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2-Phenylethanol (2PE) is a representative aromatic aroma compound in tea (Camellia sinensis) leaves. However, its formation in tea remains unexplored. In our study, feeding experiments of [2H8]L-phenylalanine (Phe), [2H5]phenylpyruvic acid (PPA), or (E/Z)-phenylacetaldoxime (PAOx) showed that three biosynthesis pathways for 2PE derived from L-Phe occurred in tea leaves, namely, pathway I (via phenylacetaldehyde (PAld)), pathway II (via PPA and PAld), and pathway III (via (E/Z)-PAOx and PAld). Furthermore, increasing temperature resulted in increased flux into the pathway for 2PE from L-Phe via PPA and PAld. In addition, tomato fruits and petunia flowers also contained the 2PE biosynthetic pathway from L-Phe via PPA and PAld and increasing temperatures led to increased flux into this pathway, suggesting that such a phenomenon might be common among most plants containing 2PE. This represents a characteristic example of changes in flux into the biosynthesis pathways of volatile compounds in plants in response to stresses.

Fragrance is an important quality index of horticultural flowers. Floral volatile formation in flowers during plant growth has been widely studied, but less is known about floral volatile formation in cut flowers and its responses to postharvest conditions. In this study, cut rose (Rosa hybrida cv. Tineke) flowers subjected to 5, 15 and 30 °C for 36 h showed increased concentrations of volatile phenylpropanoids/benzenoids (VPBs) including 2-phenylethanol (2PE), phenylacetaldehyde, benzyl alcohol, benzaldehyde, and phenethyl acetate, but a reduced 3,5-dimethoxytoluene concentration, as temperatures increased. l -[2H8]Phenylalanine (Phe) tracing in vivo suggested that phenylpyruvic acid (PPA) was involved in the increase in 2PE in response to increasing temperature. Genes for two aromatic amino acid aminotransferases (AAATs) were isolated and functionally characterized. Transient expression analyses in Nicotiana benthamiana plants provided in vivo evidence that RhAAAT2 was able to convert l -Phe into PPA, and that it was localized in the cytoplasm. These results advance our understanding of floral aroma formation in flowers after harvest.

Correction for ‘Insects (Thrips hawaiiensis (Morgan)) change the stereochemical configuration of 1-phenylethanol emitted from tea (Camellia sinensis) flowers’ by Ying Zhou et al., RSC Adv., 2017, 7, 32336–32343. DOI: 10.1039/C7RA03219F.

Metabolite formation is a biochemical and physiological feature of plants developed as an environmental response during the evolutionary process. These metabolites help defend plants against environmental stresses, but are also important quality components in crops. Utilizing the stress response to improve natural quality components in plants has attracted increasing research interest. Tea, which is processed by the tender shoots or leaves of tea plant (Camellia sinensis (L.) O. Kuntze), is the second most popular beverage worldwide after water. Aroma is an important factor affecting tea character and quality. The defense responses of tea leaves against various stresses during preharvest (tea growth process) and postharvest (tea manufacturing) processing can result in aroma formation. Herein, we summarize recent investigations into the biosyntheses of several characteristic aroma compounds prevalent in teas and derived from volatile fatty acid derivatives, terpenes, and phenylpropanoids/benzenoids. Several key aroma synthetic genes from tea leaves have been isolated, cloned, sequenced, and functionally characterized. Biotic stress (such as tea green leafhopper attack) and abiotic stress (such as light, temperature, and wounding) could enhance the expression of aroma synthetic genes, resulting in the abundant accumulation of characteristic aroma compounds in tea leaves. Understanding the specific relationships between characteristic aroma compounds and stresses is key to improving tea quality safely and effectively.

A series of imidazolium salts with various N-aryl groups were synthesized, and their catalytic activities were evaluated to investigate the contribution of the N-aryl groups to the catalytic activity in the synthesis of γ-butyrolactone through an a3→d3-umpolung addition. Imidazolylidenes with 2,6-diethylphenyl groups were effective catalysts, and several mechanistic studies, including a deuterium kinetic isotope effect study, revealed that both steric and kinetic effects were responsible for the enhanced catalytic activity.

2-Phenylethanol (2PE) is a representative aromatic aroma compound in tea (

Plants synthesize and emit a large variety of volatile organic compounds, which possess extremely important ecological functions. Rapid progress in the method and instruments available to separate and identify plant metabolites has led to intensive investigations of the biosynthetic pathways that form plant volatiles, which has resulted in insightful and useful discoveries. In this review, we summarize and present the successful techniques currently used for studying metabolites involved in the formation of the major classes of plant volatile compounds, including volatile fatty acid derivatives, volatile isoprenoids, volatile phenylpropanoids/benzenoids, carotenoid-derived aroma compounds, and glycosidically bound volatile compounds. We discuss the extraction, identification, and quantitative analysis of many of these metabolites, which are unstable and occur in plants at trace levels.

Insect attacks change the quantity of volatiles emitted from plants, but little is known about the effects on the volatiles' stereochemical configurations. 1-Phenylethanol (1PE) is a major internal volatile of tea (Camellia sinensis) flowers. In the present study, insect-damaged C. sinensis flowers in the field emitted a different (R)-/(S)-1PE pattern than that emitted from undamaged flowers. Laboratory experiments showed that changes in the (R)-/(S)-1PE ratio were caused by treatments with Thrips hawaiiensis (Morgan), a pest of cut flowers and vegetable crops. These treatments led to an increase in jasmonic acid (JA) through the activation of allene oxide synthase, which is involved in JA biosynthesis. Exposing flowers to exogenous JA induced an (R)-/(S)-1PE emission pattern that was similar to that induced by a T. hawaiiensis attack, suggesting that JA was involved in the changes in the stereochemical configuration of 1PE emitted from the flowers after insect treatments. This study provides new insights into the involvement of volatiles in interactions between plants and insects, showing that not only the quantity but also the stereochemical configuration of plant volatiles is affected by insects.

Catechins and theaflavins are important metabolites contributing to tea function and quality. Catechins are known to transform into theaflavins during the tea manufacturing process, but the same transformation in preharvest tea leaves is unknown. Herein, we determined that shade treatment (dark), an agronomic practise widely used in tea cultivation, reduced the contents of most catechins, but increased the theaflavin contents, in preharvest tea leaves (cv. Yinghong No.9). This was attributed to the activation of polyphenoloxidase (PPO) activity in darkness. Furthermore, CsPPO3 was highly expressed under darkness, and thus CsPPO3 had been cloned, sequenced, and characterization. The CsPPO3 recombinant protein exhibited PPO function. Furthermore, shade treatment also reduced the catechin contents and increased the theaflavin contents in Yabukita and Hoshinomidori, suggesting that this phenomenon might not be specific to certain tea cultivars. This information will aid in understanding of theaflavin formation and its response to environmental factors at the preharvest tea stage.

Metabolite formation is a biochemical and physiological feature of plants developed as an environmental response during the evolutionary process. These metabolites help defend plants against environmental stresses, but are also important quality components in crops. Utilizing the stress response to improve natural quality components in plants has attracted increasing research interest. Tea, which is processed by the tender shoots or leaves of tea plant (

Tea plants (Camellia sinensis) store volatile organic compounds (VOCs; monoterpene, aromatic, and aliphatic alcohols) in the leaves in the form of water-soluble diglycosides, primarily as β-primeverosides (6-O-β-d-xylopyranosyl-β-d-glucopyranosides). These VOCs play a critical role in plant defenses and tea aroma quality, yet little is known about their biosynthesis and physiological roles in planta. Here, we identified two UDP-glycosyltransferases (UGTs) from C. sinensis, UGT85K11 (CsGT1) and UGT94P1 (CsGT2), converting VOCs into β-primeverosides by sequential glucosylation and xylosylation, respectively. CsGT1 exhibits a broad substrate specificity toward monoterpene, aromatic, and aliphatic alcohols to produce the respective glucosides. On the other hand, CsGT2 specifically catalyzes the xylosylation of the 6′-hydroxy group of the sugar moiety of geranyl β-d-glucopyranoside, producing geranyl β-primeveroside. Homology modeling, followed by site-directed mutagenesis of CsGT2, identified a unique isoleucine-141 residue playing a crucial role in sugar donor specificity toward UDP-xylose. The transcripts of both CsGTs were mainly expressed in young leaves, along with β-PRIMEVEROSIDASE encoding a diglycoside-specific glycosidase. In conclusion, our findings reveal the mechanism of aroma β-primeveroside biosynthesis in C. sinensis. This information can be used to preserve tea aroma better during the manufacturing process and to investigate the mechanism of plant chemical defenses.

A previous study found that 1-phenylethanol (1PE) was a major endogenous volatile compound in tea (Camellia sinensis) flowers and can be transformed to glycosically conjugated 1PE (1PE-Gly). However, occurrences of 1PE-Gly in plants remain unknown. In this study, four 1PE-Glys have been isolated from tea flowers. Three of them were determined as (R)-1PE β-d-glucopyranoside ((R)-1PE-Glu), (S)-1PE-Glu, and (S)-1PE β-primeveroside ((S)-1PE-Pri), respectively, on the basis of NMR, MS, LC-MS, and GC-MS evidence. The other one was identified as (R)-1PE-Pri on the basis of LC-MS and GC-MS data. Moreover, these 1PE-Glys were chemically synthesized as the authentic standards to further confirm their occurrences in tea flowers. 1PE-Glu had a higher molar concentration than 1PE-Pri in each floral stage and organ. The ratio of (R) to (S) differed between 1PE-Glu and 1PE-Pri. In addition, a 1PE-Gly hydrolase β-primeverosidase recombinant protein produced in Escherichia coli exhibited high hydrolysis activity toward (R)-1PE-Pri. However, β-primeverosidase transcript level was not highly expressed in the anther part, which accumulated the highest contents of 1PE-Gly and 1PE. This suggests that 1PE-Gly may not be easily hydrolyzed to liberate 1PE in tea flowers. This study provides evidence of occurrences of 1PE-Glys in plants for the first time.

A flower-inducing compound, LDS1, was isolated from a free-floating aquatic plant, Lemna paucicostata . The chemical structure and the absolute stereochemistry of LDS1 were determined as (9 R ,13 R ,11 E ,15 Z )-9,13-dihydroxy-10-oxooctadeca-11,15-dienoic acid for its most abundant diastereomer. LDS1 was enzymatically produced when the plant was exposed to drought stress, and induced flowering at a concentration of 10 nM.

Camellia sinensis synthesizes and emits a large variety of volatile phenylpropanoids and benzenoids (VPB). To investigate the enzymes involved in the formation of these VPB compounds, a new C. sinensis short-chain dehydrogenase/reductase (CsSDR) was isolated, cloned, sequenced, and functionally characterized. The complete open reading frame of CsSDR contains 996 nucleotides with a calculated protein molecular mass of 34.5 kDa. The CsSDR recombinant protein produced in Escherichia coli exhibited dehydrogenase-reductase activity towards several major VPB compounds in C. sinensis flowers with a strong preference for NADP/NADPH co-factors, and showed affinity for (R)/(S)-1-phenylethanol (1PE), phenylacetaldehyde, benzaldehyde, and benzyl alcohol, and no affinity for acetophenone (AP) and 2-phenylethanol. CsSDR showed the highest catalytic efficiency towards (R)/(S)-1PE. Furthermore, the transient expression analysis in Nicotiana benthamiana plants validated that CsSDR could convert 1PE to AP in plants. CsSDR transcript level was not significantly affected by floral development and some jasmonic acid-related environmental stress, and CsSDR transcript accumulation was detected in most floral tissues such as receptacle and anther, which were main storage locations of VPB compounds. Our results indicate that CsSDR is expressed in C. sinensis flowers and is likely to contribute to a number of floral VPB compounds including the 1PE derivative AP.

We produced low temperature (15 degrees C) processed green tea (LTPGT) with higher aroma contents than normal green tea (Sencha). Normal temperature processed green tea (NTPGT), involved storing at 25 degrees C, and Sencha had no storing process. Sensory evaluation showed LTPGT had higher levels of floral and sweet odorants than NTPGT and Sencha. Aroma extract dilution analysis and gas chromatography-mass spectrometry-olfactometry indicated LTPGT had 12 aroma compounds with high factor dilution values (FD). Amongst LTPGT's 12 compounds, indole, jasmine lactone, cis-jasmone, coumarin, and methyl epijasmonate contributed to floral, fruity and sweet characters. In particular, indole increased initially, peaking at 16 h, then gradually decreased; Feeding experiments suggested [N-15]indole and [N-15]oxygenated indoles (OX-indoles) were produced from [N-15]anthranilic acid. We proposed the increase in indole was due to transformation of anthranilic acid during the 16 h storage and the subsequent decline in indole level was due to its conversion to OX-indoles.

Tea plant (Camellia sinensis) biosynthesizes a wide variety of specialized metabolites, including phenolic compounds such as catechins. Flavonol, one of the major flavonoid subclasses, in C. sinensis is present in the O-glycoside form, such as quercetin 3-O-β-D-glucopyranoside, kaempferol 3-O-β-D-glucopyranoside, and rutin (quercetin 3-O-β- glucopyranosyl-6-O-α-rhamnoside). These flavonol glycosides are highly accumulated, constituting up to 2-3% (w/w dry weight) of tea leaves; however, their biosynthetic machinery in C. sinensis remains elusive. Using high-throughput RNA sequencing from the fresh leaves of a cultivar (C. sinensis var sinensis cv Yabukita) and rapid amplification of cDNA ends (RACE) cloning with degenerate oligonucleotide primers, we identified a full-length cDNA of UDP-glycosyltransferase, designated as UGT73A17, and characterized the biochemical and molecular functions of UGT73A17. Recombinant UGT73A17 protein catalyzed 3-O-glucosylation of quercetin, yielding quercetin 3-O-β-D-glucopyranoside in vitro. The preferential expression of UGT73A17 gene in the mature, relative to young leaves, stems and roots, is roughly consistent with the accumulation pattern of flavonol glycosides in C. sinensis, suggesting that UGT73A17, in part, participates in the biosynthesis of flavonol glycosides in planta .

Green algae are of great economic importance. Seaweed is consumed fresh or as seasoning in Japan. The commercial value is determined by quality, color, and flavor and is also strongly influenced by the production area. Our research, based on solid phase microextraction gas chromatography mass spectrometry (SPME-GC-MS), has revealed that volatile compounds differ intensely in the four varieties of commercial green algae. Accordingly, 41 major volatile compounds were identified. Heptadecene was the most abundant compound from Okayama (Ulva prolifera), Tokushima (Ulva prolifera), and Ehime prefecture (Ulva linza). Apocarotenoids, such as ionones, and their derivatives were prominent volatiles in algae from Okayama (Ulva prolifera) and Tokushima prefecture (Ulva prolifera). Volatile, short chained apocarotenoids are among the most potent flavor components and contribute to the flavor of fresh, processed algae, and algae-based products. Benzaldehyde was predominant in seaweed from Shizuoka prefecture (Monostroma nitidum). Multivariant statistical analysis (PCA) enabled simple discrimination of the samples based on their volatile profiles. This work shows the potential of SPME-GC-MS coupled with multivariant analysis to discriminate between samples of different geographical and botanical origins and form the basis for development of authentication methods of green algae products, including seasonings.

A method for carbon isotope ratio (δ(13)C) analysis was developed for compound-specific isotope analysis of tea volatiles, and the values were compared with the δ(13)C value from bulk isotope analyses. The δ(13)C value of 2-phenylethanol liberated via enzymatic hydrolysis of the 2-phenylethyl β-primeveroside standard was examined first. Isotope fractionations for 2-phenylethyl β-primeveroside from preparative high-performance liquid chromatography (HPLC) were also analyzed. The enzymatic treatment and the preparative HPLC process did not cause carbon isotope fractionations, substantiating the strategies available for δ(13)C analysis of volatile compounds. On the basis of the gas chromatography-combustion-isotope ratio mass spectrometry data from 2-phenylethanol, it was possible to derive the conditions for enzyme treatment and preparative HPLC of the glycoconjugates of 2-phenylethanol, (Z)-3-hexenol, and benzyl alcohol isolated from green tea leaves. Larger variations in δ(13)C were found for individual volatile compounds compared with bulk analytical data from the leaves, indicating the potential to utilize this strategy in assigning the geographical origin of green tea.

Tea aroma is one of the most important factors affecting the character and quality of tea. Recent advances in methods and instruments for separating and identifying volatile compounds have led to intensive investigations of volatile compounds in tea. These studies have resulted in a number of insightful and useful discoveries. Here we summarize the recent investigations into tea volatile compounds: the volatile compounds in tea products; the metabolic pathways of volatile formation in tea plants and the glycosidically-bound volatile compounds in tea; and the techniques used for studying such compounds. Finally, we discuss practical applications for the improvement of aroma and flavor quality in teas.

The α-ketol oxylipin (12Z,15Z)-9-hydroxy-10-oxooctadeca-12,15-dienoic acid (1) reacts with epinephrine (2) to afford unique cycloadducts, KE1 (3) and KE2 (4), both of which strongly induce flowering in Lemna paucicostata. Here we isolate compound 5 from the reaction mixture and identify it as a Michael-type adduct of 1 with adrenochrome (6), an oxidized form of 2. Compound 5 was gradually converted into KEs in aqueous solution, suggesting that the compound is an intermediate of KEs. From these results, we newly propose the reaction mechanism of the cycloaddition between 1 and 2.

Changes of endogenous 9, 10-ketol-octadecadienoic acid (KODA) concentrations, which is synthesized from linolenic acid by 9-lipoxygenase, were analyzed in apple [Malus ×sylvestris (L.) Mill. var. domestica (Borkh.) Mansf.] buds. In addition, the effects of 15, 16-chloro, hydroxy-9-hydroxy-10-oxo-12(Z)-octadecenoic acid (CKODA) application, which is an analog of KODA, on flower bud formation and the expression of MdTFL1 (terminal flower 1) and MdFT1 (flowering locus t 1) genes in apple buds were investigated in heavy-crop treatment (HCT) and under shade. An increase of endogenous KODA in the buds in the fruit-thinning treatment, which resulted in a higher proportion of flower bud formation than in HCT, was observed at 63 days after full bloom, but no such increase was found in HCT. In the shade-treated and heavy-crop trees, the expression of MdTFL1 in the buds to which CKODA was applied was lower than that in untreated buds. In contrast, under shade, the expression of MdFT1 in the CKODA-treated buds was higher than that in untreated buds. These results suggest that endogenous KODA may be associated with flower bud formation, and its application may be effective at improving the proportion of flower bud formation through its effect on MdTFL1 and MdFT1.

The cyanoglycoside, prunasin, was isolated for the first time as an aroma precursor of benzaldehyde from fresh tea leaves (Camellia sinensis var. sinensis cv. Yabukita). Prunasin was readily hydrolyzed by a crude enzyme prepared from the fresh tea leaves to liberate benzaldehyde. The isomerization of prunasin under neutral conditions was also recognized.

The self-assembling gold nanoparticle (GNP)-supported l -proline derivative, which was readily synthesized in four steps from 4-hydroxy- l -proline and chloroauric acid, was used for enamine-based aldol reactions. The modified Brust-Schiffrin (BS) synthesis was favored over the ligand exchange reaction in order to develop a new and simple synthetic pathway for nanoparticle-supported catalyst. Use of immobilized organocatalyst on nanoscale solid carrier led to excellent selectivities (up to 94:6 dr and 94% ee) in asymmetric reactions of ketones and benzaldehydes. GNPs operate under pseudo-homogeneous conditions which are easily recycled and reused at least five times without significant loss of weight, activity, diastereo- and enantioselectivities.

Plants emit chemically diverse volatile compounds for attracting pollinators or putting up a chemical defense against herbivores. 2-Phenylethanol (2PE) is one of the abundantly emitted scent compounds in rose flowers. Feeding experiments with l-[2H8]phenylalanine into rose flowers and subsequent analysis using gas chromatography–mass spectrometry analysis revealed the hypothetical biosynthetic intermediates to [2H8]-2PE, and the biochemical and genetic analyses elucidated the principal pathway to [2H8]-2PE. We recently found season-specific 2PE pathway producing [2H7]-2PE from l-[2H8]phenylalanine. This is a unique example where the dominant pathway to a specific compound changes with the seasons. This review focuses on the biosynthesis of floral volatiles and their regulation to adapt to the changes in the environment.

Indole is a characteristic volatile constituent in oolong tea. Our previous study indicated that indole was mostly accumulated at the turn over stage of oolong tea manufacturing process. However, formation of indole in tea leaves remains unknown. In this study, one tryptophan synthase α-subunit (TSA) and three tryptophan synthase β-subunits (TSBs) from tea leaves were isolated, cloned, sequenced, and functionally characterized. Combination of CsTSA and CsTSB2 recombinant protein produced in Escherichia coli exhibited the ability of transformation from indole-3-glycerol phosphate to indole. CsTSB2 was highly expressed during the turn over process of oolong tea. Continuous mechanical damage, simulating the turn over process, significantly enhanced the expression level of CsTSB2 and amount of indole. These suggested that accumulation of indole in oolong tea was due to the activation of CsTSB2 by continuous wounding stress from the turn over process. Black teas contain much less indole, although wounding stress is also involved in the manufacturing process. Stable isotope labeling indicated that tea leaf cell disruption from the rolling process of black tea did not lead to the conversion of indole, but terminated the synthesis of indole. Our study provided evidence concerning formation of indole in tea leaves for the first time.

Floral volatile phenylpropanoids and benzenoids (VPBs) play important ecological functions and have potential economic applications. Little is known about how multi-factors in integration regulate the formation and emission of floral VPBs. In the present study, we investigated effects of multi factors including endogenous circadian clock, light, and temperature on the formation and emission of VPBs, which are major volatiles in flowers of Petunia× hybrida cv. 'Mitchell Diploid'. Endogenous circadian clock was proposed as the most important factor regulating rhythmic emission of VPBs and expressions of structural genes involved in the upstream biosynthetic pathway of VPBs, but did not affect expression levels of structural genes involved in the downstream pathway and VPBs-related regulators. In contrast to light, temperature was a more constant factor affecting emission of VPBs. VPBs emission could be inhibited within a short time by increasing temperature. The information will contribute to our understanding of emission mechanism of floral volatiles.

BACKGROUND Carotenoids are a major class of plant pigments and fulfill many functions in different organisms that either produce or consume them. Although the color of the stamina of tea (Camellia sinensis) flowers is clearly due to the presence of carotenoids, the carotenoid profile and content remain to be discovered. RESULTS We investigated the carotenoid profile of tea flowers and determined changes in concentrations over the floral development. The flowers contained oxygenated xanthophylls such as neoxanthin, lutein and zeaxanthin, as well as the hydrocarbons β-carotene and α-carotene. Flowers of the tea plant contain to vegetables comparable amounts of carotenoids. The content of 9′-cis-epoxycarotenoids, which serve as abscisic acid precursors, as well as changes in concentration of abscisic acid were studied. The concentrations of carotenoids decreased whereas the abscisic acid content increased over the floral development. Exogenously applied S-abscisic acid affected water uptake, flower opening and carotenoid accumulation. CONCLUSION In summary, this paper reports, for the first time, the carotenoid profile and content of tea flowers. The study revealed that carotenoids in tea flowers are an interesting target in respect of possible applications of tea flower extracts as well as biological functions of abscisic acid during floral development. © 2012 Society of Chemical Industry

We investigated the effect of 9,10-ketol-octadecadienoic acid (KODA), which is synthesized from linolenic acid by 9-lipoxygenase, on 1-aminocyclopropane-1-carboxylate (ACC) synthase and ACC oxidase gene expression, their enzyme activities, and ethylene production in apple fruit [Malus sylvestris (L.) Mill. Var. domestica (Borkh.) Mansf.]. The fruit was treated with C-KODA, which is a derivative of KODA, at 63 or 119 d after full bloom (DAFB). Endogenous KODA in the skin of the fruit did not change significantly from 48 to 119 DAFB on the tree, and during storage after harvest at 119 DAFB. However, softening of the fruit treated with C-KODA at 63 DAFB was inhibited. ACC synthase and oxidase activities, ACC and MACC concentrations and ethylene production were lower in the C-KODA-treated fruit than in the untreated control at 119 DAFB. In general, mRNA transcript levels of MdACS1 and MdACO1 in the skin of the C-KODA-treated fruit were lower than those in the untreated control fruit. In contrast, softening was promoted in the fruit stored after C-KODA treatment at 119 DAFB. ACC synthase activity, ACC and MACC concentration, ethylene production and the mRNA transcript levels of MdACS1 and MdACO1 in the skin were also increased in the C-KODA-treated skin. These results suggest that C-KODA may influence ethylene production via the expression of MdACS1 and MdACO1 in apple fruit, but that the effect is different depending on the time of treatment.

As a defense response to attacks by herbivores such as the smaller tea tortrix ( Adoxophyes honmai Yasuda), tea ( Camellia sinensis ) leaves emit numerous volatiles such as (Z)-3-hexen-1-ol, linalool, α-farnesene, benzyl nitrile, indole, nerolidol, and ocimenes in higher concentration. Attack of Kanzawa spider mites ( Tetranychus kanzawai Kishida), another major pest insect of tea crops, induced the emission of α-farnesene and ocimenes from tea leaves. The exogenous application of jasmonic acid to tea leaves induced a volatile blend that was similar, although not identical, to that induced by the smaller tea tortrix. Most of these herbivore-induced plant volatiles (HIPV) were not stored in the tea leaves but emitted after the herbivore attack. Both the adaxial and abaxial epidermal layers of tea leaves emitted blends of similar composition. Furthermore, HIPV such as α-farnesene were emitted mostly from damaged but not from undamaged leaf regions. A principal component analysis of metabolites (m/z 70-1000) in undamaged tea leaves exposed or not to HIPV suggests that external signaling via HIPV may lead to more drastic changes in the metabolite spectrum of tea leaves than internal signaling via vascular connections, although total catechin contents were slightly but not significantly increased in the external signaling via HIPV.

Purple-fleshed sweet potatoes (Ipomoea batatas L.) contain a very complex anthocyanin profile due to the presence of several non-, mono-, and diacylated glucosides of cyanidin and peonidin. In this study, the anthocyanin composition of four Japanese purple sweet potato cultivars (Chiran Murasaki, Tanegashima Murasaki, Naka Murasaki, and Purple Sweet) were investigated by HPLC-DAD and ESI-MSn analyses. The HPLC chromatograms of the different cultivars show a remarkable variation of the two major pigments, cyanidin-3-(6''-caffeoylsophoroside)-5-glucoside and peonidin-3-(6''-caffeoylsophoroside)-5-glucoside, respectively. According to this, they can be categorized into two groups on the basis of the peonidin/cyanidin ratio: the cultivars Chiran Murasaki and Purple Sweet showed a high content of peonidin derivatives (peonidin type), whereas the varieties Tanegashima Murasaki and Naka Murasaki were classified as cyanidin types. By means of high-speed countercurrent chromatography (HSCCC) the nonacylated 3-sophoroside-5-glucoside of cyanidin was isolated on a preparative scale. Furthermore, it was possible to isolate the monoacylated cyanidin-3-(6''-caffeoylsophoroside)-5-glucoside as well as three diacylated major pigments, cyanidin-3-(6'',6'''-dicaffeoylsophoroside)-5-glucoside, cyanidin-3-(6''-caffeoyl-6'''-p-hydroxy-benzoylsophoroside)-5-glucoside, and peonidin-3-(6''-caffeoyl-6'''-p-hydroxybenzoyl-sophoroside)-5-glucoside. The purity and identity of the so-obtained pigments were confirmed by NMR measurements.

Oxylipin glucosides (2–4) were isolated from Lemna paucicostata with their structures and absolute configurations elucidated by spectroscopic and chemical methods. Compounds 2–4 were glucosides of C14 oxylipin which were synthesized from α-linolenic acid via the 9-lipoxygenase pathway.

While beneficial health properties of tea leaves have been extensively studied, less attention has been given to that of flowers of the tea ( Camellia sinensis ) plant. In this work, the ethanolic extract and its ethyl acetate-soluble fraction (EEA) from the tea flowers were found to possess the potent antioxidant activity using 2, 2-diphenyl-1-picrylhydrazyl (DPPH) free radical-scavenging assay. The compounds present in EEA had comparatively strong DPPH scavenging activity and strongly contributed to the antioxidant activity of the tea flowers. From EEA, besides eight catechins, five flavonol glycosides were isolated and their structures were elucidated on the basis of mass spectrometry and nuclear magnetic resonance spectroscopy as myricetin 3- O -β- d -galactopyranoside, quercetin 3- O -β- d -galactopyranoside, kaempferol 3- O -β- d -galactopyranoside, kaempferol 3- O -β- d -glucopyranoside, and kaempferol 3- O -[α- l -rhamnopyranosyl-(1-6)-β- d -glucopyranoside]. In addition, epigallocatechin gallate and epicatechin gallate were found as the major active components responsible for the antioxidant activity of tea flowers through the use of a combination of preparative liquid chromatography separation and DPPH assay.

Conventionally, tea flavor is analyzed through the use of a combination of gas chromatography–mass spectrometry and human taste panel. These methods present time-consuming or inaccurate factor. In this work, a rapid, accurate and nondestructive approach was put forward to identify coumarin-enriched Japanese green teas and evaluate their particular flavor using electronic nose (E-nose) technique. The multivariable analyses including principal component analysis and cluster analysis were applied to distinguish the tea samples and evaluate the particular (coumarin-like) flavor of coumarin-enriched tea under different infusion conditions. A correct classification was achieved for the seven tea samples with different content of coumarin. The E-nose successfully characterized the drying temperature-dependently trend of coumarin content during the manufacture process of coumarin-enriched green tea. It also revealed that the comparatively low-infusion temperature and long-infusion time were favorable for the emission of coumarin-like flavor of the tea infusion. In addition, a comparatively newly developed “absolute value expression” (AVE) method was employed to divide the tea flavors into quality and express them numerically. Using AVE, the role of coumarin in the total flavor of coumarin-enriched green tea was elucidated. These results suggest that E-nose could be employed to identify the green teas with particular flavor and evaluate the tea flavor.

(12Z,15Z)-9-Hydroxy-10-oxooctadeca-12,15-dienoic acid (1) and norepinephrine (2) undergo cycloaddition to afford FN1 (3) and FN2 (4), both of which induce flowering in Lemna paucicostata. Although the derivatives of 1 were suggested to also yield FN-like compounds after reacting with 2, their structures have not been elucidated. In this report, we investigated the structure and stereochemistry of seven novel FN analogs. These analogs were shown to be formed in the same regio- and stereocontrolled manner as FNs. Moreover, the activity of FN analogs on flowering induction was investigated, and we determined that all analogs, except for compound 8, were effective flowering inducers for L. paucicostata.

The antioxidant properties of theaflavins and their gallate esters, namely theaflavin (TF1), theaflavin-3(3')-gallate (TF2) and theaflavin-3,3'-digallate (TF3) were investigated by comparing with epigallocatechin gallate (EGCG). The order of hydroxyl radicals-scavenging ability was TF3>TF2>TF1>EGCG. The order of 2,2-diphenyl-1-picrylhydrazyl scavenging ability was TF3>TF2>EGCG>TF1. TF1, TF2, and TF3 showed more effective effects than EGCG in protection against H2O2-mediated damage in HPF-1 cells. TF2 was the most potent accelerant of HPF-1 cell proliferation. TF1, TF2 and TF3 suppressed the accumulation of intracellular reactive species in H2O2-mediated damage HPF-1 cells. Pre-treated for 2h and eliminated from the cells, TF1 and TF3 still showed protective effects against H2O2-mediated damage in HPF-1 cells. This suggests that the protective effects of TF1 and TF3 on oxidative damage HPF-1 cells may be responsible for other mechanisms, rather than only scavenging the already formed reactive species. It remains to be determined whether TF1 and TF3 improved the normal HPF-1 cell resistive abilities toward radical-damage in pre-treatment. Further studies of the effects of theaflavins on some enzymes or signal transduction in the normal HPF-1 cells are underway.

FN1 (1) and FN2 (2), cycloadducts of α-ketol octadecadienoic acid (3) with norepinephrine (NE), induce flowering in Lemna paucicostata. In order to broaden our understanding of structural requirements of FN for flower induction, nine analogs of 3 (4–12) were synthesized and reacted with NE under basic conditions. These analogs, except for 8, 10, and 12, exhibited significant activity regarding to floral induction in L. paucicostata. Similar experiments were carried out by using 3 and epinephrine, and it was demonstrated that these products also possessed biological activity.

We have studied LED irradiation effects on plants and animals in the visible to UV region of light from GaN LEDs. The results are as follows. Blue light considers to be effective for pearl cultivation or for attraction of small fishes living in near the surface of sea such as Pompano or Sardine, white light radiation is effective for cultivation of botanical plankton for shells. Other experiments of UV light irradiation attracting effect on baby sea turtle and the germination UV effect of mushroom, green light weight enhance effect on baby pigs, light vernalization effect of vegitable and Ge far infrared therapic effect on human body are also given.

The reaction of 9-hydroxy-10-oxo-12(Z),15(Z)-octadecadienoic acid (KODA) and catecholamine in water (pH 8.0) in vitro generates many products, and we have shown that the major adduct FN1 among the reactants of KODA and norepinephrine is a potent inducer of floral differentiation in Lemna (L.) paucicostata 151 (Yokoyama et al. (2000) Plant Cell Physiol 41: 110-113, Yamaguchi et al. (2001) ibid 42: 1201-1209). Since such reactants may be involved in flower induction in vivo, it is of interest to elucidate the structure-activity relationships among the reactants to throw light on the nature of the putative active site. Here, we report two other adducts, designated as FN7 and FN10. They are structurally similar to FN1, having a tricyclic structure with 12-cis-olefin conjugated with noradrenochrome. FN10 was a more potent inducer than FN1. These results are consistent with the hypothesis that tricyclic KODA/norepinephrine adducts play an important role in flower-induction in L. paucicostata, strain 151.

A recently developed method enabled us to simultaneously characterize and quantitate glycosidically bound volatiles (GBVs) at picomole levels using liquid chromatography–mass spectrometry (LC-MS). On the basis of the analytical data it is possible to screen tea varieties most suitable for black tea processing, in which higher concentrations of primeverosides accumulate. The primeverosides decreased at the rolling step in black tea processing, whereas the glucopyranosides did not change much. The total contents of GBVs gradually increased at the withering steps and then remarkably increased after the fixing step at 230 °C, during oolong tea processing. The presence of 6′-O-malonyl ester type β-d-glucopyranosides in the tea samples suggested a contribution to the increment in glucopyranosides during oolong tea processing. The method was also used to analyze GBVs and their derivatives to understand their possible role in the metabolic pathway of tea.

We investigated the biosynthetic pathway for 2-phenylethanol, the dominant floral scent compound in roses, using enzyme assays. L-[(2)H8] Phenylalanine was converted to [(2)H8] phenylacetaldehyde and [(2)H8]-2-phenylethanol by two enzymes derived from the flower petals of R. 'Hoh-Jun,' these being identified as pyridoxal-5'-phosphate-dependent L-aromatic amino acid decarboxylase (AADC) and phenylacetaldehyde reductase (PAR). The activity of rose petal AADC to yield phenylacetaldehyde was nine times higher toward L-phenylalanine than toward its D-isomer, and this conversion was not inhibited by iproniazid, a specific inhibitor of monoamine oxidase. Under aerobic conditions, rose petal AADC stoichiometrically produced NH3 together with phenylacetaldehyde during the course of decarboxylation and oxidation, followed by the hydrolysis of L-phenylalanine. Phenylacetaldehyde was subsequently converted to 2-phenylethanol by the action of PAR. PAR showed specificity toward several volatile aldehydes.

α-Ketol of octadecadienoic acid (KODA, 1 ) has been suggested to play a role in the photoperiod-regulated flowering in Pharbitis nil . The level of 1 in cotyledons is temporarily controlled during short-day conditions. The biosynthesis of 1 is well studied in plants; however, its in vivo conversion is less understood. We have investigated this issue by studying the metabolism of exogenously-applied [U- 13 C]- 1 , [1- 14 C]- 1 , and non-labeled 1 in P. nil seedlings by the spectroscopic methods and identified six major metabolites ( 4 – 9 ). We have also found that the enantiomers of 1 are differentially metabolized in P. nil seedlings.

Body size determination is critical for multicellular organisms; however, the mechanisms remain largely unknown. Mutations that alter body size were studied to solve the mechanisms, for example, in mouse, fruit fly and the nematode Caenorhabditis elegans. In C. elegans, a large mutant and several small body size (sma) mutants are known. Of the latter, sma-2, sma-3, sma-4, sma-6, dbl-1 and daf-4 have a mutation in the components of the DBL-1/TGFbeta signal pathway, and sma-5 in a MAP kinase homologue. We have constructed double mutants carrying two of such small body size mutations, sma-5 and sma-4 or sma-2. They are much smaller than either of the parental single mutants, indicating that the sma-5 gene functions independently of the DBL-1/TGFbeta pathway. We show that their body volumes are as small as 1/10 of that of the wild-type, and that the sizes of major organs are much reduced, by the methods previously developed by us. But the numbers of cells are not changed, suggesting that the cells are very small. These results highlight surprising flexibility of body size and cell size in a multicellular organism, which will give a novel insight into the mechanisms of body size control.

Plants synthesize and emit a large variety of volatile organic compounds, which possess extremely important ecological functions. In most case, most plant volatiles are liquids, rather than gases, at room temperature. Some volatiles are emitted “on demand” when plants, especially vegetative parts, are exposed to abiotic or biotic stress. In this review, we summarize some of the highlights of plant vegetative volatile emission and functions research published during the past few years.

Regulation of plant growth and development by light wavelength has been extensively studied. Less attention has been paid to effect of light wavelength on formation of plant metabolites. The objective of this study was to investigate whether formation of volatiles in preharvest and postharvest tea (Camellia sinensis) leaves can be regulated by light wavelength. In the present study, in contrast to the natural light or dark treatment, blue light (470 nm) and red light (660 nm) significantly increased most endogenous volatiles including volatile fatty acid derivatives (VFADs), volatile phenylpropanoids/benzenoids (VPBs) and volatile terpenes (VTs) in the preharvest tea leaves. Furthermore, blue and red lights significantly up-regulated the expression levels of 9/13-lipoxygenases involved in VFADs formation, phenylalanine ammonialyase involved in VPBs formation and terpene synthases involved in VTs formation. Single light wavelength had less remarkable influences on formation of volatiles in the postharvest leaves compared with the preharvest leaves. These results suggest that blue and red lights can be promising technology for remodeling the aroma of preharvest tea leaves. Furthermore, our study provided evidence that light wavelength can activate the expression of key genes involved in formation of plant volatiles for the first time.

It was generally thought that aroma of oolong tea resulted from hydrolysis of glycosidically bound volatiles (GBVs). In this study, most GBVs showed no reduction during the oolong tea manufacturing process. β-Glycosidases either at protein or gene level were not activated during the manufacturing process. Subcellular localization of β-primeverosidase provided evidence that β-primeverosidase was located in the leaf cell wall. The cell wall remained intact during the enzyme-active manufacturing process. After the leaf cell disruption, GBV content was reduced. These findings reveal that, during the enzyme-active process of oolong tea, nondisruption of the leaf cell walls resulted in impossibility of interaction of GBVs and β-glycosidases. Indole, jasmine lactone, and trans-nerolidol were characteristic volatiles produced from the manufacturing process. Interestingly, the contents of the three volatiles was reduced after the leaf cell disruption, suggesting that mechanical damage with the cell disruption, which is similar to black tea manufacturing, did not induce accumulation of the three volatiles. In addition, 11 volatiles with flavor dilution factor ≥4(4) were identified as relatively potent odorants in the oolong tea. These results suggest that enzymatic hydrolysis of GBVs was not involved in the formation of volatiles of oolong tea, and some characteristic volatiles with potent odorants were produced from the manufacturing process.

We have analyzed the sma-5(n678) mutant in C. elegans to elucidate mechanisms controlling body size. The sma-5 mutant is very small, grows slowly and its intestinal granules look abnormal. We found a 15 kb deletion in the mutant that includes a 226 bp deletion of the 3′ end of the W06B3.2-coding sequence. Based on this result, rescue experiments, RNAi experiments and a newly isolated deletion mutant of W06B3.2, we conclude that W06B3.2 is the sma-5 gene. The sma-5 mutant has much smaller intestine, body wall muscles and hypodermis than those of the wild type. However, the number of intestinal cells or body wall muscle cells is not changed, indicating that the sma-5 mutant has much smaller cells. In relation to the smaller cell size, the amount of total protein is drastically decreased; however, the DNA content of the intestinal nuclei is unchanged in the sma-5 mutant. The sma-5 gene is expressed in intestine,excretory cell and hypodermis, and encodes homologs of a mammalian MAP kinase BMK1/ERK5/MAPK7, which was reported to control cell cycle and cell proliferation. Expression of the sma-5 gene in hypodermis is important for body size control, and it can function both organ-autonomously and non-autonomously. We propose that the sma-5 gene functions in a MAP kinase pathway to regulate body size mainly through control of cell growth.

The hydrolysis of 2-phenylethyl β- d -glucopyranoside ( 3 ) was found to be partially inhibited by feeding with 2-phenyl-N-glucosyl-acetamidiumbromide ( 8 ), a β-glucosidase inhibitor, resulting in a decrease in the diurnal emission of 2-phenylethanol ( 2 ) from Rosa damascena Mill. flowers. Detection of [1,1,2,2′,3′,4′,5′,6′-2H8]-2 and [1,2,2′,3′,4′,5′,6′-2H7]-2 from R. ‘Hoh-Jun’ flowers fed with [1,1,2,2′,3′,4′,5′,6′-2H8]-3 suggested that β-glucosidase, alcohol dehydrogenase, and reductase might be involved in scent emission. Comprehensive GC-SIM analyses revealed that [1,2,2,2′,3′,4′,5′,6′-2H8]-2 and [1,2,2,2′,3′,4′,5′,6′-2H8]-3 must be biosynthesized from [1,2,2,2′,3′,4′,5′6′-2H8] l -phenylalanine ( [2H8]-1 ) with a retention of the deuterium atom at α-position of [2H8]-1 .

The inhibition effects of tea theaflavins complex (TFs), theaflavin-3-3'-digallate (TFDG), theaflavin-3'-gallate (TF2B), and an unidentified compound (UC) on the growth of human liver cancer BEL-7402 cells, gastric cancer MKN-28 cells and acute promyelocytic leukemia LH-60 cells were investigated. TFs was obtained through the catalysis of catechins with immobilized polyphenols oxidase. TFDG, TF2B and UC were isolated from TFs with high speed countercurrent chromatography (HSCCC). The results showed that TF2B significantly inhibited the growth of all three kinds of cancer cells, TFs, TFDG and UC had some effect on BEL-7402 and MKN-28, but little activity on LH-60. The inhibition effects of TF2B, TFDG, and UC on BEL-7402 and MKN-28 were stronger than TFs. The relationship coefficients between monomer concentration and its inhibition rate against MKN-28 and BEL-7402 were 0.87 and 0.98 for TF2B, 0.96 and 0.98 for UC, respectively. The IC50 values of TFs, TF2B, and TFDG were 0.18, 0.11, and 0.16 mM on BEL-7402 cells, and 1.11, 0.22, and 0.25 mM on MKN-28 cells respectively.

GC–olfactometry was used to characterize the specific aroma of a pentane extract of snake fruit (Salacca edulis Reinw cv. Pondoh). Ten compounds, including two carboxylic acids, six methyl esters, an alcohol and a furaneol (2,5-dimethyl-4-hydroxy-3(2H)-furanone), were detected as the most characteristic odorants. Methyl esters exhibited particularly sweet and fruity odour characteristics in snake fruit at more mature stages, whereas carboxylic acids exhibited the sweaty odour that tends to prevent non-native consumers from liking the fruit. A headspace of snake fruit at different stages of maturation was subjected to analysis using two analytical systems an electronic nose equipped with 18 sensors and fingerprint mass spectrometry. Three sensors and five ions derived from esters were chosen as fingerprint parameters to discriminate between maturation levels. Both systems appeared to discriminate efficiently between the fruits at different maturation stages. Copyright © 2003 John Wiley & Sons, Ltd.