Screening of plant growth-promoting rhizobacteria and its effect on seed germination of Polygonum multiflorum / 中国中药杂志

In this study, the rhizobacteria and actinomycetes of Polygonum multiflorum were screened for the strains with indole acetic acid(IAA)-producing capacity by Salkowski method, the siderophore-producing strains by Chrome Azurol S(CAS) assay, and the strains with inorganic phosphorus-solubilizing capacity by PKO inorganic phosphorus medium. The strains were identified by morphological identification, physiological and biochemical characteristics, and 16 S rDNA sequences. Furthermore, the effect of growth-promoting strains on the seed germination and development of P. multiflorum was tested. The results showed that among 196 strains, two strains F17 and F42 were found to be capable of producing IAA and siderophore and solubilizing inorganic phosphorus simulta-neously. For F17 and F42, the results are listed below: 38.65 and 33.64 mg·L~(-1) for IAA production, 0.85 and 0.49 for siderophore-producing capacities(A_s/A_r), and 1.35 and 1.70 for inorganic phosphorus-solubilizing capacities(D/d), respectively. Comprehensive analysis revealed that strains F17 and F42 were identified as Pseudochrobactrum asacharolyticum and Bacillus aryabhattai, respectively, and both could significantly promote the seed germination of P. multiflorum.

Hepatotoxicity of Main Components of Polygonum multiflorum and Its Effects on Drug Metabolism Enzyme / 中国药房

OBJECTIVE：To study the hepatotoxicity of main components of Polygonum multiflorum ，and investigate its toxic mechanism based on metabolic enzymes. METHODS ：ADMETlab 2.0 platform was used to forecast the toxic or carcinogenic effects of emodin ，physcion，rhein，stilbene glycoside and gallic acid on liver ，skin and heart. The effects of those components on cytochrome P 450 enzyme system （CYP1A2，CYP2C9，CYP2C19，CYP2D6，CYP3A4）were evaluated. The effects of different concentrations of emodin ，rhein，stilbene glycoside and gallic acid （10，20，40，80 μmol/L）on the survival rate of normal hepatocyte L 02 were detected. The effects of major components of P. multiflorum on the activity of UGT 1A1 enzyme were studied by in vitro reaction system ，using bilirubin as substrate. RESULTS ：Main components of P. multiflorum ，ie. emodin ，physcion， rhein and gallic acid ，showed strong toxic effects on the liver ，while stilbene glycosides possessed weak toxic effects on the liver. Emodin and physcion had strong inhibitory effects on CYP 1A2 and medium inhibitory effects on CYP 2C9，CYP2D6 and CYP3A4；rhein showed medium inhibitory effects on CYP 1A2 and CYP 2C9，while stilbene glycoside and gallic acid possessed weak inhibitory effects on the above enzymes. Emodin （40，80 μmol/L）and gallic acid （40，80 μmol/L）could significantly reduce the survival rate of L 02 cells（P＜0.01）. The inhibition rate of 5，10，20，40，80 μmol/L emodin and gallic acid（except for 5 μmol/L emodin）on UGT 1A1 enzyme increased significantly （P＜0.01），and the inhibition effect of emodin on UGT 1A1 enzyme was reversible competitive inhibition. CONCLUSIONS ：The main components of P. multiflorum ，ie. emodin ，rhein and physcion ， are hepatotoxic ；the mechanism of it may be associated with inhibiting the activity of CYP 1A2 and CYP 2C9 and competitively blocking rate-limiting enzyme UGT 1A1 in the process of bilirubin metabolism.

The compound cell model-based evaluation for idiosyncratic liver injury of Cis-SG and Trans-SG / 药学学报

In this study, a composite cell model for evaluation of idiosyncratic drug-induced liver injury (IDILI) was established in vitro from the perspective of immune inflammation. And this model was used to evaluate the risk of IDILI for 2,3,5,4'-tetrahydroxy-cis-stilbene-2-O-β-glucoside (Cis-SG) and 2,3,5,4'-tetrahydroxy-trans-stilbene-2-O-β-glucoside (Trans-SG). To determine the low, medium, and high dosage of Cis-SG and Trans-SG, CellTiter-Glo® 3D Cell Viability Assay was used to detect the effects of Cis-SG and Trans-SG on cell viability of HepG2 cells in three dimensional (3D) culture, and MTT assay was used to detect the effects of Cis-SG and Trans-SG on cell viability of THP-1 derived macrophages. THP-1 derived macrophages were incubated by Cis-SG and Trans-SG directly or supernatants from HepG2 cells incubated with them. Enzyme linked immunosorbent assay (ELISA) was used to detect the levels of interleukin-1β (IL-1β) in the supernatants of the THP-1 derived macrophages. Western blot and reverse transcription-polymerase chain reaction (RT-PCR) were used to determine the expression of apoptosis-associated speck-like protein (ASC), Nod-like receptor protein 3 (NLRP3), cysteinyl aspartate specific proteinase-1 (caspase-1), and IL-1β in THP-1 derived macrophages. The results showed that there was no effect on the secretion of IL-1β in THP-1 derived macrophages incubated by Cis-SG and Trans-SG directly. However, the secretion of IL-1β, the protein and mRNA expression of ASC, NLRP3, caspase-1, and IL-1β significantly increased in THP-1 derived macrophages incubated by supernatants from HepG2 cells incubated with 1, 5, and 25 μmol·L-1 Cis-SG or 25 μmol·L-1 Trans-SG. In summary, the composite cell model for evaluation of IDILI established in vitro has been successfully applied in testing Cis-SG and Trans-SG. This composite cell model is helpful to evaluate and screen drugs with IDILI risk in vitro preliminarily, which provides methods for predicting and solving the idiosyncratic liver toxicity of drugs.

Reference gene screening for Real-time quantitative PCR in Polygonum multiflorum / 中国中药杂志

To select suitable references gene of Polygonum multiflorum for gene expression analysis in different tissues, five candidate reference genes like Actin,GAPDH,SAND,PP2A,TIP41 were selected from the transcriptome data of P. multiflorum, then the specific primers were designed. The expression stability of the five reference genes in different tissues of P. multiflorum was analyzed by Real-time quantitative PCR through avilable analysis methods such as geNorm, NormFinder, BestKeeper, Delta CT and RefFinder, to ensure the reliability of the analysis results. The results showed that there were significant differences in the expression levels and stability of candidate genes in different tissues of P. multiflorum. Ct distribution analysis of the expression levels of candidate genes showed that the expression levels of Actin and GAPDH genes were relatively high in different tissues, while the expression levels of SAND, PP2A and TIP41 were lower. The stability of each candidate gene was analyzed by different methods. The results of geNorm analysis showed that the expression of PP2A and GAPDH was the most stable, the expression stability of SAND was the worst, the stability of PP2A was the highest in both NormFinder and Delta CT, the stability of SAND was the lowest, and the stability of Actin was the most stable in BestKeeper analysis. Through the comprehensive evaluation and analysis of the stability of candidate genes by RefFinder, it is concluded that the stability of PP2A gene is the highest, followed by GAPDH, Actin, TIP41, SAND, and SAND gene is the worst. Therefore, the PP2A gene is an ideal reference gene for the analysis of gene expression in different tissues of P. multiflorum.

New phenolic constituents obtained from Polygonum multiflorum / 中草药·英文版

Objective: To isolate the phenolic compounds obtained from the dried roots of Polygonum multiflorum and investigate their pharmacological activities. Methods: The chemical constituents were isolated and purified by combining them with a macroporous resin (DM-8), MCI gel, and Sephadex LH-20 and by performing ODS column chromatography. Their structures were elucidated by 1D and 2D NMR analyses, as well as mass spectrometry. The isolated compounds were evaluated to determine their hepatoprotective and α-glucosidase inhibitory activities in vitro. Results: Two phenolic compounds, namely, polygonimitin E (1) and polygonimitin F (2), were isolated from the dried roots of P. multiflorum. Compound 2 (10 µmol/L) only showed moderate hepatoprotective activity against N-acetyl-p-aminophenol (APAP)-induced HepG2 cell damage. Unfortunately, these two compounds exhibited no α-glucosidase inhibitory activity. Conclusion: Compounds 1 and 2 were new compounds. Compound 2 could be one of the potential hepatoprotective constituents of P. multiflorum.

Study on potential hepatotoxicity of main monomers of Polygonum multiflorum based on liver micro-tissue / 中国中药杂志

In this study, we aimed to establish a rat liver micro-tissue evaluation system to evaluate the hepatotoxicity of the main monomers in Polygonum multiflorum. Rat primary hepatocytes were isolated and purified by two-step in situ perfusion method to prepare hepatic parenchymal cells. The ultra-low adsorption plate and the inverted model were used to establish an in vitro hepatotoxicity evaluation system. After the system was established, the main monomer components(monanthone with emodin type, rhein, emodin, emodin-8-O-β-D-glucopyranoside, physcion) of P. multiflorum were selected for in vitro hepatotoxicity evaluation. This study showed that the primary cells of the liver can form liver micro-tissues in the low adsorption plate method and the mold perfusion method, with good liver structure and function, which can be used to evaluate the hepatotoxicity of the drug to be tested after long-term administration. The five monomers to be tested in P. multiflorum can significantly affect the proliferation of primary liver micro-tissues in rats in a dose-and time-dependent manner. The hepatotoxic effects were as follows: monanthone with emodin type > rhein > emodin > emodin-8-O-β-D-glucopyranoside > physcion. The results suggested that the emodin-type monoterpene and rhein might be the potential hepatotoxic components, while the metabolites of emodin-8-O-β-D-glucoside and emodin methyl ether showed more toxic risks. The rat primary hepatocyte micro-tissue model system established in this experiment could be used to achieve long-term drug administration in vitro, which was consistent with the clinical features of liver injury caused by long-term use of P. multiflorum. The experimental results provided important information and reference on the clinical application and toxic component of P. multiflorum.

A new flavonostilbene glycoside from roots of Polygonum multiflorum / 中国中药杂志

Polygonflavanol B(1), a new flavonostilbene glycoside, was isolated from the roots of Polygonum multiforum(Polygonaceae) by various column chromatography methods including macroporous resin HP-20, silica gel, Sephadex LH-20, and preparative HPLC. The structure with absolute configuration of the new compound was identified by its physicochemical properties, spectroscopic data, ECD calculation, and chemical method.

Simultaneous determination of four anthraquinones in Polygonum multiflorum by QAMS / 中国中药杂志

A simple, specific and selective quantitative analysis of multi-components by single marker(QAMS) method for simultaneous determination of anthraquinones and anthraquinone glycosides in Polygonum multiflorum was developed. Four main anthraquinones and its glycosides, emodin, emodin-8-O-β-D-glucoside, physcion and physcion-8-O-β-D-glucoside were selected as analytes to evaluate the quality of P. multiflorum. Emodin was used as the internal standard, and the relative correction factors(RCFs) between emodin and the other three anthraquinones were calculated. Comparison of the contents of the four components in 30 batches of P. multiflorum from different regions and 12 batches decoction pieces from different manufacturers by QAMS and external standard method(ESM) showed that there was no significant difference between QAMS and ESM for quantification of the four main components by using relative error results, and the QAMS method was accurate and reliable, and had a good repeatability. In addition, compared with the results calculated by the difference method between total anthraquinone and free anthraquinone in the content determination of P. multiflorum in Chinese Pharmacopoeia, the results of direct determination combined anthraquinone by QAMS were very close to that by measured the external standard method. Therefore, simultaneous quantification of four main anthraquinones by using QAMS is suitable to evaluate the quality of P. multiflorum. Then the optimized assay method of the combined anthraquinone contents showed simple and feasible, which could be replaced and improved the quantification method of the combined anthraquinone in the current Chinese Pharmacopeia.

Study on Optimization of the Nine-time Repeat Steaming and Sun-drying Process Processing Technology of Polygonum multiflorum / 中国药房

OBJECTIVE：To establish methods for the content determination of stilbene glycoside ，emodin，emodin methyl ether and total polysaccharide in P. multiflorum ，and to optimize the processing technology of traditional nine-time repeat steaming and sun-drying process of P. multiflorum . METHODS ：HPLC method was adopted to determine the contents of stilbene glycoside ， emodin，and emodin methyl ether. The content of total polysaccharide was determined by UV spectrometry. With steaming time ， drying time and drying temperature as independent variables ，the contents of stilbene glycoside ，emodin，emodin methyl ether and total polysaccharide as indexes ，the processing technology of nine-time repeat steaming and sun-drying process of P. multiflorum was optimized by weighted comprehensive score method and central composite design-response surface method ，and the validation tests were conducted. RESULTS ：The linear range of sample size of stilbene glycoside ，emodin and emodin methyl ether were 0.27-2.7 µg（r＝0.997 1），0.063-0.63 µg（r＝0.999 9），0.038-0.38 µg（r＝0.990 9），respectively. The linear range of mass concentration of total polysaccharide was 2.07-12.42 µg/mL（r＝0.999 6），respectively. RSDs of precision ，stability and reproducibility tests were all lower than 2％. The recoveries were 99.43％-101.06％（RSD＝0.63％，n＝6），98.74％-120.33％ （RSD＝1.34％，n＝6），98.39％-102.44％（RSD＝1.49％，n＝6），99.51％-101.98％（RSD＝0.87％，n＝6），respectively. The optimal processing technology was steaming time for 4.5 h，drying time for 9 h，drying temperatrue for 66 ℃ and repeat processing for 9 times. Results of 3 times of validation tests showed that the comprehensive score of optimal technology was 35.69， relative error of which to predicted value （36.90）was 3.30％. CONCLUSIONS ：Established method is simple and reproducible ， and the optimal processing technology is predicive ，stable feasible.

Establishment of Fingerprint and Multivariate Statistical Analysis of Compatible Herb Pairs with Polygonum multiflorum as the Core / 中国药房

OBJECTIVE：To establish UPLC fin gerprint of 32 compatible herb pairs with Polygonum multiflorum as the core ， and to conduct multivariate statistical analysis. METHODS ：UPLC method was adopted. Using P. multiflorum and single decoction pieces of compatible herb as reference ，UPLC fingerprints of 32 compatible herb pairs of P. multiflorum were drawn. Common peaks were confirmed by relative retention time and UV absorption spectrum. Non-supervised PCA and supervised OPLS-DA were conducted by using SPSS 19.0 software and SIMCA 13.0 software. RESULTS ：There were totally 12 common peaks in UPLC fingerprints of 32 compatible herb pairs of P. multiflorum . The results of non-supervised PCA showed that the accumulative variance contribution rate of primary 6 principal components was 84.633％. The results of cluster analysis of PCA comprehensive score showed that single decoction piece of P. multiflorum ，compatible herb pairs of P. multiflorum with Lycium barbarum ，Rehmannia glutinosa，Paeonia lactiflora ，Codonopsis pilosula ，Eclipta prostrate ，Angelica sinensis ，Glycyrrhiza uralensis ，Astragalus membranaceus and Ophiopogon japonicus were clustered into one category ；others were clustered into one category. Results of supervised OPLS-DA analysis showed that eigen values of 4 principal components were 2.32，2.61，1.58 and 0.90，respectively. There were differences in the contents of 12 common components in the compatibility of P. multiflorum with tonic medicine and non-tonic medicine. The changes of the content of the components after compatibility with tonic medicine were similar. Common peak 7，4，6，3 were main reasons for the differences （variable importance projection value were all higher than 1）. CONCLUSIONS：Established fingerprint is simple in operation ，and can be combined with multivariate statistical analysis to evaluate the content changes of common components of 32 compatible herb pairs with P. multiflorum as the core.

Effect of Polygonum multiflorum-Andrographis paniculata intercropping system on rhizosphere soil actinomycetes community structure and diversity of P. multiflorum / 中国中药杂志

To investigate the effect of Polygonum multiflorum-Andrographis paniculata intercropping system on rhizosphere soil actinomycetes of P. multiflorum, the community structure and diversity of soil actinomycetes were studied by using the original soil as the control group and the rhizosphere soil actinomycetes communities of P. multiflorum under monoculture and intercropping systems as the experimental group. In this study 655 221 effective sequences were obtained with an average length of 408 bp. OTU coverage and rarefaction curve showed that the sequencing could represent the real situation of soil actinomycetes. According to the results of alpha diversity analysis, the diversity soil actinomycetes varied as follows: original soil>intercropping soil>monoculture soil. The soil actinomycetes community structure and the relative abundance of dominant genera were significantly changed by both monoculture and intercropping, especially monoculture. OTU clustering and PCA analysis of soil samples showed that all the soil samples were divided into three distinct groups and the original soil was more similar to intercropping soil. In addition, intercropping increased the relative abundance of some beneficial actinomyces, such as Kitasatospora and Mycobacterium, which was beneficial to maintain soil health and reduce the occurrence of soil-borne diseases. The results show that, P. multiflorum-A. paniculata intercropping reduced the change of community structure and the decrease of diversity of soil actinomycetes caused by P. multiflorum monoculture, and made the actinomycete community in rhizosphere soil of P. multiflorum close to the original soil.

Structural characterization of polysaccharide from Polygonum multiflorum and its immunomodulatory activity / 中草药

Objective: To extract and separate a polysaccharide from Polygonum multiflorum, characterize its structural features and study its immunomodulatory activity. Methods The polysaccharide from P. multiflorum (PMT) was isolated and purified by water extraction and ethanol precipitation following Q-Sepharose Fast Flow ion exchange chromatography column. Molecular weight of PMT was determined by high performance gel permeation chromatography-multiple angle laser light scattering (HPGPC-MALLS), and monosaccharide composition was analyzed by HPLC with PMP (1-phenyl-3-methyl-5-pyrazolone) pre-column derivatization, respectively. The structure of PMT was characterized by proton nuclear magnetic resonance spectrum (2D-NMR). The immunomodulatory activities were tested by MTT, neutral red colorimetric assay and Griess method. Results: PMT was a kind of α-1,4-glucan, and its molecular mass was 3.96 × 105. PMT promoted the proliferation and phagocytosis of RAW 264.7 cells, and significantly induced the increase of NO production in a dose-dependent manner. Conclusion: The polysaccharide from P. multiflorum is a linear α-1,4-glucan with potent immunomodulatory activity, which would be potentially developed as an effective drug.

Target screening and antihyperglycemic activity on SGLT2 and α-glucosidase in vitro and in vivo of TSG from Polygonum multiflorum / 中草药

Objective: To study the mechanism that TSG can reduce plasma glucose level by inhibiting sodium-dependent glucose cotransporters 2 (SGLT2) and α-glucosidase in vitro and in vivo. Methods: Molecular docking method was used to study the binding affinities of TSG and diabetes related targets. The structures of targets were taken from Protein Data Bank or references. 1-NBDG and PNPG were used as the substrates for the inhibition assays of TSG against SGLT2 and α-glucosidase respectively in vitro. The antihyperglycemic activity of TSG was operated by oral glucose tolerance test (OGTT) and urinary glucose excretion (UGE) test in rats. Results: TSG was identified as the inhibitors of SGLT2 with the docking score of -9.35 less than -9.79 of dapagliflozin as the positive control and α-glucosidase with the docking score of -5.44 compared to -5.58 of acarbose as the positive control. TSG showed the inhibitory rate of 21.6% at the dose of 10 μmol/L against SGLT2 and 32.5% at the dose of 100 μmol/L in vitro test. Compared with model group, the group of 120 mg/kg dose had significant difference (P < 0.05) but the overall effect was not as strong as dapagliflozin in OGTT and UGE test. The result of rat in vivo test showed that glucose inhibition rate of TSG (120 mg/kg) was (9.3 ± 1.0)%, urinary glucose content was (435.5 ± 84.0) mg/kg, which showed certain hypoglycemic effect. Conclusion: TSG exhibited antiglycemic activity through inhibiting SGLT2 and α-glucosidase, which was considered to be a new lead compound of dual target inhibitors.

Developing spectrum-toxicity relationship with rough set theory for hepatotoxicity material basis of Polygonum multiflorum / 中国中药杂志

Idiosyncratic hepatotoxicity of Polygonum multiflorum has attracted a great attention in the world. The most toxic part of idiosyncratic hepatotoxicity was screened by MTT assay and flow cytometry, which was the 50% ethanol elute by macroporous adsorptive resins from alcohol-extraction of P. multiflorum. The fingerprints were collected by HPLC from 50% ethanol elute of crude and processed P. multiflorum from different habitats, then 14 common peaks were determined. Spectrum-toxicity relationship was analyzed by rough set theory(RST). Two main chemical components were predicted for idiosyncratic hepatotoxicity, in which TSG was the greater contributor. Idiosyncratic hepatotoxicity of TSG was tested in vitro, and the results indicated that TSG was the most important constituent contributed to idiosyncratic hepatotoxicity of P. multiflorum. The study showed the discovery of the main chemical components for idiosyncratic hepatotoxicity, and RST was effective for analyzing the spectrum-toxicity relationship, which could be a new method used in the effective/toxic constituents field of traditional Chinese medicine.

TCR immune repertoire characteristics of patients with drug-induced liver injury due to Polygonum multiflorum preparations / 中国中药杂志

To explore the immune status of patients with drug-induced liver injury caused by Polygonum multiflorum preparations,and analyze their immune characteristics. Case-control design was used to collect the cases of drug-induced liver injury caused by P. multiflorum preparations through key specialized surveillance. Five matching factors,namely type of P. multiflorum preparations,gender,age,basic diseases and concomitant medication were controlled. According to the ratio of 1 ∶ 1,cases of patients who took P. multiflorum preparations but with no liver injury were monitored at prospective hospitals. The demographic information,disease information,medication information and laboratory examination information of the two groups were recorded,and venous blood was collected. The gene sequence was detected by high-throughput sequencing technology,and the characteristics of TCR immune repertoire of the two groups were analyzed. A total of 46 pairs of patients were enrolled in the study. The results showed significant differences in the number of CDR3 and clone species,the length of amino acid sequence in CDR3 region,the abundance of V gene and J gene,the cross-linking of V-J gene and the diversity of immune repertoire between patients with drug-induced liver injury and patients without liver injury. The immunohistochemical diversity and high-frequency V-J cross-linking characteristics of patients with liver injury caused by P. multiflorum preparations were found,which provided a reference for screening out drug users to reduce the occurrence of liver injury caused by P. multiflorum preparations.

Effects of Processed Polygonum multiflorum Containing Serum on the Proliferation and the Expression of ER of Human Breast Cancer T- 47D Cells / 中国药房

OBJECTIVE： To study the effects of processed Polygonum multiflorum containing serum on the proliferation and the expression of estrogen receptor （ER） of human breast cancer T-47D cells， and to investigate its phytoestrogen （PE）-like effect. METHODS： Sexually immature SD rats were randomly divided into estradiol valerate （Ev） group （positive control， 0.12 mg/kg）， processed P. multiflorum low-dose and high-dose groups （0.75， 3 g/kg， by crude drug）， low-dose and high-dose processed P. multiflorum+Ev groups （same dose as single drug group）， with 10 rats in each group. Blank group was given constant volume of water intragastrically， and administration groups were given relevant medicine intragastrically； once day and night， for consecutive 4 days. Two hours after last administration， blank serum and containing serum were prepared. T-47D cells were also randomly divided into blank group， Ev group， low-dose and high-dose processed P. multiflorum groups， low-dose and high-dose processed P. multiflorum+Ev groups， and then were cultured in medium which contained 20％ blank serum or drug containing serum. CCK-8 assay was used to detect proliferation rate （PR）. Western blotting assay and RT-PCR were used to detect the protein and mRNA expression of ER-α and ER-β. RESULTS： Compared with blank group， PR of administration groups [each administration group （24 h）， other administration groups （48， 72 h） except for high-dose processed P. multiflorum+Ev group] were increased significantly； high-dose processed P. multiflorum group （72 h） was significantly higher than Ev group， and low-dose processed P. multiflorum+Ev group （72 h） was significantly higher than the same-dose processed P. multiflorum group； high-dose processed P. multiflorum+Ev group （72 h） was significantly lower than the same-dose processed P. multiflorum group （P＜0.05 or P＜0.01）. Relative protein expression of ER-α in Ev group， high-dose processed P. multiflorum group and low-dose processed P. multiflorum+Ev group， relative mRNA expression of ER-α and protein expression of ER-β in administration groups， relative mRNA expression of ER-β in Ev group， low-dose processed P. multiflorum group and processed P. multiflorum+Ev groups were all increased significantly. Relative protein and mRNA expression of ER-α in Ev group were significantly higher than processed P. multiflorum groups and combination groups. Relative protein and mRNA expression of ER-β in Ev group were significantly lower than low-dose processed P. multiflorum+Ev group， but relative mRNA expression of ER-β was significantly higher than processed P. multiflorum groups and high-dose processed P. multiflorum+Ev group. Relative protein and mRNA expression of ER-α and ER-β in low-dose processed P. multiflorum+Ev group as well as relative mRNA expression of ER-β in high-dose processed P. multiflorum+Ev group were significantly higher than the same-dose processed P. multiflorum group. Relative protein and mRNA expression of ER-α in high-dose processed P. multiflorum+Ev group were significantly lower than the same-dose processed P. multiflorum group （P＜0.05 or P＜0.01）. CONCLUSIONS： The processed P. multiflorum containing serum can promote the proliferation of human breast cancer T-47D cells， and play the PE-like role through promoting protein and mRNA expression of ER-α and ER-β. However， the above effects are weaker than estrogen， and the combination of the two may antagonize the effect of estrogen.

Sequence analysis of psbA-trnH gene in chloroplast of Polygonum multiflorum from different producing areas / 中草药

Objective: The molecular identification method of Polygonum multiflorum from different producing areas was explored by using the sequencing of intergenic region of chloroplast genes psbA-trnH. Methods: A total of 116 samples of P. multiflorum were collected from 15 populations in seven provinces and autonomous regions. The total DNA was extracted and the sequences of psbA-trnH were amplified by PCR. The purified PCR products were sequenced and analyzed by MEGA 6.06 software. Results: The genetic distances among the populations of P.multiflorum are 0.001—0.187. In the maximum likelihood phylogenetic tree, 15 populations of P. multiflorum were clustered into two blanches. Conclusion: The genetic variation of P. multiflorum is significant and the psbA-trnH sequences of P. multiflorum can be used as germplasm source for molecular identification between Deqing regarded as geo-authentic habitat and other producing areas.

Optimization of High Pressure Steaming Processing Technology for Polygonum Multiflorum Thunb.by Box-Behnken Response Surface Methodology / 中国药师

Objective:To optimize the high pressure steaming processing technology for Polygonum multiflorum Thunb.by Box-Behnken response surface methodology , and compare with the traditional processing .Methods:The effects of factors such as steaming temperature, steaming time and drying temperature on polysaccharide content , stilbene glucoside content and normalized value in Po-lygonum multiflorumThunb.were studied by Box-Behnken response surface methodology .The content differences of polysaccharides and stilbene glucoside between the high pressure steaming processed product and the traditional processed product were evaluated .Re-sul ts:The best high pressure steaming processing conditions for Poyl gonumm luitlfo urm Thunb .were as follows:the steaming tempera -ture was 1254.℃ , the steaming time was3.1 h, and the drying temperature was 52℃.The contents of polysaccharides and stilbene glycosides in Polygonum multiflorum Thunb.processed by the high pressure steaming method were 1.24-fold and 5.26-fold higher than those processed by the traditional method .Conclusion:Box-Behnken response surface method can be used to optimize the high pres-sure steaming processing for Polygonum multiflorum Thunb., and the method is simple and predictable .

Transcriptional regulation effect of constituents in Polygonum multiflorum Thunb.based on human progesterone X receptor(hPXR)mediated CYP3A4 rapid screening system / 中国药理学与毒理学杂志

OBJECTIVE This study investigated transcriptional regulation of the main chemical con-stituents of Polygonum multiflorum Thunb. including Stilbene Glucoside (THSG) and anthraquinone constituents (Emodin, Rhein, Aloeemodin, Chrysophanol and Physcion) and six potential liver injury constituents(gallic acid,quercetin,luteolin,kaempferol,resveratrol)on mediated by PXR CYP3A4.Early establishment of pregnane X receptor mediated CYP3A4 drug induced rapid screening technique was used to determine the effects of these constituents. METHODS First,effect of constituents on the cell activity was detected by MTS cell viability assay. IC50was calculated. Second, the expression vector and reporter vector were co-transfected into Hep G2 cells,10 μmol·L-1Rifampicin as a positive control, 10 μmol·L-1Ketoconazole as a negative control. After treated with different concentrations of (the an-thraquinone constituents concentrations were 2.5,5 and 10 μmol·L-1;the concentrations of Gallic Acid, Quercetin,Luteolin,Kaempferol,Apigenin,Resveratrol concentrations were 5,10 and 20 μmol·L-1)for 24 h,the cells were tested for dual luciferase activity.RESULTS The results show that the inhibitory ef-fect of THSG,Chrysophanol,Emodin,Rhein and Aloeemodin on CYP3A4 was inhibited by co-transfec-tion of pcDNA3.1 and pGL4.17-CYP3A4. The expression of pcDNA3.14-PXR and pGL4.17-CYP3A4 was induced by the four constituents. Besides, Emodin has a directly inducing effect. Four anthraqui-none constituentscan induce the effect of CYP3A4 by PXR, but Emodin can directly induce CYP3A4. THSG can inhibit CYP3A4,but in the presence of PXR plasmid can induce CYP3A4.For the six poten-tial liver injury constituents, results show that the plasmid pcDNA3.1 was cotransfected with pGL4.17-CYP3A4 regulation of Gallic Acid and Resveratrol on CYP3A4 inhibitory effects of Quercetin,Luteolin, Kaempferol have an induce effect; after pcDNA3.14-PXR and pGL4.17-CYP3A4 cotransfected, Quercetin, Luteolin, Kaempferol, Apigenin, Resveratrol have induced effect, three constituents'induc-tion effect had significant difference.CONCLUSION 12 kinds of Polygonum multiflorum Thunb.constit-uents have inhibitory or activating effects on CYP3A4, after the participation of PXR, 9 components have induced effects on CYP3A4, and the induction effect of 6 components has significant difference. The results suggested that we should pay attention to potential drug interactions when combined with Polygonum multiflorum Thunb.,and improve safety and efficacy.

Effects of Processing Methods and Extraction Solvents on the Contents of Major Components in Polygonum multiflorum / 中国药房

OBJECTIVE:To study the effects of different processing methods and extraction solvents on the contents of major components in Polygonum multiflorum. METHODS:Decoction of black soybean and water were used to steam the raw P.multiflorum. Water,50％ ethanol,70％ ethanol and 90％ ethanol were used to extract the raw,black soybean steamed,water steamed and commercial processed P. multiflorum respectively. HPLC method was used to detect the contents of gallic acid,2,3,5, 4′-tetrahydroxystilbene-2-O-β-D-glucoside (THSG),emodin and physcion. RESULTS:The contents of 4 major components in 4 kinds of extracts from 3 kinds of processed P. multiflorum were higher than raw sample;the content of gallic acid extracted with water was the highest;the content of THSG extracted with 90％ ethanol was the lowest;the contents of emodin and physcione extracted with 50％ and 70％ ethanol were the higher. CONCLUSIONS:Different processing methods and different extraction solvents had effect on the contents of the main compounds of P. multiflorum. The contents of each components in the processing products didn't show certain regularity.