Phytochemical Composition and Antioxidant Capacity of 30 Chinese Teas

Tea has been reported to prevent and manage many chronic diseases, such as cancer, diabetes, obesity, and cardiovascular diseases, and the antioxidant capacity of tea may be responsible for these health benefits. In this study, the antioxidant capacities of fat-soluble, water-soluble, and bound-insoluble fractions of 30 Chinese teas belonging to six categories, namely green, black, oolong, dark, white, and yellow teas, were systematically evaluated, applying ferric-reducing antioxidant power and Trolox equivalent antioxidant capacity assays. In addition, total phenolic contents of teas were determined by Folin–Ciocalteu method, and the contents of 18 main phytochemical compounds in teas were measured by high-performance liquid chromatography (HPLC). The results found that several teas possessed very strong antioxidant capacity, and caffeine, theaflavine, gallic acid, chlorogenic acid, ellagic acid, and kaempferol-3-O-glucoside, as well as eight catechins, were the main antioxidant compounds in them. Thus, these teas could be good natural sources of dietary antioxidants, and their extracts might be developed as food additives, nutraceuticals, cosmetics, and pharmaceuticals.

In the present study, the antioxidant capacity of 30 tea samples, which are the best-selling and most commonly consumed teas in China, were systematically evaluated. In addition, their total phenolic The spectra were recorded between 200 and 600 nm, and the targeted compounds were identified by retention time and UV-Vis spectra in comparison with the standards and quantified based on the peak area under the maximum absorption wavelength. The value was expressed as mg/g DW of the tea.

Data Analysis
All tests were performed in triplicate, and the values were expressed as mean ± standard deviation (SD). SPSS 22 (International Business Machines Corp, Armonk, NY, USA) and Excel 2007 (Microsoft Corporation, Redmond, WA, USA) were applied for data analysis. One-way ANOVA and post hoc Tukey test were performed to compare means of more than two samples, and p value less than 0.05 was defined as statistical significance.

Ferric-Reducing Antioxidant Power (FRAP) Values of the Tested Teas
FRAP value was used as an important indicator for the antioxidant capacity with regard to reducing ferric ions to ferrous ions, and FRAP results of the 30 teas are displayed in Table 2 (Table 3), the FRAP values for the three fractions met the following order: water-soluble > bound-insoluble > fat-soluble.  and non-parametric tests (Table 3), TEAC values for the three fractions met the following order: water-soluble > bound-insoluble > fat-soluble bound. Abbreviations: DW, dry weight; SD, standard deviation; TEAC, Trolox equivalent antioxidant capacity. Different superscript lowercase letters ( a,b ) indicated statistical significance (p < 0.05).

Total Phenolic Content (TPC) of the Tested Teas
TPC was adopted to measure the total contents of phenolic compounds in the 30 Chinese teas, and the results are shown in Table 5. Briefly, the range of total TPC values was 37.25 ± 0.16 to 254.29 ± 15.51 mg GAE/g DW with a 7-fold difference. Dianqing Tea, Xihu Longjing Tea, Junshan Yinzhen Tea, Dongting Biluochun Tea, and Yuan'an Luyuan Tea possessed the top five total phenolic contents, namely 254.29 ± 15.51, 215.39 ± 11.87, 214.72 ± 3.22, 211.20 ± 2.52, and 210.05 ± 7.84 mg GAE/g DW, respectively. Tibetan Tea was observed with the lowest TPC of 37.25 ± 0.16 mg GAE/g DW. In addition, based on the statistical description and non-parametric tests (Table 3), the TPC values for the three fractions met the following order: water-soluble > bound-insoluble > fat-soluble.

Correlations among Ferric-Reducing Antioxidant Power (FRAP), Trolox Equivalent Antioxidant Capacity (TEAC), and Total Phenolic Content (TPC) Values
The correlations among FRAP, TEAC, and TPC values (based on the total values of three fractions) were determined by the simple linear regression model, and the results are presented in Figure 1. Both FRAP and TEAC values were significantly and positively correlated with TPC (R 2 = 0.883, p < 0.001 and R 2 = 0.941, p < 0.001, respectively). These results suggested that the phenolic compounds could be the main components contributing to the antioxidant activities of tea. In addition, FRAP values were positively and remarkably correlated with TEAC values (R 2 = 0.928, p < 0.001). Therefore, the antioxidants in tea could possess multiple functions regarding reducing oxidants (like Fe(III)) and scavenging free radicals (like ABTS• + ). Abbreviations: DW, dry weight; GAE, gallic acid equivalent; SD, standard deviation; TPC, total phenolic content. Different superscript lowercase letters ( a,b ) indicated statistical significance (p < 0.05).

Systematic Cluster of the Tested Teas
Based on the FRAP, TEAC, and TPC values, a systematic cluster analysis for the 30 teas was conducted with cluster numbers from 2 to 6, and the results are summarized in Figure 2. After that, the outcomes of cluster number = 4 were further analyzed using Online Analytical Processing (OLAP) accompanied with variance analysis (ANOVA), and the results are presented in Table 6. In detail, cluster 1 contained 12 teas, which were 4 black teas, 4 dark teas, 3 white teas, and 1 green tea, with the lowest values for FRAP, TEAC, and TPC (1050.03 ± 317.40 μmol Fe(II)/g DW, 600.57 ± 194.85 μmol Trolox/g DW, and 84.66 ± 27.90 mg GAE/g DW, respectively). In addition, cluster 2 comprised all the

Systematic Cluster of the Tested Teas
Based on the FRAP, TEAC, and TPC values, a systematic cluster analysis for the 30 teas was conducted with cluster numbers from 2 to 6, and the results are summarized in Figure 2. After that, the outcomes of cluster number = 4 were further analyzed using Online Analytical Processing (OLAP) accompanied with variance analysis (ANOVA), and the results are presented in Table 6. In detail, cluster 1 contained 12 teas, which were 4 black teas, 4 dark teas, 3 white teas, and 1 green tea, with the lowest values for FRAP, TEAC, and TPC (1050.03 ± 317. 40
In addition, for other phytochemical compounds besides catechins in teas, the content of gallic acid was low in all tea samples, ranging from 0.294 ± 0.021 to 3.822 ± 0.111 mg/g DW with a 13-fold difference. Huoshan Large Yellow Tea, Yichang Congou Black Tea, Fenghuang Shuixian Tea, Fuzhuan Brick Tea, and Keemun Black Tea possessed the top-five contents of gallic acid, which were 3.822 ± 0.111, 3.546 ± 0.050, 3.284 ± 0.141, 3.097 ± 0.122, and 2.706 ± 0.117 mg/g DW, respectively. Tieguanyin Tea was shown to have the lowest content of gallic acid, which was 0.294 ± 0.021 mg/g DW. Similarly, the contents of chlorogenic acid, ellagic acid, and kaempferol-3-O-glucoside were also relatively low in the tested teas.
As polyphenols were suggested as the main antioxidants in teas (Figure 1), we next analyzed the relationships of different polyphenols and antioxidant activities of teas. It was found that the content of catechins had moderate positive correlations ( Figure 4A Furthermore, Wuyi Rock Tea (oolong tea) and Fuzhuan Brick Tea (dark tea) as well as all of the 4 black teas (Yichang Congou Black Tea, Keemun Black Tea, Dianhong Congou Black Tea, and Lapsang Souchong Black Tea) were found with a spot of theaflavine, and the contents were 0.545 ± 0.011, 0.480 ± 0.008, 0.559 ± 0.018, 0.542 ± 0.010, 0.526 ± 0.019, and 0.488 ± 0.012 mg/g DW, respectively. the relationships of different polyphenols and antioxidant activities of teas. It was found that the content of catechins had moderate positive correlations ( Figure 4A,B) with FRAP values (R 2 = 0.476, p < 0.001) and TEAC values (R 2 = 0.515, p < 0.001), while the content of noncatechin polyphenols had no evident linear correlations ( Figure 4C,D) with FRAP values (R 2 = 0.001, p = 0.867) and TEAC values (R 2 = 0.002, p = 0.819). These results indicate that catechins can be one of the main antioxidants in tea, but noncatechin polyphenols should not be the main contributors.

Antioxidant Capacities of the Tested Chinese Teas
Many natural products, such as vegetables, fruits, and medicinal plants, possess rich phytochemicals, some of which have been recognized as strong antioxidants [23][24][25][26][27][28][29]. These natural antioxidants are often multifunctional, and their antioxidant capacities can be generally influenced by various factors, e.g., the extraction solvents, extraction conditions, and measurement methods, resulting in the difficulty to completely illustrate the antioxidant capacities only applying a single method [30,31]. In order to maximize the extraction yields of antioxidants from tea, 3 solvents × 2 repeated extraction were adopted in this study [32]. In addition, a reliable antioxidant assessing system demands comprehensive indices, which comprise different experiments to evaluate the antioxidant capacity with diverse mechanisms of action. The FRAP assay was set up based on the power of antioxidants to reduce ferric ions to ferrous ions [19], while the TEAC assay was established on the basis of the capacity of antioxidants to scavenge the ABTS• + free radicals [20]. These two assays are simple, fast, repeatable, and widely used for the evaluation of antioxidant capacity [33][34][35]. In this study, FRAP and TEAC assays were simultaneously used to assess the antioxidant capacities of the 30 Chinese teas.
The FRAP and TEAC values of the tested teas were extremely high compared with other natural products (Table 9). That is, the antioxidant capacities of tea were higher than those of most medicinal plants, edible macro-fungi, vegetables, fruits, fruit wastes (peels and seeds), and wild fruits, as well as edible and wild flowers [33][34][35]. This may be explained by the apparently higher content of phenolic compounds in tea, as revealed in Table 8. Therefore, teas rich in antioxidants may be important natural sources of dietary antioxidants, and their extracts can be used to produce food additives, cosmetics, nutraceuticals, and pharmaceuticals. Moreover, the FRAP and TEAC values of water-soluble fractions were remarkably higher than those of bound-insoluble fractions, which were mildly higher than those of fat-soluble fractions. These results suggested that the components responsible for the ferric-reducing power and ABTS free radical-scavenging capacity of teas were most water-soluble compounds (approximately 87-93%) with some bound-insoluble (about 5-8%) and fat-soluble (roughly 2-5%) ones.

Antioxidant Phytochemical Components of the Tested Chinese Teas
As demonstrated previously, there were significant and remarkable correlations among FRAP, TEAC, and TPC values. These results suggested that the phenolic compounds could be the major components contributing to the antioxidant capacities of tea, which possessed multiple effects to reduce oxidants and scavenge free radicals. The outcomes demonstrated above were consistent with several previous studies, which have reported that phenolic components were the main contributors responsible for the antioxidant capacities of vegetables, macro-fungi, wild fruits, and flowers [23,[37][38][39]. Moreover, many polyphenols have been detected in these natural products, e.g., gallic acid, chlorogenic acid, ferulic acid, anthocyanins, quercetin, rutin, myricetin, and kaempferol glycosides, which exhibited potent antioxidant capacities both in vitro and in vivo [23,[37][38][39]. Antioxidant action can be one of the most important mechanisms of the health benefits of these natural products [41][42][43]. As for tea, eight catechins, caffeine, theaflavine, gallic acid, chlorogenic acid, ellagic acid, and kaempferol-3-O-glucoside, could be detected. Among them, epicatechin, epigallocatechin, epicatechin gallate, and epigallocatechin gallate were the most abundant polyphenols in tea, especially in the green, yellow, and oolong teas, which generally undergo a low degree of fermentation. Though tea and other natural products contain several common antioxidants, their contents in tea are generally higher.

Comparison of Antioxidant Phytochemicals among Different Chinese Teas
In the light of the outcomes from systematic cluster analysis accompanied by OLAP and ANOVA for cluster number = 4, green tea and yellow tea possessed remarkably high antioxidant capacities and phenolic contents, but Lu'an Guapian Tea (green tea) and Huoshan Large Yellow Tea (yellow tea) were the exceptions. In addition, oolong tea was in the middle position. Meanwhile, white tea, black tea, and dark tea exerted relatively low antioxidant capacities and phenolic contents. Thus, fermentation degree can be a crucial factor that influences the antioxidant capacity and phenolic content of tea. Tea undergoing higher fermentation degree might have lower antioxidant capacity and phenolic content, since tea polyphenols, especially catechins, may oxidize and polymerize during fermentation, generating complicated tea pigments like theaflavins, thearubigins, and theabrownins [57][58][59]. Moreover, the maturity of tea leaves should also be taken into consideration, because the antioxidant capacity and phenolic content would decrease accompanied with the increase of tea leaf maturity [60], which may partially explain why white tea (made of old tea leaves) exhibited relatively low antioxidant capacity and phenolic content, although it has a low fermentation degree. On the other hand, it was reported that the bioavailability of fermented tea using microbes, such as bacteria, yeasts, and fungi, could be significantly higher compared to unfermented tea [61,62]. For example, green and black teas have been observed to improve endothelial function with equal effectiveness, although green tea possesses higher antioxidant activity and phenolic content, it has a lower bioavailability [63].

Conclusions
In conclusion, teas here studied possessed remarkably high antioxidant capacities regarding ferric-reducing and free radical-scavenging capacities. In addition, eight catechins, caffeine, theaflavine, and several other phenolic compounds, including gallic acid, chlorogenic acid, ellagic acid, and kaempferol-3-O-glucoside, were detected in these Chinese teas. Compared with dark, black, and white teas, green, yellow, and oolong teas exerted stronger antioxidant capacity and contained more polyphenols, especially catechins like epicatechin, epigallocatechin, epicatechin gallate, and epigallocatechin gallate. Overall, tea is a good natural source of dietary antioxidant phytochemicals, and can be used to produce food additives, functional foods, nutraceuticals, and cosmetics.