Antimutagenic activity of selenium-enriched green tea toward the heterocyclic amine 2-amino-3-methylimidazo[4,5-f]quinoline

Both selenium and green tea have been reported to exhibit antigenotoxic and cancer chemopreventive properties. We compared the antimutagenic activities of regular green tea and selenium-enriched green tea obtained from Hubei Province, China, toward the heterocyclic amine, 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) in the Salmonella assay. Selenium-enriched green tea obtained by foliar application of selenite exhibited concentration-dependent inhibition of IQ-induced mutagenesis in the presence of rat liver S9 and was significantly more effective than regular green tea tested under the same conditions. Analytical studies revealed no major differences in the polyphenol or caffeine content between regular green tea and selenium-enriched green tea, but the latter tea contained approximately 60-fold higher concentrations of selenium compared with regular green tea. The only soluble form of selenium was identified as selenite. The antimutagenic effects of certain individual tea constituents, such as epicatechin gallate and catechin, were enhanced by the addition of selenite to the Salmonella assay. Sodium selenite, sodium selenate, seleno-dl-cysteine, seleno-l-methionine, and l-Se-methylselenocysteine were not antimutagenic toward IQ when tested alone, but augmented significantly the inhibitory potency of green tea. The results suggested an enhancing (“coantimutagenic”) effect of selenium in combination with green tea in vitro, but in vivo studies are needed to assess whether there is a synergistic effect of tea and selenium to protect against heterocyclic amine-induced mutagenesis and carcinogenesis.     

Effect of flavan-3-ols on in vitro egg hatching, larval development and viability of infective larvae of Trichostrongylus colubriformis

The effects of flavan-3-ols (the monomer units of condensed tannins (CT)) and their galloyl derivatives on the viability of eggs, the development of first stage (L1) larvae, and the viability of the infective larvae of Trichostrongylus colubriformis were investigated under in vitro conditions. Each of the flavan-3-ol gallates showed some inhibition of egg hatching at 100 μg/ml, and 100% inhibition at 1000 μg/ml, with epigallocatechin gallate being the most effective in the egg hatch (EH) assay. In contrast, none of the flavan-3-ols were able to completely inhibit egg hatching. The flavan-3-ols and galloyl derivatives dose-dependently inhibited the development of infective larvae as assessed by the larval development (LD) assay. A larval migration inhibition (LMI) assay was used to assess the effect of flavan-3-ols and their galloyl derivatives on the motility of the infective third-stage (L3) larvae of T. colubriformis. In general, the flavan-3-ol gallates were more effective than the flavan-3-ols at immobilising the infective larvae as evidenced by their ability to inhibit more (P<0.05–0.01) larvae from passing through the LMI sieves. At 500 μg/ml, epigallocatechin gallate inhibited significantly more (P<0.1) larvae from passing through the sieves than did catechin gallate, epicatechin gallate, or gallocatechin gallate. Comparisons were made between the flavan-3-ols and their galloyl derivatives with the in vitro effects of CT extracts from several forage legumes, which have exhibited effects on parasites in vivo. The forage legumes tested at 200–500 μg/ml reduced the proportion of eggs that hatch, with comparable results to those obtained using the flavan-3-ols. The activities may be influenced by the prodelphinidin: procyanidin (PD:PC) ratios: CT extracts from Lotus pendunculatus and sainfoin have PD:PC ratios of 70:30 and 77:23, respectively, whereas the less active CT extract from Lotus corniculatus has a PD:PC ratio of 27:73. The active CT extracts from forage legumes have epigallocatechin as the dominant flavan-3-ol extender unit, and epigallocatechin is the most active flavan-3-ol in both the EH and LD assays.     

INVESTIGATION OF 4-METHYL STEROLS FROM CULTURED DINOFLAGELLATE ALGAL STRAINS

The marine dinoflagellates Prorocentrum micans, Gonyaulax polyedra, Gymnodinium sp., and Alexandrium tamarense, collected from the Adriatic Sea during red-tide blooms, were cultured to investigate the 4-methyl sterol constituents. To ascertain a possible influence of cell age on the 4-methyl sterol content, for one strain (Gymnodinium sp.)we investigated the composition of these constituents at exponential and stationary growing phases. The lipid material extracted with acetone from the lyophilized algal samples was fractionated by thin-layer chromatography. The 4-methyl sterols recovered from the layer were converted into the corresponding OTMS derivatives. Nine of 11 constituents were identified by gas chromatography and gas chromatography-mass spectrometry; only two minor constituents were characterized by their gas chromatographic parameters. All free methyl sterols identified in the algal samples had been detected previously in various dinoflagellates. The 4-methyl sterol fractions generally contained very few constituents. Except for the Gymnodinium sp. sample, collected at the exponential growing phase (GyD2 exp), which contains 4,24-dimethylcholestan-3-ol as a unique constituent, dinosterol was the major component. Moreover, 4,24-ethylcholestan-3-ol was also an important constituent of both Prorocentrum and Gonyaulax strains, whereas considerable amounts of dinostanol characterized all the Gymnodinium sp. strains. In addition, the latter contained several minor constituents such as 4-methylcholestan-3-ol, 4,24-dimethylcholesta-22-en-3-ol, and 4-methyl-24-ethylcholestan-3-ol. 4-Methyl-24-methylene-cholestan-3-ol was a constituent of the Gymnodinium sp. sample, collected at the stationary growing phase (GyD2 stat)only, whereas 4-methylgorgostanol was identified only in the Alexandrium tamarense Gt4 strain. Except for 4-methyl-24-ethylcholesta-8(14)-en-3-ol, all the methyl sterol constituents from our algae show a saturated polynuclear system. The pathways by which side-chain modifications occur in dinoflagellate 4-methyl sterols are considered, and a map of the fragmentation pattern of the trimethylsilyl-4-methyl sterols under electronic impact is also reported.     

Occurrence of Free Base of Phytosphingosine in Candida intermedia (IFO 0761)

The aroma concentrates from Vietnamese green tea and lotus tea were prepared and analyzed. Characterization of the components were carried out using coupled gas chromatography and mass spectrometry and infrared spectrometry, besides gas chromatographic retention data.

Anethole and 1,4-dimethoxybenzene have been identified for the first time as the flavor constituents in green tea. Linalool, two linalooloxides (cis and trans, five membered), 3,7-dimethyl-1,5,7-octatriene-3-ol, 2,5 (or 2,6)-dimethylpyrazine and 1-ethyl-2-formylpyrrole were the predominant components in Vietnamese green tea.

1,4-Dimethoxybenzene has been identified as the main component in lotus tea. The compound was also isolated from both dried and fresh lotus pollen.     

Anti-genotoxic effects of tea catechins against reactive oxygen species in human lymphoblastoid cells

Using the cytokinesis-block micronucleus assay in WIL2-NS cells, we investigated the effects of six tea constituents, (-)-epigallocatechin-3-O-gallate (EGCg), (-)-epicatechin-3-O-gallate (ECg), (-)-epigallocatechin (EGC), (-)-epicatechin (EC), (+)-catechin (+C) and gallic acid (GA), on chromosomal damage in two ways; induction by each component on its own and prevention against treatment of reactive oxygen species (ROS). None of the tea constituents induced chromosomal damage at <10 microM. On the other hand, EGCg, EGC, ECg, +C and GA prevented H(2)O(2)-induced chromosomal damage in a dose-dependent manner with a significant effect detected at 1 microM. Chromosomal damage induced by tert-butylhydroperoxide was apparently prevented by EGCg and ECg at 0.3 microM, but not by EGC and GA even at 10 microM, suggesting that the galloyl group linked to flavan-3-ol is needed for the observed protective effect. These results suggest that physiological concentration of tea constituents are not genotoxic but rather anti-genotoxic against ROS, although their preventive effects are slightly different depending on their chemical structure.     

Heterogeneity of the interflavanyl bond in proanthocyanidins from natural sources lacking C-4 (C-ring) deoxy flavonoid nucleophiles

The proanthocyanidin pool in the floral kingdom usually involves the presence of carbon-carbon bonds linking predominantly flavan-3-ol constituent moieties. Such an ensemble of flavan-3-ol units originates via electrophilic aromatic substitution of flavan-4-yl carbocations (or their equivalents) derived from flavan-4-ols and/or flavan-3,4-diols and the nucleophilic centers of the m-oxygenated A-rings of flavan-3-ol nucleophiles. In the absence of these potent flavan-3-ol nucleophiles with their aptitude for the formation of carbon-carbon bonds, alternative centers emerge as participants in interflavanyl bond formation. Such a phenomenon is demonstrated for the distribution of various profisetinidin-, prorobinetinidin-, proguibourtinidin-, promelacacinidin- and proteracacinidin-type pro- and leuco-anthocyanidins in several southern hemisphere heartwood species.     

Fractionation and chemistry of ethyl acetate-soluble thearubigins from black tea

Sephadex LH-20 chromatography was used to fractionate purified ethyl acetate-soluble thearubigins, prepared from an aqueous ethanolic extract of black tea. Three subfractions were so produced, each having a MW of about 1500 and each being degradable into cyanidin, delphinidin, gallic acid, the same two flavan-3-ols, and the same two flavan-3-ol gallates, though in different yield. Some evidence for the presence of benzotropolone moieties in at least one of the subfractions was obtained. Overall the ethyl acetate-soluble thearubigins are viewed as pentameric flavan-3-ols/flavan-3-ol gallates, containing both hydrolysable and non-hydrolysable interflavanoid links, as well as benzotropolone units, rather than as polymeric proanthocyanidins, a term previously used for all thearubigin subgroups.     

Three model systems measure oxidation/nitration damage caused by peroxynitrite

Diseases activate the innate immune response which causes ancillary damage to the human body. Peroxynitrite (OONO-) or its carbon dioxide derivatives cause oxidation/nitration and hence mutation to various body polymers e.g. DNA, RNA, protein, lipids and sugars. The control of the ancillary damage can come from antioxidants which inhibit control the amount of peroxynitrite available for damage. In this paper we have developed three different levels of antioxidant screening: (i) Peroxynitrite or SIN-1 reaction with luminol to produce light, and the inhibition of light by substances therefore represents antioxidation. (ii) Nicking of plasmid DNA occurs via oxidants: and is prevented by antioxidants. (iii) Detection of plasmid luciferase activity post-oxidation and infection indicates either prevention or repair of damage: via antioxidants. We found green tea and a number of its polyphenolic constituents effective only at the first level of antioxidation, while extracts of various fruit help at all levels antioxidation. In the final analysis, a combination of green tea extracts and fruits is suggested to produce more complete antioxidant protection.     

Tyrosinase catalysed biphenyl construction from flavan-3-ol substrates

Mushroom tyrosinase catalysed oxidation of three flavan-3-ols, viz. catechin, fisetinidol and mesquitol, was conducted to construct biphenyl bonds. Exposure of the flavan-3-ols to tyrosinase and subsequent trapping of the o-quinone intermediates resulted in the formation of novel flavan-3-ol derivatives, the structures of which were elucidated by mono- and two-dimensional 1H-NMR experiments. Application of the methodology resulted in the improved synthesis of the natural flavan-3-ol dimer, mesquitol-[5-->8]-catechin, previously isolated from Prosopis glandulosa.     

Synthesis and biological evaluation of flavan-3-ol derivatives as positive modulators of GABAA receptors

We herein describe the synthesis and positive modulatory activities of a small library of flavan-3-ol derivatives on α₁β₂γ_2L GABA_A receptors. Structure–activity relationships of various substituents on the A, B and C rings were evaluated in a functional electrophysiological assay. A trans configuration and a 3-acetoxy moiety are essential for activity. Substitution of the B ring appears to be well tolerated, with substituents on the A ring playing a major role in determining activity.     

Potent antimutagenic activity of white tea in comparison with green tea in the Salmonella assay.

There is growing interest in the potential health benefits of tea, including the antimutagenic properties. Four varieties of white tea, which represent the least processed form of tea, were shown to have marked antimutagenic activity in the Salmonella assay, particularly in the presence of S9. The most active of these teas, Exotica China white tea, was significantly more effective than Premium green tea (Dragonwell special grade) against 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) and four other heterocyclic amine mutagens, namely 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), 2-amino-3,4,8-trimethyl-3H-imidazo[4,5-f]quinoxaline (4,8-DiMeIQx), 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), and 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2). Mechanism studies were performed using rat liver S9 in assays for methoxyresorufin O-demethylase (MROD), a marker for the enzyme cytochrome P4501A2 that activates heterocyclic amines, as well as Salmonella assays with the direct-acting mutagen 2-hydroxyamino-3-methylimidazo[4,5-f]quinoline (N-hydroxy-IQ). White tea at low concentrations in the assay inhibited MROD activity, and attenuated the mutagenic activity of N-hydroxy-IQ in the absence of S9. Nine of the major constituents found in green tea also were detected in white tea, including high levels of epigallocatechin-3-gallate (EGCG) and several other polyphenols. When these major constituents were mixed to produce "artificial" teas, according to their relative levels in white and green teas, the complete tea exhibited higher antimutagenic potency compared with the corresponding artificial tea. The results suggest that the greater inhibitory potency of white versus green tea in the Salmonella assay might be related to the relative levels of the nine major constituents, perhaps acting synergistically with other (minor) constituents, to inhibit mutagen activation as well as "scavenging" the reactive intermediate(s).     

Cancer therapy and prevention by green tea: role of ornithine decarboxylase

Summary. Green tea which is widely consumed in China, Japan and India, contains polyphenolic compounds, which account for 30% of the dry weight of the leaves. Most of the polyphenols are flavanols, of which (−)-epigallocatechin-3-gallate (EGCG) is most abundant. Epidemiological studies revealed that the incidences of stomach and prostate cancers are the lowest in the world among a population that consumes green tea on a regular basis. It has also been reported that the quantity of green tea consumed, plays an important role in reducing cancer risk and in delaying cancer outbreak and recurrence. Various systems were used to confirm anti-cancer activities of green tea and/or EGCG. These included experimental animals in which cancer was induced chemically. Cultured cells transformed chemically or by oncogenes were also used. These studies clearly demonstrated that green tea or EGCG have anticancer and cancer preventive properties. The mechanisms of these activities have also been studied in details. It has been shown that green tea and its active components interfere with signal transduction pathways. Thus the activities of various protein kinases are inhibited, the expression of nuclear proto-oncogenes declines and the activity of ornithine decarboxylase (ODC) is reduced. ODC, which catalyzes the rate-limiting step in the biosynthesis of polyamines is closely linked with cellular proliferation and carcinogenesis. Inhibitors of ODC, like α-difluoromethylornithine (DFMO) have long been used for cancer prevention and therapy. It has been suggested that polyamine depletion by green tea could offer one explanation for its anti-cancer activities. Received July 27, 2001 Accepted September 8, 2001     

Tea polyphenols for health promotion

People have been consuming brewed tea from the leaves of the Camellia sinensis plant for almost 50 centuries. Although health benefits have been attributed to tea, especially green tea consumption since the beginning of its history, scientific investigations of this beverage and its constituents have been underway for less than three decades. Currently, tea, in the form of green or black tea, next to water, is the most widely consumed beverage in the world. In vitro and animal studies provide strong evidence that polyphenols derived from tea may possess the bioactivity to affect the pathogenesis of several chronic diseases. Among all tea polyphenols, epigallocatechin-3-gallate has been shown to be responsible for much of the health promoting ability of green tea. Tea and tea preparations have been shown to inhibit tumorigenesis in a variety of animal models of carcinogenesis. However, with increasing interest in the health promoting properties of tea and a significant rise in scientific investigation, this review covers recent findings on the medicinal properties and health benefits of tea with special reference to cancer and cardiovascular diseases.     

Green tea polyphenols: novel and potent inhibitors of squalene epoxidase

The green tea gallocatechins, (-)-epigallocatechin-3-O-gallate (EGCG) (IC(50) = 0.69 microM), (-)-gallocatechin-3-O-gallate (GCG) (IC(50) = 0.67 microM), (-)-epicatechin-3-O-gallate (ECG) (IC(50) = 1.3 microM), and theasinensin A (IC(50) = 0.13 microM), were found to be potent and selective inhibitors of rat squalene epoxidase (SE), a rate-limiting enzyme of cholesterol biogenesis. On the other hand, flavan-3-ols without galloyl group at C-3 did not show significant enzyme inhibition. It was demonstrated for the first time that the cholesterol lowering effect of green tea may be attributed to their potent SE inhibition activities. Inhibition kinetics revealed that EGCG inhibited SE in noncompetitive (K(I) = 0.74 microM), and non-time-dependent manner. The potent enzyme inhibition would be caused by specific binding to the enzyme, and by scavenging reactive oxygen species required for the monooxygenase reaction.     

Aroma Components Characteristic of Spring Green Tea

To investigate the aroma components characteristic of spring green tea, analysis of the aroma concentrates of green tea and fresh tea-leaves was accomplished. The research on the changes in aroma constituents of spring green tea and the aroma concentrate from fresh tea-leaves during storage, showed that cis-3-hexenylhexanoate and cis-3-hexenyl-trans-2-hexenoate contributed to the typical fresh aroma of spring green tea. Twelve isomers of hexenol esters were synthesized for comparison of the fresh green note.     

Characterization of new oxidation products of 9<Emphasis Type="Italic">H</Emphasis>-carbazole and structure related compounds by biphenyl-utilizing bacteria

9H-Carbazole and its derivatives are useful for versatile pharmacological applications. To obtain different derivatives of 9H-carbazole, 24 isolates of biphenyl-utilizing bacteria have been investigated regarding their ability to produce hydroxylated 9H-carbazole metabolites. Our analyses showed that 9H-carbazole was primarily converted into 9H-carbazol-1-ol (15 strains) and 9H-carbazol-3-ol (9 strains), while carbazol-9-ol was formed as a minor product (12 strains). The formation of 9H-carbazol-3-ol by the spontaneous release from the corresponding dihydrodiols was provided by the first-time detection of 3-hydroxy-1,2,3,9-tetrahydrocarbazol-4-one. The dependence of product yields on different parameters was exemplarily analyzed for Ralstonia sp. SBUG 290. Biphenyl-grown cells showed higher oxidation activities than cells cultivated with organic acids or nutrient broth, while co-cultivation of Ralstonia sp. SBUG 290 with biphenyl and 9H-carbazole led to an enhanced yield of 9H-carbazol-1-ol. The tested bacterial strains were also studied regarding their biotransformation of the two structure-related compounds 9H-fluorene and dibenzothiophene. Twenty-one strains primarily transformed 9H-fluorene into 9H-fluoren-9-ol and fluoren-9-one. Three strains accumulated benzo[c]chromen-6-one as a novel dead-end product during the incubation with 9H-fluorene, 9H-fluoren-9-ol, and fluoren-9-one. Dibenzothiophene has been mainly transformed into the dead-end product dibenzothiophene-5-oxide, while additional metabolites indicated that the transformation followed the so called Kodama pathway.     

Associations between flavan-3-ol intake and CVD risk in the Norfolk cohort of the European Prospective Investigation into Cancer (EPIC-Norfolk)

Dietary intervention studies suggest that flavan-3-ol intake can improve vascular function and reduce the risk of cardiovascular diseases (CVD). However, results from prospective studies failed to show a consistent beneficial effect. Associations between flavan-3-ol intake and CVD risk in the Norfolk arm of the European Prospective Investigation into Cancer and Nutrition (EPIC-Norfolk) were investigated. Data were available from 24,885 (11,252 men; 13,633 women) participants, recruited between 1993 and 1997 into the EPIC-Norfolk study. Flavan-3-ol intake was assessed using 7-day food diaries and the FLAVIOLA Flavanol Food Composition database. Missing data for plasma cholesterol and vitamin C were imputed using multiple imputation. Associations between flavan-3-ol intake and blood pressure at baseline were determined using linear regression models. Associations with CVD risk were estimated using Cox regression analyses. Median intake of total flavan-3-ols was 1034 mg/d (range: 0–8531 mg/d) for men and 970 mg/d (0–6695 mg/d) for women, median intake of flavan-3-ol monomers was 233 mg/d (0–3248 mg/d) for men and 217 (0–2712 mg/d) for women. There were no consistent associations between flavan-3-ol monomer intake and baseline systolic and diastolic blood pressure (BP). After 286,147 person-years of follow-up, there were 8463 cardiovascular events and 1987 CVD related deaths; no consistent association between flavan-3-ol intake and CVD risk (HR 0.93, 95% CI: 0.87; 1.00; Q1 vs Q5) or mortality was observed (HR 0.93, 95% CI: 0.84; 1.04). Flavan-3-ol intake in EPIC-Norfolk is not sufficient to achieve a statistically significant reduction in CVD risk.