1.Experimental Study of Anti-inflammation of Durian Peel Extract
Guo XIE ; Minzhi WU ; Jinle CHENG ; Ruoting ZHAN ; Weiwen CHEN
Journal of Guangzhou University of Traditional Chinese Medicine 2015;(1):130-135
Objective To investigate the anti-inflammatory effects and mechanism of durian peel extracts (DPE). Methods The in vivo anti-inflammation effects of DPE were examined by carrageenin-induced mice paw edema test and allergic contact dermatitis test induced by 2, 4-DNFB. And the in vitro anti-inflammation effects of DPE were evaluated with methyl thiazolyl tetrazolium ( MTT) assay in RAW 264.7 cell model of inflammation induced by lipopolysaccharide (LPS). Results The results of animal experiments showed that DPE groups could markedly relieve mice paw edema induced by carrageenin ( P<0.01 or P<0.001 compared with blank group). DPE could effectively inhibit the allergic contact dermatitis induced by 2, 4-DNFB in mice, showing good dose-effect relationship. The results of in-vitro test showed that DPE at the given concentrations had no influence on RAW 264.7 cell proliferation. Tumor necrosis factor alpha ( TNF-α) , interleukin 6 ( IL-6) , interleukin 1 beta (IL-1β), nitric oxide (NO) and nuclear factor kappaB (NF-кВ ) were observably inhibited, and anti-inflammatory cytokine IL-10 was enhanced by 25 and 50 mg/L of DPE. Conclusion DPE exert potential anti-inflammation effect, and the mechanism might be related to its inhibition of NF-кВsignal pathway.
2.In-vitro Antibacterial Activity of Herba Taraxaci Extract on Escherichia coli
Xiaoyu JI ; Yuanxia PENG ; Min LIU ; Hui XU ; Ruoting ZHAN
Journal of Guangzhou University of Traditional Chinese Medicine 2015;(1):116-120
Objective To evaluate the inhibitory activity of Herba Taraxaci extract on Escherichia coli DH5α (E. coli DH5α) and to investigate proteomic response of E. coli. Methods Medicinal powder of Herba Taraxaci was extracted with the solvents of different polarity ( n-hexane, ethyl acetate, distilled water) , and then the obtained 8 different extracts were subjected to thin layer chromatography ( TLC) analysis. Microdilution method was performed to detect the minimum inhibitory concentration ( MIC) of different extracts and the growth curves were described. The protein expression profiles of E . coli treated with the extracts were analyzed by sodium dodecyl sulfate polyacrylamide gel electropheresis ( SDS-PAGE) and two dimensional electrophoresis (2-DE) . Results Water decoction of Herba Taraxaci could obviously suppress the growth of E. coli with a MIC of 1.95 mg/mL. The different extractions exhibited no antibacterial activity except ethyl acetate phase 3 with a MIC of 0.13 mg/mL, which was equal to 19.23 mg/mL of crude drugs. The results of TLC analysis showed that chlorogenic acid was undetectable in n-hexane extract and ethyl acetate phase 1 extract, and ethyl acetate phase 2 and 3 extracts showed obviously increased spots. The results of SDS-PAGE and 2-DE showed that water decoction of Herba Taraxaci had inhibitory effect on the expression of functional protein. The results of 2-DE showed that after treatment with ethyl acetate phase 3 at the concentration of 2 × MIC for 21 hours, the amount of protein spots were 92 less than those of the blank control group, the spots of E. coli DH5α soluble protein with expression amount down-regulated doubly were 24, and those with expression amount up-regulated doubly were 19. Ethyl acetate phase 3 extract had an effect on down-regulating the protein expression of E. coli DH5α soluble protein pH3-10, and water decoction of Herba Taraxaci had inhibitory effect on E. coli DH5αprotein expression. Conclusion Herba Taraxaci has significant antibacterial activity on E. coli DH5α, and the water-soluble fraction of chlorogenic acid and caffeic acid might be the active components. The possible antibacterial mechanism may be related with the regulation of bacterial protein expression.
3.Study on Callus Induction in Explant of Aquiliaria sinensis Root
Yanfengyang JIANG ; Chao LIN ; Ruoting ZHAN ; Xinye MA
Journal of Guangzhou University of Traditional Chinese Medicine 2015;(4):751-754
Objective To study the conditions of callus induction with the roots of Aquilaria sinensis as explants. Methods Two sources of roots of Aquiliaria sinensis were selected as the explants. The effects of sterilization methods and the combination of different concentrations of phytohormones on callus induction were evaluated. Results When Aquiliaria sinensis root seedling was sterilized in 0.01mg/mL HgCl2 solution for 3 minutes, the sterilized effect was the best. The optimal callus induction medium was MS+0.1 mg/L 2,4-dichlorophenoxyacetic acid (2,4-D) +0.1 mg/L 6-benzylaminopurine (6-BA). Aseptic Aquiliaria sinensis root seedling cultivated in callus induction medium containing MS+1.0 mg/L naphthalene acetic acid ( NAA) +0.8 mg/L 6-BA achieved the highest callus induction rate. Conclusion Callus can be induced from two sources of Aquilaria sinensis roots. The induction rate of callus is lower when the explant root seedling is cultivated using 2,4-D alone as inducer, and is increased when used together with 6-BA.
4.Identification of Original Plants of Guanzhong (Dryopteris Species) by Chloroplast psbA-trnH Intergenic Region
Feng LIU ; Yanfengyang JIANG ; Ruoting ZHAN ; Xinye MA ; Weiwen CHEN
World Science and Technology-Modernization of Traditional Chinese Medicine 2014;(7):1482-1486
This article was aimed to study the species identification capability of psbA-trnH among Guanzhong herbs from Dryopteri. The nucleotide sequence information of psbA-trnH region was abstracted using standardized manners from 9 Dryopteri species (19 samples). The identification efficiency of psbA-trnH was analyzed based on TaxonGap among both tested materials (9 species, 19 samples) and tested materials plus GenBank data (17 species, 44 samples in total). The results showed that with the expanded species range, the discriminative efficiency of psbA-trnH de-clined from 100% (9/9) to 82.4% (14/17), while the proportion of within-specific heterogeneity larger than between-specific separability increased from 11.1% (1/9) to 52.9% (9/17). It was concluded that psbA-trnH can be recom-mended as the valuable barcode for homonym distinguishment among Guanzhong herbs, with attention of adequate sampling within species.
5.Transcriptomic Analysis and Systematic Mining of Genes Involved in Biosynthetic Pathway of Triterpenoid Saponins in Ilex Asprella
Xiasheng ZHENG ; Xiuxiu LUO ; Hui XU ; Ruoting ZHAN ; Weiwen CHEN
World Science and Technology-Modernization of Traditional Chinese Medicine 2014;(7):1505-1512
This study was aimed to screen candidate genes involved in the triterpenoid saponins biosynthetic pathway of the Ilex asprella root. The Illumina platform was applied to perform transcriptomic sequencing of I. asprella root, followed by a series of bioinformatics analysis. The results showed that a total of 272 candidate unigenes were anno-tated to be involved in the biosynthetic pathway of terpenoid in the transcriptome of I. asprella root, including 72 u-nigenes for the upstream pathway and 26 unigenes for cyclization, oxidation and glycosylation in the downstream pathway. Phylogenetic analysis was carried out to further analyze the evolution relationship of some candidate uni-genes and their homologous genes. Two genes IaA S1 and IaA S2 were proved to be mixed amyrin synthases in yeast expression system. Moreover, IaA S1 was identified to one of the rare ASs with α-amyrin as the major product. It was concluded that a series of candidate genes, which might be involved in the biosynthetic pathway of triterpenoid saponins, were screened out from the transcriptome of I. asprella root. Further investigation of these candidate genes will provide insight into their actual functions in the triterpenoid saponins biosynthetic pathway in I. asprella.
6.Analysis of Fingerprints and Formononetin Content of Caulis Spatholobi from Different Habitats by High Performance Liquid Chromatography
Hongying CHEN ; Ping YAN ; Min ZHANG ; Cuiting LIU ; Zhengzhou HAN ; Ruoting ZHAN
Journal of Guangzhou University of Traditional Chinese Medicine 2015;(5):923-928,936
Objective To establish the fingerprints and formononetin content determination method for Caulis Spatholobi from different habitats by high performance liquid chromatography ( HPLC) , thus to control the quality of Caulis Spatholobi. Methods Reversed phase-high performance liquid chromatography (RP-HPLC) for fingerprint was performed on Feini Gen RedClassical AQ-C18 column ( 4.6 mm × 250 mm, 5 μm) with acetonitrile-0.1%acetic acid solution as the mobile phase by gradient elution, and the detection wavelength was 260 nm. High performance liquid chromatography-diode array detector ( HPLC-DAD) for the determination of formononetin content was performed on AcclaimTM 120-C18 column ( 4.6 mm × 250 mm, 5 μm) with acetonitrile-water solution by isocratic elution, the detection wavelength was 254 nm, the flow rate was 1.0 mL/min and the column temperature was 25℃. Results The standard fingerprint of Caulis Spatholobi was set up through the evaluation of the fingerprints of 24 batches of Caulis Spatholobi samples from different habitats. Thirteen common peaks were identified with reference to formononetin peak, and the content of formononetin was determined by HPLC-DAD method. The similarity of the fingerprints of Caulis Spatholobi from different habitats and their formononetin content had great differences. Conclusion The established method is simple, accurate, highly sensitive, and repeatable, and can be applied for the quality control of Caulis Spatholobi.
7.Diagnosis and Comprehensive Evaluation of Soil Fertility of Good Agricultural Practices Base for Spatholobus suberectus Dunn. in Pingyuan of Guangdong Province
Jincai LU ; Zhengzhou HAN ; Zhengjun WU ; Zhiming LAI ; Rui HE ; Shourong ZHANG ; Ruoting ZHAN
Journal of Guangzhou University of Traditional Chinese Medicine 2015;(1):141-146
Objective To investigate the status of soil fertility of Good Agricultural Practices ( GAP) base for Spatholobus suberectus Dunn. (SSD) in Pingyuan county of Guangdong province, thus to provide reference for GAP research and the subsequent fertilization for SSD. Methods The deep layer and superficial layer of GAP soil were collected for the physiochemical detection and nutrient assay. Compared with the classification standard of the second national general soil investigation, single base soil fertility index was diagnosed and the comprehensive soil fertility was evaluated with modified Nemoro Index. Results The soil pH value and the contents of exchangeable calcium and magnesium were unbalanced, and the contents of macroelements of nitrogen and phosphonium, microelements, and organic matter were low. Therefore, the measures for improving the base soil fertility should be as follows: ( 1) soil amendments of bentonite, gypsum and slaked lime should be used to adjust the soil pH value; ( 2) each plant should be given 10 kg of slaked organic fertilizer as base fertilizer; ( 3) in the process of nurturing, some special micro-fertilizer solution should be used to treat the cut slips, and 5 kg of urea should be used for every 667 meter square of land; ( 4) besides compound fertilizer, every 667 meter square of land should be fertilized with 15 kg of ammonium dihydrogen phosphate for the supplement of nitrogen and phosphorus, and slaked lime and magnesium carbonate should be used for the supplement of soil moderate-quantity elements after transplantation. Conclusion The comprehensive fertility of Pingyuan GAP base for Spatholobus suberectus Dunn. is at low level, and should be improved in combination with GAP requirements.
8.Screening of Reference Genes for Real-time Fluorescence Quantitative PCR in Amomum villosum Lour
Anmin YU ; Huan WANG ; Xueying HE ; Ke DENG ; Ruoting ZHAN ; Jinfen YANG
Journal of Guangzhou University of Traditional Chinese Medicine 2014;(5):814-820
Objective To identify the reliable reference genes for gene expression analysis of the pericarp and seed of Amomum villosum Lour. by using real-time fluorescence quantitative polymerase chain reaction ( qRT-PCR). Methods Using the fruits ( separated into peels and seeds) of A. villosum at three different developmental periods as the experimental material, 5 candidate reference genes (β-actin, EF-1α, GAPDH, PGK, TUA) with steady expression were screened out by the high throughout sequencing of transcriptome and expression profile data. The qRT-PCR technique was applied to study the expression levels of 5 candidate reference genes in different samples. The stability of the candidate reference genes were evaluated by GeNorm and NormFinder software. Results The 5 reference genes had different stabilities in the pericarp and seed of A. villosum Lour. at different development periods . The order of the steadiness of reference genes showed by GeNorm was EF-1α = TUA>PGK>GAPDH>β-actin. The results of NormFinder revealed that EF-1α was the most stable, followed by TUA, and the order of the other three genes was as same as the results of GeNorm. Conclusion EF-1αand TUA could be used as double reference genes for the normalization of gene expression in A. villosum fruits at different developmental periods by using qRT-PCR.
9.The empirical research on cultivation mechanism of manufacture-learning-research cooperation for postgraduates of science of Chinese Pharmacology of Guangzhou University of Chinese Medicine
Huafeng PAN ; Sichen WU ; Xin GE ; Jinling REN ; Yichong FENG ; Ruoting ZHAN
Chinese Journal of Medical Education Research 2011;10(10):1176-1178
The paper summarized the successful experience such as preliminarily establishing cultivation mechanism of manufacture-learning-research cooperation for postgraduates of science of Chinese pharmacology of Guangzhou University of Chinese Medicine,ensuring innovation ability training for postgraduates,getting reasonable configuration for social education resources,and put forward existing problems in traditional Chinese medicine system,such as the loose management and faulty evalution system.Suggestion is to futher perfect relevant rules and regulations,clear and definite responsibility of school and enterprise,reserch regularly to impove ideological education of postgraduates,etc.In the way,we can achieve the aim of promoting high-level talents of traditional Chinese medicine industry.
10.Dynamic Study on Soluble Sugar Accumulation and Sucrose Metabolizing Enzyme Activities During Fruit Development of Amomum villosum Lour
Anmin YU ; Min ZHANG ; Jinfen YANG ; Wenhua LIN ; Ping YAN ; Jinpei LIANG ; Ruoting ZHAN
World Science and Technology-Modernization of Traditional Chinese Medicine 2014;(7):1497-1504
This study was aimed to discuss the dynamic variation of soluble sugar contents, sucrose metabolizing en-zyme activities and gene expression quantities during the fruits development of A momum villosum, in order to pro-vide the basis of improvement of the fruit yield. Fresh fruits at three different development processes (30 DAF, 60 DAF, 90 DAF) were used to investigate changes of soluble sugar components and sucrose metabolizing enzyme activ-ities by HPLC and UV spectrophotometry. Combining with the high-throughput sequencing expression profile data of three fruit development period, the trends of three key enzymes gene expressed in sugar metabolism were analyzed. The results showed that the fruit sugar components were dominated by fructose, glucose and sucrose. The concentra-tion of hexose (fructose and glucose) gradually decreased in peel. But in seeds the concentration of hexose decreased at first and then increased. The content of sucrose and the net activities of sucrose synthase (synthesizing direction minus decomposing direction) in peel and seeds were gradually increased. The expression trends of key enzyme gene in sugar metabolism examined by RNA-seq quantification showed that sucrose phosphate synthase and sucrose syn-thase gene increased and then kept constant, but the invertase gene expression trend was gradually rising. Conse-quently, sucrose synthase was the key enzyme catalyzing sucrose synthesis and decomposition. The activity of sucrose synthase and sucrose contents in peel and seeds reached the highest peak in the end of fruit mature.