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Thearubigins rich black tea fraction reveals strong antioxidant activity
Abstract
Background: Black tea contains two major polyphenols namely Theaflavins (TFs) and Thearubigins (TRs). Several reports are available on antioxidant properties of TFs. However, very little information is known about TRs. Aim: To investigate in vitro antioxidant properties of not well-known TR rich fractions in comparison with TF rich fraction. Materials and Methods: TR and TF rich fractions were checked for in vitro antioxidant activities by using standard biochemical radical scavenging assays and pulse radiolysis. These fractions were also used to evaluate the protection conferred against induced oxidative damage to biomolecules (lipids, proteins and DNA) in rat liver mitochondria. Statistical Analysis: The one-way analysis of variance test associated with the Tukey′s test was used to determine the statistical significance of the differences among experimental groups. All the statistical analyses were done using SPSS 19.0 software. Results and Conclusions: Results from this study suggest that like TF rich fraction, TR rich fraction has good antioxidant properties, which correlate to the total phenolics and flavonoids content and provide significant protection against oxidative damage to biomolecules (lipids, proteins and DNA) in rat liver mitochondria. This is the first report about the in vitro antioxidant activity of TRs rich fractions from black tea.
... Thearubigins in low concentrations showed a higher ability to scavenge hydroxyl radicals than theaflavins and EGCG [30]. Teas rich in theaflavins and thearubigins have good scavenging activity by donating hydrogen atom and teas rich in thearubigins exhibits better ability to scavenge hydroxyl radical [31]. Dried tea leaves contain up to 36% of polyphenolic compounds responsible for all antioxidant properties; tea leaves are the most prominent compound group [27]. ...
... Thearubigins in low concentrations showed a higher ability to scavenge hydroxyl radicals than theaflavins and EGCG [30]. Teas rich in theaflavins and thearubigins have good scavenging activity by donating hydrogen atom and teas rich in thearubigins exhibits better ability to scavenge hydroxyl radical [31]. ...
In this paper, the possibility of obtaining uniaxially rotomolded composite parts was discussed. The used matrix was bio-based low-density polyethylene (bioLDPE) filled with black tea waste (BTW) to prevent the thermooxidation of samples during processing. In rotational molding technology, the material is held at an elevated temperature in a molten state for a relatively long time, which can result in polymer oxidation. The Fourier transform infrared spectroscopy (FTIR) shows that adding 10 wt% of black tea waste has not led to the formation of carbonyl compounds in polyethylene, and adding 5 wt% and above prevents the appearance of the C–O stretching band connected with degradation of LDPE. The rheological analysis proved the stabilizing effect of black tea waste on the polyethylene matrix. The same temperature conditions of rotational molding did not change the chemical composition of black tea but slightly influenced the antioxidant activity of methanolic extracts; the detected changes suggest degradation is a color change, and the total color change parameter (ΔE) is 25. The oxidation level of unstabilized polyethylene measured using the carbonyl index exceeds 1.5 and gradually decreases with the addition of BTW. The BTW filler did not influence the melting properties of bioLDPE; the melting and crystallization temperature remained stable. The addition of BTW deteriorates the composite mechanical performance, including Young modulus and tensile strength, compared to the neat bioLDPE.
... Thus, both concentration and Table 3). Tea extracts with high theaflavin content would produce a yellowish-brown color and gave an astringent or bitter taste [23], while tea extracts with high thearubigins content would produce a reddish-brown color with ashy and slight astringent or slightly bitter [30]. These results in line with research of Verloop et al. ...
The problem with ready-to-drink (RTD) black tea is a long manufacturing process and contains phenolic compounds and low antioxidant activity due to the enzymatic oxidation process. Viscozyme is an enzyme that degrades plant cell walls during extraction so that it releases active components from the cell wall, such as phenolic contents and antioxidant activity. Meanwhile, tyrosinase includes the polyphenol oxidase enzyme, which can take advantage of catechins conversion into theaflavins and thearubigins. This study aimed to determine the addition of viscozyme and tyrosinase combination to increase the content of phenolic compounds, antioxidant activity, and improve the appearance of black tea (color). The study used young leaves that had their enzymes inactivated was added with viscozyme to fresh tea leaves extract. The tea extract that had been given viscozyme was then added with tyrosinase, and the incubation was carried out. The addition of tyrosinase with higher concentrations (3,571 U/mL) and longer incubation times (40 min) decreased total phenolic contents, antioxidant activity using DPPH free radical scavenging method, increased tea cream formation (precipitation that forms as the tea cools), the compound of theaflavins, thearubigins, and TF:TR ratio (comparison of theaflavins and thearubigins contents) in tea extracts that had been treated with viscozyme. The combination of viscozyme and tyrosinase gave higher total phenolic compounds, antioxidant activity, theaflavins, thearubigins content, and TF:TR ratio compared to control and could prevent the formation of tea cream. The use of viscozyme 250 µL and tyrosinase 3,571 U/mL for 20 min on ready-to-drink black tea gave the best effects in this study. HIGHLIGHTS RTD tea has an extensive manufacturing procedure and contains phenolic components. However, its antioxidant efficacy is minimal because of the enzymatic oxidation process. Viscozyme is an enzyme that breaks down the cell walls of plants during extraction to liberate active components, such as phenolic contents and antioxidant activity, from the cell wall. Tyrosinase contains the polyphenol oxidase enzyme, which can utilise catechins to produce theaflavins and thearubigins. Increasing the quantity and incubation duration of tyrosinase resulted in a reduction in both total phenolic contents and antioxidant activity as measured by DPPH. However, the concentration of theaflavins and thearubigins was elevated. The utilisation of viscozyme and tyrosinase resulted in an enhanced antioxidant profile. GRAPHICAL ABSTRACT
... These antioxidant nutrients include vitamin C, four forms of vitamin E (α-tocopherol, β-tocopherol, γ-tocopherol, and δ-tocopherol), six carotenoids (α-carotene, β-carotene, β-cryptoxanthin, lutein, lycopene, and zeaxanthin), 29 flavonoids (cyanidin, delphinidin, malvidin, pelargonidin, peonidin, petunidin, (+)-catechin, (+)-gallocatechin, (−)-epicatechin, (−)-epigallocatechin, (−)-epicatechin 3-gallate, (−)-epigallocatechin 3-gallate, theaflavin, theaflavin 3-gallate, theaflavin 3′-gallate, theaflavin 3,3′-digallate, thearubigins, eriodictyol, hesperetin, naringenin, apigenin, luteolin, isorhamnetin, kaempferol, myricetin, quercetin, daidzein, genistein, and glycitein), and two PA (dimers and trimmers). As thearubigins have been reported to be as potent antioxidants as theaflavins [26], thearubigins were assigned the same antioxidant power as theaflavins. Each participant's average intake of individual antioxidants was estimated by multiplying the contents of individual antioxidants in foods by the daily consumption of each food item. ...
Purpose:
Although evidence strongly supports that antioxidant-rich diets reduce risk of chronic disease and mortality, findings from the previous studies on the effect of individual antioxidants on mortality have been inconsistent. The aim of this study was to assess the relationship between dietary total antioxidant capacity (TAC) and all-cause and disease-specific mortality in a representative sample of the US population.
Methods:
A total of 23,595 US adults aged 30 years and older in NHANES 1988-1994 and 1999-2004 were selected for this study. Dietary TAC was calculated from 1-day 24-h diet recall data at baseline and all-cause, cancer and cardiovascular disease (CVD) mortality was assessed through December 31, 2011.
Results:
During a mean follow-up of 13 years, deaths from all-cause, cancer and CVD were 7157, 1578, and 2155, respectively. Using cause-specific Cox proportional hazards models, inverse associations and linear trends were observed between dietary TAC and all-cause mortality [highest quartile (Q4) versus Q1 ref. HR 0.78; 95% CI 0.71-0.86], cancer mortality (Q4 versus Q1 ref. HR 0.75; 95% CI 0.60-0.93), and CVD mortality (Q4 versus Q1 ref. HR 0.83; 95% CI 0.69-0.99), respectively, after adjusting for age, sex, ethnicity, and total energy intake. The inverse association and linear trend still remained between dietary TAC and all-cause mortality (Q4 versus Q1 ref. HR 0.79; 95% CI 0.71-0.87) and CVD mortality (Q4 versus Q1 ref. HR 0.74; 95% CI 0.61-0.89) when further adjusted for relevant covariates.
Conclusions:
These findings support that antioxidant-rich diets are beneficial to reducing risk of death from all-cause and CVD.
... The antioxidant capacity of individual antioxidants was expressed as vitamin C equivalents (VCE) using the 2,2 1 -azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) assay [29]. These antioxidant nutrients include 29 flavonoids (cyanidin, delphinidin, malvidin, pelargonidin, peonidin, petunidin, (+)-catechin, (+)-gallocatechin, (´)-epicatechin, (´)-epigallocatechin, (´)-epicatechin 3-gallate, (´)-epigallocatechin 3-gallate, theaflavin, theaflavin 3-gallate, theaflavin 3 1 -gallate, theaflavin 3,3 1 -digallate, thearubigins, eriodictyol, hesperetin, naringenin, apigenin, luteolin, isorhamnetin, kaempferol, myricetin, quercetin, daidzein, genistein, and glycitein), three PA (monomers, dimers, and trimers), six carotenoids (α-carotene, β-carotene, β-cryptoxanthin, lutein, lycopene, and zeaxanthin), four vitamin E (α-tocopherol, β-tocopherol, γ-tocopherol, and δ-tocopherol) and vitamin C. As thearubigins have been reported to be potent antioxidants like theaflavins [30], thearubigins were assigned the same antioxidant power as theaflavins. The individual antioxidant intake of subjects was estimated by multiplying the contents of individual antioxidants by the daily consumption of each food and supplement. ...
Evidence from epidemiologic studies has shown that total antioxidant capacity (TAC) in the diet might be inversely associated with stroke, heart failure, and inflammatory biomarkers. However, studies on the association of TAC from both diet and supplements with cardiovascular disease (CVD) risk factors in the U.S. population are lacking. This cross-sectional population-based study aimed to investigate the association of TAC with both diet and supplements with CVD risk factors among 4039 U.S. adults in National Health and Nutrition Examination Survey (NHANES) 2007-2012. TAC from both food sources and dietary supplements was estimated from two 24-h dietary recalls using the NHANES supplement ingredient database, United States Department of Agriculture (USDA) proanthocyanidin, flavonoid, and isoflavone databases. Top contributors to TAC were tea, antioxidant supplements, vegetable mixture, orange juice, berries, and wine. Antioxidant supplement users had 1.6 times higher TAC than non-users. Greater TAC was associated with reduced triglycerides (TG) (-1.39% change; 95% CI = -2.56 to -0.21), TG to high-density lipoprotein cholesterol (HDL-C) ratio (-2.03% change; 95% CI = -3.45 to -0.60), HDL-C (0.65% change; 95% CI = 0.07 to 1.23), insulin (-1.37% change; 95% CI = -2.64 to -0.09), homeostasis model assessment of insulin resistance (HOMA-IR) (-1.57% change; 95% CI = -3.02 to -0.09) and C-reactive protein (CRP) (-0.83% change; 95% CI = -1.29 to -0.38) after adjusting for potential confounders. There was no significant association between TAC and waist circumference, BMI, blood pressure, low-density lipoprotein cholesterol (LDL-C), total cholesterol (TC), and fasting glucose. The findings of this study support the hypothesis that an antioxidant-rich diet and intake of supplements are beneficial to reduce CVD risk.
... Kuhnert (2010) as well as Kuhnert, Dairpoosh, Yassin, Golon, and Jaiswal (2013) highlighted similar observations in the chromatographic behaviour when describing the so-called "thearubigin hump" in the analysis of black tea. Although the molecular weight of the thearubigins ranges from 1000 to 40,000 (Yao et al. 2006), Sinha and Ghaskadbi (2013) recently presumed strong antioxidant properties of black tea thearubigins. Here again, although thearubigins are present in large amounts in black tea, the information on formation and structure of this heterogeneous polymer is very limited. ...
Bio‐fillers are functional substances that are increasingly added to polymer compositions due to their unique properties and sustainable nature. There is a lack of a review publication that comprehensively describes bio‐fillers from different natural origins in various types of polymer, although there are many publications focusing on a narrow range of bio‐filler applications. The aim of this publication is to review the correlation between bio‐fillers and their effect on polymer properties, including mechanical and thermal properties, degradation processes etc. The scope of the work covers the analysis of cellulose bio‐fillers (nanocellulose, bacterial cellulose, plant waste raw materials), starch, protein‐based bio‐fillers (of plant and animal origin) and mineral fillers, as well as methods of their modification to improve compatibility with polymers. The work systematises the current knowledge on different types of bio‐filler used in polymers, and indicates the challenges faced by the use of bio‐fillers.
p> Black tea has lower antioxidant activity than green tea, the color is slightly darker and less stable during storage. Therefore, it is necessary to add natural antioxidants so that antioxidants increase, and the color is brighter and stable. This study aims to determine the effect of adding lime juice and Eucalyptus globulus essential oil (MEEg) on antioxidant activity and color, as well as organoleptic hedonic brewing of black tea. The research method used was an experimental method consisting of 2 stages, namely the stage of determining the length of brewing black tea and the stage of adding lime juice (0, 1, 2, 3% v/v) and MEEg (0.00, 0.05, 0). ,10, and 0.15% v/v). The results showed that the best brewing time was 8 minutes with an IC50 value of 1411.24±75.79 ppm, a pH value of 5.25±0.02, a yellow-red color (oHue 75.93±3.16), and a lightness value. 47.09±1.06. The addition of lime juice tends to decrease antioxidant activity, total phenolic, total flavonoid, and total condensed tannins, but the addition of MEEg increases antioxidant activity, total phenolic, total flavonoid, and total condensed tannins in black tea. The addition of lime juice and MEEg increased the preference for the tea color and mint flavor, but decreased the preference for the sour taste and tea aroma. The selected treatment was the one with the highest antioxidant activity and hedonic value, namely the addition of 3.0% lime and 0.15% MEEg.
Bahasa Indonesia Abstract:
Teh hitam mempunyai aktivitas antioksidan lebih rendah daripada the hijau, warna seduhannya agak gelap dan kurang stabil selama penyimpanan. Oleh karena itu perlu ditambahkan antioksidan alami agar antioksidan meningkat dan warna lebih terang dan stabil. Penelitian ini bertujuan mengetahui pengaruh penambahan perasan jeruk nipis dan minyak esensial Eucalyptus globulus (MEEg) terhadap akivitas antioksidan dan warna, serta organoleptik hedonic seduhan teh hitam. Metode penelitian yang digunakan metode eksperimen yang terdiri dari 2 tahap, yaitu tahap penentuan lama penyeduhan teh hitam dan tahap penambahan perasan jeruk nipis (0, 1, 2, 3% v/v) dan MEEg (0,00, 0,05, 0,10, dan 0,15% v/v). Hasil penelitian menunjukkan bahwa lama waktu penyeduhan terbaik adalah 8 menit dengan nilai IC50 1411,24±75,79 ppm, nilai pH 5,25±0,02, warna kuning merah (oHue 75,93± 3,16), dan nilai lightness 47,09±1,06. Penambahan perasan jeruk nipis cenderung menurunkan aktivitas antioksidan, total fenolik, total flavonoid, dan total tanin terkondensasi, tetapi penambahan MEEg meningkatkan aktivitas antioksidan, total fenolik, total flavonoid, dan total tanin terkondensasi seduhan teh hitam. Penambahan perasan jeruk nipis dan MEEg meningkatkan kesukaan terhadap warna seduhan teh dan rasa mint, tetapi menurunkan kesukaan terhadap rasa asam dan aroma teh. Perlakuan terpilih adalah yang memiliki aktivitas antioksidan dan nilai hedonik tertinggi yaitu penambahan jeruk nipis 3,0% dan MEEg 0,15%.</p
A study was carried out to quantitatively estimate the L-theanine content in 19 teas commercially available in the Kenyan market by High Performance Liquid Chromatography (HPLC). The test tea samples analyzed were green (n = 4), black (n = 8) and flavored (n = 7) teas from different origins viz., Kenya (n = 4), Uganda (n = 2), Tanzania (n = 5), Rwanda (n = 4), Cameroon (n = 1) and Sri-Lanka (n = 2) commercially available in the Kenyan market. The estimated Limit of Detection (LOD) of the current method was 0.01% L-theanine. The L-theanine content ranged from below the detection limit (<0.01% L-theanine) to 1.60% L-theanine on a dry weight (d.w) basis. Statisti- cally significant differences (p < 0.05) were observed in the L-theanine contents of black, green and flavoured teas. Rwandan green tea contained the highest L-theanine content with 1.60% d.w. whereas six of the seven flavoured teas had very low theanine levels (<0.01%) that could not be quantified by the current method.
The plant extracts of 17 commonly used Indian medicinal plants were examined for their possible regulatory effect on nitric oxide (NO) levels using sodium nitroprusside as an NO donor in vitro. Most of the plant extracts tested demonstrated direct scavenging of NO and exhibited significant activity. The potency of scavenging activity was in the following order: Alstonia scholaris > Cynodon dactylon > Morinda citrifolia > Tylophora indica > Tectona grandis > Aegle marmelos (leaf) > Momordica charantia > Phyllanthus niruri > Ocimum sanctum > Tinospora cordifolia (hexane extract) = Coleus ambonicus > Vitex negundo (alcoholic) > T cordifolia (dichloromethane extract) > T. cord folia (methanol extract) > Ipomoea digitata > V negundo (aqueous) > Boerhaavia diffusa > Eugenia jambolana (seed) > T. cord folia (aqueous extract) > V. negundo (dichloromethane/methanol extract) > Gingko biloba > Picrorrhiza kurroa > A. marmelos (fruit) > Santalum album > E. jambolana (leaf). All the extracts evaluated exhibited a dose-dependent NO scavenging activity. The A. scholaris bark showed its greatest NO scavenging effect of 81.86% at 250 mug/mL, as compared with G. biloba, where 54.9% scavenging was observed at a similar concentration. The present results suggest that these medicinal plants might be potent and novel therapeutic agents for scavenging of NO and the regulation of pathological conditions caused by excessive generation of NO and its oxidation product, peroxynitrite.
Cultivation of Spirulina platensis under salt stress conditions (0.02 M as control), 0.04 and 0.08 M NaCl led to a remarkable alteration of algal metabolism as well as an enhancement or induction of biologically active compounds. Concerning algal growth, salt stress caused a decrease in dry weight, chlorophyll a content as well as certain xanthophylls (neoxanthin and violaxanthin) while β-carotene production was stimulated especially at higher salt concentrations. Biochemical analysis of salt stressed algal revealed that lipid content was slightly increased together with certain saturated and unsaturated fatty acids especially the polyunsaturated ones (γ-inolenic acid, omega 3 fatty acid). Electrophoretic analysis of soluble protein pointed out that certain high molecular weight protein bands were not detected comparing with the protein marker. Five new protein bands of molecular weights 190, 158, 113, 77 and 28 KDa were recorded, in addition to an increase in the intensity of 6 already existing bands. Phosphate buffer and water extracts of the algal exhibited antiviral activities against both Hepatitis-A-virus-type-MBB (HAV-MBB strain, RNA virus) and Herpes simplex-virus-type-1 (HSV-1, DNA virus). Water extracts were found to be more effective than phosphate buffer extracts in inducing antiviral activities (98%) especially against HSV-1 virus. The same water extract of the salt stressed algal demonstrated higher anticoagulating activity compared with those of heparin and the positive control measured by clotting time assay. Antioxidant activity of the algal successive extracts against 2, 2 diphenyl-1-picrylhydrazyl and 2,2'- azino-bis (ethylbenzthiazoline-6- sulfonic acid) radical methods revealed moderate antioxidant activity of the non-polar algal extracts (petroleum ether) which were doubled with increasing extract concentration. The lowest activity was recorded by the partially polar (ethyl acetate) algal extract of both concentrations at all salinity levels. While the polar extracts (ethanol and water) showed higher antioxidant activities which were doubled with increasing extract concentration. Ethanolic algal extract (100 μg/ ml at 0.08 M NaCl) exhibited the highest antioxidant activity compared with those of the synthetic antioxidant butylated hydroxy anisol as standard (85.0, 89.9 and 86.0, 91.8% respectively).
Kutki or Picrorhiza kurroa is a herbal medicinal plant belonging to Scrophulariaceae family and is found in the Himalayan region in India. This herb has been traditionally used in treating liver disorders. The antioxidant properties of P. kurroa were evaluated in vitro using different radical scavenging assays. Furthermore, liver slice culture system was used to test the antioxidant activity of this extract and ethanol was used as a hepatotoxin to generate oxidative stress. Hepatotoxicity was quantified in terms of release of intracellular marker enzymes lactate dehydrogenase, glutamate oxaloacetate transaminase and glutamate pyruvate transaminase. Oxidative stress induced by ethanol and its modulation in the presence of P. kurroa extract was tested by estimating the levels of antioxidant enzymes like catalase, superoxide dismutase, glutathione peroxidase, glutathione reductase and glutathione-S-transferase, and of antioxidant molecules like uric acid and reduced glutathione that were quantitated along with lipid peroxidation. Our results clearly demonstrate that aqueous extract of P. kurroa with high antioxidant activity, as demonstrated using different radical scavenging assays, was effective in suppressing the deleterious effects of ethanol. Addition of P. kurroa aqueous extract along with ethanol restored the activities of antioxidant enzymes and significantly reduced lipid peroxidation.
Ataxia telangiectasia (A-T) and Nijmegen breakage syndrome (NBS) are recessive genetic disorders with susceptibility to cancer and similar cellular phenotypes. The protein product of the gene responsible for A-T, designated ATM, is a member of a family of kinases characterized by a carboxy-terminal phosphatidylinositol 3-kinase-like domain. The NBS1 protein is specifically mutated in patients with Nijmegen breakage syndrome and forms a complex with the DNA repair proteins Rad50 and Mre11. Here we show that phosphorylation of NBS1, induced by ionizing radiation, requires catalytically active ATM. Complexes containing ATM and NBS1 exist in vivo in both untreated cells and cells treated with ionizing radiation. We have identified two residues of NBS1, Ser 278 and Ser 343 that are phosphorylated in vitro by ATM and whose modification in vivo is essential for the cellular response to DNA damage. This response includes S-phase checkpoint activation, formation of the NBS1/Mre11/Rad50 nuclear foci and rescue of hypersensitivity to ionizing radiation. Together, these results demonstrate a biochemical link between cell-cycle checkpoints activated by DNA damage and DNA repair in two genetic diseases with overlapping phenotypes.
Glutathione peroxidase (GPX), superoxide dismutase (SOD) and catalase are the most important enzymes of the cell antioxidant defense system. However, these molecules are themselves susceptible to oxidation. The aim of this work was to estimate to what extent this system could be inactivated by its own substrates. We tested the effect of hydrogen peroxide, cumene hydroperoxide, t-butyl hydroperoxide and hydroxyl and superoxide radicals on GPX, SOD and catalase. For GPX, a 50% inactivation was observed at 10−1 M (30 min, 37°C) for hydrogen peroxide, 3 × 10−4 M (15 min, 37°C) for cumene hydroperoxide and 5 × 10−5 M (11 min, 37°C) for t-butyl hydroperoxide. Unlike the hydroxyl radicals, superoxide anions did not inactivate this enzyme. Catalase was inactivated by hydroxyl radicals and by superoxide anions but organic peroxides had no effect. SOD was inactivated by 50% by hydrogen peroxide at 4 × 10−4 M (20 min, 37°C), but organic peroxides and hydroxyl radicals were ineffective on this enzyme. Since the three enzymes of the antioxidant system are susceptible to at least one of the oxidative reactive molecules, in the case of high oxidative stresses such an inhibition could take place, leading to an irreverisble autocatalytical process in which the production rate of the oxidants will continuously increase, leading to cell death.
To investigate the protective effects of octacosanol in 6-hydroxydopamine-induced Parkinsonian rats and find whether octacosanol has effects on pro nerve growth factor (pro-NGF), NGF and the downstream effector proteins.
Behavioral tests, enzymatic assay, tyrosine hydroxylase immunohistochemistry, TUNEL and Western blot were used to investigate the effects of octacosanol in this rat model of PD.
Oral administration of octacosanol (35-70 mg/kg, po for 14 d) significantly improved the behavioral impairments in rats induced by 6-OHDA and dose-dependently preserved the free radical scavenging capability of the striatum. Octacosanol treatment also effectively ameliorated morphological appearances of TH-positive neuronal cells in nigrostriatal systems and decreased the apoptotic cells induced by 6-OHDA in striatum. In addition, octacosanol strikingly blocked the 6-OHDA-induced increased expression of proNGF-p75NTR-sortilin death signaling complex and its downstream effector proteins. Meantime, octacosanol prevented the decreased levels of NGF, its receptors TrkA and p-Akt which together mediated the cell survival pathway.
The findings implicated that the anti-parkinsonism effects afforded by octacosanol might be mediated by its neuro-microenvironment improving potency through retrieving the ratios of proNGF:NGF and the respective receptors p75NTR:TrkA in vivo. Due to its excellent tolerability and non-toxicity, octacosanol may be a promising agent for PD treatment.
Radioprotective activity of pure compounds isolated from the plant Phyllanthus amarus was studied using rat liver mitochondria and pBR322 plasmid DNA as an in vitro model system. These compounds were ellagitannins namely amariin, 1-galloyl-2,3-dehydrohexahydroxydiphenyl (DHHDP)-glucose, repandusinic acid, geraniin, corilagin, phyllanthusiin D, and flavonoids namely rutin, and quercetin 3-O-glucoside. The activity was then correlated with their hydroxyl and superoxide radical scavenging activity. Both ellagitannins and flavonoids effectively prevented lipid peroxidation and protein oxidation in mitochondria. The compounds also prevented radiation induced single strand breaks in pBR322 plasmid DNA. The radioprotective activity of ellagitannins and flavonoids could be due to their ability to scavenge different radicals more or less efficiently, relieving the oxidative stress. Protection conferred by flavonoids, rutin and quercetin 3-O-glucoside to rat liver mitochondria and plasmid pBR322 DNA from radiation induced damage was due to their strong hydroxyl radical scavenging activity. The inhibitory effect of ellagitannins on lipid peroxidation in liver mitochondria was due to their efficient superoxide radical scavenging ability. This is the first report about the radioprotective activity of pure ellagitannins from Phyllanthus amarus.
Nematodes are the dominant soil animals in Antarctic Dry Valleys and are capable of surviving desiccation and freezing in an anhydrobiotic state. Genes induced by desiccation stress have been successfully enumerated in nematodes; however we have little knowledge of gene regulation by Antarctic nematodes which can survive multiple environmental stresses. To address this problem we investigated the genetic responses of a nematode species, Plectus murrayi, that is capable of tolerating Antarctic environmental extremes, in particular desiccation and freezing. In this study, we provide the first insight into the desiccation induced transcriptome of an Antarctic nematode through cDNA library construction and suppressive subtractive hybridization.
We obtained 2,486 expressed sequence tags (ESTs) from 2,586 clones derived from the cDNA library of desiccated P. murrayi. The 2,486 ESTs formed 1,387 putative unique transcripts of which 523 (38%) had matches in the model-nematode Caenorhabditis elegans, 107 (7%) in nematodes other than C. elegans, 153 (11%) in non-nematode organisms and 605 (44%) had no significant match to any sequences in the current databases. The 1,387 unique transcripts were functionally classified by using Gene Ontology (GO) hierarchy and the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. The results indicate that the transcriptome contains a group of transcripts from diverse functional areas. The subtractive library of desiccated nematodes showed 80 transcripts differentially expressed during desiccation stress, of which 28% were metabolism related, 19% were involved in environmental information processing, 28% involved in genetic information processing and 21% were novel transcripts. Expression profiling of 14 selected genes by quantitative Real-time PCR showed 9 genes significantly up-regulated, 3 down-regulated and 2 continuously expressed in response to desiccation.
The establishment of a desiccation EST collection for Plectus murrayi, a useful model in assessing the structural, physiological, biochemical and genetic aspects of multiple stress tolerance, is an important step in understanding the genome level response of this nematode to desiccation stress. The type of transcript analysis performed in this study sets the foundation for more detailed functional and genome level analyses of the genes involved in desiccation tolerance in nematodes.
Free radicals that escape scavenging by antioxidant defense damage lipids, proteins, and DNA. Damage to DNA can be repaired. Therefore, both cells' antioxidant defense and their ability to repair oxidatively damaged DNA decide its fate to survive oxidative stress. Pancreatic islets cells with poor antioxidant defense were checked for their ability to remove oxidative damage form DNA.
For ex vivo DNA repair, assay-cultured pancreatic islets and liver slices were treated with 1 and 10 mM H2O2, respectively, for 30 minutes. After incubation for different time intervals, 8-hydroxy-2'-deoxyguanosine (8-OHdG) in DNA of these cells was estimated using monoclonal antibody raised against 8-OHdG by competitive enzyme-linked immunosorbent assay. For in vitro DNA repair assay, oxidatively damaged pBR322 was incubated with nuclear extracts of islet and liver cells, and 8-OHdG retained in the plasmid was quantitated.
Oxidative damage induced by H2O2 was removed quickly and efficiently from DNA by liver cells compared with islet cells. The repair of oxidatively damaged plasmid DNA in vitro was also performed more efficiently (P < 0.05) by nuclear extracts from liver cells compared with islet cell.
We clearly demonstrate that in addition to their low antioxidant defense, islets are very poor in rectifying the oxidative DNA damage.
An antioxidant fraction of Chinese green tea (green tea antioxidant; GTA), containing several catechins, has been previously
shown to inhibit 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced tumor promotion in mouse skin. In the present study, GTA was shown to have
antioxidative activity toward hydrogen peroxide (H2O2) and the superoxide radical (O2−). GTA also prevented oxygen radical and H2O2-induced cytotoxicity and inhibition of intercellular communication in cultured B6C3F1 mouse hepatocytes and human keratinocytes
(NHEK cells). GTA (0.05–50 μg/ml) prevented the killing of hepatocytes (measured by lactate dehydrogenase release) by paraquat
(1–10 mM) and glucose oxidase (0.8–40 μg/ml) in a concentration-dependent fashion. GTA (50μg/ml) also prevented the inhibition
of hepatocyte intercellular communication by paraquat (5 mM), glucose oxidase (0.8 μg/ml), and phenobarbital (500 μg/ml).
In addition, GTA (50 μg/ml) prevented the inhibition of intercellular communication in human keratinocytes by TPA (100 ng/ml).
Cytotoxicity and inhibition of intercellular communication, two possible mechanisms by which tumor promoters may produce their
promoting effects were therefore prevented by GTA. The inhibition of these two effects of pro-oxidant compounds may suggest
a mechanism by which GTA inhibits tumor promotion in vivo.
Nitro blue tetrazolium has been used to intercept O2⁻ generated enzymically or photochemically. The reduction of NBT by O2⁻ has been utilized as the basis of assays for superoxide dismutase, which exposes its presence by inhibiting the reduction of NBT. Superoxide dismutase could thus be assayed either in crude extracts or in purified protein fractions. The assays described are sensitive to ng/ml levels of super-oxide dismutase and were applicable in free solution or on polyacrylamide gels. The staining procedure for localizing superoxide dismutase on polyacrylamide electrophoretograms has been applied to extracts obtained from a variety of sources. E. coli has been found to contain two superoxide dismutases whereas bovine heart, brain, lung, and erthrocytes contain only one.
Butanol-soluble neutral and acidic thearubigins were prepared by a combination of solvent extraction, fractional precipitation, and Toyopearl column chromatography. These pigments showed similar properties to those of Roberts’ thearubigins on the paper chromatogram, and a convex broad band with several peaks on reversed-phase HPLC. The thearubigins were methylated, degallated, and chemically degraded with KMnO4 under alkaline conditions. From the degradation products, methyl esters of 4-methoxy benzoic acid, 3,4-dimethoxy benzoic acid, 3,4,5-trimethoxy benzoic acid, 3,4-dimethoxy-1,2-benzenedicarboxylic acid, 3,4-dimethoxy-1,5-benzenedicarboxylic acid, 3,4-dimethoxy-1,6-benzenedicarboxylic acid, 3,4,5-trimethoxy-1,2-benzenedicarboxylie acid, 4,5,6-trimethoxy-1,2,3-benzenetricarboxylic acid, 4,5,4′,5′-tetramethoxy-2,2′-diphenic acid, and 3,4,5,3′,4′,5′-hexamethoxy-2,2′-diphenic acid were detected by using GC-MS and GC-SIM. The butanol-soluble neutral thearubigins, which had been purified by Toyopearl column chromatography, were hydrolyzed by treating with HCl–BuOH. Cyanidin and delphinidin were identified in the reaction mixture. These results suggest that thearubigins are heterogeneous polymers of flavan-3-ols and flavan-3-ol gallates which have a bond at C4, C6, C8, C2′, C5′, and C6′ in the flavan-3-ol units. In addition to the C4–C8 or C6 interflavanoid linkage, a C6′–C6′ linkage was also found, and a possible partial structure for thearubigin is proposed.
Objective: To determine the modulating effect of Garcinia cambogia fruit extract on ethanol induced peroxidative damage in rats. Method: Male albino rats weighing 125 to 150g were administered ethanol (7.11g per kg body weight / day) for 45 days. Ethanol administered rats were treated concomitantly with Garcinia cambogia fruit extract (1g/kg body weight / day) orally for 45 days. After the experimental period the antioxidant enzymes, LPO, conjugated diene in the liver tissue, serum AST, ALT and alkaline phosphatase and lipid levels in both serum and liver tissue were estimated. Results: Co-treatment of the rats with Garcinia cambogia significantly inhibited the rise in lipid levels and also the peroxidative damage caused by ethanol, which is evident from the improved antioxidant status. The levels of serum AST, ALT and alkaline phosphatase were maintained at near normalcy in Garcinia cambogia treated rats. Conclusion: The imbalance in lipid metabolism could be the reason for increase in lipid peroxidation. In our present study the treatment with Garcinia cambogia fruit extract resulted in reduction of both serum and liver lipid to near normalcy. This hypolipidemic property of Garcinia cambogia in turn reduces the peroxidative damage, enhanced by ethanol.
Several of the most commonly used methods for in vitro determination of antioxidant capacity are reviewed in the present paper. The chemical principles of methods based either on biological oxidants (peroxyl radical, superoxide radical anion, hydrogen peroxide, hydroxyl radical, hypochlorous acid, singlet oxygen, nitric oxide radical, and peroxynitrite) or on non-biological assays (scavenging of 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulphonate) radical cation (TEAC assay), scavenging of 2,2-diphenyl-1-picrylhydrazyl radical (DPPH assay), ferric reducing antioxidant power (FRAP assay), Folin-Ciocalteu reducing capacity (FC assay), electrochemical total reducing capacity) are outlined and critically discussed. The scope of application, the advantages and shortcomings of each method are also highlighted.
1. Oxygen is a toxic gas - an introductionto oxygen toxicity and reactive species 2. The chemistry of free radicals and related 'reactive species' 3. Antioxidant defences Endogenous and Diet Derived 4. Cellular responses to oxidative stress: adaptation, damage, repair, senescence and death 5. Measurement of reactive species 6. Reactive species can pose special problems needing special solutions. Some examples. 7. Reactive species can be useful some more examples 8. Reactive species can be poisonous: their role in toxicology 9. Reactive species and disease: fact, fiction or filibuster? 10. Ageing, nutrition, disease, and therapy: A role for antioxidants?
In order to understand the factors responsible for the potent antioxidant and radioprotecting ability of triphala, it has been evaluated for radical scavenging ability, xanthine oxidase inhibitory activity and phytochemical (phenolics) content. The radical scavenging experiments were carried out using fast reaction kinetic tools and the reactivity of triphala towards different radicals such as hydroxyl radicals, superoxide radicals, DPPH and ABTS • •- - was determined. When triphala was tested for superoxide radical scavenging activity using xan- thine and xanthine oxidase assay, it was observed that in addition to reacting with superoxide radical, it also inhibited uric acid formation, indicative of xanthine oxidase enzyme inhibitory activity. Phytochemical ana - lysis showed that triphala is rich in phenols/polyphenols (38 ± ± 3%) and tannins (35 ± ± 3%), while flavonoids were found to be absent. HPLC analysis showed that triphala contains 73 ± ± 5 mg gallic acid per gram of triphala, which was found to increase to 150 ± ± 5 mg/g upon acid hydrolysis. Relevance of these studies to the antioxidant and radio protection properties of triphala has also been discussed.
Theaflavins (TF) and thearubigins (TR) were separated from Korean microbially fermented tea leaves. Contents of TF (74.4 μM/g) and TR (37.2%) were higher than reported for black tea fermented by oxidase. Antioxidant activities of TF, TR and EGCG were analyzed and protective effects of COS-7 cells against copper and cadmiuminduced toxicity were investigated. TF and TR exhibited good inhibition rates of about 85-90% for antioxidant and scavenging activities of free radicals and protected COS?7 cells against apoptosis or damage caused by stress, such as cadmium and copper-oxidative injury, free radicals etc. These results indicate that TF, TR and EGCG have antioxidant and scavenging activities against free radicals and protect COS?7 cells from Cu, Cd induced injury.
The antioxidant activity, expressed in mM Trolox equivalent antioxidant capacity (TEAC), of some edible Yemeni plants belonging to different families was evaluated using the ferrylmyoglobin/ABTS.+ method. Methanolic (80%) extracts of tested plants exhibited higher (p<0.05) TEAC than extracts of other solvents. Extracts of coffee beans (Coffea arabica), sweet basil (Ocimum basilicum), wild thyme (Thymus serpyllum) and sorrel (Rumex acetosa) had antioxidant activities equivalent to 8.7-47.1 mM Trolox or 7.9-42.8 mM BHA per g dry weight of plant. The TEAC of methanolic extracts for sorrel was not significantly different (p>0.05) from extracts of Leucaena leucocephala (Thailand) and Japanese green tea (Camellia sinensis). A good correlation (R2=0.937) was observed between TEAC and total phenol contents in 80% methanol extracts of tested plants.
Daily administration of ethyl alcohol (3.76 g/kg, p.o.) for 45 days resulted in significant changes in several biochemical parameters of the liver and serum of albino rats. After exposure to alcohol for 30 days when Picroliv (12 mg/kg, p.o.), an iridoid glycoside fraction of Picrorhiza kurroa, was administered for 15 days along with alcohol, the degree of change in most of the parameters was reduced.
Picroliv, the active principle an iridoid glycoside mixture isolated from the plant Picrorhiza kurrooa, showed dose-dependent (0.75–12 mg/kg × 7 days) protective activity on isolated hepatocytes (ex vivo) against paracetamol-induced hepatic damage in rats. It increased the percent viability of the hepatocytes. Picroliv also restored the normal values of enzyme (glutamic oxaloacetic transaminase [GOT], glutamic-pyruvic transaminase [GPT], and alkaline phosphatase) both in the isolated hepatocyte suspension as well as in the serum. Picroliv was found to be more potent than silymarin, a known hepatoprotective agent.
There is currently much interest in phytochemicals as bioactive components of food. The roles of fruit, vegetables and red wine in disease prevention have been attributed, in part, to the antioxidant properties of their constituent polyphenols (vitamins E and C, and the carotenoids). Recent studies have shown that many dietary polyphenolic constituents derived from plants are more effective antioxidants in vitro than vitamins E or C, and thus might contribute significantly to the protective effects in vivo. It is now possible to establish the antioxidant activities of plant-derived flavonoids in the aqueous and lipophilic phases, and to assess the extent to which the total antioxidant potentials of wine and tea can be accounted for by the activities of individual polyphenols.
The determination of the depletion pattern of catechins in black tea processing is important in achieving optimum tea quality. This study investigated catechins (unoxidised di- and trihydroxylated) depletion patterns in relation to theaflavin and thearubigin formation. It was during the process of green leaf fermentation at selected temperature and time combinations. The tea leaves were obtained from three clones (6/8, 303/577 and 311/287) within the Tea Research Foundation of Kenya. The results were showed that unequal depletion rates of di- and trihydroxylated catechins led to a decline in total theaflavin and an increase in thearubigins levels. An equitable decline in both groups of catechins corresponded to a subsequent rise in theaflavins content. The decline in the catechins levels was much faster at higher temperatures resulting in a shorter fermentation time to achieve a peak of the theaflavins content. Clone 311/287 had the highest mean theaflavins content (26.99 μmol/g) and the least mean percent thearubigins (15.02%) level. Theaflavins content correlated positively with liquor brightness determined by a spectrophotometer and tea tasters (r = 0.7221, p < .0001). The thearubigins content was however found to relate negatively with liquor brightness. It was concluded that the experimental conditions tested form a good basis for clonal specific processing conditions that can be utilised in manufacturing quality black tea.
Publisher Summary This chapter discusses the analysis of total phenols and other oxidation substrates and antioxidants by means of Folin-Ciocalteu reagent. Analyses of the Folin-Ciocalteu (FC) type are convenient, simple, and require only common equipment and have produced a large body of comparable data. Under proper conditions, the assay is inclusive of monophenols and gives predictable reactions with the types of phenols found in nature. Because different phenols react to different degrees, expression of the results as a single number—such as milligrams per liter gallic acid equivalence—is necessarily arbitrary. Because the reaction is independent, quantitative, and predictable, analysis of a mixture of phenols can be recalculated in terms of any other standard. The assay measures all compounds readily oxidizable under the reaction conditions and its very inclusiveness allows certain substances to also react that are either not phenols or seldom thought of as phenols (e.g., proteins). Judicious use of the assay—with consideration of potential interferences in particular samples and prior study if necessary—can lead to very informative results. Aggregate analysis of this type is an important supplement to and often more informative than reems of data difficult to summarize from various techniques, such as high-performance liquid chromatography (HPLC) that separate a large number of individual compounds .The predictable reaction of components in a mixture makes it possible to determine a single reactant by other means and to calculate its contribution to the total FC phenol content. Relative insensitivity of the FC analysis to many adsorbents and precipitants makes differential assay—before and after several different treatments—informative.
An alcoholic extract of Picrorrhiza kurroa was found to lower blood glucose in basal conditions and after a heavy glucose load in normal rats. Maximum reduction in serum glucose was observed after 2 h at a dose level of 75 mg extract/kg of body weight. P. kurroa extract was also found to reduce the increase of blood sugar in alloxan-induced diabetic rats (43% at 75 mg/kg body weight and 60% at 150 mg/kg body weight). Chronic administration of the extract significantly reduced the blood sugar in alloxan-induced diabetic rats for several days (10 days). The extract was also found to reduce the increased blood urea nitrogen and serum lipid peroxides in alloxan-induced diabetic animals and to inhibit the body weight reduction and leukopenia induced by alloxan administration. These results indicate that P. kurroa extracts are able to ameliorate biochemical damages induced by alloxan in diabetic rats.
Adriamycin, an anthracycline antibiotic, which is used in the treatment of various tumors, is known to cause severe cardiomyopathy. The present study examined the protective effects of Picrorhiza kurroa, an ayurvedic medicinal plant, on myocardial antioxidant defense system in adriamycin-induced cardiomyopathy in rats. Intraperitoneal administration of adriamycin (1.5 mg/kg body weight/ day, i.p. for 15 days) caused significant rise in the levels of diagnostic marker enzymes [alanine aminotransferase (ALT), aspartate aminotransferase (AST), lactate dehydrogenase (LDH) and creatine phoshokinase (CPK)] in plasma and lipid peroxidation in the heart tissue of experimental rats. Concomitant decline in the level of reduced glutathione (GSH) and the activities of glutathione-dependent antioxidant enzymes (GPx and GST) and antiperoxidative enzymes (SOD and CAT) in the myocardial tissue were also observed. Oral administration of P. kurroa extract (50 mg/kg body weight/day, for a period of 15 days) significantly prevented all these adriamycin-induced adverse effects and maintained the rats at normal status. The protective effect of P. kurroa might be ascribable to its membrane-stabilizing property and/or antioxidant nature.
Oxidative stress is well recognized to be a key step in the pathogenesis of ethanol-associated liver injury. Ethanol administration induces an increase in lipid peroxidation either by enhancing the production of oxygen reactive species and/or by decreasing the level of endogenous antioxidants. Numerous experimental studies have emphasized the role of the ethanol-inducible cytochrome P450 in the microsomes and the molybdo-flavoenzyme xanthine oxidase in the cytosol. This review shows the putative role of ethanol-induced disturbances in iron metabolism in relation to iron as a pro-oxidant factor. Ethanol administration also affects the mitochondrial free radical generation. Many previous studies suggest a role for active oxygens in ethanol-induced mitochondrial dysfunction in hepatocytes. Recent studies in our laboratory in the Department of Internal Medicine, Keio University, using a confocal laser scanning microscopic system strongly suggest that active oxidants generated during ethanol metabolism produce mitochondrial membrane permeability transition in isolated and cultured hepatocytes. In addition, acetaldehyde, ethanol consumption-associated endotoxaemia and subsequent release of inflammatory mediators may cause hepatocyte injury via both oxyradical-dependent and -independent mechanisms. These cytotoxic processes may lead to lethal hepatocyte injury. Investigations further implicate the endogenous glutathione—glutathione peroxidase system and catalase as important antioxidants and cytoprotective machinery in the hepatocytes exposed to ethanol.
We examined the effects of tea polyphenols (TP) on the body weight gain and contents of lipids and lipid peroxides in rat plasma, kidney and liver. The supplementation of TP (0.5% and 1.0%) in the diet was performed from weanling (3 weeks of age) to 19 months old. A significant suppression of body weight gain was observed only at 7–11 weeks of age, but there were no significant changes in liver and kidney weight between control group and TP-feeding group (TP group). Daily food intake of animals in two TP groups was almost the same as the control group.
In the older rats (13–19 months of age), thiobarbituric acid reactive substances (TBARS) in the plasma in 1.0% TP group were significantly lower than that of the control g roup (p<0.01). The contents of plasma lipids (triglyceride, total cholesterol, and phospholipid) in 1.0% TP group at 19 months old were significantly lower as compared with the control group (p<0.05). There was a positive correlation in an age group 13–19 months old between these lipid contents and TBARS.
These results suggest that long-term dietary supplementation of TP caused the decrease of plasma TBARS, mainly due to the hypolipidemic activity and antioxidative effect of TP in vivo.
Phenolic compounds constitute 50–70% of tea water extract and are the main quality parameters for teas. Theaflavins (TF), thearubigins (TR) and theabrownins (TB) are the major polyphenols that determine the quality of black tea. These compounds were measured in 56 leaf teas and teabags sampled from Australian supermarkets in Queensland. The various quantities of TF, ranging from 0.29% to 1.25%, indicate a quality difference that exists among the teas studied. Low TF content in black tea may be due to over-fermenting and/or long periods of storage. The solubility of TR and TB from teabags ranged from 82% to 92%, indicating that the permeability of teabags was variable. Variable quantities of TF in Australian teas show instability and a tendency of TF to oxidize during storage. Total polyphenols in green teas ranged from 14% to 34%, indicating a large variation, which was not reflected in price. The solubility of total polyphenols from teabags has been proposed as a useful quality index of the filtering paper used for the teabags. This chemical analysis of phenolic compounds in commercial teas may be a potential tool for the quality control of Australian manufactured and imported teas in Australian markets.
Glutathione peroxidase activity in the liver supernatant from rats fed a Se-deficient diet for 2 weeks was 8% of control when measured with H2O2 but 42% of control when assayed with cumene hydroperoxide. Two peaks of glutathione peroxidase activity were present in the Sephadex G-150 gel filtration chromatogram of rat liver supernatant when 1.5 mM cumene hydroperoxide was used as substrate. Only the first peak was detected when 0.25 mM H2O2 was used as substrate. The first peak was absent from chromatograms of Se-deficient rat liver supernatants; but the second peak, which eluted at a position corresponding to M.W. = 39,000, appeared unchanged. The second peak thus represents a second glutathione peroxidase activity which catalyzes the destruction of organic hydroperoxides but has little activity toward H2O2 and which persists in severe selenium deficiency.
Momordica charantia Linn. var. abbreviata Ser. (Cucurbitaceae), also known as “Shan Ku Gua”, is a wild variety of bitter melon (BM) in Taiwan. The size of its fruits is only about one-fifth of the commonly seen BM. It is commonly consumed as vegetable and also used as a popular folk medicine. In this study, the antioxidant and free radical scavenging activities of BM aqueous (BM-H2O) and ethanol (BM-EtOH) extracts were evaluated using 2,2-diphenyl-1-picrylhydrazyl (DPPH), metal chelation, cytochrome c and xanthine oxidase inhibition (XOI) assays, as well as FeCl2-ascorbic acid induced lipid peroxidation (thiobarbituric acid reactive substances, TBARS) assay in rat liver homogenates in vitro. Total flavonoid and phenol contents of BM extracts were also analyzed. Results showed that both BM-H2O (IC50=129.94 μg/ml) and BM-EtOH (IC50=156.78 μg/ml) possess potent DPPH radical scavenging activity, which was better than vitamin E (IC50=172.21 μg/ml). These extracts also showed better iron chelating activity than vitamin E. However, they were weaker than vitamin E in free radical scavenging, xanthine oxidase inhibitory and anti-lipid peroxidation activities. With the exception of XOI activity [IC50=7.90 μg/ml (BM-H2O) vs. 7.69 μg/ml (BM-EtOH)], BM-H2O showed a lower IC50 value in free radical scavenging [IC50=6.15 μg/ml (BM-H2O) vs. 7.08 μg/ml (BM-EtOH)] and anti-lipid peroxidation [IC50=53.72 μg/ml (BM-H2O) vs. 88.51 μg/ml (BM-EtOH) for liver; 82.53 μg/ml (BM-H2O) vs. 91.83 μg/ml (BM-EtOH) for brain] activities than BM-EtOH. Both BM extracts showed a weak anti-lipid peroxidation activity in plasma. BM-H2O (62.0 mg/g) possessed a significant higher concentration of total flavonoids than BM-EtOH (44.0 mg/g), but was lower in the total phenol content (BM-H2O: 51.6 mg/g vs. BM-EtOH: 68.8 mg/g). In conclusion, BM extracts possess potent antioxidant and free radical scavenging activities. These antioxidant activities could have contributed, at least partly, to the therapeutic benefits of the certain traditional claims of wild BM.
The ability of curcumin, a natural antioxidant from turmeric, to inhibit radiation-induced lipid peroxidation in rat liver microsomes was examined. Curcumin was incorporated into microsomes during ultracentrifugation. The antioxidant has significant time- and concentration-dependent inhibitory effect on lipid peroxidation induced by r-radiation. Inhibition of lipid peroxidation was also observed in microsomes samples previously saturated with N2O. Curcumin also inhibited lipid peroxidation during the post-irradiation incubation.
A simple method for rapid and reliable assessment of hepatotoxic agents is described. Liver slices from rats and mice of two age groups were incubated with the test hepatotoxins. Exposure of liver slices from 3-month-old mice to acetaminophen (6.8 mg/ml) resulted in 80% leakage of lactate dehydrogenase into the incubation medium, whereas liver slices from one-day-old mice showed only 12% leakage. Similar results were obtained with rat liver slices. The relative lack of response by livers of newborn rats was also demonstrated with carbon tetrachloride. The in vitro liver slice system has also been used to test the potency of N-acetylcysteine (NAC) as an antidote to acetaminophen toxicity. NAC protected mouse liver slices against acetaminophen toxicity in a dose-dependent manner. Addition of the antidote (10 mm) 20 min following hepatotoxin application reduced enzyme leakage by 75% as compared with the system with acetaminophen only. These findings demonstrate that the liver slice system provides the same type of information about hepatotoxins that is usually obtained by the use of acute in vivo tests on a large number of animals. It can be used for testing potential antidotes against hepatotoxins as well as for demonstration of species and age differences in the toxicity of various substances.
A simple method for the rapid screening of hepatotoxic agents is described. Liver slice systems were prepared from rats and mice, and incubated in Krebs-Ringer-Hepes medium with different concentrations of the test compounds. Hepatotoxicity was monitored by determination of liver enzymes in the slice medium. Enzyme leakage was dose- and time-dependent. Histopathological changes in the hepatotoxin-treated slices were well correlated with the extent of enzyme leakage. Species differences in susceptibility to various hepatotoxins could be easily detected by this in vitro system: the dose-toxicity curves revealed that the mouse is more vulnerable than the rat to acetaminophen and furosemide. These findings are well correlated with those of in vivo experiments. A preliminary study showed that, in the same species, the relative toxicities of various chemicals in the liver slice system were similar to those reported in vivo. In summary, these results on the tissue slice system are encouraging. However, much more work will have to be done before the system can be considered sufficiently well validated for routine use.
Thearubigins are the most abundant group of phenolic pigments found in black tea accounting for an estimated 60% of the solids in a typical black tea infusion. Fifty years ago the term thearubigins was first introduced and up to now the chemical nature of the thearubigins remains largely unresolved if not mysterious despite many efforts clarifying their structure. This paper summarizes some of our attempts to clarify and elucidate the chemical nature of the thearubigins, presenting for 15 commercially representative teas data obtained using combustion analysis, IR spectroscopy, NMR spectroscopy, Diffusion NMR spectroscopy, UV-vis spectroscopy, Circular Dichroism spectroscopy and atomic force microscopy, MALDI-TOF-MS and ESI-FT-ICR-MS. The thearubigin fractions from these 15 teas are remarkably similar with respect to their spectroscopic fingerprint. The data obtained are consistent with the thearubigins being structures of not more than 2000Da with more than 5000 individual chemical entities detected that are susceptible to concentration-driven aggregation in aqueous solution, and that retain the chiral properties of the flavanols and theaflavins. By applying petrolomics-style data interpretation strategies and by developing novel data interpretation strategies a structural model for the thearubigins was developed.
We examined whether superoxide is a factor responsible for paraquat-induced liver injury in terms of superoxide dismutase using cultured rat liver slices exposed to various concentrations of paraquat. The degree of liver injury was assessed by measurement of percentage of lactate dehydrogenase leakage into the medium and lipid hydroperoxides in the liver slices and by direct histopathological observation. Paraquat produced concentration- and time-related liver injury in the cultured rat liver slices. Notably, after exposure to 5 mmol/L paraquat, a significant increase of the percentage of lactate dehydrogenase leakage occurred from 4 hr (p less than 0.05 vs. control group), and this gradually increased up to 8 hr (p less than 0.01 and p less than 0.001 vs. control group at 6 and 8 hr, respectively). Changes in lipid hydroperoxides in the liver slices were similar to those in percentage of lactate dehydrogenase leakage (p less than 0.05 and p less than 0.01 vs. control group at 6 and 8 hr, respectively). Liver injury was located around the central vein at 6 hr and gradually spread at 8 hr. Paraquat-induced liver injury was aggravated both biochemically and histopathologically by pretreatment with 5 mmol/L diethyldithiocarbamate, an inhibitor of copper- and zinc-containing superoxide dismutase activity (percentage of lactate dehydrogenase leakage: p less than 0.01 and p less than 0.001 vs. paraquat group at 6 and 8 hr, respectively; lipid hydroperoxides: p less than 0.01 vs. paraquat group at 8 hr). Incubation of liver slices with liposome-encapsulated superoxide dismutase increased the augmentation of superoxide dismutase in the liver slices.(ABSTRACT TRUNCATED AT 250 WORDS)
The peroxidation of rat liver microsomal lipids is stimulated in the presence of iron by the addition of NADPH or ascorbate and is inhibited by the addition of glutathione (GSH). The fate of GSH and the oxidative modification of proteins under these conditions have not been well studied. Rat liver microsomes were incubated at 37 degrees C under 95% O2:5% CO2 in the presence of 10 microM ferric chloride, 400 microM ADP, and either 450 microM ascorbic acid or 400 microM NADPH. Lipid peroxidation was assessed in the presence 0, 0.2, 0.5, 1, or 5 mM GSH by measuring thiobarbituric acid reactive substance (TBARS) and oxidative modification of proteins by measuring protein thiol and carbonyl groups. GSH inhibited TBARS and protein carbonyl group formation in both ascorbate and NADPH systems in a dose-dependent manner. Heat denaturing of microsomes or treatment with trypsin resulted in the loss of this protection. The formation of protein carbonyl groups could be duplicated by incubating microsomes with 4-hydroxynonenal. Ascorbate-dependent peroxidation caused a loss of protein thiol groups which was diminished by GSH only in fresh microsomes. Both boiling and trypsin treatment significantly decreased the basal protein thiol content of microsomes and enhanced ascorbate-stimulated lipid peroxidation. Protection against protein carbonyl group formation by GSH correlated with the inhibition of lipid peroxidation and appeared not to be due to the formation of the GSH conjugate of 4-hydroxynonenal as only trace amounts of this conjugate were detected. Ninety percent of the GSH lost after 60 min of peroxidation was recoverable as borohydride reducible material in the supernatant fraction. The remaining 10% could be accounted for as GSH-bound protein mixed disulfides. However, only 75% of the GSH lost during peroxidation appeared as glutathione disulfide, suggesting that some was converted to other soluble borohydride reducible forms. These data support a role for protein thiol groups in the GSH-mediated protection of microsomes against lipid peroxidation.
The present work has investigated the correlation between short-term ethanol administration and the free radicals production in rat liver. The non-protein-SH and lipid peroxides contents, superoxide dismutase and glutathione peroxidase activities were investigated. Increased lipid peroxides concentration, decreased glutathione content, significant increase in superoxide dismutase activity and a decrease in glutathione peroxidase activity were observed.
Picroliv, the hepatoprotective principle of the plant Picrorhiza kurroa, showed a dose-dependent (1.5-12 mg/kg x 7) choleretic effect in conscious rats and anaesthetised guinea pigs. It also possessed a marked anticholestatic effect against paracetamol- and ethynylestradiol-induced cholestasis. It antagonised the changes in bile volume as well as the contents (bile salts and bile acids). Silymarin, a known hepatoprotective agent, was tested simultaneously for comparison. Picroliv was found to be a more potent choleretic and anticholestatic agent than silymarin.
From the roots of Picrorhiza kurrooa Royle and Benth., seven cucurbitacin glycosides have been isolated and structurally elucidated mainly by NMR and mass spectroscopy. Four of them (4, 5, 6, 7) are new and two, the 2-O-glycoside of cucurbitacin B (25-acetoxy-2-beta-glucosyloxy-16,20-dihydroxy-9-methyl-19-norl anosta-5, 23-diene-3,11,22-trione) and the 2-O-glucoside of 23,24 didydrocucurbitacin B (25-acetoxy-2-beta-glucosyloxy-16,20-dihydroxy-9-methyl-19-norl anost-5-ene-3, 11-22-trione) were so far not reported as constituents of this plant. The four new cucurbitacins could be identified as 2-beta-glucosyloxy-3,16,20,25-tetrahydroxy-9-methyl-19-norlanos ta-5, 23-diene-22-one, 2-beta-glucosyloxy-3,16,20,25-tetrahydroxy-9-methyl-19-norlanos t-5-ene-22-one, the 2-O-glucoside of cucurbitacin Q (25-acetoxy-2-beta-glucosyloxy-3,16,20-trihydroxy-9-methyl-19-n orlanosta-5, 23-diene-11,22-dione), and the 2-O-glucoside of deacetoxycucurbitacin B (2-beta-glucosyloxy-16,20-dihydroxy-9-methyl-19-norlanosta-5 , 24-diene-3,11,22-trione).
Plasmid expression vectors have been constructed that direct the synthesis of foreign polypeptides in Escherichia coli as fusions with the C terminus of Sj26, a 26-kDa glutathione S-transferase (GST; EC 2.5.1.18) encoded by the parasitic helminth Schistosoma japonicum. In the majority of cases, fusion proteins are soluble in aqueous solutions and can be purified from crude bacterial lysates under non-denaturing conditions by affinity chromatography on immobilised glutathione. Using batch wash procedures several fusion proteins can be purified in parallel in under 2 h with yields of up to 15 micrograms protein/ml of culture. The vectors have been engineered so that the GST carrier can be cleaved from fusion proteins by digestion with site-specific proteases such as thrombin or blood coagulation factor Xa, following which, the carrier and any uncleaved fusion protein can be removed by absorption on glutathione-agarose. This system has been used successfully for the expression and purification of more than 30 different eukaryotic polypeptides.
Lipid peroxidation in rat uterus has been studied using NADPH- and ascorbate-induced systems. Lipid peroxidation in rat uterus is low as compared to rat liver. Uterus is more sensitive to ascorbate-induced lipid peroxidation than that induced by NADPH. Uterus contains lower amounts of phospholipids and has a lesser degree of unsaturation in lipids. Co-factor studies show that Fe2+ is more important for ascorbate-induced lipid peroxidation. Endometrium is more sensitive to ascorbate-induced lipid peroxidation than myometrium. It also contains more total lipids and phospholipids besides having a higher degree of unsaturation in the lipids as compared to myometrium. Among the subcellular fractions, mitochondria are more prone to ascorbate-induced lipid peroxidation, whereas microsomes are more sensitive to NADPH-induced lipid peroxidation. Uteri from old rats (24 months) and pregnant rats are more resistant to lipid peroxidation than those from 3-month-old control rats. Uterus of pregnant rats contains more factors which inhibit lipid peroxidation and also has a lesser degree of unsaturation in lipids compared with uterus of control rats. The possible consequences of the resistance of uterus to lipid peroxidation, especially during pregnancy and senescence, are discussed.
Hydroxyl radicals, generated by reaction of an iron-EDTA complex with H2O2 in the presence of ascorbic acid, attack deoxyribose to form products that, upon heating with thiobarbituric acid at low pH, yield a pink chromogen. Added hydroxyl radical "scavengers" compete with deoxyribose for the hydroxyl radicals produced and diminish chromogen formation. A rate constant for reaction of the scavenger with hydroxyl radical can be deduced from the inhibition of color formation. For a wide range of compounds, rate constants obtained in this way are similar to those determined by pulse radiolysis. It is suggested that the deoxyribose assay is a simple and cheap alternative to pulse radiolysis for determination of rate constants for reaction of most biological molecules with hydroxyl radicals. Rate constants for reactions of ATP, ADP, and Good's buffers with hydroxyl radicals have been determined by this method.