Low-density Lipoprotein Antioxidant Research Articles

Antioxidant potential and low-density lipoprotein cholesterol (LDL-c) uptake of the black seed and honey mixture on human hepatocellular carcinoma (HepG2) cells

High lipid levels especially low-density-lipoprotein cholesterol (LDL-c) are associated with increased risk of cardiovascular (CVD) and coronary heart disease (CHD). Both black seed (Nigella sativa L.) and honey are well-known in the hypolipidemic potential and have CVD protective effects. In the present study, LDL-c uptake of the black seed and honey mixture was tested on HepG2 cells. Antioxidant activities of black seed and honey mixtures were determined through the 2, 2’-diphenyl-1-picrylhydrazyl (DPPH) assay. The anticancer potential of black seed and honey mixtures in HepG2 cells was performed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) cytotoxicity assay. Black seed possessed the highest antioxidant activities with EC50 6.54 mg/mL as compared to honey with EC50 value 9.56 mg/mL while the black seed and honey mixture have EC50 between black seed and honey. From the results obtained, no synergistic effect was observed in the mixtures as the EC50 values were within the range of black seed and honey. Furthermore, no significant difference (p>0.05) among ratios (1:1, 2:1 and 1:2). However, the decrease in cell proliferation was the highest in black seed and honey mixture at 1:1 ratio (p<0.05) than individually treated black seed and honey. Thus, the black seed and honey mixture at ratio 1:1 was the most potent anticancer agent with an IC50 value of 7.44 μg/mL. The present study illustrated that black seed and honey mixtures possess a lipid-lowering effect via LDL-c uptake in HepG2 cells (p<0.05). The highest LDL-c uptake was observed at 15 μg/mL with the treatment of black seed and honey mixture at 1:2 ratio which was 294.4%. Further studies should be conducted on primary human liver cells to further justify the correlation between the antioxidant level and LDLc uptake mechanism of black seed and honey mixtures.

Ubiquinol Ameliorates Endothelial Dysfunction in Subjects with Mild-to-Moderate Dyslipidemia: A Randomized Clinical Trial.

In this randomized, double-blind, single-center trial (ANZCTR number ACTRN12619000436178) we aimed to investigate changes in endothelium-dependent vasodilation induced by ubiquinol, the reduced form of coenzyme Q10 (CoQ10), in healthy subjects with moderate dyslipidemia. Fifty-one subjects with low-density lipoprotein (LDL) cholesterol levels of 130–200 mg/dL, not taking statins or other lipid lowering treatments, moderate (2.5%–6.0%) endothelial dysfunction as measured by flow-mediated dilation (FMD) of the brachial artery, and no clinical signs of cardiovascular disease were randomized to receive either ubiquinol (200 or 100 mg/day) or placebo for 8 weeks. The primary outcome measure was the effect of ubiquinol supplementation on FMD at the end of the study. Secondary outcomes included changes in FMD on week 4, changes in total and oxidized plasma CoQ10 on week 4 and week 8, and changes in serum nitrate and nitrite levels (NOx), and plasma LDL susceptibility to oxidation in vitro on week 8. Analysis of the data of the 48 participants who completed the study demonstrated a significantly increased FMD in both treated groups compared with the placebo group (200 mg/day, +1.28% ± 0.90%; 100 mg/day, +1.34% ± 1.44%; p < 0.001) and a marked increase in plasma CoQ10, either total (p < 0.001) and reduced (p < 0.001). Serum NOx increased significantly and dose-dependently in all treated subjects (p = 0.016), while LDL oxidation lag time improved significantly in those receiving 200 mg/day (p = 0.017). Ubiquinol significantly ameliorated dyslipidemia-related endothelial dysfunction. This effect was strongly related to increased nitric oxide bioavailability and was partly mediated by enhanced LDL antioxidant protection.

DNA scission and LDL cholesterol oxidation inhibition and antioxidant activities of Bael (Aegle marmelos) flower extracts

Phenolic compounds and other antioxidants have been implicated in protection against non-communicable diseases (NCDs) in which oxidative stress is a main contributor. The extracts of Bael (Aegle marmelos) flower were examined for their phenolic content, free radical scavenging efficacy and inhibition of low density lipoprotein (LDL) cholesterol oxidation and DNA scission activities. The extracts of Bael flowers were prepared using different solvent systems and their total phenolic content (TPC) and total flavonoid content (TFC) determined. Selected extracts which showed high TPC were subsequently used to determine their efficacy in scavenging hydroxyl, and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals, using electron paramagnetic resonance (EPR) spectroscopy. The corresponding peroxyl radical scavenging activity was measured using oxygen radical absorbance capacity (ORAC) assay. The potency of the extracts in inhibiting hydroxyl and peroxyl radical-induced supercoiled DNA scission and inhibition of LDL cholesterol oxidation was also evaluated. The chemical identity of phenolic compounds present in the extracts was tentatively unraveled using HPLC-MS. Phenolic extracts of Bael flowers effectively inhibited hydroxyl, and peroxyl radicals. Phenolic extracts demonstrated notable inhibitory activity against hydroxyl and peroxyl radical-induced DNA scission and LDL oxidation. Vanillic, p-coumaric, chlorogenic, caffeic, and gentisic acids were identified as major phenolic acids, along with flavonoids, mainly catechin, and quercetin. The knowledge gained here may help better use of Bael flower extracts as functional herbal beverage ingredients in the prevention of NCDs.

The Health Benefits of Jeju Gamgyul ( Citrus unshiu Marc.) Brandy and Wine

Beneficial health effects of fruits and vegetables in the diet have been attributed to the reduction of risk for some diseases, such as diabetes, chronic diseases, cancer, or cardiovascular diseases. Citrus and its peels have been used in Asian folk medicine due to abundant flavonoids and the fact that their pulp are usually consumed like juice or jam, although their functional effects are much higher than those of fruit peels. This investigation inquired into the effects of Gamgyul wine, which is derived from whole fruits of Citrus unshiu, with respect to antioxidant property, anti-diabetic activity, low density lipoprotein (LDL) oxidative inhibition and flavanones. In particular, antioxidant and antidiabetic activities of both Gamgyul wine and brandy were measured. However, the differences of the antioxidant and antidiabetic effects of Gamgyul wine were trifling between dynamite and microfiltration processes. In particular, the effect of antidiabetic activity was rather significant as demonstrated in the Gamgyul brandy. The results showed that α-glucosidase inhibition rates at 20 µL/mL for Gamgyul brandy and wine were 50% and 20%, respectively. Almost 100% inhibition rate was shown at 80 µL/mL treatment for all the samples. Antioxidant activities and LDL oxidative inhibition were also measured. Gamgyul wine showed increased concentration-dependent antioxidant activities, while epigallocatechin gallate (EGCG) revealed an almost same level of activities. On the other hand, Gamgyul brandy presented little activity at any treatment concentration. Gamgyul wine in 20 µL/mL inhibited LDL oxidation approximately three times more than that of the comparative negative control, D.W. Flavanone analysis showed detections of both narirutin and hesperidin, while narirutin was the most abundant flavanone in Gamgyul wine. These overall results suggest that Gamgyul wine has the potential to be used as a functional dietary supplement and is an effective α-glucosidase inhibitor, which may be beneficial only for low to moderate intake.

Low-Density Lipoprotein (LDL) Antioxidants and Antiplatelet Compounds from Selected Malaysian Plants as Leads for the Development of Cardiovascular Protective Agents

Oxidative modification of low-density lipoprotein (LDL) plays a significant role in the onset and acceleration of atherosclerotic process and the development of cardiovascular diseases. The chronic antioxidant, hypolipidemic, antithrombotic and antiplatelet activities of many phytochemicals have important roles in prevention of lipoprotein oxidation and atherosclerotic lesion development. Phytochemicals of diverse chemical structures, including phenolic compounds, xanthones, prenylflavonoids, benzophenones, alkaloids and triterpenoids isolated from Malaysian plants were investigated for their ability to inhibit arachidonic acid (AA), collagen and adenosine diphosphate (ADP) induced platelet aggregation in human whole blood using an electrical impedance method. The compounds were also investigated for their ability to inhibit human LDL oxidation using thiobarbituric acid reactive substances (TBARS) assay. Among the compounds tested, 2-(3-methylbut-2-enyl)1,3,5,-trihydroxyxanthone, 2-(3-methylbut-2-enyl)-1,3,5,6-tetrahydroxyxanthone, rubraxanthone and 3,5,3′,5′-tetrahydroxy-4methoxybenzophenone isolated from Guttiferae species showed strong inhibition on platelet aggregation. Curcumin, cardamonin, pinocembrine, 5,6-dehydrokawain and 3-deacetylcrotepoxide were the strong antiplatelet compounds identified from Zingiberaceae species. Of the alkaloids tested, (-)-N-acetylnornuciferine, (-)-N-acetylanonaine and 1-(N-acetyl-Nmethylamino)ethyl-3,4,6-trimethoxy-7-hydroxyphenanthrene from Armodendron elegans showed strong inhibition on platelet aggregation caused by all three inducers. 3,5,3′,5′-Tetrahydroxy-4-methoxybenzophenone and 1, 7-dihydroxyxanthone showed strong inhibitory activity on LDL oxidation. Curcumin, demethoxycurcumin and bisdemethoxycurcumin exhibited stronger activity than the reference, probucol, as LDL antioxidants. Xanthorrhizol, zerumbone and ar-turmerone from the oils of Curcuma species showed relatively strong antioxidant activity. 3,5,3′,5′-Tetrahydroxy-4-methoxybenzophenone and 1, 7dihydroxyxanthone also showed strong inhibitory activity on LDL oxidation. Of the triterpenoids tested, garcihombronane B from Garcinia cantleyana showed moderate activity against LDL oxidation. The compounds are new classes of natural products that can be used as leads for development of new cardiovascular protective agents.

Lipoprotein X: clinical implications

In this issue of the journal, Stepien et al. describe a case of serum lipoprotein X (Lp-X) in a patient with cholestasis and severe hyperlipidaemia. This abnormal lipoprotein is rich in phospholipid and unesterified cholesterol. The main protein component is albumin within the core and apolipoprotein C upon the surface. Unlike low-density lipoprotein (LDL), Lp-X does not contain apolipoprotein B nor is it removed by the LDL receptor, but Lp-X is cleared by the reticuloendothelial system and the kidneys. Lp-X is about 50 nm in diameter and its major component bile acid is lithocholic. Early studies reported the presence of Lp-X in the serum of patients with cholestasis. The failure of Lp-X to exert feedback inhibition on cholesterol synthesis may contribute to the mechanism of hypercholesterolaemia in obstructive jaundice. However, the presence of serum Lp-X does not facilitate the distinction between intrahepatic and extrahepatic cholestasis. Bile lipoprotein is a precursor of Lp-X and in cholestasis this refluxes into the plasma pool and binds to albumin to form Lp-X. Serum Lp-X usually disappears soon after the cholestasis is relieved. The ratio of plasma bile salts to albumin is important for the integrity of Lp-X and it is catabolized by the action of phospholipase which helps convert lecithin into lysolecithin. In graft versus host disease involving the liver and with cholestasis plasma Lp-X can result in severe hypercholesterolaemia, xanthelasma and cholesteroloma. Lp-X can also be found in cholestasis of pregnancy and primary biliary cirrhosis (PBC). The case presented by Stepien et al. confirms these earlier observations of raised serum Lp-X in cholestasis. Increased serum cholesterol concentrations occur in PBC and this is largely due to the presence of Lp-X, although it is debatable whether this confers increased cardiovascular risk. Indeed, Lp-X may not be atherogenic and data suggest it may have antioxidant LDL activity. Additionally, Lp-X is not significantly taken up by macrophages but is taken up by hepatocytes. Sometimes severe and prolonged cholestasis can result in very high serum Lp-X concentrations resulting in severe hypercholesterolaemia and tendon xanthomata. Complete regression of these xanthomata has been described by treatment with apheresis, which the authors stated should be considered possible first-choice treatment for patients with massive Lp-X accumulation due to cholestasis. Statin therapy may be ineffective in lowering Lp-X. Lecithin-cholesterol acyltransferase (LCAT) is an enzyme responsible for cholesterol esterification in plasma. Lp-X is found in LCAT deficiency which is characterized by low concentrations of high-density lipoprotein (HDL) as well as anaemia, corneal opacities, proteinuria and chronic kidney disease. The Lp-X found in LCAT deficiency may contribute to the renal disease observed in this condition by stimulating monocyte renal infiltration. Proteoglycans have an important role in regulating the uptake of Lp-X into kidney mesangial cells and this mechanism is independent of the scavenger receptor. Guerin et al. reported that in LCAT deficiency the presence of Lp-X is correlated with a progressive change in renal function. In addition to cholestasis and LCAT deficiency, Lp-X may also be found in the plasma after intravenous infusion of Intralipid emulsion such as occurs with total parenteral nutrition. Intralipid administration can elicit an increase in serum total cholesterol (mainly unesterified), phospholipid and LDL. The Lp-X associated with Intralipid infusion seems to occur when phospholipid clearance is exceeded, resulting in phospholipid accumulation with unesterified cholesterol. Pseudohyponatraemia has also been described in some patients with severe hypercholesterolaemia secondary to raised Lp-X. Thus the presence of Lp-X should be considered in pseudohyponatraemia associated with cholestasis. Sivakumar et al. described not only pseudohyponatraemia, but also pseudohypokalaemia and pseudohypochloridaemia in the presence of high concentrations of Lp-X. Various researchers have also found assay bias for serum LDL-cholesterol in the presence of Lp-X. This is an important practical point when it comes to interpreting LDL cholesterol results in severely cholestatic serum samples. Lp-X possesses strong aggregatory properties and can bind to alkaline phosphatase (ALP) producing a Lp-X-ALP complex. This complex may be found in cholestasis; typically that evoked by hepatic malignancy. The Lp-X-ALP complex is also associated with the presence of high-molecular mass ALP and the liver isoenzyme of ALP. Brocklehurst et al. described a stationary band of ALP using starch gel electrophoresis of serum samples from patients with biliary obstruction which was thought to be due to ALP binding to Lp-X. The strong aggregating ability of Lp-X may be important as it can bind and remove free cholesterol from the circulation. There are various assays available for measuring serum Lp-X. Agarose gel electrophoresis utilizes the cathodal

Correlation between Chemical Composition ofCurcuma domesticaandCurcuma xanthorrhizaand Their Antioxidant Effect on Human Low-Density Lipoprotein Oxidation

The antioxidant activity of the curcuminoids of Curcuma domestica L. and C. xanthorrhiza Roxb. and eight compounds which are prevalent constituents of their rhizome oils were investigated in an effort to correlate human low-density lipoprotein (LDL) antioxidant activity with the effect of the herbs and their components. The antioxidant activity was examined using thiobarbituric acid reactive substances (TBARSs) assay with human LDL as the oxidation substrate. The methanol extracts and rhizome oils of C. xanthorrhiza and C. domestica showed strong inhibitory activity on copper-mediated oxidation of LDL. Curcumin, demethoxycurcumin, and bisdemethoxycurcumin, isolated from the methanol extracts of both plants, exhibited stronger activity than probucol (IC50 value 0.57 μmol/L) as reference, with IC50 values ranging from 0.15 to 0.33 μmol/L. Xanthorrhizol, the most abundant component (31.9%) of the oil of C. xanthorrhiza, showed relatively strong activity with an IC50 value of 1.93 μmol/L. The major components of C. domestica, ar-turmerone (45.8%) and zerumbone (3.5%), exhibited IC50 values of 10.18 and 24.90 μmol/L, respectively. The high levels of curcuminoids in the methanol extracts and xanthorrhizol, ar-turmerone and zerumbone in the oils, and in combination with the minor components were responsible for the high LDL antioxidant activity of the herbs.

Effects of selected medicinal plants on human low-density lipoprotein oxidation, 2, 2-diphenyl-1-picrylhydrazyl (DPPH) radicals and human platelet aggregation

The effects of the methanol extracts of 20 selected medicinal plants on free radical scavenging capacity, human low-density lipoprotein (LDL) peroxidation and platelet aggregation were investigated. The antioxidant activity was evaluated using 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging capacity and thiobarbituric acid reactive substances (TBARS) assay with LDL as the oxidation substrate. The antiplatelet activity in human whole blood was investigated using an electrical impedance method. The total phenolic contents (TPC) of the extracts were determined by the Folin-Ciocalteau method. Among the extracts, Phyllanthus amarus and Labisia pumila var. alata possessed potent radical scavenging activity with IC50 values of 3.4 and 5.7 µg/ml, respectively. The extracts ofZingiber officinale, Curcuma xanthorrhiza and Curcuma domestica showed strong inhibition of LDL peroxidation with IC50 values ranging from 0.1 to 0.9 µg/ml. The extract of Z. officinalewas the most effective sample against platelet aggregation caused by arachidonic acid (AA) and adenosine diphosphate (ADP) with IC50 values of 10.9 and 7.7 µg/ml, respectively. The Pearson correlation analysis indicated that the TPC of the extracts showed significant positive correlations with DPPH scavenging activity (r = 0.846) and LDL antioxidant activity (r= 0.639) and moderate positive correlations with antiplatelet activities (AA-induced, r = 0.51; ADP-induced, r = 0.40; collagen-induced, r = 0.44). The antioxidant and antiplatelet activities of the plant extracts could partly be due to their TPC. Key words: Medicinal plants, LDL antioxidant activity, antiplatelet activity, DPPH scavenging capacity, total phenolic contents.

Inhibitory effects of the extracts of Garcinia species on human low-density lipoprotein peroxidation and platelet aggregation in relation to their total phenolic contents

Twenty-two methanol extracts from different parts of nine Garcinia species were investigated for their ability to inhibit platelet aggregation and low-density lipoprotein (LDL) peroxidation and their total phenolic contents (TPC). The antioxidant activity of the extracts was examined using thiobarbituric acid reactive substances (TBARS) assay with human LDL as the oxidation substrate and their antiplatelet activity in human whole blood was determined by using an electrical impedance method. The TPC of the extracts were measured by the Folin-Ciocalteau method. Among all samples studied, the leaf extract ofGarcinia eugenifolia Wall showed the highest inhibitory activity on LDL peroxidation with IC50 value of 12.5 µg/ml. The twig extract of Garcinia mangostana Linn. was the most effective sample against platelet aggregation caused by arachidonic acid (AA) with IC50value of 15.6 µg/ml. The TPC of the extracts varied from 4.4 to 62.8 mg of gallic acid equivalents per g (mg GAE/g). The Pearson correlation analysis revealed that TPC showed moderate positive correlations with antioxidant (r = 0.30, p < 0.05) and antiplatelet activities (AA-induced, r = 0.62, p < 0.05; ADP-induced, r = 0.42, p < 0.05; collagen-induced, r = 0.54, p < 0.05). Thus, it was concluded that the antioxidant and antiplatelet activities of theGarcinia extracts could partly be due to their total phenolic contents. Key words: Garcinia species, low-density lipoprotein, LDL antioxidant activity, antiplatelet activity, total phenolic contents.

LDL Oxidation, Antioxidant Capacity and Growth of Cultured Grey Mullet (Mugil cephalus) Fed Dietary Sorghum Distillery Residue Pretreated with Polyethylene Glycol

Dietary sorghum distillery residue (SDR) showed antioxidant and blood thinning effects on grey mullet during winter, but inhibited their growth. The objective of this study was to establish a preliminary treatment of the dietary SDR with polyethylene glycol (PEG), a tannin-binding agent, to enhance growth and blood antioxidant capacity of grey mullet ( Mugil cephalus ) feed. The feeding trial was carried out from June to November. The water temperature was between 25 and 30 degrees C; the specific growth rate of mullet was reduced significantly by feeding diet containing 20% SDR in comparison to fish fed the control diet or diet containing 20% SDR and PEG. In the period of October-November, the water temperature decreased to 19-25 degrees C; the specific growth rates of the 20% SDR-PEG group and the 20% SDR group were 0.13 and 0.19% day(-1), respectively, significantly higher than those fed the control diet (0.07% day(-1)). Feeding with 20% SDR or 20% SDR-PEG diets resulted in prolonged lag phase of low-density lipoprotein (LDL) oxidation compared to fish fed the control diet. The total antioxidant capacity of the plasma of the grey mullet fed 20% SDR-PEG was 1.24 mmol/L, significantly higher than those in the fish fed 20% SDR diet (0.84 mmol/L) or the control (0.72 mmol/L). In vivo observations found that preliminary treatment of SDR with PEG eliminated the endogenous undesirable growth inhibitory factors but maintained its protective effects against LDL oxidation in blood and improved the total antioxidant capacity and cold adaptation of grey mullet. The ethanol extract of SDR contained 31.9 +/- 7.8 mg/g gallic acids equivalent. The concentration needed to scavenge 50% of the DPPH radicals (IC(50)) was 0.86 mg/mL. Increased gallic acid equivalent and decreased IC(50) of DPPH scavenging activity of SDR fed to fish increased the total antioxidant capacity in blood plasma of grey mullet significantly.

Antioxidant effects of hormone replacement therapy in postmenopausal women.

Postmenopausal women are more likely to develop coronary artery disease than premenopausal women of the same age. Postmenopausal oral oestrogen therapy is associated with increased levels of high-density lipoprotein (HDL) cholesterol and decreased levels of lowdensity lipoprotein (LDL) cholesterol. In this study we investigated the direct contribution of hormone replacement therapy on total antioxidant capacity rather than its effects on the serum lipid profile. At the time of enrolment and after the drug delivery plasma total cholesterol, triglycerides (TG), HDL, LDL, uric acid, total bilirubin, albumin, oestradiol levels and total antioxidant capacity of plasma were assessed. Levels of plasma TG and total antioxidant capacity were significantly increased in the study group. Our results suggest that oestrogen has an antioxidant effect following 3 months of hormone replacement therapy. Progesterone in combination with oestrogen does not have this effect. Also plasma TG levels increased in those patients receiving HRT after 3 months.

Electronegative low‐density lipoprotein subfraction from type 2 diabetic subjects is proatherogenic and unrelated to glycemic control

The physicochemical and biological characteristics of electronegative low-density lipoprotein (LDL) (LDL(-)) from type 2 diabetic patients (DM2), before and after insulin therapy, were studied. Total LDL was subfractionated in LDL(+) (native LDL) and LDL(-) by anion-exchange chromatography. The proportion of LDL(-) was increased in plasma from DM2 patients compared to control subjects (13.8 +/- 4.6% versus 6.1 +/- 2.5, P < 0.05) and was not modified after glycemic optimization (14.0 +/- 5.9%). LDL(-) from DM2 patients presented similar differential characteristics versus LDL(+) than LDL(-) from controls; that is, decreased apoB and oxidizability, and increased triglyceride, nonesterified fatty acids (NEFA), apoE, apoC-III, platelet-activating factor (PAF) acetylhydrolase activity and aggregability. No difference in particle size, antioxidants, malondialdehyde (MDA), fructosamine or glycated low-density lipoprotein (gLDL) was observed between LDL subfractions. Concerning differences between LDL subfractions isolated from DM2 and from control subjects, the former showed increased MDA, fructosamine and gLDL proportion and decreased LDL size and antioxidant content. The only effect of glycemic optimization was a decrease in fructosamine and gLDL in LDL(+) from DM2 subjects. LDL(-) from DM2 patients presented low binding affinity to the low-density lipoprotein receptor (LDLr) in cultured fibroblasts compared to LDL(+) and two- to threefold increased ability to release interleukin-8 (IL-8) and monocyte chemotactic protein 1 (MCP-1) in endothelial cells. These results suggest that, although nonenzymatic glycosylation and oxidation are increased in type 2 diabetes, these features would not be directly involved in the generation of LDL(-). Moreover, LDL(-) properties suggest that the high proportion observed in plasma could promote accelerated atherosclerosis in DM2 patients through increased residence time in plasma and induction of inflammatory responses in artery wall cells.

Antioxidant flavonol glycosides in mulberry ( Morus alba L.) leaves isolated based on LDL antioxidant activity

Oxidation of low-density lipoprotein (LDL) has been implicated in atherogenesis. Antioxidants that prevent LDL from oxidation may reduce atherosclerosis. We investigated LDL antioxidant activity and extracted compounds of mulberry ( Morus alba L.) leaves. The LDL antioxidant activity of 60% ethanol extracted of mulberry leaves, which inhibits human LDL oxidation induced by copper ion, was determined on the basis of oxidation lag time and calculated as epigallocatechin 3-gallate equivalents (58.3 μmol of EGCG equivalent/g of dry weight). Three flavonol glycosides [quercetin 3-(6-malonylglucoside), rutin (quercetin 3-rutinoside) and isoquercitrin (quercetin 3-glucoside)] were identified as the major LDL antioxidant compounds by LC-MS and NMR. The amounts of these flavonol glycosides in mulberry leaves and mulberry-leaf tea were determined by HPLC. Our results showed that quercetin 3-(6-malonylglucoside) and rutin were the predominant flavonol glycosides in the mulberry leaves.

Dietary Antioxidants for Prevention of Cardiovascular Disease

The traditional Japanese diet with its high intake of fruits and vegetables that are rich in antioxidants is believed to effectively ward off cardiovascular disease. Oxidative stress, related to reactive oxygen and nitrogen species produced by aerobic organisms, is responsible for the pathogenesis of most chronic diseases. Oxidative low density lipoprotein (LDL) is thought to play a key role in the pathogenesis of early atherosclerosis. There has been increasing interest in antioxidant substances derived from edible plants. It has been suggested that the measurement of LDL antioxidant activity is physiopathologically more important and informative for screening antioxidant activity to prevent atherosclerosis than other methods. We assessed by LDL oxidation assay the antioxidant characteristics of various edible plants from rural areas. The mulberry (Morus alba L.) leaf showed comparatively high antioxidant activity. We identified the antioxidant compounds and investigated compound levels in the mulberry leaf and found the antioxidant activity of mulberry leaves to be mainly attributable to quercetin 3-(6-malonylglucoside) (Q3MG). Dietary consumption of mulberry leaves and/or Q3MG may enhance resistance to oxidative modification of LDL and attenuated atherosclerotic lesion development. However, the outcome of intervention trials suggested that a single antioxidant had little effect on the risk of developing cardiovascular disease. The synergic effect of certain combinations may determine outcome. We need to widen our understanding of the synergic effect of diets and nutrient-gene interactions related to nutrient/disease risk.

α-tocopherol is important to inhibit low-density lipoprotein oxidation in smokers

Smoking is recognized as an independent risk factor for atherosclerosis. This study was designed to study the effect of smoking on plasma and low-density lipoprotein (LDL) antioxidant concentrations and LDL oxidation susceptibility. A total of 89 subjects, consisting of 27 smokers and 62 nonsmokers, were recruited. Biochemical values and plasma ascorbic acid, α-tocopherol, and LDL α-tocopherol levels were assessed. LDL lag time, oxidation rate, and correlation between LDL α-tocopherol and LDL oxidation rate were determined. No differences were demonstrated for ascorbic acid and α-tocopherol concentrations in plasma between smokers and nonsmokers; however, LDL α-tocopherol was significantly higher for nonsmokers ( P = 0.012). Furthermore, the lag time for LDL oxidation was significantly longer for nonsmokers ( P = 0.019), whereas the LDL oxidation rate was significantly greater for smokers ( P = 0.018). Additionally, a highly significant negative correlation was demonstrated for LDL α-tocopherol concentration and LDL oxidation rate in smokers ( r = −0.440, P = 0.026), but not in nonsmokers. The results of this study suggest that LDL α-tocopherol is more susceptible to oxidative stress caused by smoking than ascorbic acid and α-tocopherol in plasma. For smokers, the presence of α-tocopherol in LDL effectively protects LDL from oxidation.

Berry anthocyanins: isolation, identification and antioxidant activities

AbstractAnthocyanins from bilberry, blackcurrant and cowberry were isolated for antioxidant evaluation. Individual compounds were identified and quantified using HPLC and HPLC/ESI–MS techniques. Antioxidant and radical‐scavenging capacities of the isolates were studied in emulsified methyl linoleate and human low‐density lipoprotein (LDL) in vitro and in the 2,2‐diphenyl‐1‐picrylhydrazyl (DPPH) test. The total anthocyanin contents in the phenolic extracts of bilberry, blackcurrant and cowberry were 6000, 2360 and 680 mg kg−1 fresh weight respectively. There were four dominant compounds in blackcurrant (glucosides and rutinosides of cyanidin and delphinidin), three in cowberry (monoglycosides of cyanidin) and 15 in bilberry (monoglycosides of cyanidin, delphinidin, malvidin, peonidin and petunidin). Quantification as cyanidin‐3‐glucoside equivalents gave markedly lower results regarding the total anthocyanin concentration and the content of individual delphinidin and malvidin compounds compared with quantification based on corresponding standard compounds. Berry anthocyanins were highly active radical scavengers in the DPPH test and effective antioxidants in emulsion and human LDL. Copyright © 2003 Society of Chemical Industry

Smoking Has No Effect on the Amino Acid Composition of Apolipoprotein B100 of LDL While Directly Influencing the Antioxidant Status

Previous studies have demonstrated increased plasma levels of oxidised low-density lipoprotein (oxLDL) in chronic smokers, which has been associated with the extent of endothelial dysfunction. In this study we examine the relationship between the amino acid composition of apolipoprotein B100 (apo B) of low-density lipoprotein (LDL), by reverse phase HPLC after precolumn derivatisation, between smokers (≥40 cigarettes/day) and nonsmokers in relation to their plasma and LDL antioxidant status. While there was a significant difference in the levels of plasma vitamin C and α-tocopherol between female smokers and nonsmokers, as well as in the levels of LDL α-tocopherol, there was no significant difference in the amino acid composition of apo B between the two groups.

The effects of lipid-lowering therapy on low-density lipoprotein auto-antibodies: relationship with low-density lipoprotein oxidation and plasma total antioxidant status.

Oxidized low-density lipoprotein (Ox-LDL) is believed to play an important role in the progression of atherosclerosis. Oxidative modification of low-density lipoprotein (LDL) is a prerequisite for rapid accumulation of LDL in macrophages and for the formation of foam cells. Because of high antioxidant levels in plasma, LDL oxidation is suggested to occur mainly in the subendothelial space of the arterial wall, where there is the concomitant presence of large amounts of reactive oxygen species generated by endothelial cells and activated leukocytes. After Ox-LDL formation, antibodies against this form of LDL may occur. Auto-antibodies against Ox-LDL (AuAb-Ox-LDL) show directly in in-vivo LDL oxidation. Many studies have indicated that the amount of antibodies in serum is positively correlated to the rate of progression of atherosclerotic plaques. In this study the effect of lipid-lowering therapy on the levels of AuAb-Ox-LDL in patients with dyslipidemia was determined using atorvastatin (10 mg/day), and the relationship between the antibodies and plasma total antioxidant status (TAS) and LDL oxidation capacity was also investigated. Serum levels of AuAb-Ox-LDL, lipids, lipoproteins, TAS and susceptibility of LDL to oxidation were determined using lag time in 44 patients with dyslipidemia (29 with hypercholesterolemia and 15 with mixed-type hyperlipidemia). After lipid-lowering therapy, serum levels of AuAb-Ox-LDL were found to be significantly decreased, by 18.7%, while lag time and plasma TAS were increased (31.3% and 7.6% respectively) in patients with dyslipidemia. The percentage change in lag time was found to be negatively correlated to the percentage change in AuAb-Ox-LDL (r = -0.31, P < 0.05). The percentage change in lag time also showed a positive correlation with the percentage change in TAS (r = 0.58, P < 0.01). AuAb-Ox-LDL levels decreased by 21.7% in patients with hypercholesterolemia and by 12.6% in patients with mixed-type hyperlipidemia. Also AuAb-Ox-LDL levels in patients with hypercholesterolemia were higher than in those with mixed-type hyperlipidemia (367 +/- 294 compared with 300 +/- 176 mU/l). It was concluded that lipid-lowering therapy may contribute to the reduction in levels of AuAb-Ox-LDL and the increase in the antioxidant capacity of plasma LDL and TAS. It was also suggested that the measurement of antibodies against Ox-LDL during lipid-lowering therapy may be used as an important marker for representing in-vivo LDL oxidation and atherosclerotic processes.

Diffusion of Nitric Oxide into Low Density Lipoprotein

A key early event in the development of atherosclerosis is the oxidation of low density lipoprotein (LDL) via different mechanisms including free radical reactions with both protein and lipid components. Nitric oxide (( small middle dot)NO) is capable of inhibiting LDL oxidation by scavenging radical species involved in oxidative chain propagation reactions. Herein, the diffusion of ( small middle dot)NO into LDL is studied by fluorescence quenching of pyrene derivatives. Selected probes 1-(pyrenyl)methyltrimethylammonium (PMTMA) and 1-(pyrenyl)-methyl-3-(9-octadecenoyloxy)-22,23-bisnor-5-cholenate (PMChO) were chosen so that they could be incorporated at different depths of the LDL particle. Indeed, PMTMA and PMChO were located in the surface and core of LDL, respectively, as indicated by changes in fluorescence spectra, fluorescence quenching studies with water-soluble quenchers and the lifetime values (tau(o)) of the excited probes. The apparent second order rate quenching constants of ( small middle dot)NO (k(NO)) for both probes were 2.6-3.8 x 10(10) m(-1) s(-1) and 1.2 x 10(10) m(-1) s(-1) in solution and native LDL, respectively, indicating that there is no significant barrier to the diffusion of ( small middle dot)NO to the surface and core of LDL. Nitric oxide was also capable of diffusing through oxidized LDL. Considering the preferential partitioning of ( small middle dot)NO in apolar milieu (6-8 for n-octanol:water) and therefore a larger ( small middle dot)NO concentration in LDL with respect to the aqueous phase, a corrected k(NO) value of approximately 0.2 x 10(10) m(-1) s(-1) can be determined, which still is sufficiently large and consistent with a facile diffusion of ( small middle dot)NO through LDL. Applying the Einstein-Smoluchowsky treatment, the apparent diffusion coefficient (D(')NO) of ( small middle dot)NO in native LDL is on average 2 x 10(-5) cm(2) s(-1), six times larger than that previously reported for erythrocyte plasma membrane. Thus, our observations support that ( small middle dot)NO readily traverses the LDL surface accessing the hydrophobic lipid core of the particle and affirm a role for ( small middle dot)NO as a major lipophilic antioxidant in LDL.

Composition of LDL as determinant of its susceptibility to in vitro oxidation in patients with well-controlled type 2 diabetes.

There is increasing evidence that oxidation of low-density lipoprotein (LDL) in the vascular wall plays an important role in the development of atherosclerosis. The present study was undertaken to characterise how different constituents of LDL contribute to its in vitro oxidisability. The LDL composition, i.e. lipids, antioxidants, fatty acids, plasmenylcholines, and baseline level of conjugated dienes (CD) of 94 well-controlled and normolipidaemic type 2 diabetic patients was measured. Two oxidisability indices were determined: the lag time, reflecting the resistance of LDL to copper-induced oxidation, and the amount of conjugated dienes formed during oxidation of LDL. The lag time was not related to the total level of saturated, monounsaturated, and polyunsaturated fatty acids, but a strong inverse relationship was observed with fatty acids with three or more double bonds (r = -0.56, p < 0.001). In addition, an inverse relation was observed between the lag time and LDL-plasmenylcholine (r = -0.35, p < 0.001). Although not related to lag time in univariate analysis, alpha-tocopherol was a significant determinant in multiple regression analysis. A multiple linear regression model with LDL polyunsaturated fatty acids with three or more double bonds, alpha-tocopherol, monounsaturated fatty acids, and plasmenylcholines as determinants explained 47% of the variation in lag time. CD production was negatively correlated to oleic acid and positively to linoleic acid (r = -0.45 and r = 0.73, respectively; p<0.001). Fatty acids with three or more double bonds were the most important predictor of LDL lag time, whereas oleic acid and linoleic acid were major determinants of the amount of CD formed during oxidation of LDL.