Chemical characterization and chemo-protective activity of cranberry phenolic powders in a model cell culture. Response of the antioxidant defenses and regulation of signaling pathways

Oxidative stress and reactive oxygen species (ROS)-mediated cell damage are implicated in various chronic pathologies. Emerging studies show that polyphenols may act by increasing endogenous antioxidant defense potential. Cranberry has one of the highest polyphenol content among commonly consumed fruits. In this study, the hepato-protective activity of a cranberry juice (CJ) and cranberry extract (CE) powders against oxidative stress was screened using HepG2 cells, looking at ROS production, intracellular non-enzymatic and enzymatic antioxidant defenses by reduced glutathione concentration (GSH), glutathione peroxidase (GPx) and glutathione reductase (GR) activity and lipid peroxidation biomarker malondialdehyde (MDA). Involvement of major protein kinase signaling pathways was also evaluated. Both powders in basal conditions did not affect cell viability but decreased ROS production and increased GPx activity, conditions that may place the cells in favorable conditions against oxidative stress. Powder pre-treatment of HepG2 cells for 20 h significantly reduced cell damage induced by 400 μM tert-butylhydroperoxide (t-BOOH) for 2 h. Both powders (5–50 μg/ml) reduced t-BOOH-induced increase of MDA by 20% (CJ) and 25% (CE), and significantly reduced over-activated GPx and GR. CE, with a significantly higher amount of polyphenols than CJ, prevented a reduction in GSH and significantly reduced ROS production. CJ reversed the t-BOOH-induced increase in phospho-c-Jun N-terminal kinase. This study demonstrates that cranberry polyphenols may help protect liver cells against oxidative insult by modulating GSH concentration, ROS and MDA generation, antioxidant enzyme activity and cell signaling pathways.     

Noni increased the systemic exposure of methotrexate in rats through inhibition on multi-drug resistance protein 2 (MRP 2) and breast cancer resistance protein (BCRP)

Methotrexate (MTX) is an important immunosuppressant with narrow therapeutic window. The transport of MTX was associated with multi-drug resistance proteins (MRPs) and breast cancer resistance protein (BCRP). Noni fruit is a phenolics rich health food. This study set out to investigate the effect of noni juice on MTX pharmacokinetics and the underlying mechanisms. Rats were administered MTX with and without single dose or the 7th dose of noni juice (6 mL/kg). The serum MTX concentrations were determined using a specific monoclonal fluorescence polarization immunoassay. The results revealed that single dose and multiple doses of noni juice significantly increased the AUC_0–t, C_max and MRT of MTX. A further inquiry into the underlying mechanism indicated that noni metabolites inhibited the activities of MRP 2 and BCRP. In conclusion, concomitant intake of noni juice significantly increased the systemic exposure of MTX through inhibition on MRP 2 and BCRP.     

Phenolic extracts of brewers’ spent grain (BSG) as functional ingredients – Assessment of their DNA protective effect against oxidant-induced DNA single strand breaks in U937 cells

Brewers’ spent grain (BSG), a by-product of the brewing industry, contains high amounts of phenolic acids, which have antioxidant effects. The present study examined the ability of BSG extracts to protect against the genotoxic effects of oxidants, hydrogen peroxide (H₂O₂), 3-morpholinosydnonimine hydrochloride (SIN-1), 4-nitroquinoline 1-oxide (4-NQO) and tert-butylhydroperoxide (t-BOOH) in U937 cells. Four pale (P1–P4) and four black (B1–B4) BSG extracts were investigated. U937 cells were pre-incubated with BSG extracts, exposed to the oxidants and the DNA damage was measured by the Comet assay. The black BSG extracts (B1–B4) significantly protected against H₂O₂-induced DNA damage. Extract B2, which had the highest phenol content, provided the greatest protection. Extracts P2, B2, B3 and B4 provided significant protection against SIN-1-induced DNA damage. None of the extracts protected against DNA damage induced by t-BOOH and 4-NQO. The DNA protective effects of the BSG phenolic extracts may be related to iron chelation.     

Dihydrocaffeic acid,a major microbial metabolite of chlorogenic acids,shows similar protective effect than a yerba mate phenolic extract against oxidative stress in HepG2 cells

The hepatoprotective effect of a yerba mate phenolic extract (YMPE), rich in chlorogenic acids, and its main circulating metabolites dihydrocaffeic (DHCA) and dihydroferulic (DHFA) acids were assessed in human hepatoma HepG2 cells subjected to oxidative damage induced by tert-butylhydroperoxide (t-BOOH). Direct treatment of HepG2 cells with realistic concentrations of YMPE (1, 10 and 50 μg/mL), DHCA or DHFA (0.2, 1, 10 μM) for 20 h was not cytotoxic and significantly decreased ROS generation. Pre-treatment with YMPE and DHCA prevented the cytotoxicity and macromolecular damage induced by t-BOOH. Moreover, decreased levels of reduced glutathione (GSH), and increased ROS levels and antioxidant enzyme activity induced by t-BOOH were dose-dependently recovered. DHFA only showed a slight protection against cell cytotoxicity, lipid oxidation and GSH depletion. In conclusion, YMPE and one of its major microbial metabolites, DHCA, confer significant protection against oxidative damage, adding evidences to the beneficial health effects associated with mate intake.     

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.     

Assessment of the ability of seaweed extracts to protect against hydrogen peroxide and tert-butyl hydroperoxide induced cellular damage in Caco-2 cells

The ability of brown seaweed extracts, Ascophyllum nodosum, Laminaria hyperborea, Pelvetia canaliculata, Fucus vesiculosus and Fucus serratus to protect against tert-butyl hydroperoxide (tert-BOOH) induced stress in Caco-2 cells was investigated. Oxidative stress was determined by measuring alteration in the enzymatic activity of catalase (CAT) and superoxide dismutases (SOD) and cellular levels of glutathione (GSH). L. hyperborea, P. canaliculata and F. serratus significantly protected against tert-BOOH induced SOD reduction but did not protect against the reduction in CAT activity or the increased cellular levels of GSH. The ability of F. serratus and F. vesiculosus to protect against H₂O₂ and tert-BOOH induced DNA damage was also assessed. The DNA protective effects of the two seaweed extracts was compared to those of three metal chelators; deferoxamine mesylate (DFO), 1,10-phenanthroline (o-phen) and 1,2-Bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid tetrakis (BAPTA-AM). F. serratus and F. vesiculosus significantly protected (P < 0.05) against H₂O₂ (50 μM) induced DNA damage but not tert-BOOH induced damage.     

Hepatoprotective effects of lycopene against carbon tetrachloride-induced acute liver injury in rats

Lycopene, the major carotenoid in tomatoes, is a known antioxidant that may lower oxidative stress biomarkers by a mechanism that is not fully elucidated. The intoxication of rats with carbon tetrachloride (CCl₄) resulted in significant histological hepatic degradation accompanied by a marked increase in reactive oxygen species (ROS) and in the number of apoptotic cells. The induced oxidative stress in turn results in a significant elevation of lipid peroxidation and H₂O₂ generation, together with a decrease in the concentration of reduced glutathione (GSH) and a significant reduction in activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and glutathione S transferase (GST). CCl₄-intoxicated rats, pre-treated with lycopene, showed strongly reduced cell damage and ROS generation. The level of markers for hepatic integrity in lycopene pre-treated rats was close to the controls in the absence of CCl₄ treatment, indicating the protective effect of lycopene pre-treatment. In the same way, lycopene pre-treated rats significantly increased SOD, CAT, GPx, GST activities and GSH level. In addition, we measured an increased lipoxygenase (LOX) activity in CCl₄-intoxicated rats. This activity was reduced in lycopene pre-treated rats to values close to those observed in the controls, suggesting a potential pharmacological application of this dietary carotenoid.     

Extraction and antioxidant property of polyhydroxylated naphthoquinone pigments from spines of purple sea urchin Strongylocentrotus nudus

Purple sea urchin (Strongylocentrotus nudus) spines were dissolved in HCl–ethanol-aqueous solution. The polyhydroxylated 1,4-naphthoquinone (PHNQ) pigments were condensed and purified by using macroporous resin in static adsorption mode. PHNQ in the extract were characterised rapidly by using an ultra-performance liquid chromatography (UPLC) coupled to hybrid quadrupole orthogonal acceleration time-of-flight mass spectrometer (Q-TOFMS). Six known compounds including spinochrome E, 2,7-dihydroxynaphthazarin, spinochrome B, spinochrome C, spinochrome A and echinochrome A, and two new compounds with molecular formula of aminopentahydroxynaphthoquinone and acetylaminotrihydroxynaphthoquinone were identified tentatively. The pigment extract was evaluated for antioxidant activity by using 1,1-diphenyl-2-picrylhydrazyl (DPPH) scavenging capacity assay, Fe²⁺ chelating assay, reducing power assay, lipid peroxidation inhibition assay and tertiary-butyl hydroperoxide (t-BOOH)-induced macrophages protection assay. In all testing methods, the extract showed excellent activity, indicating the PHNQ from S. nudus spines are potential sources of natural antioxidants.     

Antioxidant activity,anthocyanins, and phenolics of rabbiteye blueberry (Vaccinium ashei) fluid products as affected by fermentation

Frozen rabbiteye blueberries (Vaccinium ashei) were processed into juice (BJ), wines made without (BW1) or with (BW2) skin contact fermentation, and vinegars made from BW1 (JV), BW2 (WV) or blueberries (BV). Total phenolics, total anthocyanins, antioxidant activities (beta-carotene bleaching assay and ferric thiocyanate assay), and antiradical activity (DPPH radical-scavenging method) of these fluid products were determined. The differences in total anthocyanin contents of all blueberry products were significant. The BW2 had the highest content of anthocyanins and polyphenols and the highest beta-carotene bleaching activity and antiradical activity. Acetification decreased total anthocyanin content, total polyphenols and antioxidant activities. Correlations indicate that anthocyanins made significant contributions than did phenolics to antioxidant activities of products. The abilities of BJ, BW1 (wine from blueberry juice), and BW2 to inhibit linoleic acid peroxidation were high (∼95%). The abilities of vinegar products to inhibit linoleic acid peroxidation were low. The results indicate that skin-contact fermentation is a better method for obtaining higher antioxidant activity of blueberry products. Also, acetification significantly decreased anthocyanins and antioxidant activities.     

Thermal degradation kinetics of anthocyanins from Chinese red radish (Raphanus sativus L.) in various juice beverages

Thermal and storage stabilities of red radish anthocyanins (RRAs) in various juice beverages (apple, grape, peach, pear, pomegranate and lemon) were studied over temperature range 70–90 °C and 4–25 °C. RRAs degradation in all juice beverages followed first-order reaction kinetics. RRAs showed a much faster degradation rate during storage at room temperature (t _1/2 value ≤84.0 days) than did in refrigerated temperature (t _1/2 ≥value 130.9 days). The rate constant (k), E _a and Q ₁₀ values for RRAs in juice beverages varied from 1.33 to 0.33, 47.94 to 14.77 kJ mol^?1 and 1.16 to 1.89 at 70–90 °C. During heating, RRAs in peach and pomegranate showed higher stability than others at these temperatures. There was a positive correlation (R ² > 0.9128) between ascorbic acid content of juice beverages (8–36 mg/100 mg) and stability of RRAs at 70–90 °C. It was found that RRAs in apple and pear juice beverage were more stable than in other juice beverages.     

Effect of tea phenolics on iron uptake from different fortificants by Caco-2 cells

The in vitro effects of tea phenolics on Fe uptake from different fortificants (FeSO₄, FeCl₃, FeEDTA) by Caco-2 cells were compared. Cell cultures were exposed to catechin, tannic acid, green or black tea solutions, added within Fe-containing solution, or used to pre-treat cell cultures before Fe-exposure. Cell ferritin formation was used as a measure of Fe uptake. Reverse phase chromatography was used to identify specific phenolics in tea solutions, and the Fe-binding catechol and galloyl groups were determined spectrophotometrically. The results showed a positive effect of catechin on Fe uptake only from dissociable Fe sources, and a marked inhibitory effect of tannic acid regardless of the Fe source. Tea phenolics exhibit similar inhibitory patterns on Fe uptake from FeCl₃ and FeEDTA solutions; however, the Fe uptake from FeSO₄ solutions was significantly less affected. These data improve the understanding of interactions by which tea phenolics affect Fe uptake at the intestinal level.     

Antioxidant capacities of Gundelia tournefortii L. extracts and inhibition on glutathione-S-transferase activity

Gundelia tournefortii L. is an important food source and a well-known medicinal plant in Eastern Anatolia. Therapeutic effects of medicinal plants are known to be closely related to their antioxidant capacities. Antioxidant activities of G. tournefortii, both for the aerial parts and seeds, were investigated by using both 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging and lipid peroxidation inhibition methods. The seeds were found to have higher antioxidant potential than the aerial, with IC₅₀ values of 0.073 mg/mL for DPPH scavenging and 0.146 mg/mL for lipid peroxidation inhibition capacities. In addition, total phenolic contents of the Gundelia tournefortii L. extracts, especially the seed extracts correlates to its high antioxidant activity with 105.1 ± 8.7 μg gallic acid equivalents (GAEs) per mg of seed extract. Plant extracts with high phenolics content are known to have important effects on various enzymes, as well as glutathione-S-transferases, which are important detoxification enzymes in phase II systems with an important role in developing multi-drug resistance to chemotherapy in tumour cells. Consequently, the effects of G. tournefortii extracts on crude cytosolic glutathione-S-transferase was also studied and the seed extracts have shown effective inhibition of cytosolic GST activity, with an IC₅₀ of 97.51 μg/mL.     

Antioxidation activity of oil extracts prepared from various seeds

The anti-oxidative activity of ethanol and methanol extracts from various seed oils (perilla seed oil, camellia seed oil, pine seed oil, flaxseed oil, and olive oil) was investigated using DPPH free radical scavenging activity (DPPH-RSA), hydroxyl radical scavenging activity (Hydroxyl-RSA), fluorescent development, browning level development, microsomal lipid peroxidation (MLP), and t-BOOH-induced cytotoxicity assays. Both browning and fluorescence level were observed higher value in methanol extract of camellia seed oil (CSO-ME). This meant that CSO-ME had more available antioxidant matters. Radical scavenging activity of CSO-ME was the highest among all seed oil extracts, and was 61.1% in the DPPH-RSA and 64.5% in the Hydroxyl-RSA test. Based on in vitro tests, all extract treatments significantly reduced both thiobarbituric acid-reactive substrate formation and lactate dehydrogenase release, indicating high antioxidant activity. These results suggest that ethanol and methanol extracts from seed oils can be a potentially good dietary lipid supplement source against oxidative stress.     

Decalepis hamiltonii roots boost antioxidant status of rat liver and brain

BACKGROUND: Roots of Decalepis hamiltonii are traditionally consumed as pickles and juice for their health benefits. We have earlier demonstrated the antioxidant property of the root extract and identified the constituent antioxidant molecules. RESULTS: This paper reports the effect of multiple‐dose (7, 15, and 30 days) treatment of Decalepis hamiltonii aqueous root extract (DHA) (50 and 100 mg kg^?1 body weight) on the antioxidant profile of rat liver and brain. Activities of the antioxidant enzymes superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione reductase (GR), and glutathione‐S‐transferase (GST) were increased and glutathione content was elevated in both liver and brain, apart from reduction in the basal level of lipid peroxidation. DHA induced stronger antioxidant boost in brain by increasing the activities of SOD, CAT, and GPx compared to liver. CONCLUSION: As failure to grapple with oxidative stress is an important factor in the etiology of several diseases, DHA's effects on improvement of antioxidant status could provide a scientific justification for the health‐promoting properties attributed to it. Copyright © 2009 Society of Chemical Industry     

Inactivation of Escherichia coli O157:H7, Salmonella typhimurium and Listeria monocytogenes in apple juice with gaseous ozone

This research was initiated to assess the efficacy of gaseous ozone for inactivation Escherichia coli O157:H7, Salmonella typhimurium and Listeria monocytogenes in apple juice. Juice samples with solids content of 18, 36, and 72 °Brix inoculated with a culture cocktail of three foodborne pathogens were treated with gaseous ozone at a flow rate of 3.0 L/min and an ozone generation rate of 0.10, 0.90, 3.51, and 5.57 g/h for 0.5, 1, 5, and 10 min, respectively. The inactivation kinetics of gaseous ozone on foodborne pathogens conformed to the Weibull model. The time required to achieve a 5 log reduction (t_5d) was estimated using the parameters of the Weibull model. The t_5d increased with increasing solids content of apple juice. The ozone generation rate did not impart a significant effect (p > 0.05) on t_5d. Gaseous ozone is effective at inactivating foodborne pathogens in apple juice but the efficacy is dependent on the solids content of the juice sample.     

Antioxidant activity of Nyctanthes arbor-tristis leaf extract

Nyctanthes arbor-tristis (Harsingar) leaf extracts are extensively used in Indian traditional medicine. The acetone-soluble fraction of its ethyl acetate extract showed impressive antioxidant activity as revealed by several in vitro experiments, e.g., DPPH, hydroxyl and superoxide radicals, as well as H₂O₂ scavenging assays. Moreover, its preventive capacity against Fe(II)-induced lipid peroxidation of liposomes and γ-ray-induced DNA damage also confirmed this. The strong reducing power and high phenolics and flavonoids contents could be responsible for the antioxidant activity.     

Application of nanoparticle-immobilized thermostable β-glucosidase for improving the sugarcane juice properties

β-glucosidases are among the key enzymes for juice and beverage industries. They are responsible for the release of aromatic compounds in fruits and fermentation products. In this study, β-glucosidase was isolated, purified, and characterized from an indigenously developed Bacillus subtilis mutant PS-5CM-UM3. It is a 56 kDa protein monomer (isoelectric point of 5.6) belonging to 1 glycosyl hydrolase family. The purified β-glucosidase was immobilized on SiO₂ nanoparticles (with 52% efficiency and 14.1% yield) to improve the thermostability and Michaelis constant (K_m) value of β-glucosidase from 0.9 to 1.1 mM. The immobilized enzyme showed improved storage stability and was reusable for up to 10 cycles with 70% residual activity. β-glucosidase treatment in sugarcane juice elevated the phenolics content with about 2.6 folds and 2.4 folds increase in p-hydroxy benzoic acid (PHBA) and gallic acid, respectively. The results show that recyclable immobilized enzyme system is a novel green approach for improving the sugarcane juice properties.Industrial relevanceIn this study, β-glucosidase originally isolated and purified from an indigenously developed Bacillus subtilis mutant was immobilized on SiO₂ nanoparticles. The immobilization has improved the thermostability, storage stability, and Michaelis constant (K_m) value of the β-glucosidase. The immobilized β-glucosidase is now reusable for 10 cycles with 70% residual activity. Further, β-glucosidase treatment in sugarcane juice elevated the phenolics content with about 2.6 folds and 2.4 folds increase in p-hydroxy benzoic acid (PHBA) and gallic acid, respectively. Hence, this study provides a green and sustainable approach for the food industry to efficiently enhance the juice properties.     

Preparation of a resveratrol-enriched grape juice based on ultraviolet C-treated berries

Grapes are rich in bioactive phenolics. However, most of them remain in the by-product after juice processing, and only a minor part pass to the juice. Our aim was to obtain a resveratrol-enriched white grape juice based on UVC-treated berries in combination with different conditions during juice production. Postharvest UVC treatment of berries enabled the further selective stilbenes enrichment of the juice, especially resveratrol. Macerating enzymes did not modify the phenolic profile but increased juice yield up to 30%, keeping its phenolic concentration, and thus reducing the phenolics remaining in the by-product. Maceration with Na₂S₂O₅ was critical to increase the phenolic content in the juice whereas β-cyclodextrin only tended to increase the stilbenes content. Optimum conditions for juice production (maceration for 2 h at 45 °C with 0.2% Na₂S₂O₅) using UVC-treated grape berries significantly increased stilbenes concentration (up to 35-fold over the control), without affecting the sensory properties of the juice. The phenolic (resveratrol)-enriched grape juice obtained here could be an ideal alternative to wine in the search of grape-derived health benefits.Industrial relevanceThe aim of this study is to obtain a resveratrol-enriched grape juice by combining postharvest UV-C treatment of grape berries with several technological approaches in the juice processing. Results obtained indicate that maceration for 2 h at 45 °C with 0.2% Na₂S₂O₅ using UVC-treated grape berries significantly increased phenolic concentration, especially stilbenes (up to 35-fold over the control), without affecting the sensory properties of the juice. The resveratrol-enriched juice has an added-value compared to the conventional ones and could exert more health benefits due to its improved phenolic profile.     

The role of two putative nitroreductases,Frm2p and Hbn1p,in the oxidative stress response in Saccharomyces cerevisiae

The nitroreductase family is comprised of a group of FMN‐ or FAD‐dependent enzymes that are able to metabolize nitrosubstituted compounds using the reducing power of NAD(P)H. These nitroreductases can be found in bacterial species and, to a lesser extent, in eukaryotes. There is little information on the biochemical functions of nitroreductases. Some studies suggest their possible involvement in the oxidative stress response. In the yeast Saccharomyces cerevisiae, two nitroreductase proteins, Frm2p and Hbn1p, have been described. While Frm2p appears to act in the lipid signalling pathway, the function of Hbn1p is completely unknown. In order to elucidate the functions of Frm2p and Hbn1p, we evaluated the sensitivity of yeast strains, proficient and deficient in both oxidative stress proteins, for respiratory competence, antioxidant‐enzyme activities, intracellular reactive oxygen species (ROS) production and lipid peroxidation. We found reduced basal activity of superoxide dismutase (SOD), ROS production, lipid peroxidation and petite induction and higher sensitivity to 4‐nitroquinoline‐oxide (4‐NQO) and N‐nitrosodiethylamine (NDEA), as well as higher basal activity of catalase (CAT) and glutathione peroxidase (GPx) and reduced glutathione (GSH) content in the single and double mutant strains frm2Δ and frm2Δ hbn1Δ. These strains exhibited less ROS accumulation and lipid peroxidation when exposed to peroxides, H₂O₂ and t‐BOOH. In summary, the Frm1p and Hbn1p nitroreductases influence the response to oxidative stress in S. cerevisae yeast by modulating the GSH contents and antioxidant enzymatic activities, such as SOD, CAT and GPx. Copyright © 2009 John Wiley & Sons, Ltd.