A comprehensive review of polyphenol oxidase in tea (Camellia sinensis): Physiological characteristics, oxidation manufacturing, and biosynthesis of functional constituents.

Polyphenol oxidase (PPO) is a metalloenzyme with a type III copper core that is abundant in nature. As one of the most essential enzymes in the tea plant (Camellia sinensis), the further regulation of PPO is critical for enhancing defensive responses, cultivating high-quality germplasm resources of tea plants, and producing tea products that are both functional and sensory qualities. Due to their physiological and pharmacological values, the constituents from the oxidative polymerization of PPO in tea manufacturing may serve as functional foods to prevent and treat chronic non-communicable diseases. However, current knowledge of the utilization of PPO in the tea industry is only available from scattered sources, and a more comprehensive study is required to reveal the relationship between PPO and tea obviously. A more comprehensive review of the role of PPO in tea was reported for the first time, as its classification, catalytic mechanism, and utilization in modulating tea flavors, compositions, and nutrition, along with the relationships between PPO-mediated enzymatic reactions and the formation of functional constituents in tea, and the techniques for the modification and application of PPO based on modern enzymology and synthetic biology are summarized and suggested in this article.

Identification, Molecular Characteristic, and Expression Analysis of PIFs Related to Chlorophyll Metabolism in Tea Plant (Camellia sinensis).

The phytochrome-interacting factors (PIFs) proteins belong to the subfamily of basic helix-loop-helix (bHLH) transcription factors and play important roles in chloroplast development and chlorophyll biosynthesis. Currently, knowledge about the PIF gene family in Camellia sinensis remains very limited. In this study, seven PIF members were identified in the C. sinensis genome and named based on homology with AtPIF genes in Arabidopsis thaliana. All C. sinensis PIF (CsPIF) proteins have both the conserved active PHYB binding (APB) and bHLH domains. Phylogenetic analysis revealed that CsPIFs were clustered into four groups-PIF1, PIF3, PIF7, and PIF8-and most CsPIFs were clustered in pairs with their corresponding orthologs in Populus tremula. CsPIF members in the same group tended to display uniform or similar exon-intron distribution patterns and motif compositions. CsPIF genes were differentially expressed in C. sinensis with various leaf colors and strongly correlated with the expression of genes involved in the chlorophyll metabolism pathway. Promoter analysis of structural genes related to chlorophyll metabolism found DNA-binding sites of PIFs were abundant in the promoter regions. Protein-protein interaction networks of CsPIFs demonstrated a close association with phytochrome, PIF4, HY5, TOC1, COP1, and PTAC12 proteins. Additionally, subcellular localization and transcriptional activity analysis suggested that CsPIF3b was nuclear localized protein and possessed transcriptional activity. We also found that CsPIF3b could activate the transcription of CsHEMA and CsPOR in Nicotiana benthamiana leaves. This work provides comprehensive research of CsPIFs and would be helpful to further promote the regulation mechanism of PIF on chlorophyll metabolism in C. sinensis.

Neuroprotective Effects of Theaflavins Against Oxidative Stress-Induced Apoptosis in PC12 Cells.

Oxidative stress can induce neuronal apoptosis via the production of superoxide and hydroxyl radicals. This process is as a major pathogenic mechanism in neurodegenerative disorders. In this study, we aimed to clarify whether theaflavins protect PC12 cells from oxidative stress damage induced by H2O2. A cell model of PC12 cells undergoing oxidative stress was created by exposing cells to 200 µM H2O2 in the presence or absence of varying concentrations of theaflavins (5, 10, and 20 µM). Cell viability was monitored using the MTT assay and Hoechst 33258 staining, showing that 10 µM theaflavins enhanced cell survival following 200 µM H2O2 induced toxicity and increased cell viability by approximately 40 %. Additionally, we measured levels of intracellular reactive oxygen species (ROS) and antioxidant enzyme activity. This suggested that the neuroprotective effect of theaflavins against oxidative stress in PC12 cells is derived from suppression of oxidant enzyme activity. Furthermore, Western blot analyses indicated that theaflavins downregulated the ratio of pro-apoptosis/anti-apoptosis proteins Bax/Bcl-2. Theaflavins also downregulated the expression of caspase-3 compared with a H2O2-treated group that had not been treated with theaflavins. Interestingly, this is the first study to report that the four main components of theaflavins found in black tea can protect neural cells (PC12) from apoptosis induced by H2O2. These findings provide the foundations for a new field of using theaflavins or its source, black tea, in the treatment of neurodegenerative diseases caused by oxidative stress.

Review: Research progress on seasonal succession of phyllosphere microorganisms.

Phyllosphere microorganisms have recently attracted the attention of scientists studying plant microbiomes. The origin, diversity, functions, and interactions of phyllosphere microorganisms have been extensively explored. Many experiments have demonstrated seasonal cycles of phyllosphere microbes. However, a comprehensive comparison of these separate investigations to characterize seasonal trends in phyllosphere microbes of woody and herbaceous plants has not been conducted. In this review, we explored the dynamic changes of phyllosphere microorganisms in woody and non-woody plants with the passage of the season, sought to find the driving factors, summarized these texts, and thought about future research trends regarding the application of phyllosphere microorganisms in agricultural production. Seasonal trends in phyllosphere microorganisms of herbaceous and woody plants have similarities and differences, but extensive experimental validation is needed. Climate, insects, hosts, microbial interactions, and anthropogenic activities are the diverse factors that influence seasonal variation in phyllosphere microorganisms.

"Age" of tea: The impact of long-term storage on the aroma of Tuo tea and age prediction.

This study investigates the dynamic changes in the aroma profile of Tuo tea during long-term storage, a process not well understood yet critical to the formation of aged tea's unique characteristics. Aroma profiling of Tuo tea samples stored for 2 to 25 years was conducted using sensory evaluation and the HS-SPME/GC × GC-QTOFMS technique, revealing a progressive transition from fresh, fruity, and floral scents to more stale, woody, and herbal notes. Among 275 identified volatiles, 55 were correlated with storage duration (|r| > 0.8, p < 0.05), and 49 differential compounds (VIP > 1, FC > 1.2, FC < 0.833, p < 0.05) were identified across three storage stages (2-4, 5-10, and 13-25 years). Furthermore, theaspirane, eucalyptol, o-xylene, and 1-ethylidene-1H-indene were selected as potential markers of Tuo tea aging, incorporating the implementation of a Random Forest (RF) model. Additionally, our model exhibited high accuracy in predicting the age of Tuo tea within a prediction error range of -2.51 to 2.84 years. This research contributes to a comprehensive understanding of the impact of storage time on tea aroma and aids in the precise identification of tea age.

Characterization of key odorants in 'Baimaocha' black teas from different regions.

'Baimmaocha' is a distinctive resource for production of high-quality black tea, and its processed black tea has unique aroma characteristics. 190 volatile compounds were identified by comprehensive two-dimensional gas chromatography-olfactometry-quadrupole-time-of-flight mass spectrometry(GC × GC-O-Q-TOMS), and among them 23 compounds were recognized as key odorants contributing to forming different aroma characteristics in 'Baimaocha' black teas of Rucheng, Renhua, and Lingyun (RCBT, RHBT, LYBT). The odor activity value coupled with GC-O showed that methyl salicylate (RCBT), geraniol (RHBT), trans-ß-ionone and benzeneacetaldehyde (LYBT) might be the most definitive aroma compounds identified from their respective regions. Furthermore, PLS analysis revealed three odorants as significant contributors to floral characteristic, four odorants related to fruity attribute, four odorants linked to fresh attribute, and three odorants associated with roasted attribute. These results provide novel insights into sensory evaluation and chemical substances of 'Baimaocha' black tea and provide a theoretical basis for controlling and enhancement tea aroma quality.

Theanine Improves High-Dose Epigallocatechin-3-Gallate-Induced Lifespan Reduction in Caenorhabditis elegans.

Epigallocatechin-3-gallate (EGCG) is the most abundant polyphenol in green tea. Our previous report showed that induced hormesis was a critical determinant for the promotion of a healthy lifespan in Caenorhabditis elegans. In the present study, we investigated the anti-aging effects of the main active ingredients in green tea. We found that galloylated catechins (EGCG and epicatechin gallate) could extend the lifespan of C. elegans, while their metabolites (gallic acid, epicatechin, and epigallocatechin) could not. Interestingly, the combination with theanine, not caffeine, could alleviate the adverse effects induced by high-dose EGCG, including the promotion of lifespan and locomotor ability. This was due to the attenuation of the excess production of reactive oxygen species and the activation of DAF-16. These findings will facilitate further studies on the health benefits of tea active components and their interactions.

Physiological Dose of EGCG Attenuates the Health Defects of High Dose by Regulating MEMO-1 in Caenorhabditis elegans.

EGCG, as a dietary-derived antioxidant, has been extensively studied for its beneficial health effects. Nevertheless, it induces the transient increase in ROS and leads to the hormetic extension of lifespan. How exactly biology-benefiting effects with the minimum severe adverse are realized remains unclear. Here, we showed that physiological dose of EGCG could help moderate remission in health side effects exposed to high doses, including shortened lifespan, reduced body size, decreased pharyngeal pumping rate, and dysfunctional body movement in C. elegans. Furthermore, we found this result was caused by the physiological dose of EGCG to block the continued ROS accumulation and triggered acclimation responses after stressor removal. Also, in this process, we observed that EGCG downregulated the key redox protein MEMO-1 to activate the feedback loop of NADPH oxidase-mediated redox signaling. Our data indicates that the feedback signal induced by NADPH oxidase may contribute to the health-protective mechanism of dietary polyphenols in vivo.

Stretchable Multiresponsive Hydrogel with Actuatable, Shape Memory, and Self-Healing Properties.

Smart hydrogels with responsive behaviors have attracted tremendous attention. However, it is still a challenge to synthesize stretchable hydrogels capable of changing their original properties in response to multiple external stimuli. Here, integration of actuation function, shape memory, and self-healing capability in a highly stretchable hydrogel under triple external triggers is achieved by rationally engineering multiple functional moieties. The hydrogel exhibits high stretchability (average relative strain (mm/mm) is >15) and excellent fatigue resistance during 100 loading cycles of 100% strain. Incorporating a moisture-insensitive polymer film with the hydrogel, hydroactuated functionality is demonstrated. Moreover, shape memory and self-healing abilities of the hydrogel are realized by the formation of ionic crosslinking or dynamic borate ester in conditions of multivalent cations and pH, respectively. Deformable plastic flowers are displayed in this work as a proof-of-concept, and it is believed that this smart hydrogel could be used in plenty of frontier fields, such as designing electronic devices, soft robotics, and actuators.

Epigallocatechin-3-gallate promotes healthy lifespan through mitohormesis during early-to-mid adulthood in Caenorhabditis elegans.

The green tea polyphenol epigallocatechin-3-gallate (EGCG) is widely consumed as a dietary supplement. Its potential properties include slowing aging and extending lifespan, although how exactly this is achieved remains unclear. Here, we report that EGCG promoted healthy lifespan in Caenorhabditis elegans when administered throughout or only at early-to-mid adulthood. Specifically, EGCG extended lifespan in an inverted U-shaped dose-response manner. The life-extending mechanism was stimulated by EGCG-induced production of reactive oxygen species (ROS). Additionally, EGCG triggered mitochondrial biogenesis to restore mitochondrial function. The EGCG-induced increase in lifespan depends on known energy sensors such as AMPK/AAK-2, as well as SIRT1/SIR-2.1 and FOXO/DAF-16. Interestingly, aging decreased the response to EGCG and progressively neutralized its beneficial effects on longevity. Collectively, our findings link EGCG to the process of mitohormesis and suggest an inducible, AMPK/SIRT1/FOXO-dependent redox signaling module that could be invoked in different contexts to extend healthy lifespan. Its effectiveness is higher in younger adults and declines with age.

Black tea increases hypertonic stress resistance in C. elegans.

Here we identified that BTE (black tea extract), within the studied concentration range, is more effective than GTE (green tea extract) in protecting C. elegans against hypertonic stress, by enhancing survival after exposure to various salts, and alleviating suffered motility loss and body shrinkage. The mechanism of such protection may be due to the ability of black tea to induce the conserved WNK/GCK signaling pathway and down-regulation of the expression levels of nlp-29. Intriguingly, black tea does not relieve hypertonicity-induced protein damage. The findings implicate the potential health benefits of black tea consumed worldwide.

Tea polyphenol epigallocatechin gallate inhibits Escherichia coli by increasing endogenous oxidative stress.

The antibacterial effects of tea polyphenol epigallocatechin gallate (EGCG), a common phytochemical with a number of potential health benefits, are well known. However, the mechanism of its bactericidal action remains unclear. Using E. coli as a model organism, it is argued here that H2O2 synthesis by EGCG is not attributed to its inhibitory effects. In contrast, the bactericidal action of EGCG was a result of increased intracellular reactive oxygen species and blunted adaptive oxidative stress response in E. coli due to the co-administration of antioxidant N-acetylcysteine, and not on account of exogenous catalase. Furthermore, we noted a synergistic bactericidal effect for EGCG when combined with paraquat. However, under anaerobic conditions, the inhibitory effect of EGCG was prevented. In conclusion, EGCG caused an increase in endogenous oxidative stress in E. coli, thereby inhibiting its growth, and hence the use of EGCG as a prooxidant is supported by this study.

Comparative profiling of gene expression in Camellia sinensis L. cultivar AnJiBaiCha leaves during periodic albinism.

The AnJiBaiCha albino mutant tea cultivar has a reversible albino phenotype at low temperatures. Albino AnJiBaiCha leaves contain high levels of amino acids, which are important components affecting the quality of tea as a beverage. To examine the molecular mechanism of albinism and amino acid enrichment in AnJiBaiCha, we used the amplified fragment length polymorphism (cDNA-AFLP) technique to isolate genes that are differentially expressed during periodic albinism in AnJiBaiCha. A total of 127 transcript-derived fragments (TDFs) were successfully sequenced; among those, 60 TDFs showed high similarity to sequences with known functions, but 67 TDFs were not similar to any known genes. The identified transcripts include transcription factors, ubiquitination-related genes, chloroplast biogenesis genes, signal transduction genes, stress-related genes, cell cycle genes, and carbohydrate and energy metabolism genes. To validate the cDNA-AFLP results, quantitative real-time PCR was used to confirm the differential expression of six of the identified genes. The cDNA-AFLP and quantitative real-time PCR results correlated well, indicating that the cDNA-AFLP results are reliable. This study provides insights into the molecular mechanisms by which periodic albinism and amino acid accumulation take place in AnJiBaiCha.

Black tea increased survival of Caenorhabditis elegans under stress.

The present study examined the effects of black tea (Camellia sinensis) extracts (BTE) in Caenorhabditis elegans under various abiotic stressors. Results showed BTE increased nematode resistance to osmosis, heat, and UV irradiation treatments. However, BTE could not increase nematodes' lifespan under normal culture conditions and MnCl2-induced toxicity at concentrations we used. Further studies showed that BTE decreased reactive oxygen species and up-regulated some antioxidant enzymes, including GSH-PX, and genes, such as gsh-px and sod-3. However, only a slight extension in mev-1 mutants mean lifespan was observed without significance. These results indicated that the antioxidant activity of BTE might be necessary but not sufficient to protect against aging to C. elegans. Moreover, BTE increased the mRNA level of stress-response genes such as sir-2.1 and sek-1. Our finding demonstrated BTE might increase heat and UV stress resistance in a sir.2.1-dependent manner. Taken together, BTE enhanced stress resistance with multiple mechanisms in C. elegans.

Dynamic changes in catechin levels and catechin biosynthesis-related gene expression in albino tea plants (Camellia sinensis L.).

Tea (Camellia sinensis (L.) O. Kuntze) leaves are a major source of flavonoids that mainly belong to the flavan-3-ols or catechins and are implicated in a wide range of health benefits. Although the catechins in tea leaves were identified long ago, the regulatory mechanisms governing catechin biosynthesis remain unclear. In the present work, the dynamic changes of catechin levels and the expression profiles of catechin-related genes in albino tea plants were intensively examined. The amounts of most catechins decreased to their lowest levels in the albino phase, when epigallocatechingallate was the highest of the catechins compared to all catechins, and catechin the lowest. Enzyme assays indicated that phenylalanine ammonia-lyase (PAL) activity was positively correlated with the concentration of catechins (r = 0.673). Gene expression profiling by quantitative real-time reverse transcription-polymerase chain reaction showed that the transcript abundance of flavonoid biosynthetic genes followed a tightly regulated biphasic pattern, and was affected by albinism. These genes (PAL, C4H, 4CL, CHS, CHI, F3H, FLS, F3'H, F3'5'H, DFR, LAR, ANS and ANR) encode enzymes in flavonoid biosynthesis. The expression levels of PAL, F3H and FLS were correlated with the concentration of catechins and the correlation coefficients were -0.683, 0.687 and -0.602, respectively. Therefore, these results indicate that PAL might be a core regulator in the control of catechin biosynthesis in albino tea plants.