Analysis of stereochemistry and biosynthesis of epicatechin in tea plants by chiral phase high performance liquid chromatography

Qian, Yumei; Zhao, Xianqian; Жао, Леи; Cui, Lilan; Liu, Li; Jiang, Xiaolan; Liu, Yajun; Gao, Liping; Xia, Tao

doi:10.1016/j.jchromb.2015.10.024

Cited by 13 publications

(12 citation statements)

References 45 publications

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“…8 The biosynthesis of catechins in tea plants is a complex and huge pathway, which was regulated by multiple genes. 43,44 In this study, these three CsCPRs showed similar expressing trends with three CsCYP450s (CsC4H, CsF3′H, and CsF3′5′H) in different tea tissues (Figure 6). However, their expression levels did not show exactly similar variation patterns with 3′,4′,5′-catechins (or total catechins) in different tea tissues (Figure 6).…”

Section: ■ Discussionsupporting

confidence: 66%

Isolation and Functional Characterization of Multiple NADPH-Cytochrome P450 Reductase Genes from Camellia sinensis in View of Catechin Biosynthesis

Huang

Liu

et al. 2021

J. Agric. Food Chem.

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Catechins are critical constituents for the sensory quality and health-promoting benefits of tea. Cytochrome P450 monooxygenases are required for catechin biosynthesis and are dependent on NADPH-cytochrome P450 reductases (CPRs) to provide reducing equivalents for their activities. However, CPRs have not been identified in tea, and their relationship to catechin accumulation also remains unknown. Thus, three CsCPR genes were identified in this study, all of which had five CPR-related conserved domains and were targeted to the endoplasmic reticulum. These three recombinant CsCPR proteins could reduce cytochrome c using NADPH as an electron donor. Heterologous co-expression in yeast demonstrated that all the three CsCPRs could support the enzyme activities of CsC4H and CsF3′H. Correlation analysis indicated that the expression level of CsCPR1 (or CsCPR2 or CsCPR3) was positively correlated with 3′,4′,5′-catechin (or total catechins) content. Our results indicate that the CsCPRs are involved in the biosynthesis of catechins in tea leaves.

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Section: ■ Discussionsupporting

confidence: 66%

Isolation and Functional Characterization of Multiple NADPH-Cytochrome P450 Reductase Genes from Camellia sinensis in View of Catechin Biosynthesis

Huang

Liu

et al. 2021

J. Agric. Food Chem.

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“…In tea, dissociative flavan-3-ol monomers are mainly distributed in the leaves of the aerial parts of the plant, and polymeric catechins are distributed in the stems and root (Jiang et al 2013 , 2015 ). Dissociative flavon-3-ols are also called catechins and include (2R, 3S)- trans -flavan-3-ols (catechin, C, and gallocatechin, GC) and (2R, 3R)- cis -flavan-3-ols (epicatechin, EC; epigallocatechin, EGC; epicatechin gallate, ECG; and epigallocatechin gallate, EGCG) (Qian et al 2015 ). Catechins play a crucial role in the dominating astringent taste of tea and possess a number of effects that are beneficial to human health, particularly the galloylated catechins (Chung et al 2003 ; Cui et al 2016 ).…”

Section: Introductionmentioning

confidence: 99%

Evolutionary and functional characterization of leucoanthocyanidin reductases from Camellia sinensis

Wang

Zhang

Jiang

et al. 2017

Planta

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Main conclusion LARs promoted the biosynthesis of catechin monomers and inhibited their polymerization. The accumulation of catechin monomers and polymers was increased by up-regulating the expression of NtLAR and NtANR s in CsMYB5b transgenic tobacco. Tea is rich in polyphenolic compounds, and catechins are the major polyphenols in tea. The biosynthesis of polyphenols is closely related to the expression of the leucoanthocyanidin reductase (LAR) and anthocyanidin reductase (ANR) genes. In this paper, an evolutionary analysis and functional characterization of three CsLARs were performed. The phylogenetic tree showed that plant LARs could be grouped into three, including gymnosperms, monocotyledons and dicotyledons (clusters I and II). The eighth amino acid residue in a conserved LAR-specific motif is changeable due to a transversion (G → T) and transition (G → C) that occur in the corresponding codon. Therefore, plant LARs can be classified as G-type, A-type and S-type LARs due to this variable amino acid residue. Although (2R, 3S)-trans-flavan-3-ols were the products of recombinant CsLARs proteins expressed in Escherichia coli, both (2R, 3S)-trans and (2R, 3R)-cis-flavan-3-ols were detected in tobacco overexpressing CsLARs. However, a butanol/HCl hydrolysis assay indicated that overexpression of the CsLARs caused a decrease in polymerized catechins. A hybridization experiment with CsLARc + AtPAP1 also showed that no polymers other than epicatechin, catechin and glycoside were detected, although the accumulation of anthocyanins was markedly decreased. CsMYB5b promoted the biosynthesis of both flavan-3-ols and proanthocyanidins (PAs). Therefore, LARs promoted the biosynthesis of catechin monomers and inhibited their polymerization. The accumulation of catechin monomers and polymers was increased by up-regulating the expression of the NtLAR and NtANRs in CsMYB5b transgenic tobacco.Electronic supplementary materialThe online version of this article (doi:10.1007/s00425-017-2771-z) contains supplementary material, which is available to authorized users.

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“…To further confirm the chirality of enzymatic products separated by reverse phase column, the major product derived from the incubation of crude ANR extract and cyanidin was isolated on TLC plates according to our previous reports [ 9 ]. Based on the (−)-EC standard, analysis of HPLC coupled with a normal phase-chiral column identified that this metabolite was (−)-EC ( Figure 3 f,h).…”

Section: Resultsmentioning

confidence: 99%

“…The main composition of tea flavan-3-ols has been appropriately characterized by numerous investigations. Primary aglycones of flavan-3-ols include (+)-catechin [(+)-Ca)] and (−)-epicatechin [(−)-EC)] ( Figure 1 b), and (+)-gallocatechin [(+)-GC)] and (−)-epigallocatechin [(−)-EGC)] [ 9 ]. Primary conjugates are composed of (−)-epciatechin-3- O -gallate [(−)-ECG)] and (−)-epigallocatechin-3- O -gallate [(−)-EGCG)].…”

Section: Introductionmentioning

confidence: 99%

“…Since 2003, two biosynthetic pathways have been characterized to control the formation of flavan-3-ols, each with different stereo-configuration preference ( Figure 1 a) [ 9 , 10 , 11 , 12 ]. One is the anthocyanidin reductase (ANR) pathway, which starts with leucoanthocyanidins and then undergoes anthocyanidins to three stereo types of flavan-3-ols, such as (−)-Ca, (−)-EC, and (+)-EC.…”

Section: Introductionmentioning

confidence: 99%

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Metabolic Characterization of the Anthocyanidin Reductase Pathway Involved in the Biosynthesis of Flavan-3-ols in Elite Shuchazao Tea (Camellia sinensis) Cultivar in the Field

et al. 2017

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Anthocyanidin reductase (ANR) is a key enzyme in the ANR biosynthetic pathway of flavan-3-ols and proanthocyanidins (PAs) in plants. Herein, we report characterization of the ANR pathway of flavan-3-ols in Shuchazao tea (Camellia sinesis), which is an elite and widely grown cultivar in China and is rich in flavan-3-ols providing with high nutritional value to human health. In our study, metabolic profiling was preformed to identify two conjugates and four aglycones of flavan-3-ols: (−)-epigallocatechin-gallate [(−)-EGCG], (−)-epicatechin-gallate [(−)-ECG], (−)-epigallocatechin [(−)-EGC], (−)-epicatechin [(−)-EC], (+)-catechin [(+)-Ca], and (+)-gallocatechin [(+)-GC], of which (−)-EGCG, (−)-ECG, (−)-EGC, and (−)-EC accounted for 70–85% of total flavan-3-ols in different tissues. Crude ANR enzyme was extracted from young leaves. Enzymatic assays showed that crude ANR extracts catalyzed cyanidin and delphinidin to (−)-EC and (−)-Ca and (−)-EGC and (−)-GC, respectively, in which (−)-EC and (−)-EGC were major products. Moreover, two ANR cDNAs were cloned from leaves, namely CssANRa and CssANRb. His-Tag fused recombinant CssANRa and CssANRb converted cyanidin and delphinidin to (−)-EC and (−)-Ca and (−)-EGC and (−)-GC, respectively. In addition, (+)-EC was observed from the catalysis of recombinant CssANRa and CssANRb. Further overexpression of the two genes in tobacco led to the formation of PAs in flowers and the reduction of anthocyanins. Taken together, these data indicate that the majority of leaf flavan-3-ols in Shuchazao’s leaves were produced from the ANR pathway.

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Analysis of stereochemistry and biosynthesis of epicatechin in tea plants by chiral phase high performance liquid chromatography

Cited by 13 publications

References 45 publications

Isolation and Functional Characterization of Multiple NADPH-Cytochrome P450 Reductase Genes from Camellia sinensis in View of Catechin Biosynthesis

Isolation and Functional Characterization of Multiple NADPH-Cytochrome P450 Reductase Genes from Camellia sinensis in View of Catechin Biosynthesis

Evolutionary and functional characterization of leucoanthocyanidin reductases from Camellia sinensis

Metabolic Characterization of the Anthocyanidin Reductase Pathway Involved in the Biosynthesis of Flavan-3-ols in Elite Shuchazao Tea (Camellia sinensis) Cultivar in the Field

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