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Objective To establish an HPLC method for simultaneous assay of macrocyclic polyphenols from Penthorum chinense Pursh, pinocembrin-7-O-[4'', 6''-(S)-hexahydroxydiphenoyl]-β-D-glucose (PHG), pinocembrin-7-O-[3''-O-galloyl-4'', 6''-(S)-hexahydroxydiphenoyl]-β-D-glucose (PGHG) and pinocembrin dihydrochalcone-7-O-[3''-O-galloyl-4'', 6''-(S)-hexahydroxydiphenoyl]-β-D-glucoside or thonningianin A (THA), and optimize the extraction process. Methods The total extraction rate of PHG, PGHG, THA was used as an investigated index to analyze the extracts from Penthorum chinense Pursh. Orthogonal design was applied to evaluate solvent amount, extraction time, solvent concentration and extraction times as the influencing factors for the optimal extraction process of macrocyclic polyphenols from Penthorum chinense Pursh. Results When this content assay method was adopted, there were good linear relationships for PHG, PGHG, THA in the linear range. The recoveries were between 100.90% to 102.04% with the RSDs below 1.5%. The optimal extraction process was involved in cutting Penthorum chinense Pursh into 3-5 cm, adding 10 times 80% ethanol aqueous solution by volume and refluxing 2 hours twice. The extraction rate of macrocyclic polyphenols was above 90% with this process. Conclusion This assay method is accurate, stable, and repeatable. The optimized extraction process is stable and feasible for further development and utilization.
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Introduction: For the rapid and accurate genetic identification and authentication of living organisms, improved random amplified polymorphic DNA (RAPD) fragment based development of sequence-characterized amplified region (SCAR) markers is an important genetic technique. Objective: This study aimed to develop SCAR markers for perennial herb Eclipta prostrate (E. prostrate). Methods: Here the RAPD fragments by improved RAPD amplification with primers A11 and N-7 for E. prostrate were cloned into pGEX-T vector, and PCR amplification identified the positive clones. After the enzymatic digestion, they were sequenced with Sanger sequencing. Results: Two SCAR markers were developed, which were very specific to E. prostrate, not found in Penthorum chinense Pursh(P. chinense). The nucleotide sequence search by BLAST GenBank database showed that they are novel in E. prostrate, therefore they were deposited in Genbank with accession number KX671034, KX671035. The markers did not show any identity to other species. Conclusions: Thus, in this study two specific SCAR markers were developed for genetically distinguishing and identifying the plant species E. prostrate from herb P. chinense and others.
Introducción: Verificación genética del arbusto Eclipta prostrate (Asteraceae) (Para la identificación y verificación genética rápida y precisa de organismos vivos, el uso de fragmentos de ADN polimórfico amplificado aleatoriamente (RAPD) mejorado de marcadores de región amplificada caracterizada por secuencia (SCAR) es una técnica genética importante. Objetivo: Este estudio tuvo como objetivo desarrollar marcadores SCAR para la hierba perenne Eclipta postrate (E. postrate). Métodos: En este estudio os fragmentos RAPD mediante amplificación RAPD mejorada con los cebadores A11 y N-7 para E. postrate se clonaron en el vector pGEX-T, y la amplificación por PCR identificó los clones positivos. Después de la digestión enzimática, se realizó una secuenciación Sanger. Resultados: Se desarrollaron dos marcadores SCAR, muy específicos para E. postrate, que no se encuentran en Penthorum chinense Pursh (P. chinense). La búsqueda de las secuencias de nucleótidos con BLAST en GenBank mostró que son nuevos en E. postrate, por lo que fueron depositados en Genbank con los números de acceso: KX671034 y KX671035. Los marcadores no mostraron ninguna identidad a otras especies. Conclusiones: En este estudio se desarrollaron dos marcadores SCAR específicos para distinguir e identificar genéticamente la especie de planta E. postrate de la hierba P. chinense y otras.
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Objective To establish a method for the simultaneous content determination ofquercetin,kaempferol and pinocembrin in Penthorum Chinense Pursh.Methods The HPLC analysis was carried out on Alltima C18 (250 mm × 4.6 mm,5 μm) with a mixture of methanol-0.2％ phosphate (45:55) as the mobile phase.The determination wavelength was set at 270 nm with the flow rate of 1.0 ml/min.The column temperature was set at 30 ℃.Results The quercetin,kaempferol and pinocembrin showed good linearity in the range of 0.316-10.120 μg (r=0.999 9),0.012-0.397 μg (r=0.999 6) and 0.063-2.016 μg (r=0.999 8) respectively.The average recovery rates of quercetin,kaempferol and pinocembrin were 99.34％ (RSD=1.51％),99.76％ (RSD=l.96％) and 98.34％ (RSD=1.56％) respectively.Conclusions The method is accurate and sensitive,which can be used for the content determination of quercetin,kaempferol and pinocembrin in Penthorum Chinense Pursh.
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AIM:To explore the effect of Penthorum chinense Pursh and Puerariae flos-containing serum on L-02 liver cell injury induced by alcohol and its possible mechanism. METHODS:After preparing drug-containing serum, the L-02 cells cultured in vitro were divided into 6 groups:blank control group, model group, 1∶1 group, 2∶1 group and 1∶2 group of combination of Penthorum chinense Pursh and Puerariae flos, and tiopronin group. The viability of the L-02 cells was measured by MTT assay. The activity of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and superoxide dismutase (SOD), and the content of malondialdehyde ( MDA) were detected by enzyme label methods. The expression of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), nuclear factor E2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) at mRNA and protein levels was determined by real-time PCR and Western blot, respectively. RE-SULTS:Compared with control group, the levels of ALT, AST and MDA were increased significantly, and SOD was de-creased in model group ( P <0.01). Compared with model group, these indexes in all treatment groups were opposite (P<0.01). Compared with control group, the expression of TNF-α and IL-6 at mRNA and protein levels was significantly increased, the mRNA and protein expression of Nrf2 and HO-1 was decreased (P<0.01). Compared with model group, these indexes in combination groups were opposite (P<0.01). CONCLUSION:The therapeutic effects of Pentehorum chinensa Pursh and Puerariae flos-containing serum may affect the expression levels of TNF-α, IL-6, Nrf2 and HO-1, and reduce the inflammatory reaction and oxidative stress in alcohol-induced L-02 liver cells, which plays a role in attenuating alcoholic liver injury.
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Objective: To study the chemical constituents of Penthorum chinense. Methods: The compounds were isolated and repeatedly purified by column chromatography over macroperous resin, polyamide, silica gel, and Sephadex LH-20, preparative TLC, and preparative RP-HPLC. Their structures were elucidated by physicochemical properties and spectral analyses. Results: Nine flavonoids were isolated and identified as 6'-hydroxy-2'-methoxy-dihydrochalcone-4'-O-β-D-glucopyranoside (1), kaempferol (2), qercetin (3), quercitrin (4), avicularin (5), afzelin (6), helicin (7), pinocembrin-7-O-[4″,6″-(S)-hexahydroxydiphenoyl]-β-D- glucoside (8), and pinocembrin-7-O-[3″-O-galloyl-4″,6″-(S)-hexahydroxydiphenoyl]-β-D-glucoside (9). Conclusion: Compound 1 is a new dihydrochalcone and named penchinoside. Compounds 5-7 are isolated from genus Penthorum Gronov. ex L.for the first time.
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Penthorum chinense is widely distributed in eastern Asia. It has various efficacies with excellent pharmaceutical and edible prospects. It contains various types of chemical components, and main components are flavonoids, lignans, coumarins, acetophenones, tannins, triterpenes, organic acids, esters, and volatile oils. This article classifies and summarizes the chemical constituents and hopefully provides a reference for further research, development and utilization of P. chinense.
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Objective In order to obtain the reference sequences and relative expression of transcription genes and study the genetic base of active ingredients in Penthorum chinense, which were useful for researching functional gene to P. chinense. Methods In this study, by performing Illumina Hiseq 2000 and de novo assembly, the transcriptome of whole plant was sequenced, the data were filtered and assembled, and the unigene was compared and annotated. Meanwhile, the genes related to the synthesis of metabolic pathway of active ingredients in P. chinense were analyzed. Results Totally, 40 005 442 valid short sequences were obtained, and 42 306 unigenes were spliced by de novo. Also, a total of 518 open reading frames (ORF) were obtained by ORF analysis, and 75 ORF of them had transcription factor domains. In addition, by performing KEGG pathway analysis, 33, 32, 59, and 68 unigenes were found to be involved in the pathway of flavonoid biosynthesis, steroid biosynthesis, terpenoid backbone biosynthesis, and 2-oxocarboxylic acid metabolism, respectively. Conclusion The datasets provided in this study will contribute significantly to genetic improvement and study on the genes related to biosynthesis pathway of pharmaceutical active substances from P. chinense.
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Objective To establish an HPLC method for the simultaneous content determination of quercetin and kaempferol in Penthorum Chinense Pursh. Methods The HPLC analysis was carried out on Kromasil C18 (250 mm× 4.0 mm, 5.0 μm) with a mixture of methanol-0.1% phosphate (50:50) as the mobile phase. The determination wavelength was set at 360 nm with the flow rate of 1.0 mL/min. The column temperature was set at room temperature. Results The quercetin and kaempferol showed good linearity in the range of 0.105 6–2.112 0 μg and 0.023 6–0.472 μg respectively. The average recovery rates of quercetin and kaempferol were 98.46% (RSD=2.29%) and 98.17%(RSD=1.99%) respectively. Conclusion The method is accurate and sensitive, which can be used for the content determination of quercetin and kaempferol in Penthorum Chinense Pursh.
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Objective: To observe anti-aging effect and mechanism of concentrated solution of aqueous extract from Penthorum chinense. Methods: Aging mice models were caused by D-galactose, learning and memory functions were investigated by avoid dark experiment, the spleen index, activity of MAO in liver, activities of SOD and MDA in serum were recorded, and the expression of p21 and p53 protein was detected by Western blotting. Results: Concentrated solution of P. chinense by aqueous extraction increased the ability of learning and memory, improved spleen index and activity of SOD in serum, but it decreased the activity of MDA in serum and the activity of MAO in liver, and inhibited p21 and p53 protein expression in a dose-dependent manner. Conclusion: Concentrated solution of aqueous extract from P. chinense could significantly exhibit anti-aging effects, its mechanism may be related to down-regulating the p53/p21 protein expression in aging signaling pathway, enhancing immune function, and removing free radicals.
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Quality control of Chinese materia medica (CMM), a key factor in restricting the development of modernization of CMM, has always been a hot issue to traditional Chinese medicine (TCM) circles, our society, and the public. At present, determination of chemical components is the main means for quality control of vast majority of CMM, but many components lack not only specificity, but also biological activities. This is bound to greatly reduce the value of quality standard of CMM, which makes it difficult to reflect the true quality of CMM. Quality marker (Q-Marker) of CMM is a new concept that brings forward higher requirement to quality control of CMM. Based on the basic conditions of Q-Marker, Leonurus japonicus and Penthorum chinense have been investigated systematically. This paper introduces theIR research processes from five aspects: effective material basis, specificity of the isolated components, chemical structure and bioactivity, measurability, and fingerprint spectrum.
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OBJECTIVE: To develop a method of quantitative analysis of multi-components by single marker(QAMS) for simultaneous determination of five compounds in extract of Penthorum chinense Pursh. METHODS: A HPLC method was established on an Agilent ZORBAX SB-C18column(4.6 mm×250mm, 5μm). The mobile phase was acetonitrile-0.2% formic acid aqueous solution. Gradient elution was performed at a flow rate of 1.0 mL·min-1. The column temperature was 30℃ and the detection wavelength was set at 280 nm. Pinocembrin-7-0-β-D-glucoside was selected as the internal reference substance, and the relative correlation factors (RCF) of gallic acid, brevifolin carboxylic acid, 2, 6-dihydroxyacetophenone-4-0-β-D-glucoside, quercetin-3-0-β-D-xyloside and quercetin-3'-0-α-L-rhamnoside were determined by HPLC. The contents of the six components were determined by both QAMS method and external standard method and the results were compared. The accuracy and feasibility of the new method were validated by QAMSmethod and external standard method. RESULTS: No significant differences were observed in the determination results of five constituents except brevifolin carboxylicacid in 21 batches of extract of P. chinense by QAMS method and external standard method. CONCLUSION: The QAMS method is feasible and suitable to determine the contents of five constituents in the extract of P. chinense.
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This study was aimed to determinate the content of quercetin from different parts (such as flowers, fruits, stems, leaves and branches) of Penthorum chinense Pursh. from Sichuan province so as to offer reference for the comprehensivedeve lopment and utilization of P. chinense Pursh. The quantitative determination of quercetin from five groups of different parts of P. chinense Pursh. were carried out and put into comparison by the reversed-phase high performance liquid chromatography (RP-HPLC). The results showed that there were larger differences of content among different parts. The highest content of the medical parts were flowers and fruits, which was followed by the leaves, stems and branches. It was concluded that different parts (such as flowers, fruits, stems, leaves and branches) of P. chinense Pursh. from Sichuan province should be divided in the clinical and productive practice, which was to supply the scientific basis for enhancing the curative effect and reasonable exploitation and utilization of the resource of P. chinense Pursh.
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OBJECTIVE: To study the chemical constituents of Penthorum chinense Pursh. METHODS: Many chromatography means were used in separation and purification, and the structures of all compounds were identified by the means of spectroscopic analysis and physico-chemical properties. RESULTS Fourteen compounds were elucidated as: pinocembrin-7-O-β-D-glucopyranoside(1), kaempfer-ol(2), vanillic acid(3), quercetin-3-O-β-D-xylopyranosyl-(1→2) -β-D-galactopyranoside(4), kaempferol-3-O-α-L-rhamnopyranoside(5), pinocembrin(6), quercetin(7), afzelin(8), 11-O-galloylbergenin(9), bergenin(10), 4-galloylbergenin(11), protocatechuic acid(12), catechins (13), gallic acid(14). CONCLUSION: Compounds 1, 3, 6, 7, 8, 12, 13 and 14 are obtained from the title plant for the first time.
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Objective To observe the effects of serum with drug Penthorum chinense Pursh extractum on transforming growth factor (TGF)-β1 and collagen I secretions of activated hepatic stellate cells. Methods Twenty male SD rats were divided into 2 groups, and were administered with 0.9% sodium chloride solution and Penthorum chinense Pursh extraetum via gastrogavage for 3 days respectively and then sacrificed. Serum samples of these rats were collected. HSC-T6 cells were divided into the normal group and the treatment group. The cells of the normal group were incubated in Dulbecco's modified eagle medium (DMEM)with sera of normal rats, while those of the treatment group were incubated in DMEM with sera from Penthorum chinense Pursh extractum treated rats. The HSC-T6 viability was observed by AlamarBlue assay, while the toxicity of Penthorum ehinense Pursh extractum was measured by 3-(4, 5-Dimethyhhiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. The expression of collagen I and TGF-β1 mRNA were determined by real timepolymerase chain reaction (real time PCR). The protein expressions of collagen I and TGF-β1 were analyzed by Western blotting. The data were analyzed by single factor analysis of variance and pairwise comparison was done by q test. Results Different concentrations of sera from Penthorum chinense Pursh extractum treated rats could all inhibit HSC-T6 proliferation, especially when the sera concentration were 10% and the HSC-T6 cells were incubated for 24 h (P＜0.01 ). MTT assay indicated that sera from Penthorum chinense Pursh extractum treated rats showed no obvious toxicity to HSC-T6 compared with those from normal rats (P ＞0.05). After 24 h incubation, 10% sera from Penthorum chinense Pursh extractum treated rats could significantly down-regulate mRNA expression of TGF-β1 and collagen I compared with normal group (TGF-β1 2.790±0.174 vs 9. 827 ± 1.429, P＜0.01 ; collagen I 1.213 ± 0.099 vs 4.053 ± 1.005, P＜0.01 ). Mcanwhile, the protein expressions of TGF-β1 and collagen I were also obviously inhibited in drug treated group compared with normal group (P＜0.01). Conclusions Serum from Penthorum chinense Pursh extractum treated rats can significantly decrease TGF-β1 and collagen I secretions of activated hepatic stellate cells, which provides the experimental evidence for liver fibrosis treatment.