高分子抗氧化剂研究进展

过量自由基可造成人体的多种损伤,加速人的衰老.抗氧化剂可以清除或转化多余的自由基,对人体起到保护作用.本文从天然高分子及合成高分子两方面对抗氧化剂的研究进展进行了综述.在天然高分子抗氧化剂中,超氧化物歧化酶(SOD)、过氧化氢酶(CAT)及谷胱甘肽过氧化物酶等的结构与功能已经较为清晰;金属硫蛋白、血红蛋白、短肽及多糖等...     

EFFECTS OF CHRONIC EXPOSURE ULTRAVIOLET-A INCLUDING 2% ULTRAVIOLET-B ON FREE RADICAL REDUCTION SYSTEMS IN HAIRLESS MICE

Chronic ultraviolet (UV) irradiation is known to cause a variety of changes in the skin, including wrinkles, pigmented spots and carcinogenesis. To explore time dependent changes in several parameters with chronic UV irradiation, we examined the molecular changes in connective tissue, intracellular defence enzymes and free radical antioxidant substances in hairless mice skin caused by chronic exposure to UV-A including 2% UV-B. Connective tissue changes were estimated using hydroxyproline and isodesmosine assays as a measure of collagen and elastin concentrations, respectively. After 6 weeks irradiation, the insoluble collagen and elastin were both substantially elevated, as were the activities of glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD). Continued UV irradiation resulted in a steady decline in SOD and lipid soluble antioxidants, while the GSH-Px remained elevated, suggesting that SOD and lipid soluble antioxidants in the skin may be involved in protecting it from UV damage and deteriorate with chronic irradiation.     

Bucillamine prevents cisplatin-induced ototoxicity through induction of glutathione and antioxidant genes

Bucillamine is used for the treatment of rheumatoid arthritis. This study investigated the protective effects of bucillamine against cisplatin-induced damage in auditory cells, the organ of Corti from postnatal rats (P2) and adult Balb/C mice. Cisplatin increases the catalytic activity of caspase-3 and caspase-8 proteases and the production of free radicals, which were significantly suppressed by pretreatment with bucillamine. Bucillamine induces the intranuclear translocation of Nrf2 and thereby increases the expression of γ-glutamylcysteine synthetase (γ-GCS) and glutathione synthetase (GSS), which further induces intracellular antioxidant glutathione (GSH), heme oxygenase 1 (HO-1) and superoxide dismutase 2 (SOD2). However, knockdown studies of HO-1 and SOD2 suggest that the protective effect of bucillamine against cisplatin is independent of the enzymatic activity of HO-1 and SOD. Furthermore, pretreatment with bucillamine protects sensory hair cells on organ of Corti explants from cisplatin-induced cytotoxicity concomitantly with inhibition of caspase-3 activation. The auditory-brainstem-evoked response of cisplatin-injected mice shows marked increases in hearing threshold shifts, which was markedly suppressed by pretreatment with bucillamine in vivo. Taken together, bucillamine protects sensory hair cells from cisplatin through a scavenging effect on itself, as well as the induction of intracellular GSH.     

Oxidative stress and defence mechanisms of the freshwater cladoceran Daphnia longispina exposed to UV radiation

The aim of the present study is to evaluate the occurrence of oxidative stress in the cladoceran Daphnia longispina exposed to UV-A and UV-B radiation. The activity of antioxidant enzymes and lipid peroxidation markers is investigated and the protective action of ascorbic acid determined. Results show differences in the lethality radioinduced by UV-A and UV-B. Both UV-A and UV-B exposure cause an important increase in malonaldehyde (MDA) concentration and catalase activity. Ascorbic acid addition reduces the MDA concentration, indicating that the oxidative stress caused by either UV-A or UV-B radiation can be controlled by antioxidants. The increase of the antioxidant enzymes may be a response mechanism to oxidative stress.     

子宫肌瘤患者血清SOD,MDA的观察

检测了６６例子宫肌瘤患者血清中超氧化物歧化酶（ＳＯＤ）和丙二醛（ＭＤＡ）的含量,探讨其与子宫肌瘤发病的关系。结果表明,子宫肌瘤患者血清中ＳＯＤ的活性较健康妇女明显降低,而ＭＤＡ含量明显增加,两者呈负相关,提示自由基在子宫肌瘤的发病中起着重要作用．     

Effect of ascorbic acid (vitamin C) on the ESR spectra of the red and black hair: pheomelanin free radicals are not always present in red hair

Increased incidence of melanoma in the population with red hair is conditioned by synthesis of pheomelanin pigments in the skin and their phototoxic properties. The recent research has shown that free radicals of pheomelanin are produced not only by the influence of UV irradiation, but also in UV‐independent pathways of oxidative stress. It has been ascertained, that the color of the hair is not always determinant of the amount of pheolemanin radicals in red hair. Therefore, in order to evaluate the risk of melanoma in different individuals, it is necessary to define the amount of free radicals of pheomelanin in red hair using ESR spectroscopy method. Besides, it is very important to find effective antioxidant, capable of neutralizing free radicals of pheomelanin. It was proved that ascorbic acid neutralizes free radicals of pheomelanin very effectively. The main goal of our research was to define the presumably optimal concentration of ascorbic acid as an antioxidant and study the kinetics of the influence of this concentration on red and black hair. It has been found out, that ascorbic acid influences the free radicals of red and black hair, and its appropriate optimal concentration is 10 mM. The obtained results can be considered in dermatology and cosmetology. Copyright © 2015 John Wiley & Sons, Ltd.     

POTENTIATION OF OXIDATIVE DAMAGE TO PROTEINS BY ULTRAVIOLET-A AND PROTECTION BY ANTIOXIDANTS

We have studied the damage of alcohol dehydrogenase (ADH) and glyceraldehyde 3-phosphate dehydrogenase (GAPD) induced by Fe++/EDTA + H2O2 in combination with UV-A (main output at 365 nm). Enzyme inactivation, formation of hydroxyl radicals (measured in the absence of enzymes), increase in protein carbonyls, oxidation of sulfhydryl (SH) groups, loss of native protein fluorescence, and enhanced protease degradation were used to determine protein damage. Hydroxyl radical production was greatly enhanced by the combination of UV-A with Fe++/EDTA + H2O2. The combined treatment increased protein carbonyls but decreased native protein fluorescence and SH groups. The combined treatment caused turbidity in GAPD but not in ADH, whereas trypsin susceptibility was increased more in ADH than in GAPD. These measurements of protein oxidation correlated well with enzyme activities. Glyceraldehyde 3-phosphate dehydrogenase and dithiothreitol were most protective against such damage, while hydroxyl radical and singlet oxygen scavengers were partially effective. Superoxide dismutase had no effect. Thus, UV-A potentiation of protein damage induced by FE++/EDTA + H2O2 appeared to involve hydroxyl radicals and perhaps singlet oxygen but not superoxide radicals. The damage to proteins induced by combination of UV-A with physiological oxidants, iron ions and H2O2 may be relevant to UV-A-induced skin and tissue damage.     

Molecular Mechanism of Drug Photosensitization. IX. Effect of Inorganic Ions on DNA Cleavage Photosensitized by Naproxen

Abstract— Photocleavage of DNA induced by naproxen and the correlated protective effect by some inorganic ions have been considered. The presence of a DNA complex is suggested and only associated naproxen seems to be responsible for the cleavage, for which the quantum yield of single strand breaks was calculated. The inorganic ions I^-, Mn²⁺, Co²⁺ and Cu²⁺ decrease naproxen-photoin-duced DNA cleavage. Iodide acts by a heavy atom mechanism, thus inhibiting naproxen photolysis and decreasing the amount of free radicals responsible for the photocleavage both in aerobic and anaerobic conditions. Metallic ions protect only within a range of concentrations, as for higher amounts damaging processes are observed. The protective efficiency of cations decreases with the increase of free drug concentration in the bulk of the solution, due to their involvement in the scavenging of naproxen radicals generated by photolysis of the free drug. In the presence of EDTA the cations show a better protective action. The most likely hypothesis is an inhibiting effect on the damaging processes via a redox cycle. The different behaviors of copper and of the two other cations can be justified by the influence of redox potentials of free and complexed metals and by the superoxide dis-mutase-like activity of copper.     

A Concise Review of Current In Vitro Chemical and Cell-Based Antioxidant Assay Methods

Antioxidants remain interesting molecules of choice for suppression of the toxic effects of free radicals in foods and human systems. The current practice involves the use of mainly synthetic molecules as potent antioxidant agents. However, due to the potential negative impact on human health, there is an intensive effort within the research community to develop natural alternatives with similar antioxidant efficacy but without the negative side effects of synthetic molecules. Still, the successful development of new molecules depends on the use of reliable chemical or cell culture assays to screen antioxidant properties. Chemical antioxidant assays include the determination of scavenging ability against free radicals such as DPPH, superoxide anion radicals, hydroxyl radicals, hydrogen peroxide, and nitric oxide. Other antioxidant tests include the ability of compounds to bind and sequester prooxidant metal cations, reduce ferric iron, and attenuate the rate of lipid oxidation. Ex vivo tests utilize cell cultures to confirm entry of the molecules into cells and the ability to quench synthetic intracellular free radicals or to stimulate the increased biosynthesis of endogenous antioxidants. In order to assist researchers in their choice of antioxidant evaluation methods, this review presents background scientific information on some of the most commonly used antioxidant assays with a comparative discussion of the relevance of published literature data to food science and human nutrition applications.     

SPECTROSCOPIC STUDIES OF CUTANEOUS PHOTOSENSITIZING AGENTS—X. A SPIN-TRAPPING AND DIRECT ELECTRON SPIN RESONANCE STUDY OF THE PHOTOCHEMICAL PATHWAYS OF DAUNOMYCIN AND ADRIAMYCIN

Abstract— Irradiation of daunomycin (or adriamycin) and the spin trap 5,5-dimethyl-l-pyrroline-1-oxide (DMPO) at 490 nm in the presence or in the absence of air generated the hydroxyl radical adduct (DMPO-OH). The observed DMPO-OH signal was not affected by the addition of hydroxyl radical scavengers (ethanol, formate), suggesting that direct trapping of the hydroxyl radical was not involved. The DMPO-OH signal was insensitive to superoxide dismutase and catalase, which ruled out the possibility of superoxide or H₂O₂ involvement. These findings demonstrate that daunomycin (or adriamycin) does not generate hydroxyl radicals or superoxide radical anions when subjected to 490-nm excitation. However, when daunomycin (or adriamycin) was irradiated at 310 nm DMPO adducts derived from two carbon-centered radicals, superoxide and the hydroxyl radical were detected. The superoxide adduct of DMPO was abolished by the addition of SOD, providing unequivocal evidence for the generation of the superoxide anion radical. The daunomycin semiquinone radical, observed upon 310-nm irradiation of daunomycin in the absence of DMPO, appears to be the precursor of the superoxide radical anion. One of the carbon-centered radicals trapped by DMPO exhibited a unique set of hyperfine parameters and was identified as an acyl radical. This suggests that the known photochemical deacylation of daunomycin occurs via a homolytic cleavage mechanism. The free radicals generated photolytically from adriamycin and daunomycin may be involved in the etiology of the skin ulceration and inflammation caused by these drugs. A knowledge of the dependence of these photogenerated radicals on the wavelength of excitation may be important in the development of adriamycin and daunomycin for photodynamic therapy.     

Loading of free radicals on the functional graphene combined with liquid chromatography–tandem mass spectrometry screening method for the detection of radical-scavenging natural antioxidants

A novel free radical reaction combined with liquid chromatography electrospray ionization tandem mass spectrometry (FRR-LC–PDA-ESI/APCI-MS/MS) screening method was developed for the detection and identification of radical-scavenging natural antioxidants. Functionalized graphene was prepared by chemical method for loading free radicals (superoxide radical, peroxyl radical and PAHs free radical). Separation was performed with and without a preliminary exposure of the sample to specific free radicals on the functionalized graphene, which can facilitate reaction kinetics (charge transfers) between free radicals and potential antioxidants. The difference in chromatographic peak areas is used to identify potential antioxidants. The structure of the antioxidants in one sample (Swertia chirayita) is identified using MS/MS and comparison with standards. Thirteen compounds were found to possess potential antioxidant activity, and their free radical-scavenging capacities were investigated. The thirteen compounds were identified as 1,3,5-trihydroxyxanthone-8-O-β-d-glucopyranoside (PD1), norswertianin (PD2), 1,3,5,8-tetrahydroxyxanthone (PD3), 3, 3′, 4′, 5, 8-penta hydroxyflavone-6-β-d-glucopyranosiduronic acid-6′-pentopyranose-7-O-glucopyranoside (PD4), 1,5,8-trihydroxy-3-methoxyxanthone (PD5), swertiamarin (PS1), 2-C-β-d-glucopyranosyl-1,3,7-trihydroxylxanthone (PS2), 1,3,7-trihydroxylxanthone-8-O-β-d-glucopyranoside (PL1), 1,3,8-trihydroxyl xanthone-5-O-β-d-glucopyranoside (PL2), 1,3,7-trihydroxy-8-methoxyxanthone (PL3), 1,2,3-trihydroxy-7,8-dimethoxyxanthone (PL4), 1,8-dihydroxy-2,6-dimethoxy xanthone (PL5) and 1,3,5,8-tetramethoxydecussatin (PL6). The reactivity and SC₅₀ values of those compounds were investigated, respectively. PD4 showed the strongest capability for scavenging PAHs free radical; PL4 showed prominent scavenging capacities in the lipid peroxidation processes; it was found that all components in S. chirayita exhibited weak reactivity in the superoxide radical scavenging capacity. The use of the free radical reaction screening method based on LC–PDA-ESI/APCI-MS/MS would provide a new approach for rapid detection and identification of radical-scavenging natural antioxidants from complex matrices.     

Mechanisms of antioxidant action: The reaction of hindered phenols with rubber in the presence of free radicals

Simple hindered phenols, containing ortho or para methyl groups, react with natural rubber latex in the presence of oxidizing free radicals (alkoxy or acyloxy), giving up to 20% yields of bound antioxidant. This reaction is most efficient when the concentration of the phenolic antioxidant in the rubber is less than 1%. As the concentration is increased, side reactions involving both the phenol (stilbenequinone formation) and the rubber (cross-linking) supervenes. The efficiency of the bound antioxidants is much higher than conventional antioxidants under aggressive conditions of air oven ageing and solvent or detergent extraction. This effect is due primarily to the non-removal of the bound antioxidants under these conditions whereas the conventional antioxidants are removed completely.     

A Study of the Antioxidant Effect of Flavonic Compounds for Preventing Lipid Oxidation by Using Fluorescence Spectroscopy

In this paper a study of the effect of flavonic compounds in preventing and/or reducing the membrane lipid oxidation due to free radical attack was performed by using fluorescence spectroscopy techniques. Lipid peroxidation was investigated by using liposomes-artificial membrane models, which were prepared by lipid hydration method. Their oxidation was performed with a 15-W ultraviolet germicidal lamp having wavelength radiation 253.7 nm. In the first series of experiments, the protective effect of two synthetic antioxidants (quercetin and caffeic acid) against free radicals was monitored, while in the second series two 70% hydroalcoholic extracts from blueberry and blackcurrant leaves was used. It was determined that natural antioxidants have a much higher antioxidant power against free radicals than synthetic compounds but they degrade after two hours of oxidation. Liposomes are better protected when using natural antioxidants, but their degradation is completed more quickly, than in the case of synthetic antioxidants.     

芯片毛细管电泳评估脂质体介导超氧化物歧化酶减低细胞内氧化应激

超氧化物歧化酶(SOD)可用作抗氧化的药物。它能催化并清除细胞内的活性氧组分(ROS),保护细胞免受自由基的氧化破坏。但是由于SOD分子量较大,难以透过细胞膜进入细胞内,显著降低了SOD的药效。本研究用激光共聚焦荧光显微镜拍摄的荧光图像说明,纳米脂质体可介导SOD进入细胞。用芯片毛细管电泳激光诱导荧光分析法(MCE-LIF)测定单细胞中ROS和谷胱甘肽(GSH)的荧光信号强度,评估了用脂质体包裹的SOD与细胞作用的抗氧化效果。用脂质体包裹的SOD与肝癌细胞共培养2h,与直接用SOD作用于肝癌细胞相比较,细胞内ROS明显降低,GSH明显提高。实验结果说明,用脂质体包裹SOD是一种减低细胞内氧化应激的有效给药途径。     

Development of a Capillary Electrophoresis Method to Monitor Protein Oxidation and Antioxidant Protection

The development of a new capillary zone electrophoresis (CZE) method for the determination of protein oxidation by free radicals is described. The effects of various experimental conditions, such as type of capillary, pH, buffer concentration, capillary dimensions, temperature, applied voltage, and addition of some organic solvents, were investigated. The sample injection procedure was also optimized. Target proteins investigated here (lysozyme, human serum albumin, and β-lactoglobulin A) were effectively damaged by free radicals generated in the aqueous phase from 2,2′-azobis[2-amidinopropane] (AAPH). The new method allowed global protein fragmentation to be quantified as a function of time by monitoring the decrease in peak height of the native protein. Furthermore, the protective efficacy of antioxidants was assessed by concentration-inhibition curves against protein fragmentation. The method is thus suitable to screen compounds able to protect proteins against oxidative damage.     

The role of melanin as protector against free radicals in skin and its role as free radical indicator in hair

Throughout the body, melanin is a homogenous biological polymer containing a population of intrinsic, semiquinone-like radicals. Additional extrinsic free radicals are reversibly photo-generated by UV and visible light. Melanin photochemistry, particularly the formation and decay of extrinsic radicals, has been the subject of numerous electron spin resonance (ESR) spectroscopy studies. Several melanin monomers exist, and the predominant monomer in a melanin polymer depends on its location within an organism. In skin and hair, melanin differs in content of eumelanin or pheomelanin. Its bioradical character and its susceptibility to UV irradiation makes melanin an excellent indicator for UV-related processes in both skin and hair. The existence of melanin in skin is strongly correlated with the prevention against free radicals/ROS generated by UV radiation. Especially in the skin melanin (mainly eumelanin) ensures the only natural UV protection by eliminating the generated free radicals/ROS. Melanin in hair can be used as a free radical detector for evaluating the efficacy of hair care products. The aim of this study was to investigate the suitability of melanin as protector of skin against UV generated free radicals and as free radical indicator in hair.     

Physiological Responses of Scenedesmus quadricauda (Chlorophyceae) to UV-A and UV-C Light

Despite intensive research focused on the effects of UV-B, deeper metabolic responses to UV-A and UV-C are still scarce. Besides, especially microalgal species had to develop efficient protective features in comparison with tissue structure of vascular plants. We exposed axenic cultures of Scenedesmus quadricauda (Chlorophyceae) to UV-A (366 nm) and UV-C (254 nm) light over 1 h. Both wavelengths stimulated increase in soluble proteins, superoxide radical and hydrogen peroxide, but had a nonsignificant effect on cell viability. Within 17 detected free amino acids, five (including proline) increased in response to UV-A while only aspartic acid and histidine increased in UV-C treatment. Total soluble phenols and flavonoids were influenced neither by UV-A nor by UV-C while selected flavonols (quercetin and kaempferol) decreased in UV-A and were not detected in UV-C treatment. Benzoic acid derivatives increased preferentially after UV-A illumination (vanillic acid and vanillin) while cinnamic derivatives (caffeic, chlorogenic and p-coumaric acids) decreased in both UV-A and UV-C. It is concluded that UV light stimulated oxidative stress while exposure time was not sufficient to stimulate larger changes in phenolic metabolites. Present findings in the context of available data and with emphasis on phenolics in algae are discussed.     

Photochemistry and phototoxicity of fluocinolone 16,17-acetonide

Fluocinolone 16,17-acetonide is a corticosteroid used topically to treat various inflammatory skin diseases. Its photoreactivity was studied under UV-A and UV-B light in aqueous buffer in the presence of oxygen. This drug is photolabile under UV-B light and, to a lesser extent, under UV-A light, which is absorbed far less. In phosphate buffer, approximately 80% of fluocinolone acetonide decomposes after 5 J/cm2 of UV-B irradiation, whereas under 30 J/cm2 of UV-A light approximately only 20% decomposes. Both the drug and its photoproducts have been evaluated through a battery of in vitro studies and found to cause photohemolysis and induce photodamage to proteins (erythrocyte ghosts, bovine serum albumin) and linoleic acid. In addition, one of the photoproducts (the 17-hydroperoxy derivative) is highly toxic in the dark. Therefore, both loss of therapeutic activity and light-induced adverse effects may be expected when patients expose themselves to sunlight after drug administration. A major mechanism for phototoxicity involves radicals forming from drug breakdown, at least under UV-B, although reactive oxygen species may play a role, particularly under UV-A.     

Biochemical changes in hairless mouse skin collagen after chronic exposure to ultraviolet-A radiation.

Evidence is mounting that UV-B and UV-A radiation affect skin differently in responses as diverse as erythema and elastosis. We found in this study that collagen metabolism was also differentially affected. Albino hairless mice were irradiated with two UV-A sources: (1) UVASUN 3000 (340-400 nm) for cumulative exposures of 4000 and 8000 J/cm2; (2) a xenon solar simulator filtered to provide full spectrum UV-A (320-400 nm) and long wavelength UV-A (335-400 nm) for cumulative exposures of 3000 and 4000 J/cm2 respectively. Collagen was isolated from other skin proteins by acid extraction, pepsin digestion and salt precipitation. Collagen types I and III were separated by interrupted gel electrophoresis. Ultraviolet-A rendered the collagen highly resistant to pepsin digestion. In age-matched controls only 16-18% of the total collagen remained insoluble, whereas in long wavelength UV-A-irradiated skins the insoluble fraction was as high as 87%. A dose response was noted at 4000 and 8000 J/cm2 as delivered by the UVASUN. Recovery of collagen from the pepsin soluble fraction was low in all UV-A groups and the amount of type III so small that determination of ratios of type III to I collagen was unreliable. These results suggest that chronic UV-A radiation may increase cross-linking of dermal collagen.