Effects of flavonoids on the susceptibility of low-density lipoprotein to oxidative modification

Dietary flavonoid intake has been reported to be inversely associated with the incidence of coronary artery disease. To clarify the possible role of flavonoids in the prevention of atherosclerosis, we investigated the effects of some of these compounds on the susceptibility of low-density lipoprotein (LDL) to oxidative modification. In this study, six flavonoids, "apigenin, genistein, morin, naringin, pelargonidin and quercetin", were added to plasma and incubated for 3h at 37 degrees C. Then, the LDL fraction was separated by ultracentrifugation. The oxidizability of LDL was estimated by measuring conjugated diene (CD), lipid peroxides and thiobarbituric acid-reactive substances (TBARS) after cupric sulfate solution was added. We showed that among flavonoids used, quercetin and morin significantly (P<0.01 by ANOVA) and dose-dependently prolonged the lag time before initiation of oxidation reaction. Also, these two flavonoids suppressed the formation of lipid peroxides and TBARS more markedly than others. Their ability to prolong lag time and suppression of lipid peroxides and TBARS formation resulted to be in the following order: quercetin>morin>pelargonidin>genistein>naringin>apigenin. LDL exposed to flavonoids in vitro reduced oxidizability. These findings show that flavonoids may have a role in ameliorating atherosclerosis.     

Morphology and structure of lipoproteins revealed by an optimized negative-staining protocol of electron microscopy

Plasma lipoprotein levels are predictors of risk for coronary artery disease. Lipoprotein structure-function relationships provide important clues that help identify the role of lipoproteins in cardiovascular disease. The compositional and conformational heterogeneity of lipoproteins are major barriers to the identification of their structures, as discovered using traditional approaches. Although electron microscopy (EM) is an alternative approach, conventional negative staining (NS) produces rouleau artifacts. In a previous study of apolipoprotein (apo)E4-containing reconstituted HDL (rHDL) particles, we optimized the NS method in a way that eliminated rouleaux. Here we report that phosphotungstic acid at high buffer salt concentrations plays a key role in rouleau formation. We also validate our protocol for analyzing the major plasma lipoprotein classes HDL, LDL, IDL, and VLDL, as well as homogeneously prepared apoA-I-containing rHDL. High-contrast EM images revealed morphology and detailed structures of lipoproteins, especially apoA-I-containing rHDL, that are amenable to three-dimensional reconstruction by single-particle analysis and electron tomography.     

Absorption and lipoprotein transport of sphingomyelin

Dietary sphingomyelin (SM) is hydrolyzed by intestinal alkaline sphingomyelinase and neutral ceramidase to sphingosine, which is absorbed and converted to palmitic acid and acylated into chylomicron triglycerides (TGs). SM digestion is slow and is affected by luminal factors such as bile salt, cholesterol, and other lipids. In the gut, SM and its metabolites may influence TG hydrolysis, cholesterol absorption, lipoprotein formation, and mucosal growth. SM accounts for approximately 20% of the phospholipids in human plasma lipoproteins, of which two-thirds are in LDL and VLDL. It is secreted in chylomicrons and VLDL and transferred into HDL via the ABCA1 transporter. Plasma SM increases after periods of large lipid loads, during suckling, and in type II hypercholesterolemia, cholesterol-fed animals, and apolipoprotein E-deficient mice. SM is thus an important amphiphilic component when plasma lipoprotein pools expand in response to large lipid loads or metabolic abnormalities. It inhibits lipoprotein lipase and LCAT as well as the interaction of lipoproteins with receptors and counteracts LDL oxidation. The turnover of plasma SM is greater than can be accounted for by the turnover of LDL and HDL particles. Some SM must be degraded via receptor-mediated catabolism of chylomicron and VLDL remnants and by scavenger receptor class B type I receptor-mediated transfer into cells.     

Analysis of Lipoprotein Diene Formation in Human Serum Exposed to Copper

The susceptibility of low density lipoprotein (LDL) to oxidative modification can be determined by analyzing the lag phase for initiation of diene formation in isolated LDL exposed to Cu^2+. However, the applicability of this assay for clinical studies is limited by the requirement of a preparative ultracentrifugation of LDL and that the influence of water soluble antioxidants and other lipoproteins is not accounted for. The present paper describes a modification of this assay allowing determination of lag phase for lipoprotein diene formation in serum. The formation of dienes in serum exposed to Cu²⁺ begins following the consumption of serum α-tocopherol, correlates to the formation of thiobarbituric acid reactive substances (r = 0.987, n = 8), is inhibited by the addition of ascorbic acid and is absent in lipoprotein-deficient serum. It is also accompanied by an increased mobility of serum lipoproteins on agarose gel electrophoresis and with an ability of serum to displace isolated copper-oxidized LDL from binding sites mediating degradation in mouse peritoneal macrophages. The coefficient of variance of the analysis is below 3%. It is concluded that this technique allows analysis of lipoprotein oxidation susceptibility in serum samples and may prove to be useful in clinical analysis of the lipoprotein oxidation susceptibility.     

Cholesterol esterification and atherogenic index of plasma correlate with lipoprotein size and findings on coronary angiography

We examined the association between rate of cholesterol esterification in plasma depleted of apolipoprotein B-containing lipoproteins (FER(HDL)), atherogenic index of plasma (AIP) [(log (TG/HDL-C)], concentrations, and size of lipoproteins and changes in coronary artery stenosis in participants in the HDL-Atherosclerosis Treatment Study. A total of 160 patients was treated with simvastatin (S), niacin (N), antioxidants (A) and placebo (P) in four regimens. FER(HDL) was measured using a radioassay; the size and concentration of lipoprotein subclasses were determined by nuclear magnetic resonance spectroscopy. The S+N and S+N+A therapy decreased AIP and FER(HDL), reduced total VLDL (mostly the large and medium size particles), decreased total LDL particles (mostly the small size), and increased total HDL particles (mostly the large size). FER(HDL) and AIP correlated negatively with particle sizes of HDL and LDL, positively with VLDL particle size, and closely with each other (r = 0.729). Changes in the proportions of small and large lipoprotein particles, which were reflected by FER(HDL) and AIP, corresponded with findings on coronary angiography. Logistic regression analysis of the changes in the coronary stenosis showed that probability of progression was best explained by FER(HDL) (P = 0.005). FER(HDL) and AIP reflect the actual composition of the lipoprotein spectrum and thus predict both the cardiovascular risk and effectiveness of therapy. AIP is already available for use in clinical practice as it can be readily calculated from the routine lipid profile.     

A comparison of anion-exchange and steric-exclusion HPLC assays of mouse plasma lipoproteins

We compared two HPLC methods (anion exchange [AE] and steric exclusion [SE]) for analysis of mouse lipoprotein profiles by determining coefficients of variability (CVs) under varying conditions. CVs for AE and SE were comparable on fresh samples. There was an inverse relationship between subfraction curve area and CV [r = -0.65 (AE) and -0.50 (SE)], consistent with the interpretation that as curve area decreased, error variance increased and signal-to-noise ratio decreased. Sample storage did not affect SE. In contrast, with AE, alterations in measured lipoproteins were apparent after storage, including a decrease in the HDL subfraction [66.8% (baseline) vs. 15.9% (1 week); P < 0.01] and an increase in areas under LDL and VLDL peaks. Concomitant with decreasing HDL area, reproducibility deteriorated with the duration of storage. Analysis of the effects of decreasing sample injectate volume to <25 microl on SE lipoprotein subfractions revealed that areas under LDL and VLDL peaks decreased and persisted as volume was decreased further. Areas under all lipoprotein subfractions measured with either AE or SE were linearly correlated with the amount of cholesterol [r = 0.69 (AE) and 0.87 (SE)]. Both AE and SE yield reproducible, accurate, and rapid measurements of lipoproteins from small amounts of serum. AE yields more sensitive, high-amplitude, well-defined peaks that can be easily distinguished and necessitates the use of smaller sample volumes compared with SE, but sample storage causes alterations in the chromatogram. SE appears better suited to serial analyses involving stored samples.     

Levels of atherogenic lipoproteins are unexpectedly reduced in interstitial fluid from type 2 diabetes patients

At a given level of serum cholesterol, patients with T2D have an increased risk of developing atherosclerosis compared with nondiabetic subjects. We hypothesized that T2D patients have an increased interstitial fluid (IF)-to-serum gradient ratio for LDL, due to leakage over the vascular wall. Therefore, lipoprotein profiles in serum and IF from 35 T2D patients and 35 healthy controls were assayed using fast performance liquid chromatography. The IF-to-serum gradients for VLDL and LDL cholesterol, as well as for apoB, were clearly reduced in T2D patients compared with healthy controls. No such differences were observed for HDL cholesterol. Contrary to our hypothesis, the atherogenic VLDL and LDL particles were not increased in IF from diabetic patients. Instead, they were relatively sparser than in healthy controls. The most probable explanation to our unexpected finding is that these lipoproteins are more susceptible to retainment in the extravascular space of these patients, reflecting a more active uptake by, or adhesion to, tissue cells, including macrophages in the vascular wall. Further studies are warranted to further characterize the mechanisms underlying these observations, which may be highly relevant for the understanding of why the propensity to develop atherosclerosis is increased in T2D.     

Antioxidative activity on human low density lipoprotein (LDL) oxidation by pentagalloic acid

The aim of this study was to investigate the efficiency of the pentagalloic acid compound in inhibiting the metal ions and cell lines that mediate in low density lipoprotein (LDL) oxidation. Pentagalloic acid prolonged the lag time preceeding the onset of conjugated diene formation. In chemically induced LDL oxidation by Cu²⁺ plus hydrogen peroxide or peroxyl radical generated by 2, 2′-azo-bis (2-amidino propane) hydrochloride (AAPH), pentagalloic acid inhibited LDL oxidation as monitored by measuring the thiobarbituric acid reactive substances (TBARS), malondialdehyde (MDA), and gel electrophoretic mobility. The physiological relevance of the antioxidative activity was validated at the cellular level where pentagalloic acid inhibited mouse macrophage J774 and endothelial cell-mediated LDL oxidation. When compared with several other antioxidants, pentagalloic acid showed a much higher ability than naturally occuring antioxidants, α-tocopherol and ascorbic acid, and the synthetic antioxidant, probucol.     

Role of lipid peroxidation in the inhibition of mononuclear cell proliferation by normal lipoproteins

Stimulated peripheral blood mononuclear cells (PBMC) can oxidize normal lipoproteins, and sufficiently oxidized lipoproteins are cytotoxic. However, the role of lipid peroxidation in the inhibition of mitogen-stimulated PBMC proliferation by physiologic concentrations of normal lipoproteins is unclear. In the present investigation, normal low density lipoprotein (LDL) and very low density lipoprotein (VLDL) suppressed [3H]thymidine incorporation and gamma interferon production in concanavalin A-stimulated PBMC without causing cell death. This suppression was accompanied by parallel increases in lipid peroxidation products measured as thiobarbituric acid reactive substances (TBARS). In contrast, high density lipoprotein (HDL) failed to inhibit PBMC and TBARS remains low. Differences between the PBMC suppression from LDL, VLDL, and HDL were best accounted for by normalizing the lipoprotein concentrations by their total lipid content. Moreover, the antioxidants superoxide dismutase and butylated hydroxytoluene each substantially ameliorated the inhibition of PBMC caused by LDL, and reduced the levels of lipid peroxidation products that were generated. Altogether, these results suggest that reactive oxygen species generated by stimulated PMBC may cause oxidative alterations of normal lipoproteins that may, in turn, account for much of the previously reported inhibition of PBMC by normal lipoproteins.     

Atherosclerosis is enhanced by testosterone deficiency and attenuated by CETP expression in transgenic mice

In this work, we investigated the impact of testosterone deficiency and cholesteryl ester transfer protein (CETP) expression on lipoprotein metabolism and diet-induced atherosclerosis. CETP transgenic mice and nontransgenic (nTg) littermates were studied 4 weeks after bilateral orchidectomy or sham operation. Castrated mice had an increase in the LDL fraction (+36% for CETP and +79% for nTg mice), whereas the HDL fraction was reduced (-30% for CETP and -11% for nTg mice). Castrated mice presented 1.7-fold higher titers of anti-oxidized LDL (Ox-LDL) antibodies than sham-operated controls. Plasma levels of CETP, lipoprotein lipase, and hepatic lipase were not changed by castration. Kinetic studies showed no differences in VLDL secretion rate, VLDL-LDL conversion rate, or number of LDL and HDL receptors. Competition experiments showed lower affinity of LDL from castrated mice for tissue receptors. Diet-induced atherosclerosis studies showed that testosterone deficiency increased by 100%, and CETP expression reduced by 44%, the size of aortic lesion area in castrated mice. In summary, testosterone deficiency increased plasma levels of apolipoprotein B-containing lipoproteins (apoB-LPs) and anti-OxLDL antibodies, decreased LDL receptor affinity, and doubled the size of diet-induced atherosclerotic lesions. The expression of CETP led to a milder increase of apoB-LPs and reduced atherosclerotic lesion size in testosterone-deficient mice.     

Serum lipoproteins,lipid peroxides and prostacyclin biosynthesis in patients with coronary hear diseases

Serum low-density lipoproteins (LDL)_and high-density lipoproteins (HDL) were prepared by gradient ultracentrifugation and dialysis from 12 healthy subjects and 15 patients with coronary heart disease and hyperlipoproteinemia. In both lipoprotein fractions cholesterol and lipid peroxides were determined. The effect of these lipoproteins on spontaneous prostacyclin biosynthesis in rat aortic slices was studied.Serum lipoproteins were susceptible to peroxidation during the preparation procedure. LDL were more prone to peroxidation than HDL. Little lipid peroxidase were formed in lipoproteins when calcium ions had been removed by EDTA, and when butylated hydroxytoluene (BHT) was present at all stages of their preparation. LDL when prepared without these precautions either from healthy subjects or from patients with coronary heart diseases markedly suppressed prostacyclin generation by rat aortic slices. This inhibition to LDL-lipid peroxides. Peroxide-low LDL prepared from most of the healthy subjects and patients with coronary heart disease and concomitant hyperlipoproteinemia, did not inhibit prostacyclin biosynthesis. However, in one quarter of the patients. LDL was inhibitory. Consequently, in some patients with coronary heart disease, there operate unknown mechanisms which are responsible for the inhbibitory activity of LDL on prosctacylin generation.     

Plasma very-low-density lipoprotein, low-density lipoprotein, and high-density lipoprotein oxidative modification induces procoagulant profiles in endogenous hypertriglyceridemia

This study was to investigate whether oxidatively modified lipoproteins were associated with changes of pro- and anticoagulant profiles in hypertriglyceridemic subjects. Plasma VLDL, LDL, and HDL were isolated with the one-step density gradient ultracentrifugation method. The oxidation of the lipoproteins was identified. Prothrombin time (PT) and activated partial thrombplastin time (APTT), tissue plasminogen activator and plasminogen activator inhibitor-1, and platelet aggregation rate were determined with a reaction system consisting of mixed fresh normal plasma, in endogenous hypertriglyceridemic (HTG) patients, in in vitro modified lipoproteins from a normolipidemic donor, and in experimental rats. The results indicated that oxVLDL, oxLDL, and oxHDL occurred in the plasma of HTG patients. Compared with the control group, PT and APTT, incubated with plasma VLDL, LDL, or HDL from HTG patients, respectively, were significantly reduced, while platelet maximal aggregation rates were significantly higher (P < 0.05-0.01). Similar procoagulant profiles were observed in in vitro modified lipoprotein components and in rats with intrinsic hypertriglyceridemia as well. These results support our previous finding that LDL, VLDL, and HDL were all oxidatively modified in vivo in the subjects with HTG, and suggest that procoagulation state may result from the abnormal plasma lipoprotein oxidative modification in vivo.     

Lipoprotein subclass metabolism in nonalcoholic steatohepatitis

Nonalcoholic steatohepatitis (NASH) is associated with increased synthesis of triglycerides and cholesterol coupled with increased VLDL synthesis in the liver. In addition, increased cholesterol content in the liver associates with NASH. Here we study the association of lipoprotein subclass metabolism with NASH. To this aim, liver biopsies from 116 morbidly obese individuals [age 47.3 ± 8.7 (mean ± SD) years, BMI 45.1 ± 6.1 kg/m², 39 men and 77 women] were used for histological assessment. Proton NMR spectroscopy was used to measure lipid concentrations of 14 lipoprotein subclasses in native serum samples at baseline and after obesity surgery. We observed that total lipid concentration of VLDL and LDL subclasses, but not HDL subclasses, associated with NASH [false discovery rate (FDR) < 0.1]. More specifically, total lipid and cholesterol concentration of VLDL and LDL subclasses associated with inflammation, fibrosis, and cell injury (FDR < 0.1), independent of steatosis. Cholesterol concentration of all VLDL subclasses also correlated with total and free cholesterol content in the liver. All NASH-related changes in lipoprotein subclasses were reversed by obesity surgery. High total lipid and cholesterol concentration of serum VLDL and LDL subclasses are linked to cholesterol accumulation in the liver and to liver cell injury in NASH.     

Quantitative fluorescence imaging reveals point of release for lipoproteins during LDLR-dependent uptake

The LDL receptor (LDLR) supports efficient uptake of both LDL and VLDL remnants by binding lipoprotein at the cell surface, internalizing lipoprotein through coated pits, and releasing lipoprotein in endocytic compartments before returning to the surface for further rounds of uptake. While many aspects of lipoprotein binding and receptor entry are well understood, it is less clear where, when, and how the LDLR releases lipoprotein. To address these questions, the current study employed quantitative fluorescence imaging to visualize the uptake and endosomal processing of LDL and the VLDL remnant β-VLDL. We find that lipoprotein release is rapid, with most release occurring prior to entry of lipoprotein into early endosomes. Published biochemical studies have identified two mechanisms of lipoprotein release: one that involves the β-propeller module of the LDLR and a second that is independent of this module. Quantitative imaging comparing uptake supported by the normal LDLR or by an LDLR variant incapable of β-propeller-dependent release shows that the β-propeller-independent process is sufficient for release for both lipoproteins but that the β-propeller process accelerates both LDL and β-VLDL release. Together these findings define where, when, and how lipoprotein release occurs and provide a generalizable methodology for visualizing endocytic handling in situ.     

兔主动脉平滑肌细胞低密度脂蛋白受体相关蛋白(LRP)的诱导表达调节

在兔主动脉平滑肌细胞 ( SMC)培养基中分别加入正常低密度脂蛋白 ( N- LDL)、氧化低密度脂蛋白 ( ox- LDL)、正常极低密度脂蛋白 ( N- VLDL)、氧化极低密度脂蛋白 ( ox- VLDL)和 β-极低密度脂蛋白 (β- VLDL )培养 2 4 h后 ,用定量 RT- PCR和配体结合实验检测平滑肌细胞 LRP的m RNA和蛋白质水平的表达 .结果表明 :五种脂蛋白均能在转录和翻译水平诱导兔主动脉平滑肌细胞的 LRP表达 ,尤以富含胆固醇的 N- LDL ,ox- LDL和β- VLDL的刺激作用更明显 .用胆固醇单独或与脂蛋白共同温育 SMC后 ,发现胆固醇本身可促进 SMC的 LRP蛋白水平的表达 ,脂蛋白与胆固醇的共同刺激作用更为显著 .结果提示 :上述五种脂蛋白对 SMC上 LRP的表达有上调作用 ,其机制可能主要是通过其中的胆固醇来实现的 .     

Oxidative modification of lipoproteins in hypertriglyceridemic patients and hypercholesterolemic rabbits in vivo

Many lines of evidence suggest that LDL is oxidized in vivo and that Ox-LDL is present in the artery wall. But the oxidation of VLDL and HDL in vivo has not yet been reported. In this study, the oxidative modification of serum LDL, VLDL, and HDL in patients with endogenous hypertriglyceridemia (HTG) and in serum of rabbits fed on high cholesterol diet were made. The serum LDL, VLDL and HDL were isolated by the density gradient ultracentrifugation. The oxidative modification of LDL, VLDL and HDL were identified by agarose eletrophoresis, absorbance at 234 nm and fluorescence of TBARS. The results showed that serum TC, TG and TBARS in the HTG group (n= 25) and in rabbits fed with a high fat diet (for 12 weeks, n = 8) were significantly higher than those of the corresponding control groups (normal subjects, n = 25; rabbits fed with a normal diet, n = 8; p < 0.01). The electrophoretic mobilities of LDL, VLDL and HDL were increased when compared with the controls, and absorbance at 234 nm and TBARS of LDL, VLDL and HDL in the HTG group and in the high fat diet rabbits were significantly higher than those of the controls (p < 0.01). These results suggest that not only LDL but also VLDL and HDL were oxidatively modified in vivo in the patients with HTG and in the rabbits fed with a high cholesterol diet.     

A New Fluorescence Method for the Continuous Determination of Surface Lipid Oxidation in Lipoproteins and Plasma

We report on a new method for the determination of lipid oxidation in lipoproteins and plasma. The biological lipid system is preloaded with a fluorescent analog of phosphatidylcholine containing diphenylhexatriene (DPH) propionic acid covalently linked to the sn-2 position. When externally added, the respective phospholipid label (DPHPC) localizes to the surface monolayer of a lipoprotein. Under oxidative conditions (e.g. in the presence of Cu²⁺ ions) the fluorophore undergoes decomposition, resulting in a continuous decrease of fluorescence intensity which reflects the oxidation of a chemically defined phospholipid molecule with well defined localization. When incorporated into LDL particles, the kinetics of the decrease in DPHPC fluorescence intensity upon exposure to Cu²⁺ is very similar to that of conjugated diene accumulation. Furthermore, our assay can be applied to follow the oxidation of lipids in diluted serum and may also be developed into a suitable test system for clinical studies of susceptibility of plasma lipids to oxidation.     

High-fat/high-cholesterol diet promotes a S1P receptor-mediated antiapoptotic activity for VLDL

Withdrawing growth factors or serum from endothelial cells leads to the activation of effector caspases 3 and 7, resulting in apoptotic cell death. HDL protects against caspase induction through sphingosine-1-phosphate (S1P) receptors. This anti-caspase activity of HDL is antagonized by VLDL from apolipoprotein E4 (apoE4) (genotype, APOE4/4; apolipoprotein, apoE) targeted replacement (TR) mice, but not by VLDL from TR APOE3/3 mice, and requires the binding of apoE4-VLDL to an LDL receptor family member. In the absence of HDL, apoE4-VLDL and apoE3-VLDL from TR mice have limited antiapoptotic activity. In contrast, we show here that a high-fat/high-cholesterol/cholate diet (HFD) radically alters this biological activity of VLDL. On HFD, both apoE3-VLDL and apoE4-VLDL (HFD VLDL) inhibit caspase 3/7 activation initiated by serum withdrawal. This activity of HFD VLDL is independent of an LDL receptor family member but requires the activation of S1P(3) receptors, as shown by the ability of pharmacological block of S1P receptors by VPC 23019 and by small interfering RNA-mediated downregulation of S1P(3) receptors to inhibit HFD VLDL anticaspase activity.     

Role of parenchymal and non-parenchymal rat liver cells in the uptake of cholesterolester-labeled serum lipoproteins.

Very low density lipoprotein (VLDL)-remnants, prepared by extrahepatic circulation of VLDL, labeled biosynthetically in the cholesterol (ester) moiety, were injected intravenously into rats in order to determine the relative contribution of parenchymal and non-parenchymal liver cells to the hepatic uptake of VLDL-remnant cholesterol (esters). 82.7% of the injected radioactivity is present in liver, measured 30 min after injection. The non-parenchymal liver cells contain 3.1±0.1 times the amount of radioactivity per mg cell protein as compared to parenchymal cells. The hepatic uptake of biosynthetically labeled (screened) low density lipoprotein (LDL) and high density lipoprotein (HDL) cholesterolesters amounts to 26.8% and 24.4% of the injected dose, measured 6 h after injection. The non-parenchymal cells contain 4.3±0.8 and 4.1±0.7 times the amount of radioactivity per mg cell protein as compared to parenchymal cells for LDL and HDL, respectively. It is concluded that in addition to parenchymal cells, the non-parenchymal cells play an important role in the hepatic uptake of cholesterolesters from VLDL-remnants, LDL and HDL.     

Effects of some flavonoids on the susceptibility of low-density lipoprotein to oxidative modification

The effects of six flavonoids viz., apigenin, genistein, morin, naringin, pelargonidin and quercetin on the susceptibility of low-density lipoprotein (LDL) to oxidative modification were investigated. Flavonoids were added to plasma and incubated for 3 hr at 37 degrees C, and the LDL fraction was separated by ultracentrifugation. Oxidizability of LDL was estimated by measuring conjugated diene (CD), lipid peroxides and thiobarbituric acid-reactive substances (TBARS), after cupric sulfate solution was added. Quercetin and morin significantly (P<0.01 by ANOVA) prolonged the lag time before initiation of oxidation reaction in dose-dependent manner. They also suppressed the formation of lipid peroxides and TBARS more markedly than other flavonoids. The ability to prolong lag time and suppression of lipid peroxides and TBARS formation was in the following order: quercetin >morin >pelargonidin >genistein >naringin >apigenin. LDL exposed to flavonoids reduced oxidizability. These findings suggest that flavonoids may have a role in ameliorating atherosclerosis.