Hydroxycinnamates and their in vitro and in vivo antioxidant activities

Hydroxycinnamates are among the most widely distributed plant phenylpropanoids present in the free, conjugated-soluble and insoluble-bound forms. This review will focus on the occurrence, in vitro and in vivo antioxidant activities of ferulic, coumaric, caffeic and sinapic acids and their derivatives. Hydroxycinnamates are found in almost all food groups though they are abundant in cereals, legumes, oilseeds, fruits, vegetables and beverages and render antioxidant activity by scavenging hydroxyl radical, superoxide radical anion, several organic radicals, peroxyl radical, peroxinitrite and singlet oxygen, among others. Further, their antioxidant activity as chain breaking antioxidants and reducing agents is also notable. Ferulic acid and its derivatives such as ferulic acid ethyl ester, ferulic acid dehydrodimers, feruloyl glycosides and curcumin have demonstrated potent antioxidant activity in both in vitro and in vivo systems. Similarly, caffeic acid and some of its derivatives such as caffeic acid phenethyl ester, rosmarinic acid, and chlorogenic acid exhibit antioxidant activity. The highest antioxidant activity was observed for caffeic acid whereas p-coumaric acid had the least effect among major hydroxycinnamic acids. The importance of structural effects on the potency of antioxidant activity of hydroxycinnamates is discussed. While this review also shows the existence of substantial body of evidences for in vitro antioxidant activity of hydroxycinnamates, there is a clear gap for in vivo information, particularly for sinapic and p-coumaric acids and their derivatives. The role of grains, fruits, vegetables and red wine in disease risk reduction and health promotion could partly be attributed to their constituent hydroxycinnamates.     

Ferulic acid antioxidant protection against hydroxyl and peroxyl radical oxidation in synaptosomal and neuronal cell culture systems in vitro: structure-activity studies

In this study, free radical scavenging abilities of ferulic acid in relation to its structural characteristics were evaluated in solution, cultured neurons, and synaptosomal systems exposed to hydroxyl and peroxyl radicals. Cultured neuronal cells exposed to the peroxyl radical initiator AAPH die in a dose-response manner and show elevated levels of protein carbonyls. The presence of ferulic acid or similar phenolic compounds, however, greatly reduces free radical damage in neuronal cell systems without causing cell death by themselves. In addition, synaptosomal membrane systems exposed to oxidative stress by hydroxyl and peroxyl radical generators show elevated levels of oxidation as indexed by protein oxidation, lipid peroxidation, and ROS measurement. Ferulic acid greatly attenuates these changes, and its effects are far more potent than those obtained for vanillic, coumaric, and cinnamic acid treatments. Moreover, ferulic acid protects against free radical mediated changes in conformation of synaptosomal membrane proteins as monitored by EPR spin labeling techniques. The results presented in this study suggest the importance of naturally occurring antioxidants such as ferulic acid in therapeutic intervention methodology against neurodegenerative disorders such as Alzheimer's disease in which oxidative stress is implicated.     

Effect of dihydrocaffeic acid on UV irradiation of human keratinocyte HaCaT cells

Dihydrocaffeic acid, a dietary constituent and a microbial metabolite of flavonoids, is an antioxidant, but few biological effects have been examined. After its production by microflora in the colon, dihydrocaffeic acid is absorbed and found in plasma as a combination of free and metabolized forms. Excess solar UV radiation provokes damage and initiates immune response and inflammation in skin, sometimes leading to cancer. Dihydrocaffeic acid reduced the cytotoxicity and pro-inflammatory cytokine production (interleukin-6 and -8) in HaCaT cells, a keratinocyte model, following UV radiation. The effect of dihydrocaffeic acid may result from a combination of direct radical scavenging of the reactive oxygen species formed or reinforcement of the antioxidant potential of the keratinocytes, as well as a direct interference with the pathway involved in cytokine stimulation. The minimum structure required for such an effect appears to consist of a propionate side chain attached to a catechol moiety, as indicated by the efficacy of caffeic acid, but not of the methyl and glucuronide conjugates of dihydrocaffeic acid. The data obtained suggest that dihydrocaffeic acid is a potential candidate for photo-protection by interfering with the events initiated after UV exposure in keratinocytes.     

Attenuation of chronic neuroinflammation by a nitric oxide-releasing derivative of the antioxidant ferulic acid

Chronic neuroinflammation and oxidative stress contribute to the neurodegeneration associated with Alzheimer's disease and represent targets for therapy. Ferulic acid is a natural compound that expresses antioxidant and anti-inflammatory activities. Nitric oxide is also a key modulator of inflammatory responses. Grafting a nitric oxide-releasing moiety onto anti-inflammatory drugs results in enhanced anti-inflammatory activity. We compared the effectiveness of ferulic acid with a novel nitric oxide-releasing derivative of ferulic acid in an animal model of chronic neuroinflammation that reproduces many interesting features of Alzheimer's disease. Lipopolysaccharide was infused into the 4th ventricle of young rats for 14 days. Various doses of ferulic acid or its nitric oxide-releasing derivative were administered daily. Both drugs produced a dose-dependent reduction in microglia activation within the temporal lobe. However, the nitric oxide-releasing ferulic acid derivative was significantly more potent. If we delayed the initiation of therapy for 14 days, we found no reduction in microglial activation. In addition, both drugs demonstrated antioxidant and hydroxyl radical scavenging abilities in in vitro studies. Overall, our results predict that a treatment using nitric oxide-releasing ferulic acid may attenuate the processes that drive the pathology associated with Alzheimer's disease if the treatment is initiated before the neuroinflammatory processes can develop.     

In vivo protection of synaptosomes by ferulic acid ethyl ester (FAEE) from oxidative stress mediated by 2,2-azobis(2-amidino-propane)dihydrochloride (AAPH) or Fe(2+)/H(2)O(2): insight into mechanisms of neuroprotection and relevance to oxidative stress-related neurodegenerative disorders

Ferulic acid ethyl ester (FAEE) is an ester derivative of ferulic acid, the latter known for its anti-inflammatory and antioxidant properties. Previous studies from our laboratory have shown that ferulic acid protects synaptosomal membrane system and neuronal cell culture systems against hydroxyl and peroxyl radical oxidation. FAEE is lipophilic and is able to penetrate lipid bilayer. Previous studies reported that FAEE reduces Alzheimer's amyloid beta peptide Abeta(1-42)-induced oxidative stress and cytotoxicity in neuronal cell culture by direct radical scavenging and by inducing certain antioxidant proteins. In the present study we tested the hypothesis that FAEE would provide neuroprotection against free radical oxidative stress in vivo. Synaptosomes were isolated from the gerbils that were previously injected intraperitoneally (i.p.) with FAEE or DMSO and were treated with oxidants, Fe(2+)/H(2)O(2) or 2,2-azobis(2-amidino-propane)dihydrochloride (AAPH). Synaptosomes isolated from the gerbil previously injected i.p. with FAEE and treated with Fe(2+)/H(2)O(2) and AAPH showed significant reduction in reactive oxygen species (ROS), levels of protein carbonyl, protein bound 4-hydroxynonenal (HNE, a lipid peroxidation product), and 3-nitrotyrosine (3-NT, another marker of protein oxidation formed by reaction of tyrosine residues with peroxynitrite) compared to Fe(2+)/H(2)O(2) or AAPH induced oxidative stress in synapotosomes isolated from the brain of gerbils that were previously injected with DMSO. The synaptosomes isolated from gerbil pre-injected with FAEE and subsequently treated with AAPH or Fe(2+)/H(2)O(2) showed induction of heme oxygenase (HO-1) and heat shock protein 70 (HSP-70) but reduced inducible nitric oxide synthase (iNOS) levels. These results are discussed with reference to potential use of this lipophilic antioxidant phenolic compound in the treatment of oxidative stress-related neurodegenerative disorders.     

Rational discovery and development of a mitochondria-targeted antioxidant based on cinnamic acid scaffold

A novel mitochondria-targeted antioxidant (TPP-OH) was synthesized by attaching the natural hydrophilic antioxidant caffeic acid to an aliphatic lipophilic carbon chain containing a triphenylphosphonium (TPP) cation. This compound has similar antioxidant activity to caffeic acid as demonstrated by measurement of DPPH/ABTS radical quenching and redox potentials, but is significantly more hydrophobic than its precursor as indicated by the relative partition coefficients. The antioxidant activity of both compounds was intrinsic related to the ortho-catechol system, as the methoxylation of the phenolic functions, namely in TPP-OCH(3) and dimethoxycinnamic acid, gave compounds with negligible antioxidant action. The incorporation of the lipophilic TPP cation to form TTP-OH and TPP-OCH(3) allowed the cinnamic derivatives to accumulate within mitochondria in a process driven by the membrane potential. However, only TPP-OH was an effective antioxidant: TPP-OH protected cells against H(2)O(2) and linoleic acid hydroperoxide-induced oxidative stress. As mitochondrial oxidative damage is associated with a number of clinical disorders, TPP-OH may be a useful lead that could be added to the family of mitochondria-targeted antioxidants that can decrease mitochondrial oxidative damage.     

根皮素乙酰阿魏酸酯的合成及抗氧化活性研究

以阿魏酸和根皮素为原料,合成了三种根皮素乙酰阿魏酸酯,采用IR、1H NMR、13C NMR和HPLC-MS对其结构进行表征.并测定根皮素及其酯的还原能力、清除DPPH自由基及·OH自由基能力,结果表明根皮素乙酰阿魏酸酯的抗氧化活性显著优于根皮素.     

Mitochondrial electron transport-linked tocopheroxyl radical reduction

alpha-Tocopherol (vitamin E) is a lipophilic chain-breaking antioxidant which inhibits lipid peroxidation in isolated mitochondrial membranes and protects membranes from oxidative damage. The primary oxidation product of vitamin E is the tocopheroxyl radical. Reduction of the tocopheroxyl radical can occur by reactions with water-soluble anti-oxidants such as ascorbate or glutathione, resulting in the recycling of vitamin E. Physiological concentrations of vitamin E are too low to allow detection of tocopheroxyl radical by ESR. After dietary supplementation with vitamin E, a 10-20-fold increase in the rat liver mitochondrial membrane content of vitamin E was achieved and this allowed for direct detection of the tocopheroxyl radical by ESR, after treatment with an oxidizing system composed of lipoxygenase and arachidonic acid. By using submitochondrial particle membranes, it was shown that NADH, succinate, and reduced cytochrome c-linked oxidation reduce the tocopheroxyl radical, preventing both accumulation of the radical and vitamin E consumption. As the electron transport chain can reduce tocopheroxyl radical it may have an important physiological role in recycling vitamin E.     

The oxidation of endogenous ascorbic acid during histamine secretion by rat peritoneal mast cells

Endogenous ascorbic acid is oxidized to the free radical species by rat mast cells during histamine secretion. This antioxidant may function as a radical scavenger of Superoxide and other membrane-damaging radicals known to be liberated by this process. The high levels of ascorbic acid in mast cells may, therefore, function to protect the cell membrane from oxidative damage and thereby promote cell survival after an extensive secretory response.     

Lipase-catalysed synthesis of esters of ferulic acid with natural compounds and evaluation of their antioxidant properties

Lipases from Candida antarctica (Novozyme 435^®), Candida rugosa, Chromobacterium viscosum and Pseudomonas sp. were used to perform transesterifications of vinyl ferulate with hydroxyl-steroids and p-arbutin. The antioxidant activity of the products was evaluated using the free radical 2,2′-diphenyl-1-picrylhydrazyl (DPPH) and 2,2′-azinobis-(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) free radical quenching antioxidant assays, and inhibition of the oxidation of low-density lipoprotein, LDL. Arbutin ferulate was found to possess a 19% higher antiradical activity against the ABTS free radical than its precursor ferulic acid, and it also inhibited the oxidation of LDL more efficiently (by 10%) than its precursors. All of the biocatalytically synthesised products exhibited higher antioxidant activity than Trolox, the well known commercial benchmark antioxidant, and their precursor, ferulic acid.     

Structure-activity relationship analysis of antioxidant ability and neuroprotective effect of gallic acid derivatives

Gallic acid and its derivatives are a group of naturally occurring polyphenol antioxidants which have recently been shown to have potential healthy effects. In order to understand the relationship between the structures of gallic acid derivatives, their antioxidant activities, and neuroprotective effects, we examined their free radical scavenging effects in liposome and anti-apoptotic activities in human SH-SY5Y cell induced by 6-hydrodopamine autooxidation. It was found that these polyphenol antioxidants exhibited different hydrophobicity and could cross through the liposome membrane to react with 1,1-diphenyl-2-picryl-hydrazyl (DPPH) free radical in a time and dose-dependent manner. At the same time, the structure-antioxidant activity relationship of gallic acid derivatives on scavenging DPPH free radical in the liposome was also analyzed based on theoretical investigations. Analysis of cell apoptosis, intracellular GSH levels, production of ROS and the influx of Ca(2+) indicated that the protective effects of gallic acid derivatives in cell systems under oxidative stress depend on both their antioxidant capacities and hydrophobicity. However, the neuroprotective effects of gallic acid derivatives seem to depend more on their molecular polarities rather than antioxidant activities in the human SH-SY5Y cell line. In conclusion, these results reveal that compounds with high antioxidant activity and appropriate hydrophobicity are generally more effective in preventing the injury of oxidative stress in neurodegenerative diseases.     

Antioxidant Combination Inhibits Reactive Oxygen Species Mediated Damage

We examined the preventive activity of naturally occurring antioxidants against three reactive oxygen species using a protein degradation assay. The hydroxyl, hypochlorite, and peroxynitrite radicals are typical reactive oxygen species generated in human body. Previously, we found that hydrophobic botanical antioxidants exhibited specific antioxidant activity against hydroxyl radicals, whereas anserine and carnosine mixture, purified from chicken extract and vitamin C, exhibited antioxidant activities against hypochlorite and peroxynitrite radicals respectively. Since ethanol, used as a solvent in the experiments, also showed an antioxidant action against the hydroxyl radical, we re-assessed antioxidant activities using aqueous solutions of botanical antioxidants. Among the seven hydrophobic antioxidants examined, ferulic acid exhibited the strongest antioxidant activity against the hydroxyl radical. An antioxidant preparation of anserine-carnosine mixture, vitamin C, and ferulic acid prevented oxidative stress by reactive oxygen species. Loss of deformability in human erythrocytes and protein degradation caused by reactive oxygen species were completely inhibited.     

Phenolic-rich juice prevents DNA single-strand breakage and cytotoxicity caused by tert-butylhydroperoxide in U937 cells: the role of iron chelation

The antioxidant potential of phenolic compounds is generally linked to their ability to scavenge free radicals. However, in addition to their radical-scavenging activity, phenolic compounds can chelate metal ions, such as iron, to prevent their participation in Fenton-type reactions, which lead to the formation of free radicals. The aim of the present study was to evaluate the ability of a phenolic-rich juice made from grapes, cherries and berries to protect human myeloid leukemia (U937) cells from oxidative stress caused by tert-butylhydroperoxide (tB-OOH). Preincubation of cells with extracts of the phenolic-rich juice at different concentrations (0-200 microM ferulic acid equivalents) for 3 h partially prevented cell death and abolished the DNA cleavage induced by tB-OOH. Moreover, when preincubating cells with the 100-microM juice extract (the dose that diminished cell death by around 50%), the partial prevention of tB-OOH-induced formation of reactive oxygen species (ROS) and mitochondrial permeability transition pore opening was observed. The radical scavenger antioxidant N,N'-diphenyl-1,4-phenylene-diamine (DPPD) and the intracellular iron chelator o-phenanthroline (o-Phe) were also tested to know whether protective effects depended on radical-scavenging or iron-chelating activities. o-Phe prevented cell death, DNA cleavage and ROS generation, whereas DPPD only prevented cell death, suggesting that phenolics in the juice afforded protection against induced oxidative stress, most probably by means of an iron-chelating mechanism.     

Role of oxygen free radicals in carcinogenesis and brain ischemia

Even though oxygen is necessary for aerobic life, it can also participate in potentially toxic reactions involving oxygen free radicals and transition metals such as Fe that damage membranes, proteins, and nucleic acids. Oxygen free radical reactions and oxidative damage are in most cases held in check by antioxidant defense mechanisms, but where an excessive amount of oxygen free radicals are produced or defense mechanisms are impaired, oxidative damage may occur and this appears to be important in contributing to several pathological conditions including aging, carcinogenesis, and stroke. Several newer methods, such as in vivo spin-trapping, have become available to monitor oxygen free radical flux and quantitate oxidative damage. Using a combination of these newer methods collectively focused on one model, recent results show that oxidative damage plays a key role in brain injury that occurs in stroke. Subtle changes, such as oxidative damage-induced loss of glutamine synthetase activity, may be a key event in stroke-induced brain injury. Oxygen free radicals may play a key role in carcinogenesis by mediating formation of base adducts, such as 8-hydroxyguanine, which can now be quantitated to very low levels. Evidence is presented that a new class of free radical blocking agents, nitrone spin-traps, may help not only to clarify if free radical events are involved, but may help prevent the development of injury in certain pathological conditions.     

Investigations on diverse sesame ( S. indicum L.) germplasm and its wild allies reveal wide variation in antioxidant potential

Free radicals, the key mediators of a range of oxidative reactions involved in lipid oxidation are responsible for food quality deterioration leading to several health hazards. Antioxidants synthesized naturally or synthetically are capable of preventing oxidation of lipids and other related compounds. However, natural antioxidants have many benefits over synthetic ones. Sesame seeds contain large amount of natural bioactive components with high antioxidant potential. In the present study, 14 accessions of sesame containing wild species and cultivars were investigated. The antioxidant potential of sesame seed meal extract was evaluated by total phenolic content (TPC) method using Folin–Ciocalteu reagent, linoleic acid peroxidation by Ferric thiocyanate method, and free radical scavenging assay with 2,2-diphenyl-1-picryl hydrazyl radical. S. laciniatum showed highest mean values for total polyphenol content with maximum % inhibition of linoleic acid peroxidation on 10th day of course of the reaction span and highest antioxidant scavenging power. S. indicum subsp. malabaricum and S. radiatum also showed high total phenol content and radical scavenging capacity. Among the Sesamum indicum cultivars, Gujarat til 2 showed high TPC and high radical scavenging activity. Higher antioxidant property of Sesamum species in comparison to sesame cultivars highlights the need to utilize the wild genepool for the improvement of cultigens for enhanced nutraceutical value.     

Antioxidant functions of phytic acid

Phytic acid is a natural plant antioxidant constituting 1-5% of most cereals, nuts, legumes, oil seeds, pollen and spores. By virtue of forming a unique iron chelate it suppresses iron-catalyzed oxidative reactions and may serve a potent antioxidant function in the preservation of seeds. By the same mechanism dietary phytic acid may lower the incidence of colonic cancer and protect against other inflammatory bowel diseases. Its addition to foods inhibits lipid peroxidation and concomitant oxidative spoilage, such as discoloration, putrefaction, and syneresis. A multitude of other industrial applications are based on the antioxidant function of phytic acid.     

Antidepressant-like effect of a Ginkgo biloba extract (EGb761) in the mouse forced swimming test: role of oxidative stress

EGb761 is a well-defined mixture of active compounds extracted from Ginkgo biloba leaves. This extract is used clinically due to its neuroprotective effects, exerted probably via its potent antioxidant or free radical scavenger action. Previous studies suggest that oxidative stress, via free radical production, may play an important role in depression and animal models for depression-like behavior. Preclinical studies have suggested that antioxidants may have antidepressants properties. The aim of this study was to investigate the antidepressant-like of EGb761 due to its antioxidant role against oxidative stress induced in the forced swimming test, the most widely used preclinical model for assessing antidepressant-like behavior. Male BALB/c mice were pretreated with EGb761 (10 mg/kg, ip) daily for 17 days followed by the forced swimming test and spontaneous locomotor activity. Animals were sacrificed to evaluate lipid peroxidation, different antioxidant enzyme activities, serotonin and dopamine content in midbrain, hippocampus and prefrontal cortex. EGb761 significantly decreased the immobility time (39%) in the forced swimming test. This antidepressant-like effect of EGb761 was associated with a reduction in lipid peroxidation and superoxide radical production (indicated by a downregulation of Mn-superoxide dismutase activity), both of which are indicators of oxidative stress. The protective effect of EGb761 is not related to excitatory or inhibitory effects in locomotor activity, and was also associated with the modulation of serotonergic and dopaminergic neurotransmission. It is suggested that EGb761 produces an antidepressant-like effect, and that an antioxidant effect against oxidative stress may be partly responsible for its observed neuroprotective effects.     

Oxidation of syringic acid by extracellular peroxidase of white-rot fungus,Pleurotus ostreatus

The oxidative degradation of syringic acid by the extracellular peroxidase ofPleurotus ostreatus was studied. Three products formed in the oxidation of syringic acid by the peroxidase in the presence of O₂ and H₂O₂ were identified as 2,6-dimethoxyphenol, 2,6-dimethoxy-1,4-dihydroxybenzene, and 2,6-dimethoxy-1,4-benzoquinone. A free radical was detected as the reaction intermediate of the extracellular peroxidase-catalyzed oxidation of acetosyringone. These results can be explained by mechanisms involving the production of a phenoxy radical and subsequent decarboxylation. This is the first time that 2,6-dimethoxyphenol has been identified in extracellular peroxidase-catalyzed reactions.     

Scavenging of chlorpromazine cation radical by ascorbic acid or glutathione

The report presented here demonstrates that scavenging of chlorpromazine cation radical (an absorption maximum = 530 nm) by ascorbic acid or glutathione can be kinetically and stoichiometrically analyzed at pH 1.5 but not at pH 3.0 and 6.0 using a conventional absorption spectrophotometer. The cation radical decays spontaneously about 10 and 200 times faster at pH 3.0 and 6.0, respectively, than at pH 1.5.At pH 1.5, ascorbic acid scavenges the cation radical faster than glutathione does, and the following different scavenging mechanisms are postulated from the above kinetic and stoichiometric analysis. The reaction of the cation radical with ascorbic acid is second order. The ascorbic acid free radical, which decays mainly by dismutation, is generated by the bimolecular reaction. In the case of glutathione, on the other hand, about 70% of the scavenged cation radical disappears through free radical chain reactions that glutathione thiol anion and glutathione free radical probably initiate. The remaining (about 30%) disappears by conjugation with glutathione. It may be due to relative nonreactivity of ascorbic acid free radical that free radical chain reactions, found commonly in radical chemistry, do not occur in the scavenging reaction by ascorbic acid.Based on the above results, the physiological scavenging mechanisms of the cation radical by the two reducing substances are discussed briefly.     

Oxidative stress associated with exercise,psychological stress and life-style factors

Oxidative stress is a cellular or physiological condition of elevated concentrations of reactive oxygen species that cause molecular damage to vital structures and functions. Several factors influence the susceptibility to oxidative stress by affecting the antioxidant status or free oxygen radical generation. Here, we review the effect of alcohol, air pollution, cigarette smoke, diet, exercise, non-ionizing radiation (UV and microwaves) and psychological stress on the development of oxidative stress.Regular exercise and carbohydrate-rich diets seem to increase the resistance against oxidative stress. Air pollution, alcohol, cigarette smoke, non-ionizing radiation and psychological stress seem to increase oxidative stress. Alcohol in lower doses may act as an antioxidant on low density lipoproteins and thereby have an anti-atherosclerotic property.