free radical-exposed FABP was not markedly different when examined either by the non-denaturing or by denaturing PAGE, suggesting the absence of any degradation or aggregation of FABP by O₂ ^– or OH ·. It is hypothesized that O₂ ^– or OH · in physiological concentration may not alter the function of FABP markedly in the ischemic-reperfused myocardium.Abbreviations h-FABP Heart Fatty Acid Binding Protein - NEFA Non-Esterified Fatty Acids - O₂ ^– Superoxide anions - OH· hydroxyl radicals - OCI hypohalite radicals - H₂O₂ hydrogen peroxide - HPLC High Pressure Liquid Chromatography     

Significance of cytoplasmic fatty acid-binding protein for the ischemic heart

Ischemia of the heart is accompanied by the tissue accumulation of long-chain fatty acids and their metabolic derivatives such as -hydroxy fatty acids and fatty acyl-CoA and acyl-L-carnitine esters. These substances might be detrimental for proper myocardial function. Previously, it has been suggested that intracellular lipid binding proteins like cytoplasmic fatty acid-binding protein (FABP) and acyl-CoA binding protein (ACBP) may bind these accumulating fatty acyl moieties to prevent their elevated levels from potentially harmful actions. In addition, the suggestion has been made that the abundantly present FABP may scavenge free radicals which are generated during reperfusion of the ischemic heart. However, these protective actions are challenged by the continuous physico-chemical partition of fatty acyl moieties between FABP and membrane structures and by the rapid release of FABP from ischemic and reperfused cardiac muscle. Careful evaluation of the available literature data reveals that at present no definite conclusion can be drawn about the potential protective effect of FABP on the ischemic and reperfused heart. Biochem123: 167–173, 1993)Abbreviations FABP Fatty Acid-Binding Protein - ACBP Acyl-CoA Binding Protein - MDGI Mammary-Derived Growth Inhibitor - CK Creatine Kinase - LDH Lactate Dehydrogenase     

Effect of Oxygen-Derived Free Radicals and Oxidants on the Degradation in vitro of Membrane Phospholipids

The abilities of chemically generated hydroxyl radical (OH), superoxide anion (O^?) and hydrogen peroxide (H₂O₂) to degrade rat myocardial membrane phospholipids previously lableed with [1 -¹⁴C]arachidonic acid were studied. HO and H₂O₂ but not O₂^?_?, caused the degradation of phospha-tidylcholine (PC), phosphatidylethanolamine (PE), and phosphatidylinositol (PI). With OH' and H₂O₂, the loss of radiolable in PC was accompanied by an increase in the radiolabel of lysophosphatidylcholine (LPC), but not in that of free fatty acid (FFA). These results suggest the hydrolysis of l-oxygen ester bond of PC by HO' and that H₂O₂ and that HO' and H₂O₂, but not O^?, are detrimental to the structure and function of membrane phospholipids. However, since μM amounts of HO' and mM amounts of H₂O₂ were necessary to affect the membrane phospholipids, it is likely that in the reprefused myocardium only HO', but not H₂O₂, may directly cause the breakdown of membrane phospholipids.     

Formation of superoxide anion and carbon-centered radicals by photosystem II under high light and heat stress—EPR spin-trapping study

In this study, electron paramagnetic resonance spin-trapping spectroscopy was used to study the light-induced production of superoxide anion (O₂ ^?-) and carbon-centered (R^?) radicals by Photosystem II (PSII). It is evidenced here that exposure of PSII membranes to high light (2,000 μmol photons m^?2 s^?1) or heat (47 °C) treatments prior to the illumination suppressed O₂ ^?- production, while R^? was formed. Formation of R^? in the both high light- and heat-treated PSII membranes was enhanced by DCMU. Removal of molecular oxygen by glucose/glucose oxidase/catalase system and O₂ ^?- scavenging by exogenous superoxide dismutase completely suppressed carbon-centered radical formation. It is proposed here that the oxidation of polyunsaturated fatty acids and amino acids by O₂ ^?- on the electron acceptor side of PSII results in the formation of R^?, known to initiate a cascade reaction leading to the lipid peroxidation and protein degradation, respectively.     

Superoxide-dependent formation of hydroxyl radicals: Detection of hydroxyl radicals by the hydroxylation of aromatic compounds

A mixture of xanthine or hypoxanthine and xanthine oxidase generates the superoxide radical, O₂^$\text{?}$, and H₂O₂. In the presence of iron salts, O₂^$\text{?}$ and H₂O₂ can interact to produce the hydroxyl radical, OH·. Superoxide-dependent formation of OH· can be measured by its ability to hydroxylate salicylate as followed by an improved colorimetric assay described in this paper. A more accurate analysis of OH· can be obtained using its ability to hydroxylate phenol, the hydroxylated products being separated and measured after derivatization using gas-liquid chromatography and electron-capture detection. The derivatization and separation techniques are described.     

Unique antioxidant effects of herbal leaf tea and stem tea from Moringa oleifera L. especially on superoxide anion radical generation systems

ABSTRACT

This study aimed to investigate the unique antioxidative effects of Japanese moringa products, herbal leaf tea and stem tea, using established free radical assays, focusing on superoxide anion (O₂^?) radical generation systems. Hot-water extracts from moringa teas resulted in different but lower scavenging activities than Trolox in four synthetic free radical models. Interestingly, these extracts further showed higher O₂^? radical scavenging effects than Trolox in the phenazine methosulfate-NADH-nitroblue tetrazolium and xanthine oxidase assay systems. Incubating human neutrophils in the presence of these tea extracts rather than Trolox effectively suppressed cellular O₂^? radical generation. Among the eight known phenolic constituents of moringa leaves, caffeic acid and chlorogenic acid may be responsible for the O₂^–specific radical scavenging capacity stronger than that of Trolox. These results suggest that moringa herbal teas are a good source of natural antioxidants for preventing O₂^? radical-mediated disorders.

Abbreviations: O₂^?: superoxide anion; ROS: reactive oxygen species; H₂O₂: hydrogen peroxide; XOD: xanthine oxidase; DPPH: 1,1-diphenyl-2-picrylhydrazyl; ABTS⁺: 2,2′-azinobis(2-ethylbenzothiazoline-6-sulfonic acid) cation; CPZ⁺: chlorpromazine cation; PMS: phenazine methosulfate; NBT: nitroblue tetrazolium; PMA: phorbol 12-myristate 13-acetate     

Hydroxyl radical production from hydrogen peroxide and enzymatically generated paraquat radicals: Catalytic requirements and oxygen dependence

The ability of paraquat radicals (PQ^+.) generated by xanthine oxidase and glutathione reductase to give H₂O₂-dependent hydroxyl radical production was investigated. Under anaerobic conditions, paraquat radicals from each source caused chain oxidation of formate to CO₂, and oxidation of deoxyribose to thiobarbituric acid-reactive products that was inhibited by hydroxyl radical scavengers. This is in accordance with the following mechanism derived for radicals generated by γ-irradiation [H. C. Sutton and C. C. Winterbourn (1984) Arch. Biochem. Biophys.235, 106–115] PQ^+. + Fe³⁺ (chelate) → Fe²⁺ (chelate) + PQ⁺⁺ H₂O₂ + Fe²⁺ (chelate) → Fe³⁺ (chelate) + OH^? + OH.. Iron-(EDTA) and iron-(diethylenetriaminepentaacetic acid) (DTPA) were good catalysts of the reaction; iron complexed with desferrioxamine or transferrin was not. Extremely low concentrations of iron (0.03 μm) gave near-maximum yields of hydroxyl radicals. In the absence of added chelator, no formate oxidation occurred. Paraquat radicals generated from xanthine oxidase (but not by the other methods) caused H₂O₂-dependent deoxyribose oxidation. However, inhibition by scavengers was much less than expected for a reaction of hydroxyl radicals, and this deoxyribose oxidation with xanthine oxidase does not appear to be mediated by free hydroxyl radicals. With O₂ present, no hydroxyl radical production from H₂O₂ and paraquat radicals generated by radiation was detected. However, with paraquat radicals continuously generated by either enzyme, oxidation of both formate and deoxyribose was measured. Product yields decreased with increasing O₂ concentration and increased with increasing iron(DTPA). These results imply a major difference in reactivity between free and enzymatically generated paraquat radicals, and suggest that the latter could react as an enzyme-paraquat radical complex, for which the relative rate of reaction with Fe³⁺ (chelate) compared with O₂ is greater than is the case with free paraquat radicals.     

Antioxidant activities of dihydroxylated salicylic acid derivatives

SUMMARY

The ability of hydroxylated metabolites of salicylic acid to scavenge reactive oxygen species and to inhibit arachidonic acid metabolism was investigated. The tested trihydroxybenzoic acids (THBAs) were potent scavengers of hydroxyl and superoxide anion radicals produced by Fenton reaction and xanthine/xanthine oxidase system or activated macrophages respectively. In the same tests, salicylic acid possessed moderate O₂^? and low OH'scavenging activities.

Our results demonstrate that adding two hydroxyl groups to salicylic acid strongly increases the reactive oxygen species (ROS) scavenging activities. Adding two hydroxyl groups at position 4 and 5 (2,4,5-THBA) affords the most active ROS scavenging activity probably due to the ortho unsubstituted catechol moiety. In fact, we can consider that the ROS scavenging properties of salicylic acid are essentially due to its metabolic products such as 2,3- and 2,5-DHBAs, catechol and also to THBAs.     

Effects of Hyperthermic Conditions on the Reactivity of Oxygen Radicals

Generation and reactivity of superoxide (0₂^?) and hydroxyl (OH') radicals in enzymatic and radiolytic systems were investigated over the temperature range from 20^o-50^oC. The generation rate and reaction kinetics of both enzymatically and radiolytically produced superoxide radicals were determined by a cytochrome c reduction assay. For OH' radical reaction studies the degradation of hyaluronic acid was assayed. An increase in temperature leads to a greater reactivity of both radicals, but in the case of an enzymatic source a disproportionate increase in the rate of generation is observed. In the pulse radiolysis system, the reactivity of superoxide radicals was found to be stimulated 15-fold over the temperature range from 20^oC to 60^oC, although the activity of superoxide dismutase was only minimally increased (about 1.6-fold). The results are discussed with respect to the possible importance of active oxygen species to the biological effects of hyperthermia.     

Spontaneous Cleavage of Single and Double Stranded 4′-DNA Radicals Under Aerobic Conditions

Abstract

The O₂-induced strand scission of 4′-DNA radicals is initiated by a reversible O₂ addition reaction. The rate coefficient of the O₂ release from the 4′-DNA peroxyl radical is 1.00 s^?1 in single strands and 0.05 s^?1 in double strands at 20°C. Because of this reversibility, an O₂-dependent strand cleavage occurs only in the presence of H-donors which trap the 4′-DNA peroxyl radicals yielding DNA hydroperoxides. At very low H-donor concentrations the strand scission is the result of an O₂-independent, spontaneous reaction even under aerobic conditions.     

Alterations in cardiac membrane Ca2+ transport during oxidative stress

Although cardiac dysfunction due to ischemia-reperfusion injury is considered to involve oxygen free radicals, the exact manner by which this oxidative stress affects the myocardium is not clear. As the occurrence of intracellular Ca²⁺ overload has been shown to play a critical role in the genesis of cellular damage due to ischemia-reperfusion, this study was undertaken to examine whether oxygen free radicals are involved in altering the sarcolemmal Ca²⁺-transport activities due to reperfusion injury. When isolated rat hearts were made globally ischemic for 30 min and then reperfused for 5 min, the Ca²⁺ -pump and Na⁺-Ca²⁺ exchange activities were depressed in the purified sarcolemmal fraction; these alterations were prevented when a free radical scavenger enzymes (superoxide dismutase plus catalase) were added to the reperfusion medium. Both the Ca²⁺- pump and Na⁺- Ca²⁺ exchange activities in control heart sarcolemmal preparations were depressed by activated oxygen-generating systems containing xanthine plus xanthine oxidase and H₂O₂; these changes were prevented by the inclusion of superoxide dismutase and catalase in the incubation medium. These results support the view that oxidative stress during ischemia-reperfusion may contribute towards the occurrence of intracellular Ca²⁺ overload and subsequent cell damage by depressing the sarcolemmal mechanisms governing the efflux of Ca²⁺ from the cardiac cell.     

Detection and identification of oxidants formed during •NO/O2•– reaction: A multi-well plate CW-EPR spectroscopy combined with HPLC analyses

Abstract

New techniques and probes are routinely emerging for detecting short-lived free radicals such as superoxide radical anion (O₂^?–), nitric oxide (^?NO), and transient oxidants derived from peroxynitrite (ONOO^–/ONOOH). Recently, we reported the profiles of oxidation products (2-hydroxyethidium, ethidium, and various dimeric products) of the fluorogenic probe hydroethidine (HE) in the ^?NO/O₂^?– system (Zielonka et al. 2012). In this study, we used HPLC analyses of HE oxidation products in combination with continuous wave electron paramagnetic resonance (CW-EPR) spin trapping with 5-tert-butoxycarbonyl-5-methyl-1-pyrroline N-oxide (BMPO) to define the identity of the oxidizing species formed in the ^?NO/O₂^?– system. EPR spin-trapping technique is still considered as the gold standard for characterization of free radicals and their intermediates. We monitored formation of BMPO-superoxide (BMPO-^?OOH) and BMPO-hydroxyl (BMPO-^?OH) radical adducts. Simultaneous analyses of results from EPR spin-trapping and HPLC measurements are helpful in the interpretation of the mechanism of formation of products of HE oxidation.     

Modeling the scavenging activity of ellagic acid and its methyl derivatives towards hydroxyl, methoxy, and nitrogen dioxide radicals

The reaction mechanisms involved in the scavenging of hydroxyl (OH^·), methoxy (OCH₃ ^·), and nitrogen dioxide (NO₂ ^·) radicals by ellagic acid and its monomethyl and dimethyl derivatives were investigated using the transition state theory and density functional theory. The calculated Gibbs barrier energies associated with the abstraction of hydrogen from the hydroxyl groups of ellagic acid and its monomethyl and dimethyl derivatives by an OH· radical in aqueous media were all found to be negative. When NO₂ ^· was the radical involved in hydrogen abstraction, the Gibbs barrier energies were much larger than those calculated when the OH^· radical was involved. When OCH₃ ^· was the hydrogen-abstracting radical, the Gibbs barrier energies lay between those obtained with OH^· and NO₂ ^· radicals. Therefore, the scavenging efficiencies of ellagic acid and its monomethyl and dimethyl derivatives towards the three radicals decrease in the order OH^· >> OCH₃ ^· > NO₂ ^·. Our calculated rate constants are broadly in agreement with those obtained experimentally for hydrogen abstraction reactions of ellagic acid with OH· and NO₂· radicals.

Effects of fatty acids on superoxide radical generation in leukocytes

Acetylated ferricytochrome c was employed for the detection of superoxide radicals (O₂^?) generated both in intact cells and in subcellular fractions of leukocytes. Certain saturated fatty acids, myristate in particular, induced the production of O₂^? in leukocytes, suggesting a correlation between the formation of O₂^? and the hydrophobic interaction of fatty acids with the leukocyte plasma membrane. As compared with O₂^? radical generation from phagocytizing leukocytes, a greater stimulation of O_2? formation was observed in cells in which myristate was added. The enhanced activity which generated O₂^? in the cell-free system was located in a particulate fraction but not in the cytosol. The rate of O₂^? generation in the particulate fraction was higher in the presence of NADPH than in the presence of NADH. The effects of reagents such as KCN, 2,4-dichlorophenol and aminotriazole on the O₂^? generation in this fraction are examined and the nature of the O₂^? generating system is discussed.     

A reaction of the superoxide radical with tetrapyrroles

Bilirubin and biliverdin were bleached during exposure to the aerobic xanthine oxidase reaction. Enzymic scavenging of O₂^?, by Superoxide dismutase, inhibited, whereas enzymic scavenging of H₂O₂, by catalase, did not. Increasing the rate of production of O₂^? without increasing the turnover rate of xanthine oxidase, by increasing pO₂, accelerated the bleaching of the biliverdin. Moreover, a scavenger of OH·, such as benzoate, or an inactivating chelating agent for iron, such as diethylenetriamine pentaacetate or desferrioxamine mesylate, did not inhibit. It follows that O₂^? can directly attack these tetrapyrroles. Kinetic competition between Superoxide dismutase and bilirubin yielded a value for k_{bilirubin, O2^?} = 2.3 × 10⁴ M^?1s^?1 at pH 8.3 and at 23 °C. A similar experiment for biliverdin yielded a value for k_{bilirubin, O2^?} = 7 × 10⁴ M^?1s^?1.     

Studies on the antioxidant properties of some phytoestrogens

Isoflavones genistein and daidzein are nonsteroidal phytoestrogens occurring mainly in soybean foods. These phytoestrogens possess estrogenic properties and show a variety of health benefits as anti‐inflammatory agents. However, the mechanism of their action has not been identified in detail. The aim of this study is to characterize the antioxidant powers of genistein, daidzein and daidzein metabolite–equol through their activities to scavenge superoxide anion radical (O^?₂^?), hydroxyl radical (HO^?), 2,2–diphenyl–1‐picrylhydrazyl radical (DPPH^?) and hydrogen peroxide (H₂O₂) using chemiluminescence and spectrophotometry techniques. Potassium superoxide in dimethyl sulphoxide (DMSO) and 18‐crown‐6 ether were used as a source of O^?₂^?. Hydroxyl radicals were produced using the Fenton reaction. In free radical assays, genistein had the IC₅₀ values (an amount of antioxidant concentration required to decrease the initial radical concentration by 50%) 0.391 ± 0.012 mM for O^?₂^?, 0.621 ± 0.028 mM for HO^? and 1.89 ± 0.16 mM for DPPH^?. The IC₅₀ values for daidzein for these free radicals were 1.924 ± 0.011 mM, 0.702 ± 0.012 mM and 2.81 ± 0.03 mM, respectively. Equol was the most active the free radical scavenger with IC₅₀ = 0.451 ± 0.018 mM for HO^? and IC₅₀ = 1.36 ± 0.11 mM for DPPH^?. All tested compounds exerted a significant effect on the H₂O₂: IC₅₀ = 18.1 ± 1.1 μM for genistein, IC₅₀ = 2.1 ± 0.5 μM for daidzein, and IC₅₀ = 1.06 ± 0.2 μM for equol. These findings show that genistein, daidzein and equol are effective free radical scavengers and possess high antioxidant power in vitro. Copyright © 2016 John Wiley & Sons, Ltd.     

The role of interactions between O2, H2O2, .OH,e- and O2- in free radical damage to biological systems

The occurrence of the Haber-Weiss reaction and other interactions between free radicals has been investigated in the effects of mixtures of free radicals on the permeability of resealed erythrocyte ghosts and on the activity of membrane-bound glyceraldehyde-3-phosphate dehydrogenase. The following mixtures were found to induce damage greater than that which could be accounted for by the independent actions of the constituent free radicals: (i) · OH + H₂O₂, and (ii) · OH + H₂O₂ + O₂^?. In contrast, the following mixtures were found to induce less damage than that predicted on the basis of independent actions of constituent free radicals: (i) H₂O₂ + O₂^?, and (ii) oxidizing radicals ( · OH, H₂O₂) + reducing radicals (e^?, H · ). These results suggest a Haber-Weiss-like interaction between H₂O₂ and O₂^?and an interaction between H₂O₂ and · OH to produce a species more potent than either in causing increased permeability. The decrease in damage observed in the simultaneous presence of oxidizing and reducing radicals suggests an antagonistic effect by which each tends to moderate damage by the other. Inactivation of glyceraldehyde-3-phosphate dehydrogenase was found to be more sensitive to radiation than permeability by an order of magnitude, while permeability was more sensitive to the enhancement of damage by oxygen. Comparison of the effectiveness of free radical scavengers in inhibiting the increase in permeability caused by free radicals showed the following order of effectiveness, expressed in terms of percentage protection: formate (90%) > nitrogen (65%) > catalase (60%) > dismutase (32%), and with respect to enzymatic inactivation, nitrogen (100%) > formate (77%) > dismutase (48%) > catalase (44%). The relative rates observed anaerobically and aerobically in the presence and absence of the above scavengers suggest that (at least in the case of radiation damage to the membranes of erythrocyte ghost cells) the “oxygen effect” is due to the interaction of oxygen with e^? and H., producing O₂^? which aggravates damage under conditions which allow consequent Haber-Weiss-like reactions. The further increase in damage when oxygen concentration is raised yet higher is due to the interaction of oxygen with the sites of initial damage.     

Radical scavenging ability of some compounds isolated from Piper cubeba towards free radicals

The purpose of this study was to identify the antioxidant activity of 16 compounds isolated from Piper cubeba (CNCs) through the extent of their capacities to scavenge free radicals, hydroxyl radical (HO^?), superoxide anion radical () and 2,2‐diphenyl‐1‐picrylhydrazyl radical (DPPH^?), in different systems. Electron paramagnetic resonance (EPR) and 5,5‐dimethyl‐1‐pyrroline‐N‐oxide, DMPO, as the spin trap, and chemiluminescence techniques were applied. Using the Fenton‐like reaction [Fe(II) + H₂O₂], CNCs were found to inhibit DMPO? OH radical formation ranging from 5 to 57% at 1.25 mmol L^?1 concentration. The examined CNCs also showed a high DPPH antiradical activity (ranging from 15 to 99% at 5 mmol L^?1 concentration). Furthermore, the results indicated that seven of the 16 tested compounds may catalyse the conversion of superoxide radicals generated in the potassium superoxide/18‐crown‐6 ether system, thus showing superoxide dismutase‐like activity. The data obtained suggest that radical scavenging properties of CNCs might have potential application in many plant medicines. Copyright © 2010 John Wiley & Sons, Ltd.