Exploring the genes involved in biosynthesis of dihydroquercetin and dihydromyricetin in Ampelopsis grossedentata.

Dihydroquercetin (DHQ), an extremely low content compound (less than 3%) in plants, is an important component of dietary supplements and used as functional food for its antioxidant activity. Moreover, as downstream metabolites of DHQ, an extremely high content of dihydromyricetin (DHM) is up to 38.5% in Ampelopsis grossedentata. However, the mechanisms involved in the biosynthesis and regulation from DHQ to DHM in A. grossedentata remain unclear. In this study, a comparative transcriptome analysis of A. grossedentata containing extreme amounts of DHM was performed on the Illumina HiSeq 2000 sequencing platform. A total of 167,415,597 high-quality clean reads were obtained and assembled into 100,584 unigenes having an N50 value of 1489. Among these contigs, 57,016 (56.68%) were successfully annotated in seven public protein databases. From the differentially expressed gene (DEG) analysis, 926 DEGs were identified between the B group (low DHM: 210.31 mg/g) and D group (high DHM: 359.12 mg/g) libraries, including 446 up-regulated genes and 480 down-regulated genes (B vs. D). Flavonoids (DHQ, DHM)-related DEGs of ten structural enzyme genes, three myeloblastosis transcription factors (MYB TFs), one basic helix-loop-helix (bHLH) TF, and one WD40 domain-containing protein were obtained. The enzyme genes comprised three PALs, two CLs, two CHSs, one F3'H, one F3'5'H (directly converts DHQ to DHM), and one ANS. The expression profiles of randomly selected genes were consistent with the RNA-seq results. Our findings thus provide comprehensive gene expression resources for revealing the molecular mechanism from DHQ to DHM in A. grossedentata. Importantly, this work will spur further genetic studies about A. grossedentata and may eventually lead to genetic improvements of the DHQ content in this plant.

The complete chloroplast genome of Ampelopsis grossedentata (Hand.-Mazz.) W. T. Wang (Family: Vitaceae) and its phylogenetic analysis.

Ampelopsis grossedentata (Hand.-Mazz.) W. T. Wang is rich in flavonoids and also displays excellent pharmacological activities. The phylogenetic relationship between A. grossedentata and other related Vitaceae family members remains unclear. The chloroplast (cp) genome is a useful model for assessing genome evolution. In this study, we assembled the cp genome of A. grossedentata using the high-throughput Illumina pair-end sequencing data and characterized the genome to providing useful information for future genetic studies. The circular cp genome was 162,147 bp in size, including a large single-copy (LSC) region of 89,244 bp and a small single-copy (SSC) region of 18,439 bp, which were separated by two inverted repeat (IR) regions (27,232 bp each). A total of 135 genes were predicted, including 8 ribosomal RNAs (rRNAs), 37 transfer RNAs (tRNAs), and 90 protein-coding genes (PCGs). Furthermore, phylogenetic analysis revealed that A. grossedentata within Ampelopsis genus and formed a different clade from other three congeneric species. This study provides useful information for future genetic study of A. grossedentata.

GC-MS analyses of the volatiles of Houttuynia cordata Thunb.

GC-MS is the basis of analysis of plant volatiles. Several protocols employed for the assay have resulted in inconsistent results in the literature. We developed a GC-MS method, which were applied to analyze 25 volatiles (α-pinene, camphene, ß-pinene, 2-methyl-2-pentenal, myrcene, (+)-limonene, eucalyptol, trans-2-hexenal, γ-terpinene, cis-3-hexeneyl-acetate, 1-hexanol, α-pinene oxide, cis-3-hexen-1-ol, trans-2-hexen-1-ol, decanal, linalool, acetyl-borneol, ß-caryophyllene, 2-undecanone, 4-terpineol, borneol, decanol, eugenol, isophytol and phytol) of Houttuynia cordata Thunb. Linear behaviors for all analytes were observed with a linear regression relationship (r2>0.9991) at the concentrations tested. Recoveries of the 25 analytes were 98.56-103.77% with RSDs <3.0%. Solution extraction (SE), which involved addition of an internal standard, could avoid errors for factors in sample preparation by steam distillation (SD) and solidphase micro extraction (SPME). Less sample material (≍0.05g fresh leaves of H. cordata) could be used to determine the contents of 25 analytes by our proposed method and, after collection, did not affect the normal physiological activity or growth of H. cordata. This method can be used to monitor the metabolic accumulation of H. cordata volatiles.

Quality evaluation of Houttuynia cordata Thunb. by high performance liquid chromatography with photodiode-array detection (HPLC-DAD).

A new, validated method, developed for the simultaneous determination of 16 phenolics (chlorogenic acid, scopoletin, vitexin, rutin, afzelin, isoquercitrin, narirutin, kaempferitrin, quercitrin, quercetin, kaempferol, chrysosplenol D, vitexicarpin, 5-hydroxy-3,3',4',7-tetramethoxy flavonoids, 5-hydroxy-3,4',6,7-tetramethoxy flavonoids and kaempferol-3,7,4'-trimethyl ether) in Houttuynia cordata Thunb. was successfully applied to 35 batches of samples collected from different regions or at different times and their total antioxidant activities (TAAs) were investigated. The aim was to develop a quality control method to simultaneously determine the major active components in H. cordata. The HPLC-DAD method was performed using a reverse-phase C18 column with a gradient elution system (acetonitrile-methanol-water) and simultaneous detection at 345 nm. Linear behaviors of method for all the analytes were observed with linear regression relationship (r(2)>0.999) at the concentration ranges investigated. The recoveries of the 16 phenolics ranged from 98.93% to 101.26%. The samples analyzed were differentiated and classified based on the contents of the 16 characteristic compounds and the TAA using hierarchical clustering analysis (HCA) and principal component analysis (PCA). The results analyzed showed that similar chemical profiles and TAAs were divided into the same group. There was some evidence that active compounds, although they varied significantly, may possess uniform anti-oxidant activities and have potentially synergistic effects.

[Chemical constituents from Viscum nudum and their accelerating PC12 cell differentiation].

OBJECTIVE: To investigate chemical constituents of the stem of Viscum nudum and their bioacyivity. METHOD: The major chemical constituents were isolated from the AcOEt-solved part of ethanol-extract of the plant by column chromatography and the active screening test in vitro were taken out for looking for compounds to acccelerate PC12 cell differentiation. RESULT: 5 compounds were identified as eriodictyol (1), 5, 7-dihydroxy-3', 4'-dimethoxy flavanone (2), oleanolic (3), 5, 7-dihydroxychromone (4) and homeriodictyol (5) by spectral evidences, in which homeriodictyol (5) had acceleration differentiation to PC12 cell. CONCLUSION: All compounds were obtained from this plant for the first time, and bioactive constituent was observed in the AcOEt-solved part.

[Study on chemical constituents from Incarvillea arguta and their accelerating PC-12 cell differentiation].

OBJECTIVE: To study chemical constituents of Incarvillea arguta and their accelerating PC-12 cell differentiation. METHOD: The constituents were isolated and repeatedly purified on silica gel column chromatography, and were identified on the basis of physicochemical and spectroscopic analysis. The neurotrophic activity of different portion and all purified compounds from I. arguta was determined on the model of PC-12 cell. RESULT: Five compounds were isolated from BuOH portion of alcohol extraction of I. arguta. Their structures were identified as plantarenaloside (I), 5-hydroxy-4', 6 7-trimethoxy-flavone (II), 4', 5-dihydroxy-6, 7-dimethoxyflavone (III), 4', 5-dihydroxy-7-methoxyflavone (IV), 5-dydroxy-4', 7-dimethoxyflavone (V). CONCLUSION: Compound I is isolated from the plant for the first time and it has neurotrophic activity for PC-12 cell. Compounds II approximately V are isolated from the genus Incarvillea for the first time.