Increased Resistance to Oxidation of Betalain-enriched Human Low Density Lipoproteins

Betalains are natural pigments recently considered as compounds with potential antioxidative properties. In this work, ex vivo plasma spiking of pure either betanin or indicaxanthin, followed by isolation of low density lipoprotein (LDL), and measurement of its resistance to copper-induced oxidation, has been used to research if these betalains can bind to LDL and prevent oxidation of LDL lipids. When pooled human plasma from 10 healthy volunteers was incubated in the presence of 25–100?μM either betanin or indicaxanthin, incorporation of both compounds in LDL was observed, with a maximum binding of 0.52±0.08, and 0.51±0.06?nmoles of indicaxanthin and betanin, respectively, per mg LDL protein. Indicaxanthin-enriched and betanin-enriched LDL were more resistant than homologous native LDL to copper-induced oxidation, as assessed by the elongation of the induction period. The incorporated indicaxanthin, however, appeared twice as effective as betanin in increasing the length of the lag phase, while both compounds did not affect the propagation rate. Both betalains were consumed during the inhibition period of lipid oxidation, and delayed consumption of LDL-beta carotene. Indicaxanthin, but not betanin, prevented vitamin E consumption at the beginning of LDL oxidation, and prolonged the time of its utilization. The resistance of LDL to oxidation when vitamin E and indicaxanthin acted separately in a sequence, was lower than that measured when they were allowed to act in combination, indicating some synergistic interaction between the two molecules. No prooxidant effect over a large concentration range of either betanin or indicaxanthin was observed, when either betalain was added to the LDL system undergoing a copper-induced oxidation.

These results show than indicaxanthin and betanin may bind to LDL, and are highly effective in preventing copper-induced lipid oxidation. Interaction with vitamin E appears to add a remarkable potential to indicaxanthin in the protection of LDL. Although molecular mechanisms remain uncompletely understood, various aspects of the action of betanin and indicaxanthin in preventing LDL lipid oxidation are discussed.     

Cytoprotective effects of the antioxidant phytochemical indicaxanthin in beta-thalassemia red blood cells

Antioxidant phytochemicals are investigated as novel treatments for supportive therapy in β-thalassemia. The dietary indicaxanthin was assessed for its protective effects on human β-thalassemic RBCs submitted in vitro to oxidative haemolysis by cumene hydroperoxide. Indicaxanthin at 1.0-10 μM enhanced the resistance to haemolysis dose-dependently. In addition, it prevented lipid and haemoglobin (Hb) oxidation, and retarded vitamin E and GSH depletion. After ex vivo spiking of blood from thalassemia patients with indicaxanthin, the phytochemical was recovered in the soluble cell compartment of the RBCs. A spectrophotometric study showed that indicaxanthin can reduce perferryl-Hb generated in solution from met-Hb and hydrogen peroxide (H₂O₂), more effectively than either Trolox or vitamin C.

Collectively our results demonstrate that indicaxanthin can be incorporated into the redox machinery of β-thalassemic RBC and defend the cell from oxidation, possibly interfering with perferryl-Hb, a reactive intermediate in the hydroperoxide-dependent Hb degradation. Opportunities of therapeutic interest for β-thalassemia may be considered.     

Impaired resistance to oxidation of low density lipoprotein in cystic fibrosis: Improvement during vitamin E supplementation

Antioxidants such as vitamin E protect unsaturated fatty acids of LDL against oxidation. In the ex vivo model used, LDL was exposed to Cu²⁺ ions, a potent prooxidant capable of initiating the oxidation of LDL. The lag time, indicating the delay of conjugated diene formation in LDL due to antioxidant protection, was measured in 54 cystic fibrosis (CF) patients with plasma -tocopherol levels below (Group A, n = 30) or above (Group B, n = 24) 15.9 μmol/L (mean - 2 SD of Swiss population). Patients were reevaluated after 2 months on 400 IU/d of oral RRR--tocopherol. In group A, -tocopherol concentrations in LDL increased significantly from 3.2 ± 1.6 mol/mol LDL to 8.2 ± 2.8 mol/mol (P < 0.001) and lag times increased from 79 ± 33, min to 126 ± 48 min (P < 0.001), whereas in the vitamin E sufficient group B no further increase neither in LDL -tocopherol concentrations or in lag times was observed. LDL oleic acid concentrations were higher, and linoleic acid concentrations were lower in patients than in controls. After efficient vitamin E supplementation, lag times were positively related to LDL -tocopherol (P < 0.01) and negatively to LDL linoleic and arachidonic acid content (P < 0.001). The maximum rate of oxidation correlated positively with linoleic and arachidonic acid concentrations, as did the maximum conjugated diene absorbance. These results indicate that LDL resistance to oxidation is impaired in vitamin E deficient CF patients but can be normalized within 2 months when -tocopherol is given in sufficient amounts. Linoleic and arachidonic acid content exhibit a major influence on the LDL resistance to oxidation.     

Cytoprotective effects of the antioxidant phytochemical indicaxanthin in β-thalassemia red blood cells

Antioxidant phytochemicals are investigated as novel treatments for supportive therapy in β-thalassemia. The dietary indicaxanthin was assessed for its protective effects on human β-thalassemic RBCs submitted in vitro to oxidative haemolysis by cumene hydroperoxide. Indicaxanthin at 1.0–10 μM enhanced the resistance to haemolysis dose-dependently. In addition, it prevented lipid and haemoglobin (Hb) oxidation, and retarded vitamin E and GSH depletion. After ex vivo spiking of blood from thalassemia patients with indicaxanthin, the phytochemical was recovered in the soluble cell compartment of the RBCs. A spectrophotometric study showed that indicaxanthin can reduce perferryl-Hb generated in solution from met-Hb and hydrogen peroxide (H₂O₂), more effectively than either Trolox or vitamin C.

Collectively our results demonstrate that indicaxanthin can be incorporated into the redox machinery of β-thalassemic RBC and defend the cell from oxidation, possibly interfering with perferryl-Hb, a reactive intermediate in the hydroperoxide-dependent Hb degradation. Opportunities of therapeutic interest for β-thalassemia may be considered.     

Betanin inhibits the myeloperoxidase/nitrite-induced oxidation of human low-density lipoproteins

Production of nitrogen dioxide by the activity of myeloperoxidase (MPO) in the presence of nitrite is now considered a key step in the pathophysiology of low-density lipoprotein (LDL) oxidation. This study shows that betanin, a phytochemical of the betalain class, inhibits the production of lipid hydroperoxides in human LDL submitted to a MPO/nitrite-induced oxidation. Kinetic measurements including time-course of particle oxidation and betanin consumption, either in the presence or in the absence of nitrite, suggest that the antioxidant effect is possibly the result of various actions. Betanin scavenges the initiator radical nitrogen dioxide and can also act as a lipoperoxyl radical-scavenger. In addition, unidentified oxidation product(s) of betanin by MPO/nitrite inhibit(s) the MPO/nitrite-induced LDL oxidation as effectively as the parent compound. In the light of betanin bioavailability and post-absorbtion distribution in humans, present findings may suggest favourable in vivo activity of this phytochemical.     

Betanin inhibits the myeloperoxidase/nitrite-induced oxidation of human low-density lipoproteins

Production of nitrogen dioxide by the activity of myeloperoxidase (MPO) in the presence of nitrite is now considered a key step in the pathophysiology of low-density lipoprotein (LDL) oxidation. This study shows that betanin, a phytochemical of the betalain class, inhibits the production of lipid hydroperoxides in human LDL submitted to a MPO/nitrite-induced oxidation. Kinetic measurements including time-course of particle oxidation and betanin consumption, either in the presence or in the absence of nitrite, suggest that the antioxidant effect is possibly the result of various actions. Betanin scavenges the initiator radical nitrogen dioxide and can also act as a lipoperoxyl radical-scavenger. In addition, unidentified oxidation product(s) of betanin by MPO/nitrite inhibit(s) the MPO/nitrite-induced LDL oxidation as effectively as the parent compound. In the light of betanin bioavailability and post-absorbtion distribution in humans, present findings may suggest favourable in vivo activity of this phytochemical.     

The main components of St John's Wort inhibit low-density lipoprotein atherogenic modification: a beneficial "side effect" of an OTC antidepressant drug?

Hypericin and pseudohypericin are polycyclic-phenolic structurally related compounds found in Hypericum perforatum L. (St John's wort). As hypericin has been found to bind to LDL one may assume that it can act as antioxidant of LDL lipid oxidation, a property which is of prophylactic/therapeutic interest regarding atherogenesis as LDL oxidation may play a pivotal role in the onset of atherosclerosis. Therefore, in the present paper hypericin, pseudohypericin and hyperforin, an other structurally unrelated constituent in St John's wort were tested in their ability to inhibit LDL oxidation. LDL was isolated by ultracentrifugation and oxidation was initiated either by transition metal ions (copper), tyrosyl radical (myeloperoxidase/hydrogen peroxide/tyrosine) or by endothelial cells (HUVEC). LDL modification was monitored by conjugated diene and malondialdehyde formation. The data show that all compounds (hypericin, pseudohypericin and hyperforin) at doses as low as 2.5 μmol/l are potent antioxidants in the LDL oxidation systems used. The results indicate that the derivatives found in Hypericum perforatum have possible antiatherogenic potential.     

The kinetics of copper-induced LDL oxidation depend upon its lipid composition and antioxidant content

Copper promotes oxidation of human low-density lipoprotein (LDL) through molecular mechanisms that are still under investigation. We employed native human LDL, phospholipid-containing delipidated LDL ghosts, or trilinolein-reconstituted, phospholipid-containing LDL to investigate both LDL oxidation and the associated process of copper reduction. Both LDL ghosts and trilinolein-reconstituted LDL were devoid of antioxidants and were extremely susceptible to AAPH-induced oxidation but, paradoxically, were rather resistant to copper-mediated oxidation. The dynamic reduction of Cu(II) to Cu(I) was quantitatively decreased in LDL ghosts and in trilinolein-reconstituted LDL, also lacking the initial rapid reduction and the subsequent inhibition phases, due to the absence of endogenous antioxidants. Conversely, the rate of copper reduction was linear and likely due to lipid peroxides, either already present or formed during copper-induced oxidation. We suggest that copper undergoes redox transitions in LDL by utilizing reducing equivalents originating from endogenous antioxidants and/or from lipid peroxides in the LDL lipid core.     

Diphenylhexatriene as a Fluorescent Probe for Monitoring Low Density Lipoprotein Peroxidation

The use of the fluorescent probe diphenylhexatriene (DPH) for monitoring low density lipoprotein (LDL) peroxidation has been investigated. The DPH incorporation into LDL results in a high fluorescence signal which decreases with time after addition of cupric ions. A strong correlation was found between the decay of the DPH fluorescence signal and the appearance of the thiobarbituric reactive substances (TBARS). HPLC and spectrofluorometric analyses demonstrated that DPH is destroyed during the time course of the copper-induced LDL peroxidation. The decrease in DPH fluorescent signal is prevented by addition of EDTA, vitamin E and drugs which protect LDL against peroxidation such as probucol or calcium antagonists. The high fluorescence of DPH allows the use of very small quantities of LDL (less than 5 μg/ml LDL protein). We thus suggest that DPH could be of use for continuous monitoring of LDL autooxidation, especially for the in vitro testing of the protective effect of antioxidant compounds.     

The monoterpene terpinolene from the oil of Pinus mugo L. in concert with alpha-tocopherol and beta-carotene effectively prevents oxidation of LDL.

Antioxidants from several nutrients, e.g. vitamin E, beta-carotene, or flavonoids, inhibit the oxidative modification of low-density lipoproteins. This protective effect could possibly retard atherogenesis and in consequence avoid coronary heart diseases. Some studies have shown a positive effect of those antioxidants on cardiovascular disease. Another class of naturally occurring antioxidants are terpenoids, which are found in essential oils. The essential oil of Pinus mugo and the contained monoterpene terpinolene effectively prevent low-density lipoprotein (LDL)-oxidation. In order to test the mechanism by which terpinolene protects LDL from oxidation, LDL from human blood plasma enriched in terpinolene was isolated. In this preparation not only the lipid part of LDL is protected against copper-induced oxidation--as proven by following the formation of conjugated dienes, but also the oxidation of the protein part is inhibited, since loss of tryptophan fluorescence is strongly delayed. This inhibition is due to a retarded oxidation of intrinsic carotenoids of LDL, and not, as in the case of some flavonoids, attributable to a protection of intrinsic alpha-tocopherol. These results are in agreement with our previous results, which showed the same effects for a monoterpene from lemon oil, i.e. gamma-terpinene.     

The Chemiluminescence Assay of Lipid Peroxidation Products in Human Blood Plasma Lipoproteins

A chemiluminescence (CL) flash kinetics on the addition of Fe²⁺ ions into oxidized low density lipoprotein (LDL) suspension has been studied. LDL oxidation was carried out at 37°C without and in the presence of 5 or 50 μM of Cu.²⁺ It has been found that under certain experimental conditions (the addition of excess iron ions, more than 1 mM) the amplitude of CL flash depended almost linearly (1) on the concentration of oxidized LDL and (2) on the extent of LDL oxidation measured as diene conjugates (DC) and 2-thiobarbituric acid-reactive substance (TBARS) accumulation. The corresponding correlation coefficients were: for TBARS - 0.94 and for DC - 0.97, in the case of LDL autooxidation; 0.72 and 0.98, in the case of copper-induced LDL oxidation. A sensitivity of the CL method was shown to be significantly enhanced (by more than two orders) in the presence of CL sensitizer - 2, 3,5, 6-lH,4H-tetrahydro-9-(2' -benzoimidazolyl)-quinolizin-(9, 9a, 1 -gh)coumarin.     

Vitamin C protects low-density lipoprotein from homocysteine-mediated oxidation

Homocysteine, an atherogenic amino acid, promotes iron-dependent oxidation of low-density lipoprotein (LDL). We investigated whether vitamin C, a physiological antioxidant, could protect LDL from homocysteine-mediated oxidation. LDL (0.2 mg of protein/ml) was incubated at 37 degrees C with homocysteine (1000 microM) and ferric iron (10-100 microM) in either the absence (control) or presence of vitamin C (5-250 microM). Under these conditions, vitamin C protected LDL from oxidation as evidenced by an increased lag time preceding lipid diene formation (> or = 5 vs. 2.5 h for control), decreased thiobarbituric acid-reactive substances accumulation (< or = 19 +/- 1 nmol/mg when vitamin C > or = 10 microM vs. 32 +/- 3 nmol/mg for control, p <.01), and decreased lipoprotein anodic electrophoretic mobility. Near-maximal protection was observed at vitamin C concentrations similar to those in human blood (50-100 microM); also, some protection was observed even at low concentrations (5-10 microM). This effect resulted neither from altered iron redox chemistry nor enhanced recycling of vitamin E in LDL. Instead, similar to previous reports for copper-dependent LDL oxidation, we found that vitamin C protected LDL from homocysteine-mediated oxidation through covalent lipoprotein modification involving dehydroascorbic acid. Protection of LDL from homocysteine-mediated oxidation by vitamin C may have implications for the prevention of cardiovascular disease.     

Radical Exchange Reactions between Vitamin E, Vitamin C and Phospholipids in Autoxidizing Polyunsaturated Lipids

Antioxidant reactions of mixtures of vitamin E, vitamin C and phospholipids in autoxidizing lipids at 90°C have been studied by ESR spectroscopy. When the phospholipid contained a tertiary amine (e.g. phosphatidylcholine), the vitamin C and the vitamin E radicals were successively observed as these two vitamins were sequentially oxidised during lipid oxidation. In the presence of the primary amine contained in phosphatidylserine, the vitamin E oxidation was delayed for a few hours. In this case neither the vitamin C, nor the vitamin E radicals but a nitroxide radical derived from the phospholipid was observed. Similar results to those obtained with PS were obtained in the presence of either phospha-tidylethanolamine or soybean lecithin. The participation in the radical reactions of phospholipids possessing a primary amine can therefore explain the synergistic effect of these phospholipids in a mixture of vitamins E and C.     

Oxidation of low-density lipoproteins: effect of antioxidant content,fatty acid composition and intrinsic phospholipase activity on susceptibility to metal ion-induced oxidation

The oxidative modification of low-density lipoprotein (LDL) may play an important role in atherogenesis. Our understanding of the mechanism of LDL oxidation and the factors that determine its susceptibility to oxidation is still incomplete. We have isolated LDL from 45 healthy individuals and studied the relationship between LDL fatty acid, vitamin E and β-carotene composition, intrinsic phospholipase A₂-like activity and parameters of LDL oxidation. LDL was exposed to a copper ion-dependent oxidising system and the kinetics of oxidation studied by monitoring formation of fatty acid conjugated dienes. The length of the lag phase of inhibited lipid peroxidation was measured as well as the rate of lipid peroxidation during the propagation phase. There was no significant correlation between LDL antioxidant vitamin or fatty acid composition and lag time to LDL oxidation. Oleic acid was negatively correlated with the rate of LDL oxidation (r = −0.41, P < 0.01) whilst linoleic acid was significantly correlated with the extent of LDL oxidation measured by the production of total dienes (r = 0.34, P < 0.05). Interestingly, LDL vitamin E content was positively correlated with both the rate (r = 0.28, P < 0.05) and extent of LDL oxidation (r = 0.43, P < 0.01). LDL isolated from this group of subjects showed significant intrinsic phospholipase-like activity. The phospholipase activity, whilst not correlated with lag time, was significantly correlated with both rate (r = 0.43, P < 0.01) and total diene production (r = 0.44, P < 0.01) of LDL oxidation. We conclude that antioxidant content, fatty acid composition and intrinsic phospholipase activity have little influence on the lag time of Cu-induced LDL oxidation. These components do however, significantly influence both the rate and extent of LDL oxidation, with increased vitamin E, linoleic acid content and phospholipase activity associated with faster and more extensive oxidation. The possible pro-oxidant effect of vitamin E has interesting implications for the postulated ‘protective’ effects of vitamin E on atherogenesis.     

Effect of oral supplementation with D-alpha-tocopherol on the vitamin E content of human low density lipoproteins and resistance to oxidation.

Twelve clinically healthy subjects participated in a vitamin E supplementation study. Eight were given daily dosages of 150, 225, 800, or 1200 IU RRR-alpha-tocopherol for 21 days (two persons per dose) and four received placebo. Prior, during, and after the supplementation period, alpha-tocopherol, gamma-tocopherol, and carotenoids were determined in plasma and low density lipoprotein (LDL). The maximum levels of alpha-tocopherol were 1.7- to 2.5-times the baseline values in plasma and 1.7- to 3.1-times in LDL. A high correlation existed between alpha-tocopherol in plasma and LDL. gamma-Tocopherol significantly decreased in plasma and LDL during vitamin E supplementation. No significant influence on the lipoprotein and lipid status and carotenoid levels of the participants occurred throughout the supplementation. The resistance of LDL against copper-mediated oxidation was also measured. The oxidation resistance of LDL was significantly higher during vitamin E supplementation. However, the efficacy of vitamin E in protecting LDL varied from person to person. The statistical evaluation of all data gave a correlation of r2 = 0.51 between alpha-tocopherol in LDL and the oxidation resistance as measured by the length of the lag-phase preceding the oxidation of LDL. No association was seen between levels of carotenoids and vitamin E in plasma and LDL. The present study clearly shows that in humans the oxidation resistance of LDL can be increased by vitamin E supplementation.     

Molecular action of vitamin E in lipoprotein oxidation: implications for atherosclerosis

The oxidation theory of atherosclerosis proposes that the oxidative modification of low-density lipoproteins (LDL) plays a central role in the disease. Although a direct causative role of LDL oxidation for atherogenesis has not been established, oxidized lipoproteins are detected in atherosclerotic lesions, and in vitro oxidized LDL exhibits putative pro-atherogenic activities. alpha-Tocopherol (alpha-TOH; vitamin E), the major lipid-soluble antioxidant present in lipoproteins, is thought to be antiatherogenic. However, results of vitamin E interventions on atherosclerosis in experimental animals and cardiovascular disease in humans have been inconclusive. Also, recent mechanistic studies demonstrate that the role of alpha-TOH during the early stages of lipoprotein lipid peroxidation is complex and that the vitamin does not act as a chain-breaking antioxidant. In the absence of co-antioxidants, compounds capable of reducing the alpha-TOH radical and exporting the radical from the lipoprotein particle, alpha-TOH exhibits anti- or pro-oxidant activity for lipoprotein lipids depending on the degree of radical flux and reactivity of the oxidant. The model of tocopherol-mediated peroxidation (TMP) explains the complex molecular action of alpha-TOH during lipoprotein lipid peroxidation and antioxidation. This article outlines the salient features of TMP, comments on whether TMP is relevant for in vivo lipoprotein lipid oxidation, and discusses how co-antioxidants may be required to attenuate lipoprotein lipid oxidation in vivo and perhaps atherosclerosis.     

The main components of St John's Wort inhibit low-density lipoprotein atherogenic modification: A beneficial “side effect” of an OTC antidepressant drug?

Hypericin and pseudohypericin are polycyclic–phenolic structurally related compounds found in Hypericum perforatum L. (St John's wort). As hypericin has been found to bind to LDL one may assume that it can act as antioxidant of LDL lipid oxidation, a property which is of prophylactic/therapeutic interest regarding atherogenesis as LDL oxidation may play a pivotal role in the onset of atherosclerosis. Therefore, in the present paper hypericin, pseudohypericin and hyperforin, an other structurally unrelated constituent in St John's wort were tested in their ability to inhibit LDL oxidation. LDL was isolated by ultracentrifugation and oxidation was initiated either by transition metal ions (copper), tyrosyl radical (myeloperoxidase/hydrogen peroxide/tyrosine) or by endothelial cells (HUVEC). LDL modification was monitored by conjugated diene and malondialdehyde formation. The data show that all compounds (hypericin, pseudohypericin and hyperforin) at doses as low as 2.5 μmol/l are potent antioxidants in the LDL oxidation systems used. The results indicate that the derivatives found in Hypericum perforatum have possible antiatherogenic potential.     

Serum ex vivo lipoprotein oxidizability in patients with ischemic heart disease supplemented with vitamin E

The decreased oxidizability of plasma lipoproteins is related to the increased vitamin E intake and its association with a relatively lower incidence of coronary heart disease has been proposed. We investigated the effect of the in vivo vitamin E supplementation on the oxidizability of serum lipids in patients with ischemic heart disease and a moderate hypercholesterolemia. Thirty-two patients (16 males and 16 postmenopausal women) participated in this placebo-controlled, randomized trial. They were treated with 400 mg vitamin E/day for 6 weeks. The copper-induced serum lipid oxidizability ex vivo was assessed by measuring conjugated diene formation at 245 nm. We also measured vitamin E, malondialdehyde (MDA) and uric acid concentrations in the plasma. Because of observed significant differences in parameters of serum lipid oxidizability (lag time and maximal rate of oxidation), plasma alpha-tocopherol and MDA levels between male patients and postmenopausal women supplemented with vitamin E, the results were compared between both genders. Six weeks of vitamin E supplementation significantly increased plasma vitamin E levels (by 87 %) in male patients but in postmenopausal women only by 34 %. Concomitantly with increased plasma levels of vitamin E the decrease in plasma MDA levels was observed in male patients (decrease by 20 %; p=0.008), but in postmenopausal women the decrease did not attain statistical significance. Plasma uric acid levels were not apparently changed in placebo or vitamin E supplemented groups of patients. The changes in ex vivo serum lipid oxidizability after vitamin E, supplementation have shown a significantly prolonged lag time (by 11 %; p=0.048) and lowered rate of lipid oxidation (by 21 %; p=0.004) in male patients in comparison with postmenopausal women. Linear regression analysis revealed a significant correlation between plasma vitamin E levels and the lag time (r=0.77; p=0.03) and the maximal rate of serum lipid oxidation (r=-0.70; p=0.05) in male patients. However, in postmenopausal women the correlations were not significant. We conclude that 400 mg vitamin E/day supplementation in patients with ischemic heart disease and a moderate hypercholesterolemia influenced favorably ex vivo serum lipid oxidation of male patients when compared with postmenopausal women. The observed differences between both genders could be useful in the selection of the effective vitamin E doses in the prevention of coronary heart disease.     

Betacyanins as phenol antioxidants. Chemistry and mechanistic aspects of the lipoperoxyl radical-scavenging activity in solution and liposomes

Reaction kinetics of betanin and its aglycone betanidin towards peroxyl radicals generated from the azo-initiated oxidation of methyl linoleate in methanol and of a heterogeneous aqueous/soybean phosphatidylcholine liposomal system were studied by monitoring formation of linoleic acid hydroperoxides and consumption of the pigments. Betanin was a weak retarder in methanol and an effective chain breaking antioxidant in the liposomal model, indicating that kinetic solvent effects and partition in lipid bilayers may affect its activity. Betanidin behaved as a chain terminating antioxidant in both models. Kinetic parameters characterizing peroxyl radical-scavenging activity showed that betanidin was more effective than betanin, in terms of both radical-scavenging rate constant and stoichiometric factor, with effectiveness of the same order as vitamin E under comparable conditions. Products identified by spectrophotometric and HPLC techniques indicated reaction of the glucose-substituted monophenol and ortho-diphenol moieties of betanin and betanidin, respectively, and suggested mechanisms of the antioxidant activity. Either betanin or betanidin incorporated in liposomes with α-tocopherol had additive effects, supporting partition of the pigments in the bilayer and lipoperoxyl radical reduction.     

Effect of dietary oils on lipid peroxidation and on antioxidant parameters of rat plasma and lipoprotein fractions.

In order to investigate the influence of fatty acid pattern and antioxidants other than vitamin E on lipid peroxidation and antioxidant levels of plasma very low density and low density lipoproteins (VLDL + LDL), the effects of three diets (equalized for vitamin E) containing soybean oil, olive oil, or an oleate-rich mixture of triglycerides (triolein) were studied in rats. A significantly lower concentration of thiobarbituric acid-reactive substances (TBA-RS) in plasma and lipoproteins was found after the olive oil diet (soybean oil, 3.7 +/- 0.4 nmol/ml; triolein, 2.1 +/- 0.5 nmol/ml; olive oil, 1.5 +/- 0.3 nmol/ml, in plasma) (soybean oil, 0.99 +/- 0.16 nmol/ml; triolein, 0.96 +/- 0.13 nmol/ml; olive oil, 0.38 +/- 0.12 nmol/ml, in the VLDL + LDL fraction). Furthermore, the results from in vitro copper-induced lipid peroxidation, expressed in terms of conjugated dienes, lipid hydroperoxides, and TBA-RS content, showed that VLDL + LDL particles from olive olive oil-fed rats were remarkably resistant to oxidative modification. The results suggest that the fatty acid unsaturation of dietary oils is not the only determining factor of the antioxidant capacity of lipoproteins in this animal model. The maximal protection observed after the olive oil diet may be explained by the presence of other unidentified antioxidants in addition to vitamin E, derived from oil intake. Therefore, the optimal balance between the content of unsaturated fatty acids and natural antioxidants in dietary oils appears to be of major importance.