Assessment of pro-oxidant activity of foods by kinetic analysis of crocin bleaching.

The pro-oxidant activity of potent oxidants and foods was determined using the kinetic analysis of crocin bleaching. In its reduced form, crocin has an absorption band at 443 nm, which disappears upon oxidation by a generic radical species. Hydroxyl radicals generated by hydrogen peroxide, peroxyl radicals from ABAP, and the stable free radical DPPH(*) were allowed to react with crocin in an aqueous solution at 40 degrees C. Pro-oxidant activity was taken as the ratio between the decrease in crocin absorbance at 5 min and the relevant oxidant concentration. The test proposed was used to evaluate the pro-oxidant activity of widely consumed foods such as pasteurized skim milk and bread. They both exerted significant pro-oxidant activities, which were attributed to the early nonenzymatic browning products formed upon heat treatment.

Identification of radical scavengers in sweet grass (Hierochloe odorata).

Extracts from aerial parts of sweet grass (Hierochloe odorata) were active DPPH free radical scavengers. The active compounds were detected in extract fractions using HPLC with on-line radical scavenging detection. After multistep fractionation of the extract, two new natural products possessing radical scavenging activity were isolated, and their structures were elucidated by NMR and MS. They were identified as 5,8-dihydroxybenzopyranone and 5-hydroxy-8-O-beta-D-glucopyranosyl-benzopyranone. Activities of the compounds isolated were tested by DPPH and ABTS free radical scavenging assays, and compared with the known natural antioxidant rosmarinic acid and Trolox.

Antioxidative activity and safety of the 50 ethanolic extract from red bean fermented by Bacillus subtilis IMR-NK1.

This study aimed at evaluating the antioxidative activities and the safety of 50% ethanolic extract from red bean fermented by Bacillus subtillis IMR-NK1. The antioxidative activities, including alpha,alpha-diphenyl-beta-picryl-hydrazyl (DPPH) radicals scavenging effects, Fe(2+)-chelating ability, and reducing power, were studied in vitro. It was found that the antioxidative activity increased with the concentrations of the extract to a certain extent and then leveled off as the concentration further increased. As compared to the commercial antioxidants, the fermented red bean extract showed less scavenging effect on the DPPH radical and reducing power than alpha-tocopherol and BHT, but better Fe(2+)-chelating ability. No mutagenicity or toxicity effect toward all tester strains was found in the 50% ethanolic extract of fermented red bean by means of the Ames test. The results suggested that the 50% ethanolic extract was safe in genotoxicity.

Comparison of radical scavenging effect, inhibition of microsomal oxygen free radical generation, and serum lipoprotein oxidation of several natural antioxidants.

Typical components of the Mediterranean diet, such as olive oil and red wine, contain high concentrations of complex phenols, which have been suggested to have an important antioxidant role. The aim of the present work was to determine the inhibitory potency of compounds such as oleuropein, hydroxytyrosol, and other structurally related compounds, such as gallic acid, toward reactive oxygen species generation and free radical scavenging ability. The potency of these compounds was also examined with respect to protecting in vitro low-density lipoprotein oxidation. These studies indicate that complex phenols, such as hydroxytyrosol, and gallic acid both inhibit free radical generation and act as free radical scavengers. The use of three different approaches to determine antioxidant potency demonstrates that activity in one test does not necessarily correlate with activity in another. It was also demonstrated that the presence of two phenolic groups is not always associated with antioxidant activity.

Antioxidants from the bark of Burkea africana, an African medicinal plant.

The bark of the tree Burkea africana is used medicinally in large areas of sub-Saharan Africa. The constituents responsible for its putative activity are not well known. We have investigated the bark of B. africana for antioxidant and radical scavenging activity. A hydroethanol bark extract showed high activity, and most of this activity was located in semipolar fractions of the extract. From chromatographic purification and spectroscopical structure studies, we conclude that the active constituents are proanthocyanidins. Two major components appear to be fisetinidol-(4alpha- --> 8)-catechin 3-gallate and bis-fisetinidol-(4alpha- --> 6, 4alpha- --> 8)-catechin 3-gallate. The latter compound is a new natural product. Smaller amounts of monomeric flavan-3-ols (catechin, epicatechin and fisetinidol) were also found.

On-line HPLC-DPPH screening method for evaluation of radical scavenging phenols extracted from apples (Malus domestica L.).

An on-line HPLC-DPPH screening method for phenolic antioxidants in apple methanol/water (80:20, v/v) extracts was applied. The determination of antioxidants was based on a decrease in absorbance at 515 nm after postcolumn reaction of HPLC-separated antioxidants with the 2,2'-diphenyl-1-picrylhydrazyl radicals (DPPH*). Each of the antioxidants separated by the HPLC column was observed as a negative peak corresponding to its antioxidative activity. The on-line method was applied for quantitative analysis of the antioxidants. A linear dependence of negative peak area on concentration of the reference antioxidants was observed. For validation of the on-line method the limit of detection, LOD (microg/mL), and the limit of quantification, LOQ (microg/mL), of the phenolic compounds were determined. Comparison of the UV and DPPH radical quenching chromatograms with authentic compounds identified catechin, chlorogenic acid, caffeic acid, epicatechin, and phloridzin in the apple cultivars (Lobo, Golden Delicious, and Boskoop), and the distribution of total antioxidant activity was calculated.

Antioxidant functions of selected allium thiosulfinates and S-alk(en)yl-L-cysteine sulfoxides.

Pure thiosulfinates, R-S(O)S-R (2), where R = Me (2a), Pr (2b), or All (2c), at levels up to 4 mM were not capable of scavenging hydrogen peroxide or superoxide anion. Relative to standard antioxidants (ascorbic acid, n-propyl gallate, butylated hydroxytoluene, Trolox, and reduced glutathione), these thiosulfinates were 1-3 orders of magnitude less efficient at reducing 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical, 0.5-2 orders of magnitude less efficient at quenching singlet oxygen, and about equally effective at scavenging hydroxyl radical. Generally, AllS(O)SAll (2c) was the most effective and PrS(O)SPr (2b) was the least effective thiosulfinate in these assays, except that MeS(O)SMe (2a) exhibited no quenching effect toward singlet oxygen. These thiosulfinates were also incapable at levels up to 0.1 mM (where they were toxic) of in vitro induction of quinone reductase (QR) in murine hepatoma (hepa 1c1c7) cells. However, S-1-propenyl-L-cysteine sulfoxide (isoalliin, 1a) and cycloalliin (3) induced QR in this system at 2 mM and 1 mM, respectively, although doubling of QR required levels of 10-15 mM.

Free radical scavenging properties of wheat extracts.

Three hard winter wheat varieties (Akron, Trego, and Platte) were examined and compared for their free radical scavenging properties and total phenolic contents (TPC). Free radical scavenging properties of wheat grain extracts were evaluated by spectrophotometric and electron spin resonance (ESR) spectrometry methods against stable 2,2-diphenyl-1-picryhydrazyl radical (DPPH*) and radical cation ABTS*+ (2,2'-azino-di[3-ethylbenzthiazoline sulfonate]). The results showed that the three wheat extracts differed in their capacities to quench or inhibit DPPH* and ABTS*+. Akron showed the greatest activity to quench DPPH radicals, while Platte had the highest capacity against ABTS*+. The ED50 values of wheat extracts against DPPH radicals were 0.60 mg/mL for Akron, 7.1 mg/mL for Trego, and 0.95 mg/mL for Platte under the experimental conditions. The trolox equivalents against ABTS*+ were 1.31 +/- 0.44, 1.08 +/- 0.05, and 1.91 +/- 0.06 micromol/g of grain for Akron, Trego, and Platte wheat, respectively. ESR results confirmed that wheat extracts directly reacted with and quenched free radicals. The TPC were 487.9 +/- 927.8 microg gallic acid equivalents/g of grain. No correlation was observed between TPC and radical scavenging capacities for DPPH* and ABTS*+ (p = 0.15 and p > 0.5, respectively).

Antioxidant properties of ferulic acid and its related compounds.

Antioxidant activity of 24 ferulic acid related compounds together with 6 gallic acid related compounds was evaluated using several different physical systems as well as their radical scavenging activity. The radical scavenging activity on 1,1-diphenyl-2-picrylhydrazyl (DPPH) decreased in the order caffeic acid > sinapic acid > ferulic acid > ferulic acid esters > p-coumaric acid. In bulk methyl linoleate, test hydroxycinnamic acids and ferulic acid esters showed antioxidant activity in parallel with their radical scavenging activity. In an ethanol-buffer solution of linoleic acid, the activity of test compounds was not always associated with their radical scavenging activity. Ferulic acid was most effective among the tested phenolic acids. Esterification of ferulic acid resulted in increasing activity. The activity of alkyl ferulates was somewhat influenced by the chain length of alcohol moiety. When the inhibitory effects of alkyl ferulates against oxidation of liposome induced by AAPH were tested, hexyl, octyl, and 2-ethyl-1-hexyl ferulates were more active than the other alkyl ferulates. Furthermore, lauryl gallate is most effective among the tested alkyl gallates. These results indicated that not only the radical scavenging activity of antioxidants, but also their affinity with lipid substrates, might be important factors in their activity.

Free radical studies of ellagic acid, a natural phenolic antioxidant.

Ellagic acid, a plant-derived polyphenol, inhibits gamma-radiation (hydroxyl radical) induced lipid peroxidation in rat liver microsomes in a dose- and concentration-dependent manner. Its antioxidant capacity has been estimated using the 1,1-diphenyl-2-picrylhydrazyl radical assay. To understand the actual mechanisms involved in antioxidant activity and the free radical scavenging ability,a nanosecond pulse radiolysis technique has been employed. The rate constants for the reactions of several reactive oxygen species and reactive nitrogen species such as hydroxyl, peroxyl, and nitrogen dioxide radicals have been found to be in the range of 10(6)-10(9) M(-1) s(-1). The ellagic acid radicals have been characterized by the absorption spectra and decay kinetics. Studies on the reactions of ellagic acid with the 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonate) radical and the radicals of ellagic acid with ascorbate have been used to estimate its one-electron reduction potential. Ellagic acid has also been found to be a good scavenger of peroxynitrite. Using stopped-flow reaction analyzer with absorption detection, the rate constant for this reaction has been determined to be 3.7 x 10(3) M(-1) s (-1). The electron spin resonance spectra of the oxidized ellagic acid radicals have been recorded by horseradish peroxidase and hydrogen peroxide method.

Free radical scavenging activity of red ginseng aqueous extracts.

This study was performed to investigate the free radical scavenging activity of Panax red ginseng C.A. Meyer aqueous extract on 1,1-dipheny-2-picrylhydrazyl (DPPH), carbon-centered radical, hydroxyl and superoxide radicals using Electron Spin Resonance (ESR) spectrometer and spin-trapping techniques. Two different Red ginseng aqueous extracts prepared by boiling water or room temperature extraction exhibited no significant difference in free radical scavenging activity. Ginseng extracts completely eliminated DPPH radical at 2 mg/ml. About 0.5 mg/ml ginseng extracts quenched 80% carbon-centered free radicals generated from 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH). Hydroxyl radical and superoxide radical were generated by UV irradiation and trapped by 5,5-dimethyl-l-pyrroline-N oxide (DMPO). Ginseng extracts scavenged 40% of hydroxyl radical at 0.1 mg/ml. Two mg/ml ginseng extracts completely scavenged superoxide radical. Ginseng extracts did not scavenge nitric oxide. The ESR data demonstrate that red ginseng aqueous extract is not a strong free radical scavenger.

Antioxidant effects of 1,5-anhydro-D-fructose, a new natural sugar, in vitro.

The antioxidant effects of 1,5-anhydro-D-fructose (1,5-AF), a unique anhydrohexulose, were studied in 1,1-diphenyl-2-picrylhydrazyl (DPPH) solution, in human cells along with lipid peroxidation of low-density lipoprotein (LDL). We have confirmed that 1,5-AF scavenges DPPH radicals directly in solution and inhibits the formation of hydrogen peroxide and superoxide anion, typical reactive oxygen species (ROS), induced by phorbol myristate acetate (PMA) in a dose-dependent manner in THP-1 cells. We also observed the dose-dependent antioxidant effects of 1,5-AF on copper-mediated LDL oxidation. These findings suggest that 1,5-AF might play a role in reducing the risk of atherosclerosis and may help prevent coronary heart disease.

Antioxidant capacities of ten edible North American plants.

The EtOAc extract obtained from ten edible North American plants, Acorus calamus, Clintonia borealis, Gaultheria shallon, Juniperus osteosperma, Opuntia polyacantha, Prunus americana, Prunus virginiana, Sambucus cerulea, Sorbus americana and Vaccinium parvifolium, were tested in the 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical assay. High antioxidant activity was obtained from the extracts of three fruits, Gaultheria shallon, Sambucus cerulea and Prunus americana and one extracted rhizome, Acorus calamus. Catechin and epicatechin, potent polyphenolic antioxidants, were identified in the EtOAc extracts of Gaultheria shallon and Sambucus cerulea by reversed-phase thin-layer chromatography (TLC) and reversed-phase high-performance liquid chromatography (HPLC).

Antioxidant enzymes and DPPH-radical scavenging activity in chilled and heat-shocked rice (Oryza sativa L.) seedlings radicles.

Chilling whole rice seedlings at 5 degrees C significantly increased the time needed to recover linear growth and reduced the subsequent linear rate of radicle growth. Subjecting nonchilled seedlings to a 45 degrees C heat shock for up to 20 min did not alter subsequent growth, whereas a 3 min heat shock was optimal in reducing growth inhibition caused by 2 days of chilling. The activity of five antioxidant enzymes [superoxide dismutase (EC 1.15.1.1), catalase (CAT; EC 1.11.1.6), ascorbate peroxidase (APX; EC 1.11.1.11), glutathione reductase (GR; EC 1.6.4.2), and guaiacol peroxidase (GPX; EC 1.11.1.7)] and DPPH (1,1-diphenyl-2-picrylhydrazyl)-radical scavenging activity were measured in heat-shocked and/or chilled radicles. Heat shock slightly increased the activity of CAT, APX, and GR and suppressed the increase of GR and GPX activity during recovery from chilling. Increased CAT, APX, GR, and DPPH-radical scavenging activity and protection of CAT activity during chilling appear to be correlated with heat shock-induced chilling tolerance.

Antioxidant properties of methanolic extracts from several ear mushrooms.

Five kinds of ear mushrooms are commercially available in Taiwan, including black, red, jin, snow, and silver ears. Methanolic extracts were prepared from these ear mushrooms, and their antioxidant properties were studied. For all methanolic extracts from ear mushrooms, the antioxidant activities in the 1,3-diethyl-2-thiobarbituric acid method were moderate (38.6 approximately 74.6%) at 1.0-5.0 mg/mL. Methanolic extracts from red, jin, and snow ears showed excellent antioxidant activities in the conjugated diene method at 5.0 mg/mL. At 5.0 mg/mL, reducing powers of methanolic extracts were in the descending order of snow > black approximately red approximately jin > silver ears. The scavenging effect of methanolic extracts from ear mushrooms on 1,1-diphenyl-2-picrylhydrazyl radicals was excellent except for that from silver ears. Ear mushroom extracts were not good scavengers for hydroxyl free radicals but were good chelators for ferrous ions. Naturally occurring antioxidants, including ascorbic acid, tocopherols, and total phenols, were found in the methanolic extracts. However, beta-carotene was not detected. Total antioxidant components were 15.69, 30.09, 27.83, 49.17, and 31.70 mg/g for black, red, jin, snow, and silver ears, respectively.

Antioxidative components from the aerial parts of Lactuca scariola L.

The antioxidant activity of Lactuca scariola (Compositae) was investigated by measuring the radical scavenging effect on DPPH (1,1-diphenyl-2-picrylhydrazyl) radical. The methanolic extract of the aerial parts of Lactuca scariola showed strong radical scavenging activity. The EtOAc soluble fraction exhibited a stronger activity than the others, and was purified by silica gel and Sephadex LH-20 column chromatography. Quercetin-3-O-beta-D-glucopyranoside, luteolin-7-O-beta-D-glucopyranoside, luteolin, quercetin and kaempferol, together with 1beta,13-dihydrolactucin were isolated from the EtOAc soluble fraction as active ingredients.

Electron spin resonance assessment of the antioxidant potential of medicinal plants. Part I. Contribution of anthocyanosides and flavonoids to the radical scavenging ability of fruit and herbal teas.

Radical scavenging properties of the extracts of some fruits and flowers, as well as of their complex formulations used as fruit teas, were tested on DPPH radical using electron spin resonance spectroscopy. The contents of anthocyanosides and flavonoids in plant materials were determined with spectrophotometric method. The most effective DPPH radical scavengers were extracts from Fructus Aroniae, Fructus Myrtilli and Fructus Rosae and the fruit teas, including them as main ingredients. No simple correlation was found between the scavenging activity and the content of anthocyanosides and flavonoids. The results can be rationalised by taking into account the presence of catechins and ascorbic acid.

Vasorelaxing and antioxidant constituents from Hernandia nymphaeifolia.

Three new alkaloids, (+)-nymphaedaline (1), oxo-O-methylbulbocapnine (2), and (+)-laetine (3), have been isolated from the trunk bark of Hernandia nymphaeifolia. The structures of these new compounds were elucidated by spectroscopic analysis. Among the isolates of this plant obtained till now, sixteen compounds show effective inhibitory activities on the contraction of vascular smooth muscles induced by high K+ (80 mM) or norepinephrine (3 microM). In addition, eight compounds showed effective antioxidant activities in scavenging the stable free radical, diphenyl-picryl-hydrazyl (DPPH).

Identification and antioxidant activity of novel chlorogenic acid derivatives from bamboo (Phyllostachys edulis).

One known and two novel antioxidant compounds have been isolated from bamboo (Phyllostachys edulis). The butanol-soluble extract of the bamboo leaves was found to have a significant antioxidant activity, as measured by scavenging the stable 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical and the superoxide anion radical (O(2)(-)) in the xanthine/xanthine oxidase assay system. Antioxidant activity-directed fractionation of the extract led to the isolation and characterization of three structural isomeric chlorogenic acid derivatives: 3-O-(3'-methylcaffeoyl)quinic acid (1), 5-O-caffeoyl-4-methylquinic acid (2), and 3-O-caffeoyl-1-methylquinic acid (3). Compounds 2 and 3 were isolated and characterized for the first time from the natural products. In the DPPH scavenging assay as well as in the iron-induced rat microsomal lipid peroxidation system, compounds 2 (IC(50) = 8.8 and 19.2 microM) and 3 (IC(50) = 6.9 and 14.6 microM) showed approximately 2-4 times higher antioxidant activity than did chlorogenic acid (IC(50) = 12.3 and 28.3 microM) and other related hydroxycinnamates such as caffeic acid (IC(50) =13.7 and 25.5 microM) and ferulic acid (IC(50) = 36.5 and 56.9 microM). Among the three compounds, compound 1 yielded the weakest antioxidant activity, and the DPPH scavenging and lipid peroxidation inhibitory activity (IC(50) = 16.0 and 29.8 microM) was lower than those of chlorogenic and caffeic acids. All three compounds exhibited both superoxide scavenging activities and inhibitory effects on xanthine oxidase. Their superoxide anion (O(2)(-)) scavenging activities (IC(50) = 1, 4.3 microM; 2, 2.8 microM; and 3, 1.2 microM) were markedly stronger than those of ascorbic acid (IC(50) = 56.0 microM), alpha-tocopherol (IC(50) > 100 microM), and other test compounds, although their inhibition effects on xanthine oxidase may contribute to the potent scavenging activity. alpha-Tocopherol exerted a significant inhibitory effect (65.5% of the control) on superoxide generation in 12-O-tetradecanoylphorbol-13-acetate-induced human promyelocytic leukemia HL-60 cells, and compound 3 showed moderate activity (36.0%). On the other hand, other compounds including 1, 2, chlorogenic acid, and other antioxidants were weakly active (24.8-10.1%) in the suppression of superoxide generation.

Antioxidant activities of dioscorin, the storage protein of yam (Dioscorea batatas Decne) tuber.

Dioscorin, the storage protein of yam (Dioscorea batatas Decne) tuber (which is different from dioscorine found in tubers of Dioscorea hirsuta), was purified to homogeneity after DE-52 ion exchange column according to the methods of Hou et al. (J. Agric. Food Chem. 1999, 47, 2168-2172). A single band of 32 kDa dioscorin was obtained on a sodium dodecyl sulfate-polyacrylamide gel electrophoresis gel with 2-mercaptoethanol treatment. This purified dioscorin was shown by spectrophotometric method to have scavenging activity against 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical in a pH-dependent manner. There is a positive correlation between scavenging effects against DPPH (8-46%) and amounts of 32 kDa dioscorin (5.97-47.80 nmol) added in Tris-HCl buffer (pH 7.9), which are comparable to those of glutathione at the same concentrations. Using electron paramagnetic resonance (EPR) spectrometry for DPPH radical detection, it was found that the intensities of the EPR signal were decreased by 28.6 and 57 nmol of 32 kDa dioscorin in Tris-HCl buffer (pH 7.9) more than in distilled water compared to controls. EPR spectrometry was also used for hydroxyl radical detection. It was found that 32 kDa dioscorin could capture hydroxyl radical, and the intensities of the EPR signal were significantly decreased dose-dependently by 1.79-14.32 nmol of 32 kDa dioscorin (r = 0.975) compared to the control. It is suggested that 32 kDa dioscorin, the storage protein of yam tuber, may play a role as antioxidant in tubers and may be beneficial for health when people take it as a food additive or consume yam tubers.