Revealing the variances in color formation and bioactivities of seven catechin monomers throughout the enzymatic reaction by colorimetric and mass spectrometry.

The capacity differences of seven catechin monomers to produce colors after treating with catechin-free extract were investigated. After 240-min reaction, only (-)-epicatechin (EC) and (+)-catechin (C) presented obvious luminous red color with L* values of 63.32-71.73, a* values of 37.13-46.44, and b* values of 65.64-69.99. Meanwhile, the decrease rate of EC and C was 43.52 %-50.35 %, which were significantly lower than those of other catechin monomers (85.91 %-100 %). The oxidized products of catechin monomers were analyzed by ultra-high performance liquid chromatography-quadrupole-time of flight-mass spectrometry coupled with diode array detector, wherein dehydro-dimers and -trimers (oxidative coupling products of catechins' A-B ring) were found to be the major chromogenic compounds of EC and C. Additionally, the antioxidant capacity of catechin monomers only decreased after 30-min reaction, while along with further enzymatic reaction, catechin monomers presented comparable oxyradical scavenging ability (e.g., the DPPH inhibitory rates of catechin monomers were in the range of 24.42 %-50.77 %) to vitamin C (positive control, DPPH inhibitory rate was 27.66 %). Meanwhile, the inhibitory effects of most catechin monomers on α-glucosidase were enhanced in different degrees. These results provided basis for the development of enzymatically-oxidized catechin monomers as functional food color additives.

Chemical, sensory and biological variations of black tea under different drying temperatures.

As the final processing step, drying temperature between 90 and 140 â„ƒ is usually applied to terminate enzymatic activities and improve sensory characteristics of black tea. Liquid chromatography tandem mass spectrometry (LC-MS) based non-targeted and targeted metabolomics analyses combined in vitro biological assays were adopted to investigate the chemical and biological variations after drying. Fifty-nine differentially expressed metabolites including several hydroxycinnamic acid derivatives and pyroglutamic acid-glucose Amadori rearrangement products (ARPs) were identified, the latter of which was correspondingly accumulated with increasing temperature. The levels of theaflavins (TFs), thearubigins (TRs), monosaccharides and free amino acids gradually decreased with increasing temperature. Furthermore, the bioassays of black tea showed that drying under 110 â„ƒ provided the highest antioxidant capacities, but the inhibitory effects on α-glucosidase and α-amylase were decreasing along with increasing drying temperature. These results are valuable for optimizing drying process to obtain superior sensory properties and preserve bioactivities of black tea.

Formation of Furan from Linoleic Acid Thermal Oxidation: (E,E)-2,4-Decadienal as a Critical Intermediate Product.

The linoleic acid reaction models were set at 150 °C for 120 min, and its oxidation process was monitored by nuclear magnetic resonance (NMR) and gas chromatography-mass spectrometry (GC-MS). Results showed that no furan was formed from linoleic acid without heating, while furan accumulated throughout the heating process. Linoleic acid ran out within 30 min, which indicated that furan was formed mainly from the intermediate oxidation products of linoleic acid after 30 min. It should be noticed that the content of (E,E)-2,4-decadienal reached maximum once the linoleic acid ran out and then decreased with the formation of furan. Multivariate statistical analysis suggested that (E,E)-2,4-decadienal was the most important aldehyde related to furan formation during linoleic acid oxidation. To prove this assumption, the variation of furan from (E,E)-2,4-decadienal reaction models heating at 150 °C for 60 min was also studied. Results showed that the content of furan increased with the oxidation of (E,E)-2,4-decadienal. Furthermore, NMR and GC-MS data proved that (E,E)-2,4-decadienal could be oxidized to 4,5-epoxy-(E)-2-decenal. In conclusion, our results supported (E,E)-2,4-decadienal and trans-4,5-epoxy-(E)-2-decenal as critical intermediate products of furan formation from linoleic acid oxidation.

Comprehensive applications of metabolomics on tea science and technology: Opportunities, hurdles, and perspectives.

With the development of metabolomics analytical techniques, relevant studies have increased in recent decades. The procedures of metabolomics analysis mainly include sample preparation, data acquisition and pre-processing, multivariate statistical analysis, as well as maker compounds' identification. In the present review, we summarized the published articles of tea metabolomics regarding different analytical tools, such as mass spectrometry, nuclear magnetic resonance, ultraviolet-visible spectrometry, and Fourier transform infrared spectrometry. The metabolite variation of fresh tea leaves with different treatments, such as biotic/abiotic stress, horticultural measures, and nutritional supplies was reviewed. Furthermore, the changes of chemical composition of processed tea samples under different processing technologies were also profiled. Since the identification of critical or marker metabolites is a complicated task, we also discussed the procedure of metabolite identification to clarify the importance of omics data analysis. The present review provides a workflow diagram for tea metabolomics research and also the perspectives of related studies in the future.

Novel Oxidation Oligomer of Chlorogenic Acid and (-)-Epigallocatechin and Its Quantitative Analysis during the Processing of Keemun Black Tea.

Not only do flavan-3-ols participate in the formation of chromogenic oxidation products such as theaflavins, but chlorogenic acid (3-caffeoylquinic acid, CQA) is also involved in the enzymatic oxidation during black tea processing. The critical oxidation product of CQA and (-)-epigallocatechin (EGC) were identified as an adduct containing benzobicyclo[3.2.2]nonenone structure, which was named as the dichlorogeniccatechin (DCGC) oligomer. It was composed of two molecules of CQA and one molecule of EGC. The effects of the initial reactant ratio and reaction time on the generation of DCGC were also analyzed. A high proportion of CQA promoted the production of DCGC, but a high proportion of EGC inhibited the DCGC formation. In addition, the content of DCGC in Keemun black tea during processing was determined. The content of DCGC highly increased after withering but decreased after drying. This study provides a new perspective for the investigation of other oxidation oligomers in black tea.

Clinical efficacy of weight loss herbal intervention therapy and lifestyle modifications on obesity and its association with distinct gut microbiome: A randomized double-blind phase 2 study.

Goals: To assess the efficacy and safety of Chinese Medicine Prescription "W-LHIT" in subjects with simple obesity, and to explore its potential mechanism of action. Methods: Thirty-seven patients aged 18 to 60 from Wei-En hospital (Weifang City, Shandong, China), participated in a double blinded, placebo-controlled study. Subjects were randomly divided into 2 groups, 18 in treatment and 19 in placebo group. The treatment group took the "W-LHIT" capsules for two months, while the control group received placebo capsules. Both groups accepted healthy lifestyle education materials. After a 2-month treatment, the placebo group transferred to open-label treatment after unblinding. Results: 72.22% participants in the treatment group lost more than 5% of their body weight, compared with 36.84% in the placebo group (p < 0.001). Body weight loss and body mass index reduction of the treatment group were also significantly higher than those of the placebo group (p < 0.05). These changes were accompanied by increased abundance of Akkermansia muciniphila and Enterococcus faecium, and decreased abundance of Proteobacteria in gut microbiota. Furthermore, the treatment group also showed improvement in obesity-related comorbidities such as hypertension and elevation of liver enzymes. No serious adverse reactions were found during the study period. Weight did not rebound at a follow-up visit 2 months after treatment. Conclusion: W-LHIT significantly improved body weight and comorbid conditions without obvious adverse reaction or rebound weight gain. These effects were associated with increased abundance of probiotics in gut microbiota. W-LHIT may have a potential for treating obesity in conjunction with healthy lifestyle modifications.

Amadori Reaction Products of Theanine and Glucose: Formation, Structure, and Analysis in Tea.

Amadori rearrangement products (ARPs) derived from the Maillard reaction between theanine and glucose (ARP 1), as well as pyroglutamic acid and glucose (ARP 2), were identified by liquid chromatograph tandem mass spectroscopy methods. The effects of initial reactant ratio, temperature, pH, and heating time on ARP generation were analyzed. The formation of both ARPs was most favored under 100 °C, while an alkaline environment slightly promoted the generation of ARP 1 and acidic conditions contributed more to ARP 2 formation. The decomposition of ARP 1 was suggested to be the predominant formation mechanism of ARP 2. Preparation, purification, and structure identification of ARP 1 were conducted, with its structure confirmed as 1-deoxy-1-l-theanino-d-fructose. The contents of ARP 1 in green, black, dark, white, yellow, and Oolong teas were quantitatively determined, of which black teas contained the highest levels of ARP 1, possibly due to the high glucose content and processing techniques.

Comparison of the chemical composition and antioxidant, anti-inflammatory, α-amylase and α-glycosidase inhibitory activities of the supernatant and cream from black tea infusion.

Tea cream is a kind of turbid substance commonly existing in tea infusion and tea beverage upon cooling. Herein, a comparative study was conducted on the supernatant and cream from black tea infusion in terms of antioxidant, anti-inflammatory and enzyme inhibitory activities, and chemical composition. Ultraviolet-visible (UV-vis) spectrometry and high-performance liquid chromatography (HPLC) analysis showed that the contents of protein, polyphenols, theaflavins, thearubigins, theabrownins, and caffeine in cream were significantly higher than those in the supernatant. The contents of Al, Ca, Cu, and Fe elements in cream were higher than those in the supernatant. However, higher levels of monosaccharides and free amino acids were detected in the supernatant compared with cream. The ultra-performance liquid chromatography quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) based metabolomics analysis revealed that the main marker compounds between the supernatant and the cream were organic acids, phenolic acids, and flavan-3-ols and their oxidation products, flavonol glycosides and amino acids. The cream showed better antioxidant and anti-inflammatory, as well as α-amylase and α-glycosidase inhibitory activities than the supernatant, because it contained higher contents of polyphenols than the supernatant. The present study expanded the new vision towards the cream of black tea infusion.

The Oxidation Mechanism of Flavan-3-ols by an Enzymatic Reaction Using Liquid Chromatography-Mass Spectrometry-Based Metabolomics Combined with Captured o-Quinone Intermediates of Flavan-3-ols by o-Phenylenediamine.

During the enzymatic oxidation of black tea, flavan-3-ols undergo a complicated chemical transformation and generate theaflavins and thearubigins. So far, the oxidation mechanism of flavan-3-ols has not been clarified. Liquid chromatography-tandem mass spectrometry-based metabolomics combined with o-quinone intermediates captured by o-phenylenediamine was developed and successfully applied in the liquid incubation of fresh tea homogenates. During the oxidation, the contents of catechins continuously decreased, while theaflavins increased first but decreased subsequently at the end of incubation. Meanwhile, the content of thearubigins greatly increased at the late stage of incubation. Dehydrotheasinensins were accumulated at the end of oxidation along with the decrease of theasinensins. Through o-phenylenediamine derivation, several adducts of (-)-epigallocatechin gallate, (-)-epigallocatechin, theasinensins A, B, C, and D, and corresponding dehydrotheasinensins were identified, which were considered as the substrates of thearubigins. These results suggested that theaflavins and these oxidation products contributed to the formation of thearubigins.

Free, soluble conjugated and insoluble bonded phenolic acids in Keemun black tea: From UPLC-QQQ-MS/MS method development to chemical shifts monitoring during processing.

Phenolic acids, including benzoic acid and hydroxycinnamic acid derivatives, are the main compounds of black tea. An efficient and accurate analytical method to quantify ten phenolic acids was established and validated by ultra-performance liquid chromatography coupled with triple quadrupole mass spectrometry (UPLC-QQQ-MS/MS). The chemical shifts during the processing of Keemun black tea were analyzed and the phenolic acids were quantified. Compared with fresh tea leaves, after processing, the contents of free phenolic acids, including gallic acid, salicylic acid, p-coumaric acid, and ferulic acid, increased markedly. Still, the contents of soluble conjugated phenolic acids, including p-coumaroylquinic acid isomers and chlorogenic acid isomers, decreased. Furthermore, the total contents of lignin, and insoluble bonded phenolic acids decreased. The adduct of (-)-epigallocatechin gallate and 3-caffeoylquinic acid was detected in tea samples, and its content increased highly after fermentation. The developed and validated analytical method can be used to monitor the manufacturing process of black tea.

Chemical Variation of Chenpi (Citrus Peels) and Corresponding Correlated Bioactive Compounds by LC-MS Metabolomics and Multibioassay Analysis.

The peel of Citrus reticulata "Chachi" (CP) possesses various health-promoting benefits and is not only one of the most famous Chinese herbal medicine, but also an ingredient in fermented foods. In the present study, the effects of storage years (1-, 3-, 4-, 5-, 6-, and 11-years) on the chemical profiling and potential bioactive compounds of CP were compared by metabolomics and in vitro bioactivity analysis. With the increase of storage time, the content of hesperidin significantly decreased, but nobiletin, 3,5,6,7,8,3',4'-heptamethoxyflavone, and tangeretin were increased. Meanwhile, the antioxidant activity of CP was enhanced. Phenolic acids, flavonol glycosides, fatty acids, and alkyl glycosides were marker compounds that were responsible for distinguishing the storage time of CP. Correlation analysis suggested that some polyphenols including quercetin-glucoside, quinic acid, trihydroxydimethoxyflavone, and rutin were potential antioxidant compounds in CP. The dichloromethane and n-butanol fractions showed the better antioxidant capacity and inhibitory effects on glucose-hydrolysis enzymes. They mainly contained ferulic acid, nobiletin, 3,5,6,7,8,3',4'-heptamethoxyflavone, kaempferol, and hesperidin.

Identification of low-molecular-weight color contributors of black tea infusion by metabolomics analysis based on UV-visible spectroscopy and mass spectrometry.

Nine black tea samples with different color intensity were firstly determined by chromatic difference analyzer. The color characteristics were secondly quantitatively described by UV-visible spectroscopy. Thirdly, liquid chromatography tandem mass spectrometry (LC-MS) based metabolomics analysis was applied in low-molecular-weight compounds. Finally, the color contributors were identified by the correlation analysis of color, spectrometry and mass data. UV-visible based metabolomics analysis revealed that the wavelength at 380-520 nm (VIP > 1.50) was the critical absorbance band for distinguishing different color of BT infusions, while LC-MS based metabolomics analysis indicated that there were 48 main marker compounds responsible for the classification of different BT infusions. Correlation analysis results showed that the coefficients of theaflavins, thearubigins, theabrownins, flavonoid glycosides, and some hydroxycinnamoyl acids were > 0.7, which suggested they were main color contributors of BT infusion. The present study expanded a new vision on the color analysis of BT infusion.

Study on In Vitro Preparation and Taste Properties of N-Ethyl-2-Pyrrolidinone-Substituted Flavan-3-Ols.

N-ethyl-2-pyrrolidinone-substituted flavan-3-ols (EPSFs) were prepared by an in vitro model reaction, and the taste thresholds of EPSFs and their dose-over-threshold factors in large-leaf yellow tea (LYT) were investigated. The effects of initial reactant ratios, reaction temperatures and time, pH values, and water addition on the yield of EPSFs were explored. The contents of EPSFs during roasting were determined by liquid chromatography quadrupole time-of-flight mass spectrometry (LC-Q-TOF-MS). When the initial ratio of (-)-epigallocatechin gallate (EGCG) to theanine was 1:2 and roasted under 120 °C for 120 min, the contents of EPSFs were the highest. The bitterness and astringency thresholds of four EPSF isomers were measured by the half-tongue method, of which EPSF2 and EPSF3 had higher thresholds than EGCG. In LYT, four EPSFs had lower bitterness and astringency dose-over-threshold factors than EGCG. This study suggested that the reduction of bitterness and astringency of tea after roasting may be mainly due to the formation of EPSFs.

From the forest to the plate - Hemicelluloses, galactoglucomannan, glucuronoxylan, and phenolic-rich extracts from unconventional sources as functional food ingredients.

This study aimed to characterise pressurised hot water (PHW) extracts from nonconventional sources of functional carbohydrates and phenolic compounds in terms of antioxidant capacity, antiviral activity, toxicity, and human erythrocytes' protection antidiabetic potential. PHW extracts of Norway spruce bark (E1 + E2) and Birch sawdust (E3 + E4) contained mostly galactoglucomannan and glucuronoxylan. In contrast, samples E5 to E9 PHW extracted from Norway spruce, and Scots pine bark are rich sources of phenolic compounds. Overall, phenolic-rich extracts presented the highest inhibition of α-amylase and α-glucosidase and protection against stable non-enveloped enteroviruses. Additionally, all extracts protected human erythrocytes from hemolysis. Cell-based experiments using human cell lines (IMR90 and A549) showed extracts' non-toxicin vitroprofile. Considering the relative toxicological safety of extracts from these unconventional sources, functional carbohydrates and polyphenol-rich extracts can be obtained and further used in food models.

LC-MS based metabolomics and sensory evaluation reveal the critical compounds of different grades of Huangshan Maofeng green tea.

Six grades of Huangshan Maofeng (HSMF) green tea were studied by LC-MS based metabolomics combined with sensory evaluation on bitterness, astringency and sweet aftertaste. Although there was no significant correlation (p > 0.05) between tea grades and the contents of total polyphenols and flavonoids, non-targeted metabolomics revealed that all grades of tea could be classified into two groups, group 1 (T1, T2) and group 2 (T3, 1, 2, 3). The main marker compounds responsible for distinguishing the two groups were procyanidins, flavonoid glycosides, and four hydrolysable tannins, including monogalloyl glucose, digalloyl glucose, trigalloyl glucose and galloyl-hexahydroxydiphenoyl-glucose. The Pearson correlation coefficients of these hydrolysable tannins with HSMF green tea grades were between 0.82 and 0.95. Furthermore, their Pearson correlation coefficients regarding sweet aftertaste were in the range of 0.73-0.83. This study suggested combination of metabolomics and sensory evaluation could provide an insight in searching for more potential taste-active components.

Purple tea (Camellia sinensis var. assamica) leaves as a potential functional ingredient: From extraction of phenolic compounds to cell-based antioxidant/biological activities.

A statistical optimization study was used to maximize the extraction of bioactive compounds and antioxidant activity from green tea derived from purple leaves of Camellia sinensis var. assamica. Simultaneous optimization was applied, and a combination of 60 °C, 15 min, and a mass-solvent ratio of 1 g of dehydrated purple leaves to 62.3 mL of an ethanol/citric acid solution, were determined as the ideal extraction conditions. The optimized extract of purple tea leaves (OEPL) contained showed stability in relation to variations in pH, and lyophilized OEPL exerted cytotoxic and antiproliferative effects against cancerous cells (A549 and HCT8), demonstrated antimicrobial activity towards Listeria monocytogenes (ATCC 7644), Staphylococcus aureus (ATCC 13565) and Staphylococcus epidermidis (ATCC 12288), inhibition of α-amylase and α-glycosidase enzymes and reduced the release of pro-inflammatory cytokines (TNF-α, CXCL2/MIP-2, and IL-6) in lipopolysaccharides-stimulated RAW 264.7 macrophages. Thus, our results provide a broad assessment of the bioactivity of "green" extracts obtained by a simple and low-cost process using non-toxic solvents, and they have the potential to be used for technological applications.

Identification of 4-O-p-coumaroylquinic acid as astringent compound of Keemun black tea by efficient integrated approaches of mass spectrometry, turbidity analysis and sensory evaluation.

Hydroxycinnamoyl quinic acids are important phenolic acids in tea, particularly fermented teas. However, there have been fewer studies that have confirmed their taste properties. The aim of this study was to investigate the astringent compounds in Keemun congou black tea (KBT) using a combination of mass spectrometry, turbidity analysis, and sensory evaluation. Turbidity analysis determined that p-coumaroylquinic acids were the astringent contributing compounds in KBT. Moreover, the separated compound D16 was identified as trans-4-O-p-coumaroylquinic acid (trans-4-O-pCoQA) by nuclear magnetic resonance spectroscopy and first confirmed to be the astringent contributing compound in KBT by sensory evaluation. Its astringent threshold concentration was tested to be 38 µM. The trans-4-O-pCoQA content in eight KBT samples of various grades ranged from 40.20 ± 0.15 ~ 65.53 ± 0.22 µM. Turbidity analysis combined with sensory evaluation could be a powerful tool for identifying critical compounds responsible for the astringent taste.

Model Studies on the Reaction Products Formed at Roasting Temperatures from either Catechin or Tea Powder in the Presence of Glucose.

During tea processing, roasting significantly affects the transformation pathway of catechins. When (-)-epigallocatechin gallate (EGCG) and glucose were roasted at different pH values, the degree of degradation and isomerization of EGCG was the lowest at pH 7 and the highest at pH 8. Thirty-five products were found in the model reaction of EGCG and glucose under high temperatures, of which four EGCG-glucose adducts were identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and nuclear magnetic resonance (NMR). In addition, catechins, gallic acid, and theanine in tea with added glucose were significantly reduced during roasting. The contents of four EGCG-glucose adducts were increased significantly at 150 °C after 30 min and dropped gradually after 60 min. Therefore, based on the present study, EGCG could form crosslinks with glucose under high temperatures in a short time, which provides insight for tea processing and synthesis of catechin-sugar adducts.

Quantitative changes in monosaccharides of Keemun black tea and qualitative analysis of theaflavins-glucose adducts during processing.

Monosaccharides of Keemun black tea were quantitatively determined by high performance liquid chromatography coupled with 3-methyl-1-phenyl-2-pyrazolin-5-one (PMP) pre-column derivatization. The methodology of developed analytical method was established with good linearity, recovery, repeatability and precision. The quantitative results showed that D-mannose, D-glucuronic acid, D-glucose, D-galactose and L-arabinose were detected in Keemun black tea samples. D-glucose was the predominant monosaccharide in black tea, and its concentration was continuously increased from fresh tea leaves to fermentation, but after drying its concentration was significantly decreased. Meanwhile, theaflavins' concentrations were obviously decreased after drying. When theaflavins were heated with D-glucose, the loss of theaflavins was increased. Correspondingly, theaflavins also prevented the caramelization of D-glucose and restored the loss of D-glucose during heating. Through the liquid chromatography/electrospray tandem mass spectrometry some theaflavins glucose adducts were identified.

Toxicological and bioactivity evaluation of blackcurrant press cake, sea buckthorn leaves and bark from Scots pine and Norway spruce extracts under a green integrated approach.

Aqueous extracts from blackcurrant press cake (BC), Norway spruce bark (NS), Scots pine bark (SP), and sea buckthorn leaves (SB) were obtained using maceration and pressurized hot water and tested for their bioactivities. Maceration provided the extraction of higher dry matter contents, including total phenolics (TPC), anthocyanins, and condensed tannins, which also impacted higher antioxidant activity. NS and SB extracts presented the highest mean values of TPC and antioxidant activity. Individually, NS extract presented high contents of proanthocyanidins, resveratrol, and some phenolic acids. In contrast, SB contained a high concentration of ellagitannins, ellagic acid, and quercetin, explaining the antioxidant activity and antibacterial effects. SP and BC extracts had the lowest TPC and antioxidant activity. However, BC had strong antiviral efficacy, whereas SP can be considered a potential ingredient to inhibit α-amylase. Except for BC, the other extracts decreased reactive oxygen species (ROS) generation in HCT8 and A549 cells. Extracts did not inhibit the production of TNF-alpha in lipopolysaccharide-stimulated THP-1 macrophages but inhibited the ROS generation during the THP-1 cell respiratory burst. The recovery of antioxidant compounds from these by-products is incentivized for high value-added applications.