On 'The thiobarbituric acid reaction and the autoxidations of polyunsaturated fatty acid methyl esters' by Leland K.Dahle, Eldon G.Hill and Ralph T.Holman.

This commentary describes a highly cited paper by Dahle, Hill, and Holman, Arch Biochem Biophys. 1962; 98: 253-261. They showed that the oxidation products of polyunsaturated fatty acids reacted with thiobarbituric acid to give a colored product, which might be used to assess lipid oxidation.

Trans-unsaturated fatty acid activates NLRP3 inflammasome in macrophages and exacerbates intestinal inflammation in mice.

Higher consumption of trans fatty acid (TFA) is a risk factor for several inflammatory diseases including inflammatory bowel disease (IBD). However, the detailed mechanisms by which TFA intake affects IBD pathology remain unclear. We demonstrate here that elaidate, a trans-isomer of oleate, enhances interleukin (IL)-1ß production through the activation of NLRP3 inflammasome in mouse bone marrow-derived macrophages (BMDMs). Oleate has no effect on IL-1ß production. Elaidate also induces oxidative stress and activates endoplasmic reticulum stress in BMDMs. The elaidate-induced IL-1ß production is suppressed by co-treatments with antioxidants and a chemical chaperone. Furthermore, we analyze the effects of elaidate administration on intestinal inflammation using 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis model in mice. Increased colonic damage and myeloperoxidase activity after TNBS treatment are elevated by elaidate administration. Also, TNBS treatment induces IL-1ß production in colonic mucosa; elaidate administration enhances the induction. We believe that these data reveal some mechanisms by which the TFA intake is associated with increased risk for IBD.

Isomer distribution of hydroxyoctadecadienoates (HODE) and hydroxyeicosatetraenoates (HETE) produced in the plasma oxidation mediated by peroxyl radical, peroxynitrite, hypochlorite, 15-lipoxygenase, and singlet oxygen.

Free and ester forms of unsaturated fatty acids and cholesterol are oxidized in vivo by multiple oxidants to give diverse products. Some lipid oxidation is mediated by enzymes to selectively give specific products, while others proceed randomly to produce mixtures of many kinds of regioisomers and stereoisomers. The efficacy of antioxidants against lipid oxidation depends on the nature of the oxidants and therefore the identification of oxidant is important for understanding the roles and effects of lipid oxidation and antioxidants in vivo. In the present study, the isomer distribution of hydro(pero)xyoctadecadienoates (H(p)ODEs) and hydro(pero)xyeicosatetraenoates (H(p)ETEs), the most abundant lipid oxidation products found in human plasma, produced in the oxidation of plasma by peroxyl radicals, peroxynitrite, hypochlorite, 15-lipoxygenase, and singlet oxygen were examined. It was shown that 9- and 13-(E,E)-HODEs, 13(S)-(Z,E)-HODE, and 10- and 12-(Z,E)-HODEs were specific lipid oxidation products by free radical, 15-lipoxygenase, and singlet oxygen, respectively. The isomer distribution of HODEs produced by peroxynitrite was similar to that by peroxyl radical, suggesting that the peroxynitrite mediated lipid oxidation proceeds by free radical mechanisms. The production of HODEs and HETEs by hypochlorite was very small. HODEs may be a better biomarker than HETEs since linoleates are oxidized by simpler mechanisms than arachidonates and all the HODEs isomers can be quantified more easily. These products may be used as specific biomarkers for the identification of responsible oxidants and for the assessment of oxidant-specific lipid oxidation levels and effects of antioxidants in vivo.

Oxidative stress and antioxidants: Distress or eustress?

There is a growing consensus that reactive oxygen species (ROS) are not just associated with various pathologies, but that they act as physiological redox signaling messenger with important regulatory functions. It is sometimes stated that "if ROS is a physiological signaling messenger, then removal of ROS by antioxidants such as vitamins E and C may not be good for human health." However, it should be noted that ROS acting as physiological signaling messenger and ROS removed by antioxidants are not the same. The lipid peroxidation products of polyunsaturated fatty acids and cholesterol induce adaptive response and enhance defense capacity against subsequent oxidative insults, but it is unlikely that these lipid peroxidation products are physiological signaling messenger produced on purpose. The removal of ROS and inhibition of lipid peroxidation by antioxidants should be beneficial for human health, although it has to be noted also that they may not be an effective inhibitor of oxidative damage mediated by non-radical oxidants. The term ROS is vague and, as there are many ROS and antioxidants which are different in chemistry, it is imperative to explicitly specify ROS and antioxidant to understand the effects and role of oxidative stress and antioxidants properly.

Inhibition of plasma lipid oxidation induced by peroxyl radicals, peroxynitrite, hypochlorite, 15-lipoxygenase, and singlet oxygen by clinical drugs.

With increasing evidence showing the involvement of oxidative stress in the pathogenesis of various diseases, the effects of clinical drugs possessing antioxidant functions have received much attention. The unregulated oxidative modification of biological molecules leading to diseases is mediated by multiple oxidants including free radicals, peroxynitrite, hypochlorite, lipoxygenase, and singlet oxygen. The capacity of antioxidants to scavenge or quench oxidants depends on the nature of oxidants. In the present study, the antioxidant effects of several clinical drugs against plasma lipid oxidation induced by the aforementioned five kinds of oxidants were investigated from the production of lipid hydroperoxides, which have been implicated in the pathogenesis of various diseases. Troglitazone acted as a potent peroxyl radical scavenger, whereas probucol and edaravone showed only moderate reactivity and carvedilol, pentoxifylline, and ebselen did not act as radical scavenger. Probucol and edaravone suppressed plasma oxidation mediated by peroxynitrite and hypochlorite. Troglitazone and edaravone inhibited 15-lipoxygenase mediated plasma lipid oxidation, the IC50 being 20 and 34µM respectively. None of the drugs used in this study suppressed plasma lipid oxidation by singlet oxygen. This study shows that the antioxidant effects of drugs depend on the nature of oxidants and that antioxidants against multiple oxidants are required to cope with oxidative stress in vivo.

Rapid assessment of singlet oxygen-induced plasma lipid oxidation and its inhibition by antioxidants with diphenyl-1-pyrenylphosphine (DPPP).

Recent studies suggesting the involvement of singlet oxygen in the pathogenesis of multiple diseases have attracted renewed attention to lipid oxidation mediated by singlet oxygen. Although the rate constants for singlet oxygen quenching by antioxidants have been measured extensively, the inhibition of lipid oxidation mediated by singlet oxygen has received relatively less attention, partly because a convenient method for measuring the rate of lipid oxidation is not available. The objective of this study was to develop a convenient method to measure plasma lipid oxidation mediated by singlet oxygen which may be applied to a rapid assessment of the antioxidant capacity to inhibit this oxidation using a conventional microplate reader. Singlet oxygen was produced from naphthalene endoperoxide, and lipid hydroperoxide production was followed by using diphenyl-1-pyrenylphosphine (DPPP). Non-fluorescent DPPP reacts stoichiometrically with lipid hydroperoxides to give highly fluorescent DPPP oxide. It was found that plasma oxidation by singlet oxygen increased the fluorescence intensity of DPPP oxide, which was suppressed by antioxidants. Fucoxanthin suppressed the oxidation more efficiently than ß-carotene and α-tocopherol, while ascorbic acid and Trolox were not effective. The present method may be useful for monitoring lipid oxidation and also for rapid screening of the capacity of dietary antioxidants and natural products to inhibit lipid oxidation in a biologically relevant system.

Enhancement of lipid peroxidation and its amelioration by vitamin E in a subject with mutations in the SBP2 gene.

Selenocysteine (Sec) insertion sequence-binding protein 2 (SBP2) is essential for the biosynthesis of Sec-containing proteins, termed selenoproteins. Subjects with mutations in the SBP2 gene have decreased levels of several selenoproteins, resulting in a complex phenotype. Selenoproteins play a significant role in antioxidative defense, and deficiencies in these proteins can lead to increased oxidative stress. However, lipid peroxidation and the effects of antioxidants in subjects with SBP2 gene mutations have not been studied. In the present study, we evaluated the lipid peroxidation products in the blood of a subject (the proband) with mutations in the SBP2 gene. We found that the proband had higher levels of free radical-mediated lipid peroxidation products, such as 7ß-hydroxycholesterol, than the control subjects. Treatment of the proband with vitamin E (α-tocopherol acetate, 100 mg/day), a lipid-soluble antioxidant, for 2 years reduced lipid peroxidation product levels to those of control subjects. Withdrawal of vitamin E treatment for 7 months resulted in an increase in lipid peroxidation products. Collectively, these results clearly indicate that free radical-mediated oxidative stress is increased in the subject with SBP2 gene mutations and that vitamin E treatment effectively inhibits the generation of lipid peroxidation products.

Biomarkers of lipid peroxidation in clinical material.

BACKGROUND: Free radical-mediated lipid peroxidation has been implicated in a number of human diseases. Diverse methods have been developed and applied to measure lipid peroxidation products as potential biomarkers to assess oxidative stress status in vivo, discover early indication of disease, diagnose progression of disease, and evaluate the effectiveness of drugs and antioxidants for treatment of disease and maintenance of health, respectively. However, standardized methods are not yet established. SCOPE OF REVIEW: Characteristics of various lipid peroxidation products as biomarkers are reviewed on the basis of mechanisms and dynamics of their formation and metabolism and also on the methods of measurement, with an emphasis on the advantages and limitations. MAJOR CONCLUSIONS: Lipid hydroxides such as hydroxyoctadecadienoic acids (HODE), hydroxyeicosatetraenoic acids (HETE), and hydroxycholesterols may be recommended as reliable biomarkers. Notably, the four HODEs, 9-cis,trans, 9-trans,trans, 13-cis,trans, and 13-trans,trans-HODE, can be measured separately by LC-MS/MS and the trans,trans-forms are specific marker of free radical mediated lipid peroxidation. Further, isoprostanes and neuroprostanes are useful biomarker of lipid peroxidation. It is important to examine the distribution and temporal change of these biomarkers. GENERAL SIGNIFICANCE: Despite the fact that lipid peroxidation products are non-specific biomarkers, they will enable to assess oxidative stress status, disease state, and effects of drugs and antioxidants. This article is part of a Special Issue entitled Current methods to study reactive oxygen species - pros and cons and biophysics of membrane proteins. Guest Editor: Christine Winterbourn.

Vitamin E nomenclature. Is RRR-α-tocopherol the only vitamin E?

It has generally been accepted that vitamin E refers to a group of tocochromanols, α-, ß-, γ-, and δ-tocopherols and the corresponding four tocotrienols. Recently, Azzi and colleagues proposed to restrict the term vitamin E only to RRR-α-tocopherol, not to other tocopherols and tocotrienols (Azzi A et al. Free Radic Biol Med. 2023; 207:178-180. doi: 10.1016/j.freeradbiomed.2023.06.029). The aim of this paper is to express our opinion on the nomenclature of vitamin E based on available scientific data. In our opinion, it would be inappropriate to exclude all the tocochromanols other than RRR-α-tocopherol from the vitamin E group at this stage when the molecular mechanisms showing how vitamin E deficiency causes diseases such as ataxia and how vitamin E prevents/reverses such diseases are not elucidated. Understanding of whole functions of tocochromanols including underlying mechanisms and dynamics is essential before revision of currently accepted definition of vitamin E. The potential roles of γ-tocopherol and tocotrienols are discussed despite whether they are vitamin function should be clarified in the future studies.

Cholesterol is more readily oxidized than phospholipid linoleates in cell membranes to produce cholesterol hydroperoxides.

Cholesterol is an essential component of cell membranes and serves as an important precursor of steroidal hormones and bile acids, but elevated levels of cholesterol and its oxidation products have been accepted as a risk factor for maintenance of health. The free and ester forms of cholesterol and fatty acids are the two major biological lipids. The aim of this hypothesis paper is to address the long-standing dogma that cholesterol is less susceptible to free radical peroxidation than polyunsaturated fatty acids (PUFAs). It has been observed that cholesterol is peroxidized much slower than PUFAs in plasma but that, contrary to expectations from chemical reactivity toward peroxyl radicals, cholesterol appears to be more readily autoxidized than linoleates in cell membranes. The levels of oxidation products of cholesterol and linoleates observed in humans support this notion. It is speculated that this discrepancy is ascribed to the fact that cholesterol and phospholipids bearing PUFAs are localized apart in raft and non-raft domains of cell membranes respectively and that the antioxidant vitamin E distributed predominantly in the non-raft domains cannot suppress the oxidation of cholesterol lying in raft domains which are relatively deficient in antioxidant.

Chromium(III) oxide nanoparticles induced remarkable oxidative stress and apoptosis on culture cells.

Chromium(III) oxide (Cr(2)O(3)) is used for industrial applications such as catalysts and pigments. In the classical form, namely the fine particle, Cr(2)O(3) is insoluble and chemically stable. It is classified as a low-toxicity chromium compound. Recently, industrial application of nanoparticles (a new form composed of small particles with a diameter of ≤100 nm, in at least one dimension) has been increasing. Cellular effects induced by Cr(2)O(3) nanoparticles are not known. To shed light upon this, the release of soluble chromium from Cr(2)O(3) nano- and fine-particles in culture medium was compared. Fine Cr(2)O(3) particles were insoluble in the culture medium; on the contrary, Cr(2)O(3) nanoparticles released soluble hexavalent chromium into the culture medium. Cr(2)O(3) nanoparticles showed severe cytotoxicity. The effect of Cr(2)O(3) nanoparticles on cell viability was higher than that of fine particles. Cr(2)O(3) nanoparticles showed cytotoxicity equal to that of hexavalent chromium (K(2)Cr(2)O(7)). Human lung carcinoma A549 cells and human keratinocyte HaCaT cells showed an increase in intracellular reactive oxygen species (ROS) level and activation of antioxidant defense systems on exposure to Cr(2)O(3) nanoparticles. Exposure of Cr(2)O(3) nanoparticles led to caspase-3 activation, showing that the decrease in cell viability by exposure to Cr(2)O(3) nanoparticles was caused by apoptosis. Cellular responses were stronger in the Cr(2)O(3) nanoparticles-exposed cells than in fine Cr(2)O(3) - and CrCl(3) -exposed cells. Cellular uptake of Cr(2)O(3) particles were observed in nano- and fine-particles. The cellular influence of the extracellular soluble trivalent chromium was lower than that of Cr(2)O(3) nanoparticles. Cr(2)O(3) nanoparticles showed cytotoxicity by hexavalent chromium released at outside and inside of cells. The cellular influences of Cr(2)O(3) nanoparticles matched those of hexavalent chromium. In conclusion, Cr(2)O(3) nanoparticles have a high cytotoxic potential.

Actions of Thiols, Persulfides, and Polysulfides as Free Radical Scavenging Antioxidants.

Significance: The essential roles of thiol compounds as redox signaling mediators and protectors have been established. Recently, the roles of persulfides and polysulfides as mediators involved in numerous physiological processes have been revealed. Recent Advances: Recently, it became possible to detect and measure persulfides and polysulfides in human fluids and tissues and their physiological functions, including cellular signaling and protection against oxidative stress, have been reported, but the underlying mechanisms and dynamics remain elusive. Critical Issues: Physiological functions of thiol compounds have been studied, focusing primarily on two-electron redox reactions. In contrast, the contribution of one-electron redox mechanisms, that is, free radical-mediated oxidation and antioxidation, has received much less attention. Considering the important effects of free radical-mediated oxidation of biological molecules on pathophysiology, the antioxidant functions of thiol compounds as free radical scavengers are challenging issues. Future Directions: The antioxidant actions and dynamics of thiols, hydropersulfides, and hydropolysulfides as free radical scavenging antioxidants and their physiological significance remain to be established. Antioxid. Redox Signal. 39, 728-743.

Comparison of acute oxidative stress on rat lung induced by nano and fine-scale, soluble and insoluble metal oxide particles: NiO and TiO2.

The aim of the present study is to understand the association between metal ion release from nickel oxide (NiO) nanoparticles and induction of oxidative stress in the lung. NiO nanoparticles have cytotoxic activity through nickel ion release and subsequent oxidative stress. However, the interaction of oxidative stress and nickel ion release in vivo is still unclear. In the present study, we examined the effect of metal ion release on oxidative stress induced by NiO nanoparticles. Additionally, nano and fine TiO(2) particles as insoluble particles were also examined. Rat lung was exposed to NiO and TiO(2) nanoparticles by intratracheal instillation. The NiO nanoparticles released Ni(2+) in dispersion. Bronchoalveolar lavage fluid (BALF) was collected at 1, 24, 72 h and 1 week after instillation. The lactate dehydrogenase (LDH) and HO-1 levels were elevated at 24 and 72 h after instillation in the animals exposed to the NiO nanoparticles. On the other hand, total hydroxyoctadecadienoic acid (tHODE), which is an oxidative product of linoleic acid, as well as SP-D and α-tochopherol levels were increased at 72 h and 1 week after instillation. Fine NiO particles, and nano and fine TiO(2) particles did not show lung injury or oxidative stress from 1 h to 1 week after instillation. These results suggest that Ni(2+) release is involved in the induction of oxidative stress by NiO nanoparticles in the lung. Ni(2+) release from NiO nanoparticles is an important factor inoxidative stress-related toxicity, not only in vitro but also in vivo.

A history of vitamin E.

Vitamin E (α-tocopherol) was discovered nearly 100 years ago because it was required to prevent fetal resorption in pregnant, vitamin E-deficient rats fed lard-containing diets that were easily oxidizable. The human diet contains eight different vitamin E-related molecules synthesized by plants; despite the fact that all of these molecules are peroxyl radical scavengers, the human body prefers α-tocopherol. The biological activity of vitamin E is highly dependent upon regulatory mechanisms that serve to retain α-tocopherol and excrete the non-α-tocopherol forms. This preference is dependent upon the combination of the function of α-tocopherol transfer protein (α-TTP) to enrich the plasma with α-tocopherol and the metabolism of non-α-tocopherols. α-TTP is critical for human health because mutations in this protein lead to severe vitamin E deficiency characterized by neurologic abnormalities, especially ataxia and eventually death if vitamin E is not provided in large quantities to overcome the lack of α-TTP. α-Tocopherol serves as a peroxyl radical scavenger that protects polyunsaturated fatty acids in membranes and lipoproteins. Although specific pathways and specific molecular targets have been sought in a variety of studies, the most likely explanation as to why humans require vitamin E is that it is a fat-soluble antioxidant.

Antioxidant action of persulfides and polysulfides against free radical-mediated lipid peroxidation.

Hydrogen sulfide, hydropersulfides, and hydropolysulfides have been revealed to play important physiological roles such as cell signaling and protection against oxidative stress, but the underlying mechanisms and dynamics of action remain elusive. It is generally accepted that these species act by two-electron redox mechanisms, while the involvement of one-electron redox chemistry has received less attention. In this study, the radical-scavenging activity of hydrogen persulfide, hydrogen polysulfides (HSnH n = 2-4), and diallyl- or dialkyl-sulfides (RSnR, n = 1-4) was measured. Furthermore, their antioxidant effects against free radical-mediated human plasma lipid peroxidation were assessed by measuring lipid hydroperoxides. It was found that disodium disulfide, trisulfide, and tetrasulfide acted as potent peroxyl radical scavengers, the rate constant for scavenging peroxyl radical being 3.5 × 105, 4.0 × 105, and 6.0 × 105 M-1 s-1 in PBS pH 7.4 at 37 °C respectively and that they inhibited plasma lipid peroxidation efficiently, the efficacy is increased with the catenation number. Disodium tetrasulfide was 1.5 times as reactive as Trolox toward peroxyl radical and inhibited plasma lipid peroxidation more efficiently than ascorbate and Trolox. On the other hand, diallyl- and dialkyl-sulfides did not exert significant radical-scavenging activity, nor did they inhibit lipid peroxidation efficiently, except for diallyl tetrasulfide, which suppressed plasma lipid peroxidation, despite less significantly than disodium tetrasulfide. Collectively, this study shows that hydrogen persulfide and hydrogen polysulfides act as potent radical-scavenging antioxidants and that, in addition to two-electron redox mechanisms, one electron redox reaction may also play important role in the in vivo defense against deleterious oxidative stress.

Lipid oxidation that is, and is not, inhibited by vitamin E: Consideration about physiological functions of vitamin E.

Lipids are oxidized in vivo by multiple oxidizing species with different properties, some by regulated manner to produce physiological mediators, while others by random mechanisms to give detrimental products. Vitamin E plays an important role as a physiologically essential antioxidant to inhibit unregulated lipid peroxidation by scavenging lipid peroxyl radicals to break chain propagation independent of the type of free radicals which induce chain initiation. Kinetic data suggest that vitamin E does not act as an efficient scavenger of nitrogen dioxide radical, carbonate anion radical, and hypochlorite. The analysis of regio- and stereo-isomer distribution of the lipid oxidation products shows that, apart from lipid oxidation by CYP enzymes, the free radical-mediated lipid peroxidation is the major pathway of lipid oxidation taking place in humans. Compared with healthy subjects, the levels of racemic and trans,trans-hydro (pero)xyoctadecadienoates, specific biomarker of free radical lipid oxidation, are elevated in the plasma of patients including atherosclerosis and non-alcoholic fatty liver diseases. α-Tocopherol acts as a major antioxidant, while γ-tocopherol scavenges nitrogen dioxide radical, which induces lipid peroxidation, nitration of aromatic compounds and unsaturated fatty acids, and isomerization of cis-fatty acids to trans-fatty acids. It is essential to appreciate that the antioxidant effects of vitamin E depend on the nature of both oxidants and substrates being oxidized. Vitamin E, together with other antioxidants such as vitamin C, contributes to the inhibition of detrimental oxidation of biological molecules and thereby to the maintenance of human health and prevention of diseases.

Antioxidant action of vitamin E in vivo as assessed from its reaction products with multiple biological oxidants.

Vitamin E acts as essential antioxidant against detrimental oxidation of biological molecules induced by multiple reactive species. To gain more insight into the physiological role of vitamin E, the levels of its oxidation products in humans under normal and pathological conditions were compared. α-Tocopherol quinone (α-TQ) and 5-nitro-γ-tocopherol (5-NgT) were focused. α-TQ is produced by multiple oxidants including oxygen radicals, peroxynitrite, hypochlorite, singlet oxygen, and ozone, while 5-NgT is produced by nitrogen dioxide radical derived from peroxynitrite and the reaction of nitrite and hypochlorite. The reported concentrations of α-TQ and 5-NgT in healthy human plasma are highly variable ranging from 15 to 360 and 4 to 170 nM, respectively. In general, the molar ratio 5-NgT/γ-tocopherol was higher than the ratio α-TQ/α-tocopherol. Both absolute concentrations of α-TQ and 5-NgT and the molar ratios to the parent tocopherols were elevated significantly in the plasma of patients with various diseases compared with healthy subjects except neurological diseases. The molar ratios of the products to the respective parent compounds decreased in the order of 5-NgT/γ-tocopherol > α-TQ/α-tocopherol > hydroxyoctadecadienoate/linoleate > 3-nitrotyrosine/tyrosine > isoprostane/arachidonate. The molar ratios of nitrated products to the respective parent compounds in human plasma are approximately 10-2 for 5-NgT and 10-5 for 3-nitrotyrosine, nitro-oleic acid, and 8-nitroguaine. These data indicate that vitamin E acts as an important physiological antioxidant and that α-TQ and 5-NgT represent biomarker for oxidative stress and nitrative stress respectively.

Ex vivo oxidation in tissue and plasma assays of hydroxyoctadecadienoates: Z,E/E,E stereoisomer ratios.

The primary products from peroxidation of linoleate in biological tissues and fluids are the hydroperoxy octadecadienoates, and the products normally assayed, after reduction of the hydroperoxides, are the corresponding hydroxy octadecadienoates (HODEs). The HODEs are found in tissues and fluids as a mixture of Z,E and E,E stereoisomers. Two regioisomeric sets of Z,E and E,E stereoisomers are normally observed with substitution at the 9- and 13-positions of the 18-carbon chain. The Z,E/E,E product ratio has proved to be a useful means for assessing the reducing capacity of the medium undergoing peroxidation. The HODE Z,E/E,E product ratios previously reported for tissues such as liver and brain vary from 0.5 to 2.0, and plasma ratios are somewhat higher, between 2.0 and 3.0. The reported literature protocols for HODE assay in tissues involve homogenization, reduction with sodium borohydride in the presence of BHT, and ester hydrolysis with KOH to give the free HODEs. This is followed by either reverse-phase HPLC of the free acid HODEs or by conversion to TMS derivatives and GC-MS. When sodium borohydride is replaced in the protocol by triphenylphosphine, a gentler reducing agent, HODE Z,E/E,E product ratios are much higher, and lower total HODE levels of are found. It is proposed that inclusion of sodium borohydride in the isolation procedures leads to ex vivo reactions that are avoided if triphenylphosphine is used as the reducing agent. Modified protocols for HODE analyses (tissue and plasma methods #2) are described that should be used for assays of tissues and fluids.

Hydroxyoctadecadienoic acid as a potential biomarker for oxidative stress in patients with chronic hepatitis C.

BACKGROUND AND AIM: The possible involvement of oxidative stress in hepatitis C virus (HCV)-induced liver damage and hepatocarcinogenesis has been reported. We have recently developed a novel method to measure total hydroxyoctadecadienoic acid (tHODE) and have proposed its usefulness as a biomarker for lipid peroxidation. The present study was undertaken to evaluate oxidative stress in HCV-infected liver diseases by several potential oxidative stress markers including tHODE and further to validate the biomarkers for evaluating the efficacy of iron reduction therapy. METHODS: Total hydroxyoctadecadienoic acid, total 8-iso-prostagrandin F(2alpha) (t8-iso-PGF(2alpha)), selenoprotein P and other antioxidant compounds were measured in the plasma and erythrocytes obtained from 42 healthy controls and 78 HCV patients. Plasma levels of biomarkers and antioxidants were also assessed during the iron reduction therapy for 16 weeks in 12 HCV patients. RESULTS: The concentrations of tHODE in the plasma and erythrocytes and t8-iso-PGF(2alpha) in the plasma of chronic HCV-infected patients were significantly higher than those of healthy controls. Plasma levels of vitamin E and vitamin C of HCV-infected patients were lower than those of the controls. Furthermore, the plasma tHODE significantly correlated with serum aminotransferases and type IV collagen-7S domain in chronic HCV-infected patients. During the iron reduction therapy, the plasma levels of tHODE but not t8-iso-PGF(2alpha) decreased and inversely its stereo-isomer ratio (ZE/EE) increased in parallel with the decreases of serum alanine aminotransferase, ferritin and alpha-fetoprotein. CONCLUSION: The levels of tHODE in chronic HCV-infected patients can be a useful biomarker for the evaluation of oxidative stress in chronic hepatitis C.

Assessment of the antioxidant capacity of natural fruit extracts by inhibition of probe decay and plasma lipid peroxidation.

The oxidation of lipids, proteins, and DNA induced by reactive oxygen species has been implicated in the development of various diseases, and the role of antioxidants has received much attention. Free-radical scavenging antioxidants play an important role in the defense network in vivo, and assessment of the capacity of antioxidants has been the subject of extensive studies and controversy, but there is no universal method by which antioxidant capacity can be measured accurately. In the present study, the assessment of the antioxidant capacity of natural fruit extracts was examined for radical scavenging and inhibition of lipid peroxidation. It was found that the capacity of fruit extracts for scavenging of both hydrophilic and lipophilic free radicals and for antioxidation can be assessed from the effect on the probe decay and the inhibition of plasma lipid peroxidation respectively. The importance of these two factors in the assessment of antioxidant capacity is discussed.