Identifying the temporal contributors and their interactions during dynamic formation of black tea cream.

Tea cream formed in hot and strong tea infusion while cooling deteriorates quality and health benefits of tea. However, the interactions among temporal contributors during dynamic formation of tea cream are still elusive. Here, by deletional recombination experiments and molecular dynamics simulation, it was found that proteins, caffeine (CAF), and phenolics played a dominant role throughout the cream formation, and the contribution of amino acids was highlighted in the early stage. Furthermore, CAF was prominent due to its extensive binding capacity and the filling complex voids property, and caffeine-theaflavins (TFs) complexation may be the core skeleton of the growing particles in black tea infusion. In addition to TFs, the unidentified phenolic oxidation-derived products (PODP) were confirmed to contribute greatly to the cream formation.

Analysis of volatile metabolite variations in strip green tea during processing and effect of rubbing degree using untargeted and targeted metabolomics.

Strip green tea (SGT) is widely distributed in China owing to its unique appearance and aroma but the evolution and formation mechanisms of volatile metabolites (VMs) during SGT processing, and especially in the unique process of rubbing, remain unclear. In this study, based on untargeted metabolomics, 217 VMs (8 categories) were identified, and fixation and rubbing processes were found to be key for SGT aroma formation. Moreover, targeted metabolomics was applied to obtain 38 differential VMs and their related substances, of which fatty acid-derived volatiles (14 VMs) and glycoside-derived volatiles (8 VMs) showed significant contributions to SGT aroma, and their derivation laws during SGT manufacturing were clarified. Furthermore, the effect of rubbing degree on volatile metabolite formation was explored, and 11 key differential VMs were screened by variable importance in projection, and odor activity value analyses. Appropriate rubbing promoted the loss of grassy VMs (such as 1-octanol and 2-pentyl-furan) and enrichment of floral/fruity VMs (such as trans-ß-ionone, nonanal, geraniol, citral, (Z)-3,7-dimethyl-2,6-octadien-1-ol, and (Z)-hexanoic acid, 3-hexenyl ester). Our study not only enriches the chemical theory of green tea processing but also provides technical support for the precision directional processing of high-quality SGT.

First Report of Alternaria alternata Inciting Leaf Spot of Dangshen (Codonopsis pilosula) in China.

As a commonly traditional Chinese medicine, the perennial herb Dangshen (Codonopsis pilosula) has superior curative effects including regulating immunity, strengthening the spleen, and tonifying lungs (Bai et al. 2020). To imitate natural ecological conditions, plants were grown on hillside fields with stems prostrate on the ground, tangle-up with each other. In August 2020, leaf spots were observed on C. pilosula in Wutai county, Shanxi province, China, and indicated a high disease incidence (70%-80%) in investigated fields (6.67 ha). Small brown necrotic spots, occasionally enclosed by chlorotic halos, were observed on leaves, stems, and sepals. For identification of the pathogen, 15 small pieces (5×5 mm) of symptomatic tissues from 5 randomly-collected diseased plants were surface sterilized, placed on potato dextrose agar plates, and incubated for 4 d in darkness at 25 °C to obtain the colonies. Cultures were purified by single spore isolation from these colonies. A total of 15 isolates named as Dcp-3, and Dcp-5~Dcp-18 were recovered. They produced ovoid or obclavate spores with 15.9-57.5×9.1-20.1 µm in size, 1-6 transverse septa, and 0-4 longitudinal septa. The conidial chains with 4 to 6 spores had numerous secondary and occasionally tertiary chains on potato carrot agar plates. Because all isolates had identical morphological traits, five genes from the representative isolate Dcp-3, actin (ACT), Alternaria major allergen (Alt a1), plasma membrane ATPase (ATP), histone 3 (H3), and rDNA ITS, were amplified with primer pairs ACTDF1/ACTDR1, Alt-for/Alt-rev, ATPDF1/ATPDR1, H3-1a/H3-1b, and ITS1/ITS4, respectively (Hong et al. 2005; Lawrence et al. 2013; Ma et al. 2020). BLASTn searches indicated species of Dcp-3 could not be accurately confirmed by rDNA ITS, ATP, ACT, and Alt a1 (GenBank accession nos. OM334894, OM362504, OM326344, OM362500). Phylogenetic analysis showed it was most closely related to Alternaria alternata, A. arborescens, and A. tenuissima based on concatenated sequences of above four genes. The H3 sequence (OM362508) shared 100% homology with that of A. alternata (MN481948). The phylogenetic tree using H3 also confirmed Dcp-3 as A. alternata. Heathly, two-year-old C. pilosula were transplanted to a greenhouse. A surface-sterilized leaf was sprayed with 50 µL spore suspension (106 conidia/mL) of Dcp-3. A leaf sprayed with isometric sterile water was used as controls. Each treatment used six plants (five leaves per plant). Plants were covered with sterilized plastic bags and incubated at 22 ℃. The test was repeated twice. A week later, control leaves were healthy, but brown necrotic spots similar to field symptoms emerged on treated leaves. The A. alternata isolates were re-isolated from the border of lesions, and confirmed by morphological and molecular characteristics mentioned above, fulfilling Koch's postulates. Leaf spot of C. pilosula caused by Septoria codonopsidis has been reported in China (Wang et al. 2011). However, to our knowledge, this is the first report of A. alternata inciting leaf spot of C. pilosula in China. Our report would promote growers to enhance the field management and consider associated strategies on controlling Alternaria leaf spot of C. pilosula.

The efficacy and safety of auricular point pressing therapy for knee osteoarthritis: A protocol for systematic review and meta-analysis.

BACKGROUND: Knee osteoarthritis (KOA) is one of the leading causes of disability. The effectiveness of auricular point pressing therapy for treating KOA remains controversial. This protocol describes the method of a systematic review and meta-analysis evaluating the effectiveness and safety of auricular point pressing therapy for treating KOA. METHODS: Four English databases (PubMed, Embase, Cochrane Library databases and Web of Science) and 4 Chinese databases (China National Knowledge Infrastructure, Chinese Biomedical Literature Database, VIP Database for Chinese Technical Periodicals, and Wanfang) will be searched. All randomized controlled trials related to auricular point pressing therapy for KOA will be included. Extracted data will include publication details, basic information, demographic data, intervention details and patient outcomes. The primary outcome will be Western Ontario and McMaster Universities Osteoarthritis Index and visual analogue scale. Risk of bias will be assessed using the Cochrane Collaboration tool for assessing risk of bias. Article selection, data extraction and risk of bias assessment will be performed by 2 independent reviewers. If the meta-analysis is precluded, we will conduct a descriptive synthesis using a best-evidence synthesis approach. The strength of recommendations and quality of evidence will be assessed using the Grading of Recommendations Assessment Development. RESULTS: The systematic review will provide evidence to assess the efficacy and safety of auricular point pressing therapy on KOA. CONCLUSION: The systematic review will provide evidence to assess the efficacy and safety of auricular point pressing therapy for KOA patients. ETHICS AND DISSEMINATION: For this review, ethical approval is not required. Patients will not be involved. The findings will be published in a peer-reviewed journal. INPLASY REGISTRATION NUMBER: INPLASY 202220077.

New insights into the effect of fermentation temperature and duration on catechins conversion and formation of tea pigments and theasinensins in black tea.

BACKGROUND: The phenol oxidative pathway during fermentation remains unclear. To elucidate the effect of fermentation on phenol conversion, we investigated the effects of fermentation temperature and duration on the conversion of catechins and the formation of theasinensins (TSs), theaflavins (TFs), thearubigins (TRs), and theabrownins (TBs). RESULTS: During fermentation, TSs formation increased initially and then decreased. Long fermentation durations were unfavorable for liquor brightness (LB) and resulted in the production of large amounts of TRs and TBs. Low fermentation temperatures (20 °C and 25 °C) favored the maintenance of polyphenol oxidase activity and the continuous formation of TFs, TSs, and TRSI (a TRs fraction), resulting in better LB and liquor color. Higher temperatures (30 °C, 35 °C, and 40 °C) resulted in higher peroxidase activity, higher oxidative depletion rates of catechins, and excessive production of TRSII (a TRs fraction) and TBs. Analysis of the conversion pathway of polyphenolic compounds during fermentation showed that, during early fermentation, large amounts of catechins were oxidized and converted to TFs and theasinensin B. As fermentation progressed, considerable amounts of theaflavin-3'-gallate, theasinensin A, theaflavin-3-gallate, theaflavin-3,3'-digallate, and theasinensin C were produced and then converted to TRSI; in the final stage, TRSII and TBs were converted continuously. CONCLUSION: Different fermentation temperature and duration combinations directly affected the type and composition of phenolic compounds. The key conditions for controlling phenolic compound conversion and fermentation direction were 60 or 90 min and 25 or 30 °C. Our study provides insights into the regulation of phenolic compound conversion during black tea fermentation. © 2021 Society of Chemical Industry.

Widely targeted metabolomic analysis reveals dynamic changes in non-volatile and volatile metabolites during green tea processing.

Non-volatile metabolites significantly influence the color, taste, and aromatic qualities of green tea. However, the evolutionary trajectories of non-volatile metabolites, and their transformational relationship with volatile metabolites during processing, remain unclear. In this study, ultra-performance liquid chromatography-tandem mass spectrometry and gas chromatography-tandem mass spectrometry were used to analyze a widely targeted metabolome during green tea processing. In total, 527 non-volatile metabolites, covering 11 subclasses, were identified, along with 184 volatile metabolites, covering 8 subclasses. Significant variations in metabolites were observed during processing, especially in the fixation stage, and the conversion intensity of non-volatile metabolites was consistent with the law of "Fixation > Drying > Rolling." A total of 153 non-volatile metabolites were screened out, and amino acids and esters were found to be closely associated with volatile metabolite formation. The results of the present study provide a theoretical basis that could guide green tea processing based on desired quality and components.

Influence of fixation methods on the chestnut-like aroma of green tea and dynamics of key aroma substances.

Fixation is the key process to ensure green tea quality; however, the effect of various fixation methods on the formation of green tea with a chestnut-like aroma and the evolution of key volatile compounds has not been assessed to date. In this study, we compared four types of fixation methods for green tea: roller-hot air-steam, roller-hot air, roller-steam, and single roller. Infrared-assisted headspace solid-phase microextraction and gas chromatography-tandem dual mass spectrometry technology were used to detect the volatile compounds of green tea samples during processing. Partial least-squares discriminant analysis (PLS-DA), multiple experiment viewer (MEV), odor activity value (OAV), and least-significant difference analyzes were then applied to clarify the best fixation method for forming a chestnut-like aroma and associated compounds, and to explore the change law of key volatile compounds using different green tea fixation processes. One hundred and eighty-four volatile compounds were detected in the processed samples, with roller-hot air fixation found as the optimal method for generating an intense and long-lasting chestnut-like aroma and floral taste, based on sensory evaluation. The PLS-DA model clearly distinguished the four kinds of fixation samples and obtained 32 differential volatile compounds. Combining OAVs with screening by MEV analysis, 2,6,10,10-tetramethyl-1-oxaspiro [4.5] dec-6-ene, linalool, cedrol, 3-methyl-butanal, trans-ß-ionone, and τ-cadinol emerged as key differential volatile compounds between green teas with and without a chestnut-like aroma. The evolution of these six differential volatile compounds throughout the tea-making process confirmed that rolling-hot air coupling treatment is most conducive to produce a chestnut-like aroma, which is beneficial to form and transform 2,6,10,10-tetramethyl-1-oxaspiro[4.5] dec-6-ene, 3-methyl-butanal, and τ-cadinol with baking aromas and fruity substances. These results provide a theoretical basis and technical guidance for the precise and directional processing of high-quality green tea with a chestnut-like aroma.

Different DHA or EPA production responses to nutrient stress in the marine microalga Tisochrysis lutea and the freshwater microalga Monodus subterraneus.

This study investigated the effect of nitrogen (N) and phosphorous (P) stress on the production of DHA or EPA and total fatty acids (TFAs) in the marine microalga Tisochrysis lutea and the freshwater microalga Monodus subterraneus. Five N or P starvation/limitation conditions (N sufficient and P limited, N sufficient and P starved, N starved and P sufficient, N starved and P limited, and N and P starved) and one N and P sufficient condition (control) were studied. The results demonstrated that the proportion of DHA or EPA among TFAs and production in the microalgae suspensions decreased (57%, 73% for N stress and 18%, 51% for P stress, respectively) under N or P stress in both microalgae compared with the N and P sufficient group. Differently, DHA dry weight content of T. lutea decreased significantly, and EPA dry weight content of M. subterraneus decreased slightly under N starved conditions. Clear differences in TFA content/production and the relationship between TFA and DHA or EPA production/content and CO2 fixation were observed between the two microalgae. These results give a new sight on the difference between marine microalgae and freshwater microalgae. Meanwhile, it gave a potential application to produce DHA or EPA and TFA combining with CO2 fixation by these microalgae.