Epicatechin-3-O-(3″-O-methyl)-gallate content in various tea cultivars (Camellia sinensis L.) and its in vitro inhibitory effect on histamine release

It has been reported that epigallocatechin-3-O-(3″-O-methyl)-gallate (EGCG3″Me) and the EGCG3″Me-rich green tea ( Camellia sinensis L.) cultivar 'Benifuuki' exhibit antiallergic effects. The objective of this study was to investigate the effect of various tea leaf catechins on histamine release from murine bone marrow mast cells (BMMC). At a dose of 50 μg/mL, the rank order of histamine release inhibition was observed to be epicatechin-3-O-(3″-O-methyl)-gallate (ECG3″Me) > gallocatechin-3-O-(3″-O-methyl)-gallate (GCG3″Me) > EGCG3″Me > gallocatechin-gallate (GCG) > catechin-gallate (CG) > EGCG > epicatechin-gallate (ECG) > epigallocatechin (EGC) > gallocatechin (GC). Of the various tea cultivars analyzed by HPLC, the greatest content of ECG3″Me was found in the third crop of 'Benifuuki' (1.05% dry weight). Moreover, ECG3″Me content was positively correlated with EGCG3″Me content in 'Benifuuki' tea leaves. In an assay of mixtures of ECG3″Me and EGCG3″Me, inhibitory activity (50 μg/mL in total) was increased as the content of ECG3″Me increased. This suggests that ECG3″Me might link to the antiallergic effect of 'Benifuuki' tea, as has been reported for EGCG3″Me.     

Determination of tea components with antioxidant activity

Levels of essential elements with antioxidant activity, as well as catechins, gallic acid, and caffeine levels, in a total of 45 samples of different teas commercialized in Spain have been evaluated. Chromium, manganese, selenium, and zinc were determined in the samples mineralized with HNO(3) and V(2)O(5), using ETAAS as the analytical technique. The reliability of the procedure was checked by analysis of a certified reference material. Large variations in the trace element composition of teas were observed. The levels ranged from 50.6 to 371.4 ng/g for Cr, from 76.1 to 987.6 microg/g for Mn, from 48.5 to 114.6 ng/g for Se, and from 56.3 to 78.6 ng/g for Zn. The four major catechins [(-)-epigallocatechin gallate (EGCG), (-)-epigallocatechin (EGC), (-)-epicatechin gallate (ECG), and (-)-epicatechin (EC)], gallic acid (GA), and caffeine were simultaneously determined by a simple and fast HPLC method using a photodiode array detector. In all analyzed samples, EGCG ranged from 1.4 to 103.5 mg/g, EGC from 3.9 to 45.3 mg/g, ECG from 0.2 to 45.6 mg/g, and EC ranged from 0.6 to 21.2 mg/g. These results indicated that green tea has a higher content of catechins than both oolong and fermented teas (red and black teas); the fermentation process during tea manufacturing reduces the levels of catechins significantly. Gallic acid content ranged from 0.039 to 6.7 mg/g; the fermentation process also elevated remarkably gallic acid levels in black teas (mean level of 3.9 +/- 1.5 mg/g). The amount of caffeine in the analyzed samples ranged from 7.5 to 86.6 mg/g, and the lower values were detected in green and oolong teas. This study will be useful for the appraisal of trace elements and antioxidant components in various teas, and it will also be of interest for people who like drinking this beverage.     

Metabolic dependence of green tea on plucking positions revisited: a metabolomic study

The dependence of global green tea metabolome on plucking positions was investigated through (1)H nuclear magnetic resonance (NMR) analysis coupled with multivariate statistical data set. Pattern recognition methods, such as principal component analysis (PCA) and orthogonal projection on latent structure-discriminant analysis (OPLS-DA), were employed for a finding metabolic discrimination among fresh green tea leaves plucked at different positions from young to old leaves. In addition to clear metabolic discrimination among green tea leaves, elevations in theanine, caffeine, and gallic acid levels but reductions in catechins, such as epicatechin (EC), epigallocatechin (EGC), epicatechin-3-gallate (ECG), and epigallocatechin-3-gallate (EGCG), glucose, and sucrose levels were observed, as the green tea plant grows up. On the other hand, the younger the green tea leaf is, the more theanine, caffeine, and gallic acid but the lesser catechins accumlated in the green tea leaf, revealing a reverse assocation between theanine and catechins levels due to incorporaton of theanine into catechins with growing up green tea plant. Moreover, as compared to the tea leaf, the observation of marked high levels of theanine and low levels of catechins in green tea stems exhibited a distinct tea plant metabolism between the tea leaf and the stem. This metabolomic approach highlights taking insight to global metabolic dependence of green tea leaf on plucking position, thereby providing distinct information on green tea production with specific tea quality.     

Degradation of green tea catechins in tea drinks

Green tea cateachins (GTC). namely (-) epicatechin (EC), (-) epicatechin gallate (ECG), (-) epigallocatechin (EGC), and (-) epigallocatechin gallate (EGCG), have been studied extensively for their wide-ranging biological activities. The goal of the present study was to examine the stability of GTC as a mixture under various processing conditions. The stability study demonstrated that GTC was stable in water at room temperature. When it was brewed at 98 degrees C for 7 h, longjing GTC degraded by 20%. When longjing GTC and pure EGCG were autoclaved at 120 degrees C for 20 min, the epimerization of EGCG to (-) gallocatechin gallate (GCG) was observed. The relatively high amount of GCG found in some tea drinks was most likely the epimerization product of EGCG during autoclaving. If other ingredients were absent, the GTC in aqueous solutions was pH-sensitive: the lower the pH, the more stable the GTC during storage. When it was added into commercially available soft drinks or sucrose solutions containing citric acid and ascorbic acid, longjing GTC exhibited varying stability irrespective of low pH value. This suggested that other ingredients used in production of tea drinks might interact with GTC and affect its stability. When canned and bottled tea drinks are produced, stored, and transported, the degradation of GTC must be taken into consideration.     

茶叶添加方式对酿造啤酒理化特性及感官品质的影响

在啤酒酿造工艺的煮沸和主发酵阶段分别添加乌龙茶,探讨茶叶不同添加方式对啤酒理化特性、抗氧化能力、啤酒贮存稳定性以及感官特性的影响。结果显示,与未添加茶叶的啤酒产品(对照组CG)相比,在煮沸阶段(煮沸添加组BG)和主发酵阶段(主发酵添加组MG)分别添加0.3g/L的茶叶均提高了酵母发酵速率;添加茶叶的两个茶啤酒产品(煮沸添加组和主发酵添加组)的DPPH自由基清除能力分别提升至82.74%和89.21%、ABTS+自由基清除能力分别提升至41.53%和51.49%、铁离子还原能力分别提升至36.49和43.83 mg/L,且成品茶啤酒贮藏期间的抗老化能力提高;茶啤酒中总酚以及儿茶素(EGC、C、EGCG、EC、GCG、ECG)和咖啡碱(CAF)含量升高,其中,主发酵添加组茶啤酒样品总酚和EGCG含量显著高于对照组和煮沸添加组啤酒(P0.05),分别达到734.40和8.43 mg/L。进一步的感官品评结果显示添加茶叶提高了啤酒产品的茶香气和茶滋味,其中主发酵添加组啤酒的茶香气、茶滋味及酒体协调性最好。由此可知,在啤酒酿造工艺中添加茶叶提高了酵母发酵速率,增强了啤酒中酚类物质含量及其抗氧化和抗老化性能,同时也为啤酒增添了新的茶风味产品。     

β-carotene radical cation addition to green tea polyphenols. Mechanism of antioxidant antagonism in peroxidizing liposomes

Green tea polyphenols, (-)-epicatechin (EC), (-)-epigallocatechin (EGC), (-)-epicatechin gallate (ECG), and (-)-epigallocatechin gallate (EGCG), all showed antioxidative effect in liposomes for lipid oxidation initiated in the lipid phase (antioxidant efficiency EC > EGCG > ECG > EGC) or in the aqueous phase (EC ? EGC > EGCG > ECG) as monitored by the formation of conjugated dienes. For initiation in the lipid phase, β-carotene, itself active as an antioxidant, showed antagonism with the polyphenols (EC > ECG > EGCG > EGC). The Trolox equivalent antioxidant capacity (TEAC EGC > EGCG > ECG > EC) correlates with the lowest phenol O-H bond dissociation enthalpy (BDE) as calculated by density functional theory (DFT). Surface-enhanced Raman spectroscopy (SERS) was used to assess the reducing power of the phenolic hydroxyls in corroboration with DFT calculations. For homogeneous (1:9 v/v methanol/chloroform) solution, the β-carotene radical cation reacted readily with each of the polyphenol monoanions (but not with the neutral polyphenols) with a rate approaching the diffusion limit for EC as studied by laser flash photolysis at 25 °C monitoring the radical cation at 950 nm. The rate constant did not correlate with polyphenol HOMO/LUMO energy gap (DFT calculations), and β-carotene was not regenerated by an electron transfer reaction (monitored at 500 nm). It is suggested that the β-carotene radical cation is rather reacting with the tea polyphenols through addition, as further evidenced by steady-state absorption spectroscopy and liquid chromatography-mass spectroscopy (LC-MS), in effect preventing regeneration of β-carotene as an active lipid phase antioxidant and leading to the observed antagonism.     

Stopped-flow kinetic study of the aroxyl radical-scavenging action of catechins and vitamin C in ethanol and micellar solutions

Kinetic study of the aroxyl radical-scavenging action of catechins (epicatechin (EC), epicatechin gallate (ECG), epigallocatechin (EGC), and epigallocatechin gallate (EGCG)) and related compounds (methyl gallate (MG), 4-methylcatechol (MC), and 5-methoxyresorcinol (MR)) has been performed. The second-order rate constant ( k s) for the reaction of these antioxidants with aroxyl radical has been measured in ethanol and aqueous Triton X-100 micellar solution (5.0 wt %). The k s values decreased in the order of EGCG > EGC > MC > ECG > EC > MG > MR in ethanol, indicating that the reactivity of the OH groups in catechins decreased in the order of pyrogallol B-ring > catechol B-ring > gallate G-ring > resorcinol A-ring. The structure-activity relationship in the free radical-scavenging reaction by catechins has been clarified by the detailed analyses of the pH dependence of k s values. From the results, the p K a values have been determined for catechins. The monoanion form at catechol B- and resorcinol A-rings and dianion form at pyrogallol B- and gallate G-rings show the highest activity for free radical scavenging. It was found that the free radical-scavenging activities of catechins are 3.2-13 times larger than that of vitamin C at pH 7.0.     

Effects of heat processing and storage on flavanols and sensory qualities of green tea beverage

This research was conducted to understand the effects of heat processing and storage on flavanols and sensory qualities of green tea extract. Fresh tea leaves were processed into steamed and roasted green teas by commercial methods and then extracted with hot water (80 degrees C) at 1:160 ratio (tea leaves/water by weight). Green tea extracts were heat processed at 121 degrees C for 1 min and then stored at 50 degrees C to accelerate chemical reactions. Changes in flavanol composition and sensory qualities of green tea extracts during processing and storage were measured. Eight major flavanols (catechin, epicatechin, gallocatechin, epigallocatechin, epicatechin gallate, catechin gallate, epigallocatechin gallate, and gallocatechin gallate) were identified in the processed tea extract. Among them, epigallocatechin gallate and epigallocatechin appeared to play the key role in the changes of sensory qualities of processed green tea beverage. The steamed tea leaves produced a more desirable quality of processed green tea beverage than the roasted ones.     

Stability of tea catechins in the breadmaking process

A green tea extract (GTE) was incorporated into bread as a source of tea catechins. The stability of tea catechins in the breadmaking process including unfrozen and frozen dough was studied. A method was developed for the separation and quantification of tea catechins in GTE, dough, and bread samples using a RP-HPLC system. The separation system consisted of a C18 reversed-phase column, a gradient elution system of water/methanol and formic acid, and a photodiode array UV detector. Tea catechins were detected at 275 nm. GTEs at 50, 100, and 150 mg per 100 g of flour were formulated. The results obtained showed that green tea catechins were relatively stable in dough during freezing and frozen storage at -20 degrees C for up to 9 weeks. There were no further detectable losses of tea catechins in bread during a storage of 4 days at room temperature. It was also revealed that (-)-epigallocatechin gallate (EGCG) and (-)-epigallocatechin (EGC) were more susceptible to degradation than (-)-epicatechin gallate (ECG) and (-)-epicatechin (EC). (-)-EGCG and (-)-ECG were normally selected as the quality indices of green tea catechins, and their retention levels in freshly baked bread were ca. 83 and 91%, respectively. One piece of bread (53 g) containing 150 mg of GTE/100 g of flour will provide 28 mg of tea catechins, which is approximately 35% of those infused from one green tea bag (2 g).     

基于高光谱成像的茶叶中EGCG分布可视化

针对目前关于表没食子儿茶素没食子酸酯(epigallocatechin gallate,EGCG)在茶叶中的分布缺乏可视化表达问题,该文采用高光谱成像技术以实现EGCG在茶叶中的分布可视化。通过高光谱成像仪采集茶叶的光谱信息,按照标准方法 HPLC(high performance liquid chromatography)法测量茶叶的EGCG浓度。运用化学计量学方法建立光谱与EGCG浓度之间的回归模型。为寻求相对较优的模型效果,对光谱进行不同的预处理,以确定最优的预处理方法;采用4种建模方法建立回归模型,以确定最优的建模方法;对光谱进行特征波段选择,以降低数据冗余提高模型的稳定性和运算速度。最后,将高光谱图像中像素点对应的光谱变量导入模型,从而生成EGCG浓度分布图。结果表明:可见-近红外光谱与EGCG浓度之间具有很强的相关性,其回归模型的决定系数达到0.905,利用高光谱成像技术对茶叶中EGCG分布进行可视化可行。通过对不同品种、叶位的茶叶中EGCG浓度分布进行可视化,能够为高EGCG浓度茶树品种的培育、EGCG代谢规律的分析以及茶树采摘部位的识别提供有效手段。     

The apoptotic effect of green tea (-)-epigallocatechin gallate on 3T3-L1 preadipocytes depends on the Cdk2 pathway

This study was designed to investigate the effect of green tea catechins, especially (-)-epigallocatechin gallate (EGCG), on the apoptosis of 3T3-L1 preadipocytes. Preadipocyte apoptosis as indicated by formation of DNA fragments was induced by EGCG in dose-dependent manners. While EGCG was demonstrated to decrease Cdk2 expression and activity and increase caspase-3 activity, overexpression of Cdk2 and treatment with the caspase-3 inhibitor respectively prevented preadipocytes from induction of DNA fragmentation and caspase-3 activity by doses of 100-400 muM of EGCG. This suggests the Cdk2- and caspase-3-dependent apoptotic effects of EGCG. Moreover, EGCG was more effective than EC, ECG, and EGC in changing the apoptotic signals. Results of this study may relate to the mechanism by which EGCG modulates body weight.     

A new LC/MS/MS rapid and sensitive method for the determination of green tea catechins and their metabolites in biological samples

A new rapid and sensitive method has been developed, using liquid chromatography in tandem mass spectrometry (LC-ESI-MS/MS) to identify green tea catechin metabolites in plasma and urine after oral intake of a green tea extract. (-)-Epigallocatechin-3-gallate (EGCG), (-)-epicatechin-3-gallate (ECG), (-)-epigallocatechin (EGC)-glucuronide, (-)-epicatechin (EC)-glucuronide, and EC-sulfate were identified in plasma, whereas in urine only the conjugated catechins were detected (EGC-glucuronide, EGC-sulfate, EC-glucuronide, and EC-sulfate). Standard calibration curves prepared in plasma were found to be linear in the range of 10.9-1379.3 nmol/L for EGCG, EGC, ECG, and EC. The accuracy and precision of this assay showed a coefficient of variation of <15%. The method allowed the detection and quantification limits (for 20 microL injection) from 1.1 to 2.6 nmol/L and 3.8-8.7 nmol/L, respectively, in plasma and 0.8-1.8 nmol/L and 2.6-6.0 nmol/L, respectively, in urine. This method can be applied for future clinical and epidemiological studies, allowing the identification of the active metabolites that will reach the target tissues.     

Catechin and caffeine content of green tea dietary supplements and correlation with antioxidant capacity

The health benefits associated with tea consumption have resulted in the wide inclusion of green tea extracts in botanical dietary supplements, which are widely consumed as adjuvants for complementary and alternative medicines. Tea contains polyphenols such as catechins or flavan-3-ols including epicatechin, epigallocatechin, epicatechin gallate, and epigallocatechin gallate (EGCG), as well as the alkaloid, caffeine. Polyphenols are antioxidants, and EGCG, due to its high levels, is widely accepted as the major antioxidant in green tea. Therefore, commercial green tea dietary supplements (GTDS) may be chemically standardized to EGCG levels and/or biologically standardized to antioxidant capacity. However, label claims on GTDS may not correlate with actual phytochemical content or antioxidant capacity nor provide information about the presence and levels of caffeine. In the current study, 19 commonly available GTDS were evaluated for catechin and caffeine content (using high-performance liquid chromatography) and for antioxidative activity [using trolox equivalent antioxidant capacity (TEAC) and oxygen radical antioxidant capacity (ORAC) assays]. Product labels varied in the information provided and were inconsistent with actual phytochemical contents. Only seven of the GTDS studied made label claims of caffeine content, 11 made claims of EGCG content, and five specified total polyphenol content. Caffeine, EGCG, and total polyphenol contents in the GTDS varied from 28 to 183, 12-143, and 14-36% tablet or capsule weight, respectively. TEAC and ORAC values for GTDS ranged from 187 to 15340 and from 166 to 13690 mumol Trolox/g for tablet or capsule, respectively. The antioxidant activities for GTDS determined by TEAC and ORAC were well-correlated with each other and with the total polyphenol content. Reliable labeling information and standardized manufacturing practices, based on both chemical standardization and biological assays, are recommended for the quality control of botanical dietary supplements.     

Electrofocusing of methanolic extracts for identification of individual flavonol biomolecules in Camellia species

An effort has been made to isolate individual catechin compounds from green tea leaves in their pure form by electrophoresis. In the present study total polyphenol extraction was carried out initially and estimated through spectrophotometric and HPLC methods. Extracted polyphenol was separated on 0.7% agarose gel and visualized at 360 nm. Fragmented individual compounds were gel eluted with methanol and confirmed as (-)-epigallocatechin (EGC), (-)-epicatechin (EC), (-)-epicatechin gallate (ECG), and (-)-epigallocatechin gallate (EGCG) by HPLC. The method developed describes a suitable method for the isolation of valuable molecules in tea.     

HPLC analysis of naturally occurring methylated catechins, 3' '- and 4' '-methyl-epigallocatechin gallate, in various fresh tea leaves and commercial teas and their potent inhibitory effects on inducible nitric oxide synthase in macrophages

(-)-Epigallocatechin-3-gallate (EGCG), a major polyphenol of green tea, undergoes substantial biotransformation to species that includes the methylated compounds. Recent studies have demonstrated that the methylated EGCG has many biological activities. In this study, we have investigated the composition of the three O-methylated EGCG derivatives, (-)-epigallocatechin-3-O-(3-O-methyl)gallate (3' '-Me-EGCG), (-)-epigallocatechin-3-O-(4-O-methyl)gallate (4' '-Me-EGCG) and (-)-4'-methyl epigallocatechin-3-O-(4-O-methyl)gallate (4',4' '-di-Me-EGCG) in tea leaves which were picked from various species and at various seasons, ages of leaves, locations, and fermentation levels. Higher levels of 3' '-Me-EGCG and 4' '-Me-EGCG were detected in Chinshin-Kanzai (a species of Camellia sinensis) cultivated in the mountain area of Sansia, Taipei County, Taiwan. Also, these O-methylated EGCG levels were found to be higher in autumn and winter than in spring and summer. The young leaves were found to be richer in the O-methylated compounds than old leaves and the amount of O-methylated EGCG was higher in unfermented longjin green tea than in semifermented oolong tea. However, the fermented black tea and puerh tea did not contain these compounds. 4',4' '-diMe-EGCG could not be detected in either fresh tea leaves or commercial tea leaves. We also found that 3' '-Me-EGCG has a higher inhibitory effect on the nitric oxide generation and inducible nitric oxide synthase (iNOS) expression as compared with EGCG, while 4' '-Me-EGCG and 4',4' '-di-Me-EGCG were less effective.     

Total phenol, catechin, and caffeine contents of teas commonly consumed in the United kingdom.

Levels of total phenol, catechins, and caffeine in teas commonly consumed in the United Kingdom have been determined using reversed phase high-performance liquid chromatography. Tea bags or tea leaves were purchased from local supermarkets and extracted in boiling water for 5 min. The resulting data showed considerable variability in both total phenols [80.5-134.9 mg/g of dry matter (DM) in black teas and 87-106.2 mg/g of DM in green teas] and catechins (5.6-47.5, 51.5-84.3, and 8.5-13.9 mg/g of DM in black, green, and fruit teas, respectively); this was most probably a result of differing agronomic conditions, leaf age, and storage during and after transport, as well as the degree of fermentation. Caffeine contents of black teas (22-28 mg/g of DM) were significantly higher than in less fermented green teas (11-20 mg/g of DM). The relative concentration of the five major tea catechins ranked EGCG > ECG > EC > EGC > C. The estimated U.K. dietary intakes of total tea catechins, calculated on the basis of an average tea consumption of three cups of tea (200 mL cup, 1% tea leaves w/v), were 61.5, 92.7, and 405.5 mg/day from fruit teas, black teas, and green teas, respectively. The coefficients of variation were 19.4, 88.6, and 17.3%, respectively, indicating the wide variation in these intakes. The calculated caffeine intake ranged between 92 and 146 mg/day. In addition, many individuals will consume much larger quantities of tea, of various strengths (as determined by the brewing conditions employed). This broad spread of U.K. daily intakes further emphasizes the need for additional research to relate intake and effect in various population groups.     

Inhibitory effects of tea catechins and O-methylated derivatives of (-)-epigallocatechin-3-O-gallate on mouse type IV allergy

The inhibitory effects of tea catechins, the O-methylated derivatives of (-)-epigallocatechin-3-O-gallate (EGCG), and the polyphenol extracts from tea leaves (Camellia sinensis L.) on oxazolone-induced type IV allergy in male ICR mice were investigated. Four major tea catechins and two O-methylated derivatives, (-)-epigallocatechin-3-O-(3-O-methyl)gallate (EGCG3' 'Me) and (-)-epigallocatechin-3-O-(4-O-methyl)gallate (EGCG4' 'Me), showed significant inhibitory effects on mouse type IV allergy after a percutaneous administration at a dose of 0.13 mg/ear. Among tea catechins, the compounds including galloyl moieties, such as EGCG and (-)-epicatechin-3-O-gallate (ECG), showed the strongest inhibitory activities on mouse type IV allergy. The inhibitory activities of EGCG3' 'Me and EGCG4' 'Me were higher than that of EGCG at a dose of 0.05 mg/ear. Polyphenol extract from tea leaves of Benihomare cultivar, which includes EGCG3' 'Me, strongly inhibited mouse type IV allergy after percutaneous administration in comparison with that from Yabukita cultivar, which does not include EGCG3' 'Me, at doses of 0.05 and 0.13 mg/ear. EGCG3' 'Me is thought to contribute, at least in part, to the inhibitory ability of Benihomare tea leaves on mouse type IV allergy. EGCG and the polyphenol extracts from Benihomare and Yabukita tea leaves also inhibited mouse type IV allergy by oral administration at 1 h before the sensitization and at 1 h before the challenge with oxazolone. Therefore, daily intake of tea drinks could have potential to prevent type IV allergy.     

Identification of oxidation products of (-)-epigallocatechin gallate and (-)-epigallocatechin with H(2)O(2)

(-)-Epigallocatechin gallate (EGCG) and (-)-epigallocatechin (EGC) are two important antioxidants in tea. They also display some antitumor activities, and these activities are believed to be mainly due to their antioxidative effects. However, the specific mechanisms of antioxidant action of tea catechins remain unclear. In this study are isolated and identified two novel reaction products of EGCG and one product of EGC when they were reacted separately with H(2)O(2). These products are formed by the oxidation and decarboxylation of the A ring in the catechin molecule. This study provides unequivocal proof that the A ring of EGCG and EGC may also be an antioxidant site. This study also indicates an additional reaction pathway for the oxidation chemistry of tea catechins.     

Different effects of five catechins on 6-hydroxydopamine-induced apoptosis in PC12 cells.

Five catechins [(-)-epigallocatechins gallate (EGCG), (-)-epicatechin gallate (ECG), (-)-epigallocatechin (EGC), (-)-epicatechin (EC), and (+)-catechin (C)] were compared with regard to their effects on 6-hydroxydopamine (OHDA)-induced apoptosis in PC12 cells--the vitro model of Parkinson's disease. Measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, 6-OHDA inhibited cell viability in a time- and concentration-dependent manner. When PC12 cells were pretreated with the five catechins for 30 min before exposure to 250 microM 6-OHDA, MTT results showed that the five catechins had different effects: EGCG and ECG had obvious concentration-dependent protective effects at 50-400 microM; EC and (+)-C had almost no effects; and EGC especially decreased cell viability. Catechins also had different effects on apoptotic morphology. Only 200-400 microM EGCG and ECG kept cells adhering well. When pretreated with other catechins at any concentration, PC12 cells became round and some of them were detached as when treated with 6-OHDA. In addition, typical apoptotic characteristics of PC12 cells were determined by fluorescence microscopy, flow cytometry, and DNA fragment electrophoresis after the cells were treated with 250 microM 6-OHDA for 24 h or pretreated with catechins before it. Preincubation with 200-400 microM EGCG and ECG led to significant inhibitory effects against PC12 cell apoptosis, as shown by flow cytometry. The other catechins have little protective effect. Therefore, at 200-400 microM, the classified protective effects of the five catechins were in the order ECG > EGCG > EC > (+)-C > EGC. The data also indicated that EGCG and ECG might be potent neuroprotective agents for Parkinson's disease. The results of fluorescence microscopy and DNA fragment analysis supported the conclusion.     

Catechin degradation with concurrent formation of homo- and heterocatechin dimers during in vitro digestion

Catechins were subjected to in vitro gastric and small intestinal digestion. EGCG, EGC, and ECG were significantly degraded at all concentrations tested, with losses of 71-91, 72-100, and 60-61%, respectively. EC and C were comparatively stable, with losses of 8-11 and 7-8%, respectively. HLPC-ESI-MS/MS indicated that EGCG degradation under simulated digestion resulted in production of theasinensins (THSNs) A and D (m/z 913) and P-2 (m/z 883), its autoxidation homodimers. EGC dimerization produced the homodimers THSN C and E (m/z 609) and homodimers analogous to P-2 (m/z 579). ECG homodimers were not observed. EGCG and EGC formed heterodimers analogous to the THSNs (m/z 761) and P-2 (m/z 731). EGCG and ECG formed homodimers analogous to the THSNs (m/z 897). This study provides an expanded profile of catechin dimers of digestive origin that may potentially form following consumption of catechins. These data provide a logical basis for initial screening to detect catechin digestive products in vivo.