Systematic Analysis of the R2R3-MYB Family in Camellia sinensis: Evidence for Galloylated Catechins Biosynthesis Regulation

The R2R3-MYB transcription factor (TF) family regulates metabolism of phenylpropanoids in various plant lineages. Species-expanded or specific MYB TFs may regulate species-specific metabolite biosynthesis including phenylpropanoid-derived bioactive products. Camellia sinensis produces an abundance of specialized metabolites, which makes it an excellent model for digging into the genetic regulation of plant-specific metabolite biosynthesis. The most abundant health-promoting metabolites in tea are galloylated catechins, and the most bioactive of the galloylated catechins, epigallocatechin gallate (EGCG), is specifically relative abundant in C. sinensis. However, the transcriptional regulation of galloylated catechin biosynthesis remains elusive. This study mined the R2R3-MYB TFs associated with galloylated catechin biosynthesis in C. sinensis. A total of 118 R2R3-MYB proteins, classified into 38 subgroups, were identified. R2R3-MYB subgroups specific to or expanded in C. sinensis were hypothesized to be essential to evolutionary diversification of tea-specialized metabolites. Notably, nine of these R2R3-MYB genes were expressed preferentially in apical buds (ABs) and young leaves, exactly where galloylated catechins accumulate. Three putative R2R3-MYB genes displayed strong correlation with key galloylated catechin biosynthesis genes, suggesting a role in regulating biosynthesis of epicatechin gallate (ECG) and EGCG. Overall, this study paves the way to reveal the transcriptional regulation of galloylated catechins in C. sinensis.

Matcha Green Tea Exhibits Bactericidal Activity against Streptococcus pneumoniae and Inhibits Functional Pneumolysin

Streptococcus pneumoniae is a causative pathogen of several human infectious diseases including community-acquired pneumonia. Pneumolysin (PLY), a pore-forming toxin, plays an important role in the pathogenesis of pneumococcal pneumonia. In recent years, the use of traditional natural substances for prevention has drawn attention because of the increasing antibacterial drug resistance of S. pneumoniae. According to some studies, green tea exhibits antibacterial and antitoxin activities. The polyphenols, namely the catechins epigallocatechin gallate (EGCG), epigallocatechin (EGC), epicatechin gallate (ECG), and epicatechin (EC) are largely responsible for these activities. Although matcha green tea provides more polyphenols than green tea infusions, its relationship with pneumococcal pneumonia remains unclear. In this study, we found that treatment with 20 mg/mL matcha supernatant exhibited significant antibacterial activity against S. pneumoniae regardless of antimicrobial resistance. In addition, the matcha supernatant suppressed PLY-mediated hemolysis and cytolysis by inhibiting PLY oligomerization. Moreover, the matcha supernatant and catechins inhibited PLY-mediated neutrophil death and the release of neutrophil elastase. These findings suggest that matcha green tea reduces the virulence of S. pneumoniae in vitro and may be a promising agent for the treatment of pneumococcal infections.

Nondestructive Testing and Visualization of Catechin Content in Black Tea Fermentation Using Hyperspectral Imaging

Catechin is a major reactive substance involved in black tea fermentation. It has a determinant effect on the final quality and taste of made teas. In this study, we applied hyperspectral technology with the chemometrics method and used different pretreatment and variable filtering algorithms to reduce noise interference. After reduction of the spectral data dimensions by principal component analysis (PCA), an optimal prediction model for catechin content was constructed, followed by visual analysis of catechin content when fermenting leaves for different periods of time. The results showed that zero mean normalization (Z-score), multiplicative scatter correction (MSC), and standard normal variate (SNV) can effectively improve model accuracy; while the shuffled frog leaping algorithm (SFLA), the variable combination population analysis genetic algorithm (VCPA-GA), and variable combination population analysis iteratively retaining informative variables (VCPA-IRIV) can significantly reduce spectral data and enhance the calculation speed of the model. We found that nonlinear models performed better than linear ones. The prediction accuracy for the total amount of catechins and for epicatechin gallate (ECG) of the extreme learning machine (ELM), based on optimal variables, reached 0.989 and 0.994, respectively, and the prediction accuracy for EGC, C, EC, and EGCG of the content support vector regression (SVR) models reached 0.972, 0.993, 0.990, and 0.994, respectively. The optimal model offers accurate prediction, and visual analysis can determine the distribution of the catechin content when fermenting leaves for different fermentation periods. The findings provide significant reference material for intelligent digital assessment of black tea during processing.

Development of HPLC Method for Catechins and Related Compounds Determination and Standardization in Miang (Traditional Lanna Fermented Tea Leaf in Northern Thailand)

High performance liquid chromatography (HPLC) for catechins and related compounds in Miang (traditional Lanna fermented tea leaf) was developed to overcome the matrices during the fermentation process. We investigated a variety of columns and elution conditions to determine seven catechins, namely (+)-catechin, (−)-gallocatechin, (−)-epigallocatechin, (−)-epicatechin, (−)-epigallocatechin gallate, (−)-gallocatechin gallate, (−)-epicatechin gallate, as well as gallic acid and caffeine, resulting in the development of reproducible systems for analyses that overcome sample matrices. Among the three reversed-phase columns, column C (deactivated, with extra dense bonding, double endcapped monomeric C18, high-purity silica at 3.0 mm × 250 mm and a 5 µm particle size) significantly improved the separation between Miang catechins in the presence of acid in the mobile phase within a shorter analysis time. The validation method showed effective linearity, precision, accuracy, and limits of detection and quantitation. The validated system was adequate for the qualitative and quantitative measurement of seven active catechins, including gallic acid and caffeine in Miang, during the fermentation process and standardization of Miang extracts. The latter contain catechins and related compounds that are further developed into natural active pharmaceutical ingredients (natural APIs) for cosmeceutical and nutraceutical products.

Epicatechin Gallate as Xanthine Oxidase Inhibitor: Inhibitory Kinetics, Binding Characteristics, Synergistic Inhibition, and Action Mechanism

Epicatechin gallate (ECG) is one of the main components of catechins and has multiple bioactivities. In this work, the inhibitory ability and molecular mechanism of ECG on XO were investigated systematically. ECG was determined as a mixed xanthine oxidase (XO) inhibitor with an IC50 value of 19.33 ± 0.45 μM. The promotion of reduced XO and the inhibition of the formation of uric acid by ECG led to a decrease in O2− radical. The stable ECG–XO complex was formed by hydrogen bonds and van der Waals forces, with the binding constant of the magnitude of 104 L mol−1, and ECG influenced the stability of the polypeptide skeleton and resulted in a more compact conformation of XO. Computational simulations further characterized the binding characteristics and revealed that the inhibitory mechanism of ECG on XO was likely that ECG bound to the vicinity of flavin adenine dinucleotide (FAD) and altered the conformation of XO, hindering the entry of substrate and the diffusion of catalytic products. ECG and allopurinol bound to different active sites of XO and exerted a synergistic inhibitory effect through enhancing their binding stability with XO and changing the target amino acid residues of XO. These findings may provide a theoretical basis for the further application of ECG in the fields of food nutrition and functional foods.

Effect of particle size on phytochemical composition and antioxidant properties of Sargassum cristaefolium ethanol extract

Sample particle size is an important parameter in the solid–liquid extraction system of natural products for obtaining their bioactive compounds. This study evaluates the effect of sample particle size on the phytochemical composition and antioxidant activity of brown macroalgae Sargassum cristaefolium. The crude ethanol extract was extracted from dried powders of S.cristeafolium with various particle sizes (> 4000 µm, > 250 µm, > 125 µm, > 45 µm, and < 45 µm). The ethanolic extracts of S.cristaefolium were analysed for Total Phenolic Content (TPC), Total Flavonoid Content (TFC), phenolic compound concentration and antioxidant activities. The extract yield and phytochemical composition were more abundant in smaller particle sizes. Furthermore, the TPC (14.19 ± 2.08 mg GAE/g extract to 43.27 ± 2.56 mg GAE/g extract) and TFC (9.6 ± 1.8 mg QE/g extract to 70.27 ± 3.59 mg QE/g extract) values also significantly increased as particle sizes decreased. In addition, phenolic compounds epicatechin (EC), epicatechin gallate (ECG), epigallocatechin (EGC), and Epigallocatechin gallate (EGCG) concentration were frequently increased in samples of smaller particle sizes based on two-way ANOVA and Tukey’s multiple comparison analysis. These results correlate with the significantly stronger antioxidant activity in samples with smaller particle sizes. The smallest particle size (< 45 µm) demonstrated the strongest antioxidant activity based on DPPH, ABTS, hydroxyl assay and FRAP. In addition, ramp function graph evaluates the desired particle size for maximum phytochemical composition and antioxidant activity is 44 µm. In conclusion, current results show the importance of particle size reduction of macroalgae samples to increase the effectivity of its biological activity.

Reynoutria Rhizomes as a Natural Source of SARS-CoV-2 Mpro Inhibitors–Molecular Docking and In Vitro Study

More than a year has passed since the world began to fight the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) responsible for the Coronavirus disease 2019 (COVID-19) pandemic, and still it spreads around the world, mutating at the same time. One of the sources of compounds with potential antiviral activity is Traditional Chinese Medicinal (TCM) plants used in China in the supportive treatment of COVID-19. Reynoutria japonica is important part of the Shu Feng Jie Du Granule/Capsule-TCM herbal formula, recommended by China Food and Drug Administration (CFDA) for treatment of patients with H1N1- and H5N9-induced acute lung injury and is also used in China to treat COVID-19, mainly combined with other remedies. In our study, 25 compounds from rhizomes of R. japonica and Reynoutria sachalinensis (related species), were docked into the binding site of SARS-CoV-2 main protease. Next, 11 of them (vanicoside A, vanicoside B, resveratrol, piceid, emodin, epicatechin, epicatechin gallate, epigallocatechin gallate, procyanidin B2, procyanidin C1, procyanidin B2 3,3’-di-O-gallate) as well as extracts and fractions from rhizomes of R. japonica and R. sachalinensis were tested in vitro using a fluorescent peptide substrate. Among the tested phytochemicals the best results were achieved for vanicoside A and vanicoside B with moderate inhibition of SARS-CoV-2 Mpro, IC50 = 23.10 µM and 43.59 µM, respectively. The butanol fractions of plants showed the strongest inhibition of SARS-CoV-2 Mpro (IC50 = 4.031 µg/mL for R. sachalinensis and IC50 = 7.877 µg/mL for R. japonica). As the main constituents of butanol fractions, besides the phenylpropanoid disaccharide esters (e.g., vanicosides), are highly polymerized procyanidins, we suppose that they could be responsible for their strong inhibitory properties. As inhibition of SARS-CoV-2 main protease could prevent the replication of the virus our research provides data that may explain the beneficial effects of R. japonica on COVID-19 and identify the most active compounds worthy of more extensive research.

The catechins profile and antioxidant activity in different types of green tea bags

Tea and mainly green tea as a rich source of antioxidants has been widely known for some time. The antioxidant capacity of tea including green tea is mainly due to its catechins content. The objective of this study was to determine the relationship between the ORAC value and the catechins content. Regular and decaffeinated commercial green tea bags commonly consumed in the United Kingdom (UK) have been examined using Reversed-Phase High-pressure Liquid Chromatography (RP-HPLC). Teabags were purchased from different local supermarkets in the UK and extracted with natural mineral water at temperature 100°C for 9 minutes at the pre-adjusted pH 4. The level of four catechins (Epi-structured) for the thirteen types of green tea were separated and determined by HPLC analysis, i.e. ()-epigallocatechin (EGC), ()-epicatechin (EC), ()-epigallocatechin-3-gallate (EGCG) and ()-epicatechin gallate (ECG). The standard graphs were validated using certified reference catechins supplied by the Laboratory of the Government Chemist (LGC). The levels of total catechins and oxygen radical absorbance capacity (ORAC) values varied from 34.61to 204.55 mg/g, 830.19- 4197.81 Trolox equivalents/g tea bags for thirteen types of green teas respectively. It was clear from the results of this study that there was a significant linear and positive correlation (r = 0.951, df = 12, p < 0.05) is found to exist between the total catechins contents and ORAC values. It can be concluded that the results of catechins measurements coupled to this; the well-known ORAC assay was successfully modified to measure the antioxidant capacity of the green tea extracts throughout this study. Furthermore, the higher the level of catechins the higher is the antioxidant capacity of the tea. This may stimulus consumers in selecting the type of tea and tea brewing times, exhibiting more health benefits. Nevertheless, the differences between the studied brands are owing to shelf life, production and storage conditions.

Green tea phytocompounds targets Lansterol 14-α demethylase against ergosterol biosynthesis in Candida glabrata

Green tea is credited as one of the world’s healthiest drinks with enriched antioxidants. It is known for its multi-beneficial health benefits against diabetes, blood pressure, hypertension, gastro-intestinal upset and is bestowed with significant antimicrobial potential. There are previous scientific evidence highlighting the antifungal potential of green tea and has identified it as a potential inhibitor of non-albicans Candida species. Lansterol 14-α demethylase (Erg 11) or CYP51 protein belongs to the cytochrome P450 monooxygenase (CYP) superfamily. Erg 11 is involved in ergosterol biosynthesis and has a significant role in azole drug resistance in Candida glabrata. The present study attempted to identify the inhibitory potential of green tea phytocompounds against inhibition of Erg 11 in Candida glabrata using bioinformatics tool viz., autodock vina software. Out of 15 green tea phytocompounds investigated, the study identified, Rutin (-10.5 kcal) Kaempferitrin (-9.4kcal), Epigallocatechin gallate (-10kcal), Epicatechin gallate (-8.7kcal), and Coumaroylquinic acid (-8.6kcal) acid as the potent phytocompounds which showed significant molecular interaction with Erg 11 in Candida glabrata. In attribution to the constant emergence of azole-resistant isolates, this preliminary analysis therefore, indicated the potential of green tea phytocompounds against inhibition of non-albicans Candida specific candidiasis. However, further, in vitro antimicrobial efficacy of these phytocompounds, the dose regime, drug likeliness, and cytotoxic analysis are required to be investigated and validated.