A metabonomic strategy for the detection of the metabolic effects of chamomile (Matricaria recutita L.) ingestion

A metabonomic strategy, utilizing high-resolution 1H NMR spectroscopy in conjunction with chemometric methods (discriminant analysis with orthogonal signal correction), has been applied to the study of human biological responses to chamomile tea ingestion. Daily urine samples were collected from volunteers during a 6-week period incorporating a 2-week baseline period, 2 weeks of daily chamomile tea ingestion, and a 2-week post-treatment phase. Although strong intersubject variation in metabolite profiles was observed, clear differentiation between the samples obtained before and after chamomile ingestion was achieved on the basis of increased urinary excretion of hippurate and glycine with depleted creatinine concentration. Samples obtained up to 2 weeks after daily chamomile intake formed an isolated cluster in the discriminant analysis map, from which it was inferred that the metabolic effects of chamomile ingestion were prolonged during the 2-week postdosing period. This study highlights the potential for metabonomic technology in the assessment of nutritional interventions, despite the high degree of variation from genetic and environmental sources.     

Characterization of tea cultivated at four different altitudes using 1H NMR analysis coupled with multivariate statistics

The taste of black tea differs according to the different areas in which the tea is grown, even for the same species of tea. A combination of (1)H NMR spectroscopy and partial least-squares discriminate analysis (PLS-DA) was used to assess the quality differences of tea leaves from four cultivation areas with different elevations, RAN > 1800 m, UDA = 1200 m, MEDA = 600 m, and YATA < 300 m, in Sri Lanka. As a result of a statistical analysis, PLS-DA showed a separation between high- and low-quality black teas derived from the four different tea cultivation areas. RAN from the highest elevation showed characteristic trends in the levels of theaflavin and theaflavin 3,3'-digallate that were found only in RAN, and the levels of theanine and caffeine were higher, and the levels of thearubigins, especially thearubigin 3,3'-digallate, were lower in RAN than in UDA, MEDA, and YATA. The structures of these components were determined by 1D and 2D NMR analyses. These results demonstrate that this method can be used to evaluate black tea quality according to the chemical composition or metabolites, which are characteristic of the tea leaves cultivated in four regions with different elevations in Sri Lanka.     

Discrimination of three Pegaga (Centella) varieties and determination of growth-lighting effects on metabolites content based on the chemometry of 1H nuclear magnetic resonance spectroscopy

The metabolites of three species of Apiaceae, also known as Pegaga, were analyzed utilizing (1)H NMR spectroscopy and multivariate data analysis. Principal component analysis (PCA) and hierarchical cluster analysis (HCA) resolved the species, Centella asiatica, Hydrocotyle bonariensis, and Hydrocotyle sibthorpioides, into three clusters. The saponins, asiaticoside and madecassoside, along with chlorogenic acids were the metabolites that contributed most to the separation. Furthermore, the effects of growth-lighting condition to metabolite contents were also investigated. The extracts of C. asiatica grown in full-day light exposure exhibited a stronger radical scavenging activity and contained more triterpenes (asiaticoside and madecassoside), flavonoids, and chlorogenic acids as compared to plants grown in 50% shade. This study established the potential of using a combination of (1)H NMR spectroscopy and multivariate data analyses in differentiating three closely related species and the effects of growth lighting, based on their metabolite contents and identification of the markers contributing to their differences.     

Metabonomics approach to determine metabolic differences between green tea and black tea consumption

The purpose of this study was to compare the effects of black and green tea consumption on human metabolism. Seventeen healthy male volunteers consumed black tea, green tea, or caffeine in a randomized crossover study. Twenty-four-hour urine and blood plasma samples were analyzed by NMR-based metabonomics, that is, high-resolution 1H NMR metabolic profiling combined with multivariate statistics. Green and black tea consumption resulted in similar increases in urinary excretion of hippuric acid and 1,3-dihydroxyphenyl-2-O-sulfate, both of which are end products of tea flavonoid degradation by colonic bacteria. Several unidentified aromatic metabolites were detected in urine specifically after green tea intake. Interestingly, green and black tea intake also had a different impact on endogenous metabolites in urine and plasma. Green tea intake caused a stronger increase in urinary excretion of several citric acid cycle intermediates, which suggests an effect of green tea flavanols on human oxidative energy metabolism and/or biosynthetic pathways.     

NMR-based metabonomic studies on the biochemical effects of epicatechin in the rat

Flavonoid consumption via tea drinking has been attributed a number of potential health benefits including cancer prevention, anti-inflammatory action, and cardioprotectant activity. Although the predominant flavonoids in fresh leaf and green tea are known to be flavan-3-ols and flavan-3-O-gallates ("the catechins"), the biochemical effects of tea polyphenol consumption on living systems are generally poorly understood. Metabonomic methods utilizing (1)H NMR spectroscopy of biofluids and principal component analysis (PCA) have been applied to investigate the bioavailability and metabolic responses of rats to a single dose of 22 mg of epicatechin (EC) dissolved in water. Urine samples were collected twice daily (0-8 and 8-24 h) from male Sprague-Dawley rats (n = 10) prior to dosing and for 2 days after dosing. A series of subtle urinary biochemical effects were evident from the (1)H NMR spectra showing that EC was both bioavailable and biochemically active. The identifiable biochemical effects associated with EC dosing included decreased urinary concentrations of taurine, citrate, dimethylamine, and 2-oxoglutarate. These effects were predominately seen within the first 8 h after dosing. EC metabolites were also observed in the urine during this time period. PCA of later time points after dosing (24-32 and 32-48 h) showed that the effects of EC were reversible. This is the first in vivo study demonstrating the overall endogenous metabolic effects of EC consumption and shows the bioavailability of EC via metabolic effects and excretion of EC metabolites.     

Preparative separation of polyphenols from tea by high-speed countercurrent chromatography

High-speed countercurrent chromatography (HSCCC) was applied to the separation of polyphenols from tea leaves (Camellia sinensis L.). The capability of HSCCC to isolate pure tea polyphenols from complex mixtures on a preparative scale was demonstrated for catechins, flavonol glycosides, proanthocyanidins, and strictinin from green and black tea. The purity and identity of isolated compounds was confirmed by (1)H NMR and HPLC-ESI-MS/MS. Gram quantities of polyphenols from tea can be isolated with the procedure described.     

New phenolic components and chromatographic profiles of green and fermented teas

A standardized profiling method based on liquid chromatography with diode array and electrospray ionization mass spectrometric detection (LC-DAD-ESI/MS) was applied to establish the phenolic profiles of 41 green teas and 25 fermented teas. More than 96 phenolic compounds were identified that allowed the teas to be organized into five groups. Epigallocatechin gallate (EGCG) was the major phenolic component of green tea made from mature leaves (group 2), while green tea made from the younger buds and leaves (group 1) contained lower flavonoid concentrations. Partially fermented teas (group 3) contained one-half the EGCG content of the green tea. Fully fermented black teas (group 4) had a trace of EGCG, but contained theaflavins. Highly overfermented black tea (group 5) contained only trace amounts of flavonol glycosides and theaflavins. Over 30 phenolics are new for tea, and this is the first phenolic profile to simultaneously detect C- and O-glycosylated flavonoids, catechins, proanthocyanidins, phenolic acid derivatives, and purine alkaloids.     

1H NMR based metabolic profiling in the evaluation of Japanese green tea quality

Classification of tea quality is now mainly performed according to the sensory results by professional tea tasters. However, this evaluation method is inconsistent in differentiating their qualities. A combination of a (1)H NMR technique and a multivariate analysis was introduced to the quality evaluation of green tea by means of a metabolomic technique. A broad range of metabolites were detected by (1)H NMR spectrometry. The principal component analysis (PCA) was used to reduce the complexity of the (1)H NMR spectra data set and provided the quality discrimination result. It offered an extensive clue for classification and quality assessment without any prepurification method. A set of green teas from a Japanese tea contest were analyzed by (1)H NMR to classify the quality with respect to that judged by tea tasters and to conceive a quality prediction model. Metabolic profiling and fingerprinting of (1)H NMR spectra of green teas with different quality were studied. PCA showed a separation between the high- and the low-quality green teas. The taste marker compounds contributing to the discrimination of tea quality were identified. Reliable prediction models were obtained by the partial least-squares projection to latent structure (PLS) analysis together with a preprocessing filter of both orthogonal signal correction (OSC) and a combination between OSC and wavelet transform algorithms.     

Structure-activity relationships of tea compounds against human cancer cells

The content of the biologically active amino acid theanine in 15 commercial black, green, specialty, and herbal tea leaves was determined as the 2,4-dinitrophenyltheanine derivative (DNP-theanine) by a validated HPLC method. To define relative anticarcinogenic potencies of tea compounds and teas, nine green tea catechins, three black tea theaflavins, and theanine as well as aqueous and 80% ethanol/water extracts of the same tea leaves were evaluated for their ability to induce cell death in human cancer and normal cells using a tetrazolium microculture (MTT) assay. Compared to untreated controls, most catechins, theaflavins, theanine, and all tea extracts reduced the numbers of the following human cancer cell lines: breast (MCF-7), colon (HT-29), hepatoma (liver) (HepG2), and prostate (PC-3) as well as normal human liver cells (Chang). The growth of normal human lung (HEL299) cells was not inhibited. The destruction of cancer cells was also observed visually by reverse phase microscopy. Statistical analysis of the data showed that (a) the anticarcinogenic effects of tea compounds and of tea leaf extracts varied widely and were concentration dependent over the ranges from 50 to 400 microg/mL of tea compound and from 50 to 400 microg/g of tea solids; (b) the different cancer cells varied in their susceptibilities to destruction; (c) 80% ethanol/water extracts with higher levels of flavonoids determined by HPLC were in most cases more active than the corresponding water extracts; and (d) flavonoid levels of the teas did not directly correlate with anticarcinogenic activities. The findings extend related observations on the anticarcinogenic potential of tea ingredients and suggest that consumers may benefit more by drinking both green and black teas.     

Interactions between flavonoids and proteins: effect on the total antioxidant capacity

Flavonoids are potent antioxidants. It is also known that flavonoids bind to proteins. The effect of the interaction between tea flavonoids and proteins on the antioxidant capacity was examined. Their separate and combined antioxidant capacities were measured with the Trolox equivalent antioxidant capacity (TEAC) assay. It was observed that the antioxidant capacity of several components of green and black tea with alpha-, beta-, and kappa-casein or albumin is not additive; that is, a part of the total antioxidant capacity is masked by the interaction. This masking depends on both the protein and the flavonoid used. Components in green and black tea, which show the highest masking in combination with beta-casein, are epigallocatechin gallate and gallic acid. The results demonstrate that the matrix influences the efficacy of an antioxidant.     

Nitrogen application influences quality,pharmacological activities and metabolite profiles of Athrixia phylicoides DC. (Bush tea) cultivated under greenhouse and field conditions

Bush tea (Athrixia phylicoides DC.) is a popular aromatic plant, indigenous to South Africa. For many decades, indigenous people have widely consumed the tea as a healthy beverage and a medicine. The objective of the study was to investigate the influence of nitrogen fertiliser application on the quality, pharmacological activities and metabolite profiles of the leaves and twigs of bush tea cultivated under greenhouse and field conditions. Treatments consisted of 0, 75, 150, 225, 300 and 375?kg/ha N, arranged in a randomised complete block design with five replicates. Parameters measured were leaf tissue nitrogen, total polyphenols, total flavonoids, total tannins, total antioxidants, antimicrobial activity and cytotoxicity. In addition, metabolites were identified and measured using gas chromatography linked to mass spectrometry (GC-MS) and liquid chromatography linked to mass spectrometry (LC-MS). The results of this study showed a significant quadratic response of total polyphenols, total flavonoids, total tannins and total antioxidants of bush tea leaves and twigs, irrespective of the growth environment ranging from 150 to 225?kg/ha N. The results further suggested that, despite the growing environment, nitrogen treatments did not improve the minimum inhibition concentration and minimum microbicide concentration of bush tea with extract concentrations ranging from 25.0 to 3.1?mg/g. Bush tea was found to be non-toxic (IC₅₀ above 100) under both growing conditions, irrespective of nitrogen treatments. Despite trials being conducted under greenhouse and field conditions and with different nitrogen treatments, there were no distinct differences regarding the gas chromatography linked to mass spectrometry and liquid chromatography linked to mass spectrometry compounds. The future aims of the study are to investigate the effects of nitrogen fertilisation, the timing of nitrogen fertilisation and the planting density on the yield, quality, plant metabolites and pharmacological activity of field-grown bush tea.     

Inhibitory effect(s) of polymeric black tea polyphenol fractions on the formation of [(3)H]-B(a)P-derived DNA adducts

Five polymeric black tea polyphenol fractions (PBP-1-5) were isolated from a popular brand of black tea. The effect of these PBPs and epigallocatechin gallate (EGCG), a major green tea polyphenol, was studied on the formation of [(3)H]-B(a)P-derived DNA adducts in vitro, employing rat liver microsomes. PBP-1-3 inhibited microsome-catalyzed [(3)H]-B(a)P-derived DNA adduct formation in vitro in a dose-dependent manner. This inhibition was further enhanced on preincubation of microsomes with each of the PBPs. PBP-4 was not effective per se and required preincubation with microsomes to exhibit its inhibitory effect, whereas PBP-5 remained ineffective with or without preincubation with microsomes. Further investigations revealed that the observed decrease in [(3)H]-B(a)P-DNA adduct formation was due to inhibition of isozymes of CYP450s by PBPs. Overall, results suggest that polymeric black tea polyphenol fractions retain one of the chemopreventive effects exhibited by the monomeric green tea polyphenol EGCG in vitro.     

红茶与绿茶感官品质与其化学组分的相关性

对茶叶感官审评分属性与其化学组分的相关性进行探讨,该文以不同等级、产地红茶和绿茶为代表,利用传统感官审评方法鉴定茶叶品质（外形、香气、汤色、滋味和叶底）,并对各品质指标进行相关性分析,筛选出对茶叶品质影响最重要的品质指标,即滋味。在利用尺度评价法对茶叶滋味品质的10个分属性（浓度、厚度、甘度、鲜度、醇度、涩度、嫩度、陈味、纯正度、火工度）进行更为细致的评定和对茶叶中主要呈味物质—茶多酚、咖啡碱、茶氨酸进行定量分析的基础上,对不同等级、不同产地茶叶的滋味分属性进行差异分析,并对化学组分与滋味分属性进行相关性分析,结果表明,各种茶样之间的产地和等级间差异主要是鲜度和涩度,而与之相关的组分主要是茶氨酸和咖啡碱,茶多酚的影响还有待进一步考察。该研究实现了茶叶感官品质的定量分析,提高了感官审评结果的客观性和确定性,为茶叶原产地保护和分等分级提供核心技术和标准支撑。     

基于机器视觉形状特征参数的祁门红茶等级识别

外形是评价茶叶质量的重要指标之一,目前主要依赖人工审评的方法,客观、准确的评价外形指标对茶叶加工、销售有重要的意义。该研究提出了一种基于形状特征直方图结合支持向量机的茶叶等级识别方法。以7个等级的祁门工夫红茶为研究对象,构建图像采集系统,标定相机参数,采集各等级的茶叶图像。对图像进行灰度化、二值化处理,提取叶片的6个绝对形状特征:长度、宽度、面积、周长、最小外接矩长、宽,在此基础上计算狭长度,矩形度2个相对形状特征,生成形状特征的直方图。以直方图分布为特征向量,构建了基于BP神经网络,极限学习机(extreme learning machine,ELM),支持向量机(support vector machine,SVM),最小二乘支持向量机(least squares support vector machine,LS-SVM)的等级识别模型,并对比了不同模型的识别效果。结果表明,该文构建的图像采集系统测量精度0.3 mm,能够准确提取形状特征参数;基于形状特征直方图的LS-SVM模型识别效果最好,识别精度为95.71%,测试集决定系数为96.2%,具有算法复杂度低,易于求解的优点。研究结果为实现茶叶的客观、数字化等级鉴定,提供了试验数据和参考方法。     

Antioxidant activity and bioactive compounds of tea seed (Camellia oleifera Abel.) oil

The oil of tea seed (Camellia oleifera Abel.) is used extensively in China as cooking oil. The objectives of this study were to investigate the antioxidant activity of tea seed oil and its active compounds. Of the five solvent extracts, methanol extract of tea seed oil exhibited the highest yield and the strongest antioxidant activity as determined by DPPH scavenging activity and Trolox equivalent antioxidant capacity (TEAC). Two peaks separated from the methanol extract by HPLC contributed the most significant antioxidant activity. These two peaks were further identified as sesamin and a novel compound: 2,5-bis-benzo[1,3]dioxol-5-yl-tetrahydro-furo [3,4-d][1,3]dioxine (named compound B) by UV absorption and characterized by MS, IR, 1H NMR, and 13C NMR techniques. Sesamin and compound B decreased H2O2-mediated formation of reactive oxygen species in red blood cells (RBCs), inhibited RBCs hemolysis induced by AAPH, and increased the lag time of conjugated dienes formation in human low-density lipoprotein. The results indicate that both compounds isolated from tea seed oil exhibit remarkable antioxidant activity. Apart from the traditional pharmacological effects of Camellia oleifera, the oil of tea seed may also act as a prophylactic agent to prevent free radical related diseases.     

Role of polyphenol oxidase and peroxidase in the generation of black tea theaflavins.

It has been reported earlier that when macerated tea leaf is fermented at lower pH, the resultant black tea contains increased levels of theaflavin, an important quality marker in black tea. In an attempt to investigate the biochemistry and chemistry underlying this observation, in vitro oxidation experiments using polyphenol oxidase (PPO) from fresh tea leaves, horseradish peroxidase (POD), and tea catechins, precursors for theaflavins, were carried out. In vitro oxidation experiments using crude tea PPO resulted in higher content of theaflavins at pH 4.5 in comparison with pH 5.5, the normal pH of the macerated tea leaf. When purified PPO was used in the in vitro system, surprisingly a reversal of this trend was observed, with more theaflavins being formed at the higher pH. A combination of pure tea PPO and POD led to an observation similar to that with the crude enzyme preparation, suggesting a possible role for POD in the formation or degradation of theaflavin. POD was observed to oxidize theaflavins in the presence of H(2)O(2), leading to the formation of thearubigin, another black tea pigment. This paper demonstrates that tea PPO, while oxidizing catechins, generates H(2)O(2). The amount of H(2)O(2) produced is greater at pH 5.5, the optimum pH for PPO activity, than at pH 4.5. Hence, an observed increase of theaflavins in black teas fermented at pH 4.5 appears to be due to lower turnover of formed theaflavins into thearubigins.     

Preservation of kombucha tea-effect of temperature on tea components and free radical scavenging properties

Kombucha tea is sugared black tea fermented with a consortium of acetic acid bacteria and yeasts (tea fungus) for 14 days. The tea tastes slightly sweet and acidic. The formation of tea fungal biofilms during storage is a big problem when kombucha tea is being stored and commercialized. Various thermal treatments have been tried for long-term storage of kombucha tea. The present study revealed the influence of heat on the biochemical constituents and the free radical scavenging properties of kombucha tea. Heat treatment at 60, 65, and 68 degrees C for 1 min controlled biofilm formation in kombucha tea without changing its clarity, taste, and flavor. However, tea polyphenols and black tea quality parameters showed varying stability during the storage period. A decrease in free radical scavenging properties was also found during the storage period. Because the biological activities of kombucha tea depended on the biochemical constituents, it was concluded that heat treatment was not a suitable method for kombucha tea preservation.     

Relationship between the metabolite profile and technological properties of bovine milk from two dairy breeds elucidated by NMR-based metabolomics

The aim of the present study was to investigate the relationship between the metabolite profile of milk and important technological properties by using nuclear magnetic resonance (NMR)-based metabolomics. The metabolomics approach was introduced for the metabolic profiling of a set of milk samples from two dairy breeds representing a wide span in coagulation properties. The milk metabolite profiles obtained by proton and carbon NMR spectroscopy could be correlated to breed and, more interestingly, also with the coagulation profile, as established by traditional methods by using principal component analysis (PCA). The metabolites responsible for the separation into breed could mainly be ascribed to carnitine and lactose, whereas the metabolites varying in the samples with respect to coagulation properties included citrate, choline, carnitine, and lactose. The results found in the present study demonstrated a promising potential of NMR-based metabolomics for a rapid analysis and classification of milk samples, both of which are useful for the dairy industry.     

A multianalytical approach for determining the geographical origin of ginseng using strontium isotopes, multielements, and 1H NMR analysis

Asian ginseng (Panax ginseng C.A. Meyer) is widely used as an Oriental medicine in the East Asian regions, particularly Korea and China. In the study, the strontium isotope ratios ((87)Sr/(86)Sr), multielements, and metabolite profiles of 35 ginseng samples collected from Korea and China were examined in an attempt to develop a method to distinguish the origin of ginsengs from the two countries. A multivariate statistical approach was performed to analyze the multielements and the (1)H nuclear magnetic resonance (NMR) data. Results of a t-test for Mg, Fe, Al, and Sc showed significant variation between Korean and Chinese ginsengs, indicating potential tracers for discriminating them. Discriminating between the ginsengs from the two countries was generally successful when both the (87)Sr/(86)Sr ratios and rare earth element (REE) contents were used together. Moreover, principal component analysis (PCA) derived from the (1)H NMR data revealed a significant separation between the ginsengs originating from the two countries. The major metabolites responsible for differentiation were sugars such as glucose, xylose, and sucrose. The results suggest that this multiplatform approach offers a comprehensive method to distinguish the origin of ginsengs.