Quercetin attenuates ethanol-derived microsomal oxidative stress: Implication of haem oxygenase-1 induction

As the main source of reactive oxygen species (ROS), hepatic microsomes are susceptible to ROS attack, especially upon CYP2E1 activation by ethanol. The objective of this study was to evaluate the hepatoprotective effect of quercetin, by inducing haem oxygenase-1 (HO-1), on ethanol-induced microsomal oxidative stress. Chronic alcohol administration to adult rats (4.0 g/kg for 90 days) resulted in microsomal redox disturbance and liver dysfunction, accompanying CYP2E1 upregulation and HO-1 downregulation of both protein expression and enzymatic activity. Quercetin (100 mg/kg) induced HO-1, which was not completely suppressed by ethanol. Moreover, quercetin pretreatment to ethanol-fed rats lowered CYP2E1 induction, partially normalised ethanol-overwhelmed microsomal antioxidative system, decreased ROS level and lipid peroxidation, and alleviated the leakage of transaminases. Given the beneficial effect of HO-1, its induction by quercetin may contribute to the protective role against CYP2E1-mediated oxidative stress on hepatic microsomes.     

Hydroxytyrosyl acetate contributes to the protective effects against oxidative stress of virgin olive oil

The effects of virgin olive oil phenols, hydroxytyrosyl acetate (HTy-Ac) and hydroxytyrosol (HTy), on cell integrity and steady-state values of cellular redox status were assessed in HepG2 cells, as well as their potential protective effects against oxidative stress induced by tert-butyl hydroperoxide (t-BOOH). Direct treatment for 20 h with 0.5-10 μM HTy or HTy-Ac reduced ROS generation and increased glutathione peroxidase activity at the higher concentrations. Furthermore, after t-BOOH exposure, pretreatment with HTy-Ac and HTy for 2 or 20 h counteracted cell viability damage from 1 μM, counterbalanced reduced glutathione levels from 0.5 μM, protected against lipid peroxidation from 0.5 μM, decreased ROS generation from 1 μM as well as antioxidant enzyme activities from 1 μM. All these changes were statistically significant.HTy-Ac presents antioxidative stress protective effects at physiological concentrations similar to or even slightly higher than that of HTy, thus contributing to the protective role of virgin olive oil.     

Isolation and purification of a novel antioxidant protein from the water extract of Sundakai (Solanum torvum) seeds

Reactive oxygen species (ROS) at physiological concentrations may be required for normal cell function. Excessive production of ROS can be detrimental to cells, because ROS can cause oxidative damage to lipids, proteins, and DNA. Herein, we describe the isolation and purification of a novel antioxidant protein the water extract of dried, powdered Sundakai (Solanum torvum [Solanaceae]) seeds. Sundakai belongs to the Solanaceae family, a small shrub, which is distributed widely in India, Malaya, China, Phillipines and tropical America. Fifty percent of ammonium sulphate-precipitated crude water extract was fractionated on a Sephadex G100 column, which yielded two peaks, PI and PII. Peaks PI and PII inhibited lipid peroxidation up to 40% and 89%, respectively in linolenic acid micelles. Rechromatographing of peak PII on Sephadex G100 yielded a single peak, indicating the homogeneity of the purified protein. SDS–PAGE analysis indicated the molecular weight of the purified protein to be ∼28 kDa. The purified protein, at 0.8 μM, inhibited deoxyribose degradation induced by generation of hydroxyl radicals by 90% and scavenged DPPH (1,1-diphenyl-2-picrylhydrazyl) radicals by 76%. The reducing power and chelating power of the purified protein, at 0.8 μM, were found to be 72% and 85%, respectively. The protein, at 0.8 μM, also offered significant protection to calf thymus DNA damage induced by H₂O₂ (1 mM). Therefore, the present study demonstrates, for the first time, a novel protein from the water extract of Sundakai seeds as an excellent antioxidant.     

姜黄素抑制Cu~(2+)诱导的转APP_(695)基因SH-SY5Y细胞氧化损伤和凋亡作用

目的:研究Cu~(2+)诱导的转β-淀粉样前体蛋白(amyloid-β precursor protein,APP)基因SH-SY5Y(SHSY5Y-APP_(695))细胞氧化损伤、凋亡及姜黄素的抑制作用。方法:在姜黄素预保护和无姜黄素预保护条件下,50 μmol/L Cu~(2+)处理细胞24 h,测定细胞存活率、胞外乳酸脱氢酶(lactate dehydrogenase,LDH)水平、胞内活性氧(reactive oxygen,ROS)水平、线粒体膜电位、半胱氨酸天冬氨酸蛋白酶(caspase)-3、caspase-8和caspase-9活力,蛋白免疫印迹法测定核转录NF-E2相关因子2(NF-E2-related factor 2,Nrf2)Ser40位点磷酸化(p Ser40-Nrf2)水平以及血红素加氧酶(heme oxygenase,HO)-1蛋白表达水平。结果:与空白对照相比,Cu~(2+)损伤组细胞存活率降低,胞内ROS水平和胞外LDH活性升高,线粒体膜电位下降,caspase-9和caspase-3活性明显升高,p Ser40-Nrf2和HO-1含量增加,而姜黄素保护组细胞存活率升高,胞内ROS水平和LDH释放水平降低,线粒体膜电位恢复升高,caspase-9和caspase-3活性明显降低,p Ser40-Nrf2和HO-1表达量减少。结论:姜黄素在一定程度上减弱了Cu~(2+)诱导的氧化损伤和细胞凋亡作用。     

Diallyl disulphide depletes glutathione in Candida albicans: oxidative stress-mediated cell death studied by two-photon microscopy

Using two-photon scanning laser microscopy, we investigated the effect of an Allium sativum (garlic) constituent, diallyl disulphide (DADS), on key physiological functions of the opportunistic pathogen Candida albicans. A short 30 min exposure to 0.5 mM DADS followed by removal induced 70% cell death (50% necrotic, 20% apoptotic) within 2 h, increasing to 75% after 4 h. The early intracellular events associated with DADS-induced cell death were monitored with two-photon fluorescence microscopy to track mitochondrial membrane potential (Deltapsi(m)), reactive oxygen species (ROS) and NADH or reduced glutathione (GSH) under aerobic conditions. DADS treatment decreased intracellular GSH and elevated intracellular ROS levels. Additionally, DADS induced a marked decrease of Deltapsi(m) and lowered respiration in cell suspensions and isolated mitochondria. In vitro kinetic experiments in cell-free extracts suggest that glutathione-S-transferase (GST) is one of the intracellular targets of DADS. Additional targets were also identified, including inhibition of a site or sites between complexes II-IV in the electron transport chain, as well as the mitochondrial ATP-synthase. The results indicate that DADS is an effective antifungal agent able to trigger cell death in Candida, most probably by eliciting oxidative stress as a consequence of thiol depletion and impaired mitochondrial function.     

Antioxidant effect derived from bioaccessible fractions of fruit beverages against H2O2-induced oxidative stress in Caco-2 cells

This work evaluates the effect of bioaccessible fractions from fruit beverages against oxidative stress (OS) in Caco-2 cells. A fruit beverage (grape + orange + apricot) (with/without milk and/or iron/zinc) was subjected to in vitro gastrointestinal digestion, and bioaccessible fractions were incubated with Caco-2 cell cultures. Following preincubation, OS was induced with 5 mM H₂O₂. Intracellular reactive oxygen species (ROS), mitochondrial potential (Δψ_m), mitochondrial metabolism (MTT test), intracellular reduced glutathione (GSH) and superoxide dismutase activity (SOD) were measured. The data evidenced viable cultures with increased mitochondrial metabolism and GSH-Rd activities, without alteration in SOD activity. Accordingly, more preserved mitochondrial integrity was also evidenced, allowing the action of antioxidant systems in preincubated cultures. Based on these data, we can conclude that a cytoprotective effect is derived from bioaccessible fractions of fruit beverages, though this effect failed to prevent intracellular ROS accumulation in Caco-2 cell cultures exposed to 5 mM H₂O₂.     

沙棘花青素提取物对双氧水诱导的H1299细胞损伤的保护作用及Nrf2/HO-1通路的影响

目的：研究沙棘花青素提取物（The anthocyanidin extract of Hippophae rhamnoides L.,HRAE）对双氧水诱导H1299细胞氧化损伤的保护作用及其机制。方法：利用双氧水诱导H1299细胞氧化损伤模型,采用MTT法检测HRAE对细胞活力的影响,采用荧光探针DCFH-DA检测细胞内活性氧（Reactive oxygen species,ROS）的含量;采用试剂盒分别测定超氧化物歧化酶（Superoxide dismutase,SOD）、过氧化氢酶（Catalase,CAT）、谷胱甘肽过氧化物酶（Glutathion peroxidase,GSH-Px）及丙二醛（Malondialdehyde,MDA）的含量;采用Western Blot法检测血红素氧合酶-1(Heme oxygenase-1,HO-1)、醌氧化还原酶-1(NAD(P)H quinine oxidoreductase 1,NQO1)、Kelch样环氧氯丙烷相关蛋白1(Kelch like ECH-associated protein 1,Keap1)和核转录因子E2相关因子（Nu...     

Protective effect of two edible mushrooms against oxidative cell damage and their phenolic composition

The aim of the study was to investigate phenolic composition, antioxidative, protective and cytotoxic effects of Pleurotus eryngii and Auricularia auricula-judae. Analysis of phenolic compounds in these edible mushrooms species has been carried out by high-performance liquid chromatography (HPLC). Protective effect of these mushrooms on H₂O₂ induced oxidative cell damage was determined by using MTT (3-(4,5-Dimethylthiazole-2-yl)-2,5-diphenyltetrazolium bromide, a yellow tetrazole) assay. Antioxidant activities of the mushrooms extracts were evaluated by using complementary in vitro assays. In addition, the measurement of total antioxidant compounds in the extracts was carried out. All the extracts exhibited protective effect against H₂O₂ induced oxidative cell damage but the highest activity was observed for A. auricula-judae aqueous extract (89.5 ± 1.8% cell viability at 0.1 mg/ml). P. eryngii methanolic extract showed the highest ferrous iron chelating ability (IC₅₀ = 0.42 ± 0.03 mg/ml). A. auricula-judae extracts (at concentration of 0.025–0.100 mg/ml) were not toxic to baby hamster kidney fibroblast cell line (BHK 21). These results suggest that these mushrooms may be used as a potential source of natural antioxidants for food supplementation or in the development of nutraceuticals.     

Carnosic acid protects against ROS/RNS-induced protein damage and upregulates HO-1 expression in RAW264.7 macrophages

The overproduction of reactive oxygen species (ROS) and reactive nitrogen species (RNS) is a deleterious process that can be an important mediator of damage to cell structures including protein, lipid and DNA, and consequently leads to various disease states and senescence. In the present study, the protective effects of carnosic acid (CA) on ROS/RNS-induced protein damages were examined. CA dose-dependently decreased the fragmentation, carbonylation, and nitration of bovine serum albumin (BSA) in AAPH and Hemin/nitrite/H₂O₂ incubation systems, respectively. Moreover, CA effectively attenuated protein carbonylations in the radical-treated RAW264.7 cells. Furthermore, after pretreatments of RAW264.7 macrophages with CA, the generation of ROS and NO induced by AAPH and H₂O₂ or LPS were significantly suppressed, and heme oxygenase-1 (HO-1) protein expression was time- and dose-dependently upregulated. Hence, our results indicated that CA might be beneficial for cellular proteins in oxidative stress or inflammation conditions by alleviating ROS/RNS generation and inducing the expression of antioxidant enzyme HO-1.     

Determination of selenium in garlic (Allium sativum) and onion (Allium cepa) by electro thermal atomic absorption spectrometry

A separation/enrichment procedure has been developed for the determination of selenium in garlic and onion samples by electrothermal atomic absorption spectrometry (ET-AAS) as a slurry sampling after preconcentration with 3,3-diaminobenzidine (DAB) reagent on the activated carbon. The influences of pH, time, amount of carbon and complexing reagent were outlined. The effect of acids used in the digestion of samples was also studied and compared. Selenium level was found to be 0.024 μg g⁻¹ for onion (n = 5; LOD – 0.5 μg L⁻¹; LOQ – 1.7 μg L⁻¹) and 0.015 μg g⁻¹ for garlic (n = 5; LOD – 1.3 μg L⁻¹; LOQ – 3.3 μg L⁻¹). Three different samples of garlic were analyzed by k₀-instrumental neutron activation analysis (k₀-INAA) at the Jozef Stefan Institute (JSI). The data obtained by k₀-INAA show that the content of selenium overlapped the results obtained by ET-AAS.     

Acteoside inhibits irradiation-mediated decreases in the viability and DNA synthesis of MC3T3-E1 cells

Therapeutic irradiation can cause bone loss, whereas antioxidant supplementation is considered to attenuate irradiation-mediated damages. This study examined whether or not acteoside inhibits irradiation-mediated changes in viability and proliferation of MC3T3-E1 cells. X-ray radiation at >4 Gy not only decreased cell viability and DNA synthesis in the cells, but also increased intracellular levels of reactive oxygen species (ROS) and phosphorylated p66^Shc protein. Irradiation at 8Gy also decreased intracellular levels of reduced glutathione (GSH) and induced G₁ phase arrest of cell cycle progression with the attendant increase of p21 induction. Pretreatment with acteoside inhibited the irradiation-mediated decreases in viability and DNA synthesis by restoring the radiation-mediated changes in the levels of ROS, GSH, p21, and p-p66^Shc to the untreated control levels. These inhibitory activities of acteoside were greater than that of a synthetic antioxidant compound or N-acetyl cysteine did. Collectively, acteoside treatment may prevent irradiation-induced oxidative damages to osteoblasts.     

Dietary inclusion of diallyl disulfide, yucca powder, calcium fumarate, an extruded linseed product, or medium-chain fatty acids does not affect methane production in lactating dairy cows

Two similar experiments were conducted to assess the effect of diallyl disulfide (DADS), yucca powder (YP), calcium fumarate (CAFU), an extruded linseed product (UNSAT), or a mixture of capric and caprylic acid (MCFA) on methane production, energy balance, and dairy cow performance. In experiment 1, a control diet (CON1) and diets supplemented with 56 mg of DADS/kg of dry matter (DM), 3 g of YP/kg of DM, or 25 g of CAFU/kg of DM were evaluated. In experiment 2, an inert saturated fat source in the control diet (CON2) was exchanged isolipidically for an extruded linseed source (100 g/kg of DM; UNSAT) or a mixture of C8:0 and C10:0 (MCFA; 20.3 g/kg of DM). In experiment 2, a higher inclusion level of DADS (200 mg/kg of DM) was also tested. Both experiments were conducted using 40 lactating Holstein-Friesian dairy cows. Cows were adapted to the diet for 12 d and were subsequently kept in respiration chambers for 5 d to evaluate methane production, diet digestibility, energy balance, and animal performance. Feed intake was restricted to avoid confounding effects of possible differences in ad libitum feed intake on methane production. Feed intake was, on average, 17.5 and 16.6 kg of DM/d in experiments 1 and 2, respectively. None of the additives reduced methane production in vivo. Methane production in experiment 1 was 450, 453, 446, and 423 g/d for CON1 and the diets supplemented with DADS, YP, and CAFU, respectively. In experiment 2, methane production was 371, 394, 388, and 386 g/d for CON2 and the diets supplemented with UNSAT, MCFA, and DADS, respectively. No effects of the additives on energy balance or neutral detergent fiber digestibility were observed. The addition of MCFA increased milk fat content (5.38% vs. 4.82% for control) and fat digestibility (78.5% vs. 59.8% for control), but did not affect milk yield or other milk components. The other products did not affect milk yield or composition. Results from these experiments emphasize the need to confirm methane reductions observed in vitro with in vivo data.     

Antioxidant and antimicrobial effects of garlic in chicken sausage

The antioxidant and antimicrobial effects of equivalent concentrations of fresh garlic (FG), garlic powder (GP) and garlic oil (GO) were investigated against lipid oxidation and microbial growth in raw chicken sausage during storage at 3°C. The antioxidant activities were compared to that of a standard synthetic antioxidant; butylated hydroxyanisole (BHA). The initial mean levels of thiobarbituric acid (TBA) value and peroxide value (POV) were 0.140 and 6.32, respectively. However after 21 days of storage, TBA and POV ranged from 0.151 to 4.92, respectively, in FG (50 g/kg) formulated samples to 0.214 and 8.64, respectively, in GO (0.06 g/kg) formulation. Addition of either garlic or BHA (0.1 g/kg) significantly delayed lipid oxidation when compared with control. The antioxidant activities of the various materials added followed the order FG>GP>BHA>GO. On the other hand, the initial aerobic plate count (APC) in the samples was 4.41 log₁₀ CFU/g. Addition of FG (30 g/kg) or GP (9 g/kg) significantly reduced the APC and, subsequently, the shelf-life of the product was extended to 21 days. However, addition of GO or BHA resulted in no significant difference in APC when compared with control. Sensory analysis indicated that FG had a significant stronger flavor than the other sausage formulations. The results suggest that fresh garlic and garlic powder, through their combined antioxidant and antimicrobial effects, are potentially useful in preserving meat products.     

Dietary spices protect against hydrogen peroxide-induced DNA damage and inhibit nicotine-induced cancer cell migration

Spices are rich sources of antioxidants due to the presence of phenols and flavonoids. In this study, the DNA protecting activity and inhibition of nicotine-induced cancer cell migration of 9 spices were analysed. Murine fibroblasts (3T3-L1) and human breast cancer (MCF-7) cells were pre-treated with spice extracts and then exposed to H₂O₂ and nicotine. The comet assay was used to analyse the DNA damage. Among the 9 spices, ginger, at 50 μg/ml protected against 68% of DNA damage in 3T3-L1 cells. Caraway, cumin and fennel showed statistically significant (p < 0.05) DNA protecting activity. Treatment of MCF-7 cells with nicotine induced cell migration, whereas pre-treatment with spices reduced this migration. Pepper, long pepper and ginger exhibited a high rate of inhibition of cell migration. The results of this study prove that spices protect DNA and inhibit cancer cell migration.     

Short communication: Canola meal as a substitute for cottonseed meal in diet of midlactation Holsteins

The growing demand by humans for monounsaturated vegetable oils has provided canola meal (CM) for use in dairy diets because it possesses an excellent nitrogen profile for rumen microbes. Six midlactation cows were used in a replicated 3 × 3 Latin square design with 3 periods of 20 d each. Treatments included diets with 1) CM, 2) 50% CM + 50% cottonseed meal (CSM), and 3) CSM. Total crude protein (CP), nonprotein nitrogen, and rapidly degradable true protein (% of CP) were greater in CM than in CSM. The neutral and acid detergent fibers, slowly degradable true protein, and unavailable CP were lower in CM than in CSM. Daily feeding of 3.4 kg of CM instead of 5.6 kg of CSM enhanced milk percentage of protein and SNF, and improved total tract digestibility of dry matter and CP. Therefore, CM offers an economical substitute for CSM in midlactation diets when commercial access, cost, and quality of CSM are variable.     

Inhibitory effect of wheat bran feruloyl oligosaccharides on oxidative DNA damage in human lymphocytes

The present work assessed the protective effect of feruloyl oligosaccharides (FOs), the ferulic acid ester of oligosaccharides from wheat bran, against oxidative DNA damage in normal human peripheral blood lymphocytes induced by hydrogen peroxide (H₂O₂). The DNA damage was measured by using the single cell gel electrophoresis assay (comet assay). Lymphocytes were subjected to DNA damage by exposure to a range of H₂O₂ concentrations (10–200 μmol/l). H₂O₂, at a concentration of 200 μmol/l, resulted in nearly all cells being highly damaged. FOs showed no cytotoxicity and genotoxicity to normal human lymphocytes at the tested concentrations (10–500 μmol/l). In addition, DNA damage in human lymphocytes induced by 100 μmol/l H₂O₂ was inhibited by FOs in a concentration-dependent fashion with 91.1% inhibition of lymphocyte DNA damage at 500 μmol/l as compared with the control. The results suggest that water-soluble FOs from wheat bran are able to enhance the ability of human lymphocytes to resist H₂O₂ induced oxidative damage.     

Cytoprotective effect of a bilberry extract against oxidative damage of rat hepatocytes

The effect of a bilberry extract (BE, 25% anthocyanins) against oxidative damage in primary cultures of rat hepatocytes, induced by tert-butyl hydroperoxide and allyl alcohol, was investigated. BE displayed cytoprotective effects at 100 and 500 μg/ml in the MTT viability test. It protected the cells against lactate dehydrogenase leakage and lipoperoxidation products formation. Maximum protection (58%) was noted using 500 μg/ml of BE and intoxication by allyl alcohol. The observed cytoprotective effect is probably due to the antioxidant properties of its constituents, mainly anthocyanins. BE scavenged DPPH (IC₅₀ 3.99 ± 0.14 μg/ml) and enzymatically generated superoxide radical with an activity equivalent to 108 ± 7.2 U of superoxide dismutase per mg of extract. Our results support the use of bilberry and bilberry extracts in functional foods and food supplements designed for the prevention of chronic diseases associated with oxidative stress.     

Napiergrass (Pennisetum purpureum S.) protects oxidative damage of biomolecules and modulates antioxidant enzyme activity

The effects of water extract of napiergrass (Pennisetum purpureum S.) (WEN) on oxidative damage of biomolecules and modulation of antioxidant enzyme activity were investigated. The results showed that WEN displayed marked free radical scavenging, reducing power, as well as ferrous ions chelating effects. WEN has a dose-dependent response for protective action on oxidation of phospholipid, deoxyribose and low-density lipoprotein (LDL) in the range of 0–0.5 mg/ml, indicating that WEN had in vitro protective action on oxidative damage of biomolecules. Oxidative stress induced by H₂O₂ significantly decreased the viability of BNL cells. However, addition of WEN in the medium protected cells from H₂O₂-induced cytotoxicity. Furthermore, treatment of cells with WEN in the range of 0–0.2 mg/ml displayed protective effect from H₂O₂ induced oxidation in a concentration dependent manner. With respect to the effect of WEN on antioxidant enzymes, the results showed the WEN at 0.2 mg/ml enhanced activities of glutathione peroxidase (GPX), glutathione reductase (GR), glutathione transferase (GST) and catalase (CAT) in BNL cells by 2.93-, 35.8-, 4.23-, and 2.78-fold, respectively, compared to the control; WEN increased the GSH content by 3.2-fold, implying that WEN may up-regulate the levels of GSH and antioxidant enzymes in BNL cells. WEN scavenged NO generated by a NO donor, sodium nitroprusside (SNP) and suppressed NO production in lipopolysaccharide (LPS)-activated macrophage RAW 264.7 cells. The determination of ascorbic acid and total anthocyanins as well as HPLC analysis revealed that ascorbic acid, rutin, epicatechin, anthocyanins, p-coumaric acid, quercetin and catechin were present in WEN, which function as in vitro antioxidants by virtue of their ability to scavenge ROS and RNS. Overall, the results obtained showed that WEN is rich in antioxidant components and they can serve as an excellent potential for use as a natural phytochemicals source.     

Introducing of green garlic plant as a new source of allicin

The presence of allicin in green garlic plant extracts was investigated. Allicin in aqueous extracts from green garlic leaf, shoot and young bulbs were determined by HPLC. Allicin was present at highest level in extracts from whole green garlic plant at 0.48 ± 0.01 mg/mL, followed by that in shoot and leaf extracts at 0.44 ± 0.00 and 0.26 ± 0.01 mg/mL, respectively. The results obtained in this study offer green garlic as a new source of allicin, as green garlic plant is used as a favourite vegetable in many countries.     

Characterisation of antioxidant and antiproliferative acidic polysaccharides from Chinese wolfberry fruits

Wolfberry fruit polysaccharides (WFPs) were isolated by hot-water extraction and ethanol precipitation. With HPLC analysis, WFPs were for the first time identified as acidic polysaccharides with galacturonic acid being the main component monosaccharide (24.9%), followed by galactose (21.3%), arabinose (18.5%), and glucose (15.9%), accounting for up to 80.6% of the molar percentage. WFPs exhibited a high antioxidant activity and a dose-dependent antiproliferative effect, with IC₅₀ values of 134.9, 70.1 and 138.4 μg/mL against A549, MCF-7 and LoVo cancer cells after 48 h of incubation as estimated by MTT assay, respectively. Flow cytometry analysis showed that WFPs exerted a stimulatory effect on apoptosis of MCF-7 cells, and induced the cell-cycle arrest at the G0/G1 phase, with the observation of intracellular ROS production and DNA damage. The present study demonstrated that these polysaccharides might have the potential to provide significant natural defence against human cancer.