Antioxidative activity of green tea polyphenol in cholesterol-fed rats

This study investigated the effects of green tea polyphenol on the serum antioxidative activity and cholesterol levels of cholesterol-fed rats and compared them with those of probucol, an antioxidant hypocholesterolemic agent. To evaluate the antioxidative activity, the susceptibility to oxidative modification of low-density lipoprotein (LDL) isolated from the serum of cholesterol-fed rats was measured, as was the serum antioxidative activity using the spontaneous autoxidation system of brain homogenate. Administration of green tea polyphenol effectively inhibited LDL oxidation and elevated serum antioxidative activity to the same degree as probucol. However, higher amounts of polyphenol than probucol needed to be administered to reduce the total, free, and LDL cholesterol levels. Furthermore, green tea polyphenol increased the levels of high-density lipoprotein (HDL) cholesterol, leading to dose-dependent improvement of the atherogenic index, an effect that was not seen with probucol. Thus, green tea polyphenol may exert an antiatherosclerotic action by virtue of its antioxidant properties and by increasing HDL cholesterol levels.     

Antioxidative activities of volatile extracts from green tea, oolong tea, and black tea

Antioxidative activities of volatile extracts from six teas (one green tea, one oolong tea, one roasted green tea, and three black teas) were investigated using an aldehyde/carboxylic acid assay and a conjugated diene assay. The samples were tested at levels of 20, 50, 100, and 200 micrograms/mL of dichloromethane. The results obtained from the two assays were consistent. All extracts except roasted green tea exhibited dose-dependent inhibitory activity in the aldehyde/carboxylic acid assay. A volatile extract from green tea exhibited the most potent activity in both assays among the six extracts. It inhibited hexanal oxidation by almost 100% over 40 days at the level of 200 micrograms/mL. The extract from oolong tea inhibited hexanal oxidation by 50% in 15 days. In the case of the extract from roasted green tea, the lowest antioxidative activity was obtained at the level of 200 micrograms/mL, suggesting that the extract from roasted green tea contained some pro-oxidants. The extracts from the three black teas showed slight anti- or proactivities in both assays. The major volatile constituents of green tea and roasted green tea extracts, which exhibited significant antioxidative activities, were analyzed using gas chromatography-mass spectrometry. The major volatile chemicals with possible antioxidative activity identified were alkyl compounds with double bond(s), such as 3,7-dimethyl-1,6-octadien-3-ol (8.04 mg/kg), in the extract from green tea and heterocyclic compounds, such as furfural (7.67 mg/kg), in the extract from roasted green tea. Benzyl alcohol, which was proved to be an antioxidant, was identified both in a green tea extract (4.67 mg/kg) and in a roasted tea extract (1.35 mg/kg).     

Antioxidant activity of tocopherols, tocotrienols, and gamma-oryzanol components from rice bran against cholesterol oxidation accelerated by 2,2'-azobis(2-methylpropionamidine) dihydrochloride

The antioxidant activities of vitamin E (alpha-tocopherol, alpha-tocotrienol, gamma-tocopherol, and gamma-tocotrienol) and gamma-oryzanol components (cycloartenyl ferulate, 24-methylenecycloartanyl ferulate, and campesteryl ferulate) purified from rice bran were investigated in a cholesterol oxidation system accelerated by 2,2'-azobis(2-methylpropionamidine) dihydrochloride. All components exhibited significant antioxidant activity in the inhibition of cholesterol oxidation. The highest antioxidant activity was found for 24-methylenecycloartanyl ferulate, and all three gamma-oryzanol components had activities higher than that of any of the four vitamin E components. Because the quantity of gamma-oryzanol is up to 10 times higher than that of vitamin E in rice bran, gamma-oryzanol may be a more important antioxidant of rice bran in the reduction of cholesterol oxidation than vitamin E, which has been considered to be the major antioxidant in rice bran. The antioxidant function of these components against cholesterol oxidation may contribute to the potential hypocholesterolemic property of rice bran.     

Measuring antioxidant efficiency of wort, malt, and hops against the 2,2'-azobis(2-amidinopropane) dihydrochloride-induced oxidation of an aqueous dispersion of linoleic acid

This paper presents a simple, convenient method for determining the efficiency of antioxidants in aqueous systems. Production of conjugated diene hydroperoxide by oxidation of linoleic acid in an aqueous dispersion is monitored at 234 nm. 2, 2'-Azobis(2-amidinopropane) dihydrochloride is used as a free radical initiator. Among 12 antioxidants tested, phenolic compounds proved to be the most efficient, both kinetically and in terms of the inhibition time (T(inh)). Applied to wort, malt, and hops, the method confirmed a significant antioxidant activity in such products, especially hops. This assay can be used to follow oxidative changes throughout the brewing process and to understand the contribution of each raw material.     

Primary aminophospholipids in the external layer of liposomes protect their component polyunsaturated fatty acids from 2,2'-azobis(2-amidinopropane)- dihydrochloride-mediated lipid peroxidation

We showed in our previous study that docosahexaenoic acid-rich phosphatidylethanolamine in the external layer of small-size liposomes, as a model for biomembranes, protected its docosahexaenoic acid from 2,2'-azobis(2-amidinopropane)dihydrochloride- (AAPH-) mediated lipid peroxidation in vitro. Besides phosphatidylethanolamine, both phosphatidylserine and an alkenyl-acyl analogue of phosphatidylethanolamine, phosphatidylethanolamine plasmalogen, are reported to possess characteristic antioxidant activities. However, there are few reports about the relationship between the protective activity of phosphatidylethanolamine plasmalogen and/or phosphatidylserine against lipid peroxidation and their distribution in a phospholipid bilayer. Furthermore, it is unclear whether phosphatidylethanolamine plasmalogen and/or phosphatidylserine protect their component polyunsaturated fatty acids (PUFAs) from lipid peroxidation. In the present study, we examined the relationship between the transbilayer distribution of aminophospholipids, such as phosphatidylethanolamine rich in arachidonic acid, phosphatidylethanolamine plasmalogen, and phosphatidylserine, and the oxidative stability of their component PUFAs. The transbilayer distribution of these aminophospholipids in liposomes was modulated by coexisting phosphatidylcholine bearing two types of acyl chain: dipalmitoyl or dioleoyl. The amounts of these primary aminophospholipids in the external layer became significantly higher in liposomes containing dioleoylphosphatidylcholine than in those containing dipalmitoylphosphatidylcholine. Phosphatidylethanolamine rich in arachidonic acid, phosphatidylethanolamine plasmalogen or phosphatidylserine in the external layer of liposomes, as well as external docosahexaenoic acid-rich phosphatidylethanolamine, were able to protect their component PUFAs from AAPH-mediated lipid peroxidation.     

pH-Dependent radical scavenging capacity of green tea catechins

The effect of pH on the radical scavenging capacity of green tea catechins was investigated using experimental as well as theoretical methods. It was shown that the radical scavenging capacity of the catechins, quantified by the TEAC value, increases with increasing pH of the medium. Comparison of the pKa values to theoretically calculated parameters for the neutral and deprotonated forms indicates that the pH-dependent increase in radical scavenging activity of the catechins is due to an increase of electron-donating ability upon deprotonation. The data also reveal that the radical scavenging activity of the catechins containing the pyrogallol (or catechol) and the galloyl moiety over the whole pH range is due to an additive effect of these two independent radical scavenging structural elements. Altogether, the results obtained provide better insight into the factors determining the radical scavenging activity of the catechins and reveal that the biological activity of green tea catechins will be influenced by the pH of the surrounding medium or tissues.     

Immunostimulating activity of a crude polysaccharide derived from green tea (Camellia sinensis) extract

Green tea extract is well-known to reduce the risk of a variety of diseases. Here, we investigated the immunostimulating activity of tea polysaccharide (TPS), one of the main components in green tea extract. The water extracts from mature or immature tea leaves were precipitated by using ethanol at room temperature. The sediment was washed with ethanol and acetone alternately and then dried. We used the phagocytic activity of macrophage-like cells as an indicator of immune function activation. Chemical components were analyzed by HPLC. The immunostimulating activity of TPS from immature leaves extract was higher than that of TPS from mature leaves, and its activities were dependent on the content of strictinin in the leaf extract. Futhermore, a mixture of catechin and TPS that removed polyphenols did not increase the immunostimulating activity. These results suggest that the catechin-polysaccharide complex is a very important molecule in the immunomodulating activity of tea extracts.     

Anticarcinogenic activity of selenium-enriched green tea extracts in vivo

Both selenium and green tea have been shown to have potential antitumor effects. Here we have investigated the anticarcinogenic effect of the selenium-enriched green tea extract (Se-TE) in a Kunming mice model transplanted with human hepatoma cells HepG2. Mice were assigned to 8 groups consisting of 10 mice each after tumor cell inoculation. The control group received only water, whereas the remaining groups received regular green tea extract (RT), Se-TE which was produced by fertilization with selenite on tea leaves, selenite, and RT + selenite. After the mice were fed intragastrically with these agents for 8 days, tumor growth in RT-, Se-TE-, and selenite-fed mice was significantly suppressed, compared with that in control mice (P < 0.001). Supplementation with Se-TEs and selenite was able to elevate mice blood and liver Se concentrations, but did not significantly enhance selenoprotein glutathione peroxidase and other antioxidant enzyme superoxide dismutase activity in mice blood and liver. These results suggest that the antitumor function of Se-TEs may be attributed to the oxidative stress induced by selenium and green tea components in a suitable selenium supplementation pathway.     

Inhibition of gastrointestinal lipolysis by green tea, coffee, and gomchui (Ligularia fischeri) tea polyphenols during simulated digestion

Green tea, coffee, and gomchui (Ligularia fischeri) tea, which are rich in polyphenols, may exhibit antiobesity effects by inhibiting pancreatic lipase. However, the bioavailability of some polyphenols is poor due to either degradation or absorption difficulties in the gastrointestinal tract, thus making their beneficial effects doubtful. This study was conducted to evaluate the inhibitory effect of three beverages on lipolysis and the contribution of their major polyphenols during simulated digestion. During simulated digestion, gomchui tea was the most potent at inhibiting gastrointestinal lipolysis, whereas green tea was the least potent. The strongest lipase inhibitor among purified major polyphenols was a green tea polyphenol, (-)-epigallocatechin gallate (EGCG, IC(50) = 1.8 ± 0.57 μM), followed by di-O-caffeoylquinic acid isomers (DCQA, IC(50) from 12.7 ± 4.5 to 40.4 ± 2.3 μM), which are gomchui tea polyphenols. However, the stability of DCQA was greater than that of EGCG when subjected to simulated digestion. Taken together, gomchui tea, which has DCQA as the major polyphenol, showed stronger lipolysis inhibitory activity during simulated digestion compared to both green tea and coffee.     

Identification of potent odorants in Japanese green tea (Sen-cha)

Application of aroma extract dilution analysis using the volatile fraction of a Japanese green tea (Sen-cha) sample resulted in the detection of 36 odor-active peaks with flavor dilution (FD) factors between 10 and 5000. Thirty-six potent odorants were identified from 36 odor-active peaks by gas chromatography/mass spectrometry (GC/MS) and/or the multidimensional GC/MS (MDGC/MS) system. Among these components, 4-methoxy-2-methyl-2-butanethiol (meaty), (Z)-1, 5-octadien-3-one (metallic), 4-mercapto-4-methyl-2-pentanone (meaty), (E,E)-2,4-decadienal (fatty), beta-damascone (honey-like), beta-damascenone (honey-like), (Z)-methyl jasmonate (floral), and indole (animal-like) showed the highest FD factors. Therefore, these odorants were the most important components of the Japanese green tea odor. In addition, 4-methoxy-2-methyl-2-butanethiol, 4-mercapto-4-methyl-2-pentanone, methional, 2-ethyl-3, 5-dimethylpyrazine, (Z)-4-decenal, beta-damascone, maltol, 5-octanolide, 2-methoxy-4-vinylphenol, and 2-aminoacetophenone were newly identified compounds in the green tea.     

Antioxidative properties of cardoon (Cynara cardunculus L.) infusion against superoxide radical,hydroxyl radical,and hypochlorous acid

Polyphenols are able to act as antioxidants by virtue of their hydrogen-donating and metal-chelating capacities. Cardoon (Cynara cardunculus L.) is a species containing considerable amounts of polyphenolic compounds, namely flavonoids and phenolic acids. This study examined the antioxidant activity of cardoon lyophilized infusion against superoxide radical, hydroxyl radical, and hypochlorous acid. Superoxide radical was generated either in an enzymatic system or nonenzymatically, and the scavenging ability was assessed by the inhibition of superoxide radical-induced reduction of nitroblue tetrazolium. Hydroxyl radical was generated by the Fe3+-EDTA/ascorbate Fenton system, and scavenging capacity was estimated by evaluating the inhibition of hydroxyl radical-induced deoxyribose degradation into thiobarbituric acid-reactive substances. Inhibition of hypochlorous acid-induced 5-thio-2-nitrobenzoic acid oxidation to 5,5'-dithiobis(2-nitrobenzoic acid) was used in order to test the hypochlorous acid scavenging activity.     

Metabolic transformation of sesamol and ex vivo effect on 2,2'-azo-bis(2-amidinopropane)dihydrochloride-induced hemolysis

Sesamol (3,4-methylenedioxyphenol), a phenolic constituent in roasted sesame, was reported to exhibit various beneficial activities. To understand the metabolic transformation of sesamol in vivo, rats were given sesamol intravenously and orally. The blood samples were withdrawn via cardiopuncture at specific time points. The serum samples were assayed by high-performance liquid chromatography method before and after hydrolysis with sulfatase and beta-glucuronidase. Our results indicated that following either intravenous or oral administration, sesamol declined rapidly and the sulfate/glucuronide of sesamol emerged instantaneously. The peak serum concentration and systemic exposure of sesamol were markedly lower than sesamol sulfate/glucuronide. Ex vivo evaluation revealed that sesamol exerted profoundly higher capability against 2,2'-azo-bis(2-amidinopropane)dihydrochloride-induced hemolysis than the serum metabolites. In conclusion, sulfate and glucuronide of sesamol were the principle metabolites of sesamol in the bloodstream of rats. The conjugated metabolites of sesamol warrant more bioactivity investigations to understand the in vivo effect of sesamol.     

Modulation of cholesterol metabolism by the green tea polyphenol (-)-epigallocatechin gallate in cultured human liver (HepG2) cells

Epidemiological and animal studies have found that green tea is associated with lower plasma cholesterol. This study aimed to further elucidate how green tea modulates cholesterol metabolism. When HepG2 cells were incubated with the main green tea constituents, the catechins, epigallocatechin gallate (EGCG) was the only catechin to increase LDL receptor binding activity (3-fold) and protein (2.5-fold) above controls. EGCG increased the conversion of sterol regulatory element binding protein-1 (SREBP-1) to its active form (+56%) and lowered the cellular cholesterol concentration (-28%). At 50 microM, EGCG significantly lowered cellular cholesterol synthesis, explaining the reduction in cellular cholesterol. At 200 microM EGCG, cholesterol synthesis was significantly increased even though cellular cholesterol was lower, but there was a significant increase seen in medium cholesterol. This indicates that, at 200 microM, EGCG increases cellular cholesterol efflux. This study provides mechanisms by which green tea modulates cholesterol metabolism and indicates that EGCG might be its active constituent.     

Protective activity of green tea against free radical- and glucose-mediated protein damage

Protein oxidation and glycation are posttranslational modifications that are implicated in the pathological development of many age-related disease processes. This study investigated the effects of green tea extract, and a green tea tannin mixture and its components, on protein damage induced by 2,2'-azobis(2-amidinopropane) dihydrochloride (a free radical generator) and glucose in in vitro assay systems. We found that green tea extract can effectively protect against protein damage, and showed that its action is mainly due to tannin. In addition, it was shown that the chemical structures of tannin components are also involved in this activity, suggesting that the presence of the gallate group at the 3 position plays the most important role in the protective activity against protein oxidation and glycation, and that there is also a contribution by the hydroxyl group at the 5' position in the B ring and the sterical structure. These findings demonstrate the mechanisms of the usefulness of green tea in protein oxidation- and glycation-associated diseases.     

Enzymatic preparation of kaempferol from green tea seed and its antioxidant activity

Among the flavonols in green tea, kaempferol has many biological activities but kaempferol of plant origin is too expensive to be used in commercial products. Recently, we confirmed that green tea seed (GTS) contained a reasonable amount of kaempferol glycoside. After conducting structure analysis, two kaempferol glycosides were identified, kaempferol-3-O-[2-O-beta-D-galactopyranosyl-6-O-alpha-L-rhamnopyranosyl]-beta-D-glucopyranoside (compound 1) and kaempferol-3-O-[2-O-beta-D-xylopyranosyl-6-O-alpha-L-rhamnopyranosyl]-beta-D-glucopyranoside (compound 2), respectively. Also, a commercially useful method for kaempferol preparation was suggested by enzymatic hydrolysis using these two flavonoids. After several enzyme reactions were performed for the complete bioconversion of compounds 1 and 2 to kaempferol, we found that the optimum enzyme combination was reaction with beta-galactosidase and hesperidinase. Finally, we produced pure kaempferol with over 95% purity. We also compared the antioxidant effect of these two GTS flavonoids and its aglycone, kaempferol. Kaempferol is a more efficient scavenger of 1,1-diphenyl-2-picrylhydrazyl radicals and a better inhibitor of xanthine/xanthine oxidase than the two glycosides.     

Characterization of the constituents and antioxidant activity of Brazilian green tea (Camellia sinensis var. assamica IAC-259 cultivar) extracts

Freeze-dried extracts from Camellia sinensis var. assamica IAC-259 cultivar named Brazilian green tea were prepared by hot water and ultrasound-assisted extractions using leaves harvested in spring and summer. Their caffeine and catechin contents were measured by high performance liquid chromatography-diode array detector. The antioxidant activity of the major green tea compounds and Brazilian green tea extracts was evaluated using a 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay. The levels of caffeine were higher in the summer samples (p < 0.05); otherwise, there were no significant variations related to the catechin contents between spring and summer samples. The sonication method using water/acetone as solvent had a high efficiency to extract not only epigallocatechin gallate but also epicatechin gallate (p < 0.05). Antioxidant activities of the Brazilian green tea extracts were not significantly different among seasons and extraction systems. The antioxidant data (IC50) of the Brazilian green tea extracts showed a significant correlation with their epigallocatechin gallate and epicatechin gallate contents (p < 0.05).     

Effect of far-infrared irradiation on catechins and nitrite scavenging activity of green tea

The processed green tea leaves were irradiated by far-infrared (FIR) at eight temperatures (80, 90, 100, 110, 120, 130, 140, and 150 degrees C) for 10 min. After FIR irradiation, green teas were prepared by soaking the leaves in boiling water, and the physicochemical characteristics of the green tea were determined. FIR irradiation at 90 degrees C increased total phenol contents of green tea from 244.7 to 368.5 mg/g and total flavanol contents from 122.0 to 178.7 mg/g, compared with non-irradiated control. FIR irradiation also significantly affected the amounts of epigallocatechin and epigallocatechin gallate. Nitrite scavenging activity also increased with increasing FIR irradiation until the temperature reached 110 degrees C. However, the overall color changes of green tea irradiated with FIR at 90 and 100 degrees C were negligible. These results indicate that the chemical quality of green tea is significantly affected by FIR irradiation temperature of the green tea leaves.     

Kinetics of reduction of ferrylmyoglobin by (-)-epigallocatechin gallate and green tea extract

The hypervalent heme pigment ferrylmyoglobin, a potential prooxidant in muscle tissue and meat, is efficiently reduced by epigallocatechin gallate (EGCG) from green tea and by green tea polyphenol extract (GTP) in neutral or moderately acidic aqueous solution (0.16 M NaCl) to yield metmyoglobin in two parallel processes. The second-order rate constant for direct reduction at pH 7.4 and 25 degrees C was found to have the value 1170 +/- 83 M(-1).s(-1) and activation parameters DeltaH(#) = 70.6 +/- 7.2 kJ.mol(-1) and DeltaS(#) = 50.7 +/- 24.1 J.mol(-1).K(-1) for EGCG and the value 2300 +/- 77 M(-1).s(-1) and parameters DeltaH(#) = 60.6 +/- 2.6 kJ.mol(-1) and DeltaS(#) = 23 +/- 9 J.mol(-1).K(-1) for GTP (based on EGCG concentration). For decreasing pH, the rate increased moderately due to a parallel reduction of protonated ferrylmyoglobin. At physiological pH, EGCG is more efficient in deactivating ferrylmyoglobin than other plant phenols investigated, and the relatively high enthalpy and positive entropy of activation suggest an outer-sphere electron transfer mechanism. The interaction between EGCG and other tea catechins in GTP could be responsible for the even stronger ability for GTP to deactivate ferrylmyoglobin.     

Metabolite profiling using (1)H NMR spectroscopy for quality assessment of green tea, Camellia sinensis (L.)

A set of 191 green teas from different countries was collected and analyzed by (1)H NMR. It was proposed to establish if the teas could be discriminated according to the country of origin or with respect to quality. Both principal component analysis (PCA) and cluster analysis were applied to the data. Some separation of Chinese and non-Chinese teas was observed. The present results did not allow allocation of samples to individual countries, but cluster analysis suggested that it might be possible with an augmented sample set. The PCA did show a separation between the Longjing type (highest quality Chinese tea) and most other Chinese teas and indicated some metabolites that could be responsible for the difference. Longjing teas showed higher levels of theanine, gallic acid, caffeine, epigallocatechin gallate, and epicatechin gallate and lower levels of epigallocatechin when compared with other teas. These compounds have been mentioned previously in connection with quality, but it was also shown that higher levels of theogallin (5-galloyl quinic acid), theobromine, 2-O-(beta-l-arabinopyranosyl)-myo-inositol and some minor sugar-containing compounds were found in Longjing teas while higher levels of fatty acids and sucrose were found in the other teas. These new markers could prove to be useful for the authentication of bulk tea.     

Inhibition of pathogenic bacterial adhesion by acidic polysaccharide from green tea (Camellia sinensis)

An acidic polysaccharide CS-F2 from Camellia sinensis was examined to characterize its anti-adhesive effects against pathogenic bacteria, most notably Helicobacter pylori, Propionibacterium acnes, and Staphylococcus aureus. CS-F2 showed marked inhibitory activity against the pathogen-mediated hemagglutination with a minimum inhibitory concentration (MIC) between 0.01 and 0.1 mg/mL, which is lower than the previously reported MIC values for Panax ginseng and Artemisia capillaris. The inhibitory effects of CS-F2 on the adhesion of H. pylori to AGS adenocarcinoma gastric epithelial cells, or P. acnes and S. aureus to NIH 3T3 fibroblast cells, were further assessed resulting in MIC values between 0.063 and 0.13 mg/mL. Importantly, CS-F2 showed no inhibitory effects against Lactobacillus acidophilus, Escherichia coli, or Staphylococcus epidermidis. Our results suggest that CS-F2, which is a pectin-type polysaccharide with a molecular weight of approximately 8.0 x 10(4) Da, may exert a selective anti-adhesive effect against certain pathogenic bacteria, while exerting no effects against beneficial and commensal bacteria.