ABSTRACT
A new phloroglucinol was isolated from 50% ethanol extract of Dryopteris fragrans by silica gel column chromatography, Sephadex LH-20 gel column chromatography, thin-layer chromatography(TLC), and preparative liquid column chromatography. On the basis of MS, ~1H-NMR, ~(13)C-NMR, and reference materials, compound 1 was identified as 2,5-cyclohexadien-1-one, 2-{[2,6-dihydroxy-4-methoxy-3-methyl-5-(1-isobutyl)phenyl]methyl}-3,5-dihydroxy-4,4-dimethyl-6-(1-oxobutyl)(1), and named disaspidin BB. Compound 1 was evaluated for its antibacterial activity. The experimental results showed that compared with the commonly used topical antibiotics erythromycin or mupirocin, disaspidin BB exhibited significant antibacterial activities against Staphylococcus epidermidis(SEP), S. haemolyticus(SHA), and methicillin-resistant S. aureus(MRSA)(P<0.05). Additionally, disaspidin BB was sensitive to ceftazidime-resistant SEP1-SEP4, SHA5-SHA7, MRSA8, and MRSA9. The MIC values of disaspidin BB against SEP and SHA were 1.67-2.71 μg·mL~(-1) and 10.00-33.33 μg·mL~(-1) respectively. Disaspidin BB has good antibacterial activities and deserves development as a new anti-infective drug for external use.
Subject(s)
Anti-Bacterial Agents/pharmacology , Dryopteris , Methicillin-Resistant Staphylococcus aureus , Microbial Sensitivity Tests , Phloroglucinol/pharmacology , Plant Extracts/pharmacologyABSTRACT
Two phloroglucinol compounds(1-2) were isolated and purified from 95% ethanol extract of Dryopteris fragrans through various column chromatographies on silica gel, Sephadex LH-20, medium pressure column chromatography, and preparative HPLC. Their structures were elucidated as 2',4',6'-trihydroxy-5'-methyl acetate-3'-methyl-1'-butyrophenone(1) and aspidinol B(2) based on their chemical and physicochemical methods and spectroscopic data. Compound 1 is a new phloroglucinol compound named "dryofraginol".
Subject(s)
Chromatography, High Pressure Liquid , Dryopteris , Ethanol , Phloroglucinol , Plant ExtractsABSTRACT
Objective: To establish quantitative analysis of multi-components with single marker (QAMS) for determination of 10 phloroglucinol contents in effective fraction of Dryopteris fragrans. Methods: The relative correction factors of nine phloroglucinol (aspidin PB, aspidin AB, flavaspidic acid BB, saroaspidin A, flavaspidic acid PB, disflavapidic acid PB, flavaspidic acid AB, compound VI, and aspidinol B) were determined by HPLC method with the aspidinn BB as the internal standard, which were to calculate the content of each. At the same time, external standard method (ESM) was used to determine the contents of 10 components in effective fraction, and the differences between the two methods were compared to verify the feasibility and accuracy of QAMS method. Results: The relative correction factor (RCF) was good. There was no significant difference between the quantitative results with the two methods in the 12 batches of 10 phloroglucinols. Conclusion: The established QAMS method can be used for quantitative analysis of D. fragrans with aspidinn BB as the internal standard in the absence of reference substances, and provided a reference for the multi-index quality evaluation in effective fraction of D. fragrans.
ABSTRACT
Objectives Aspidin BB, a typical phloroglucinol derivative from Dryopteris fragrans, possesses significant antifungal property. This study aimed to investigate potential mechanism of antifungal activity of Aspidin BB against Trichophyton rubrum which is the most common pathogens responsible for chronic dermatophytosis. Methods The minimum inhibitory concentration (MIC) of Aspidin BB against strains was determined by broth microdilution. The effects of Aspidin BB on ergosterol biosynthesis were investigated by content determination based on UPLC method. Besides, the effects of drugs on squalene epoxidase (SE) in T. rubrum cell membrane were analyzed. Results MIC value of Aspidin BB against T. rubrum was 25.0 μg/mL. Aspidin BB reduced ergosterol content significantly, but no notable effect on squalene epoxidase activity. Conclusion The results suggested that Aspidin BB inhibited ergosterol biosynthesis. However, it was not squalene epoxidase but other components may sever as possible targets in ergosterol biosynthesis pathway.
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Objective: To study the chemical constituents and antifungal activitiy of phloroglucinol derivatives from Dryopteris fragrans. Methods: The phloroglucinol derivatives were isolated by silica gel column chromatography, Sephadex LH-20 column chromatography, and pre-HPLC. The chemical structures were elucidated on the basis of physicochemical and spectral properties. The antifungal activities against Trichophyton rubrum and Microsporum gypseum were tested by CLSI M38-A2 broth microdilution methods. Results: Two phloroglucinol derivatives were isolated and identified as disflavaspidic acid PB (1) and flavaspidic acid PB (2). The minimum inhibitory concentration (MIC) of compounds 1 and 2 were 13.33 and 1.875 μg/mL against T. rubrum respectively, 10 and 1.325 μg/mL against M. gypseum. Conclusion: Compound 1 is a new compound named disflavaspidic acid PB. Compound 2 is isolated from D. fragrans for the first time. Compounds 1 and 2 are a pair of isomers with promising antifungal activity.
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Objective: Expression and analysis of 4CL gene of ferns, which lays the theoretical foundation for the accumulation and regulation of secondary metabolites of ferns. Methods: Cloned 4CL gene (Df4CL) sequence from Dryopteris fragrans was analyzed by CHIPS, CUSP, and Codon W programs online, and then compared with Escherichia coli, yeast, Arabidopsis thaliana, and Nicotiana tabacum L. It is important to identify the codon usage of Df4CL gene and select appropriate expression systems. Results: Df4CL gene had not the bias toward the synonymous codons with ATGC at the third codon position, but had the preference consistency with Physcomitrella patens codon usage. The differences in codon usage frequency between Df4CL gene and A. thaliana were less than those in N. tobacum, E. coli, and yeast. Based on 4CL gene codon usage bias of cluster analysis, it was shown that D. fragrans, P. patens, and A. thaliana were classed into one group. Conclusion: The A. thaliana expression system may be more suitable for the exogenous expression of Df4CL gene of D. fragrans.
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Objective:To isolate and prepare the reference substances of phloroglucinol derivatives from Dryopteris fragrans by semi-preparation HPLC. Methods:After reflux extraction of Dryopteris fragrans with petroleum ether,the extracting solution was con-centrated and separated by silica gel column chromatography,and then isolated by semi-preparation HPLC. The isocratic elution was carried out using acetonitrile as the mobile phase at 3. 0 ml·min-1 and the injection volume was 0. 5 ml. Two phloroglucinol deriva-tives were isolated. Results:The chemical structure of the two phloroglucinol derivatives respectively was aspidin BB with the purity of 98. 81% and aspidin PB with the purity of 98. 57% by ultra high performance liquid chromatography. Conclusion:The isolation of as-pidin BB and aspidin PB by semi-preparation HPLC is simple and fast with the purity over 98%,which can be used to prepare the ref-erence substances.
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Objective: To study the chemical constituents from the whole plants of Dryopteris fragrans. Methods: The chemical constituents were isolated by various column chromatographic methods. The structures of the compounds were elucidated on the basis of physiochemical properties and spectral analyses. Results: Three compounds named 2-isopropyl-5, 7-dihydroxy-1'-O-β-D-glucopyranosyl chromone (1), (6R, 9R)-3-oxo-α-ionol-9-O-β-D-glucopyranoside (2), and taxicatin (3) were isolated and identified. Conclusion: Compound 1 is a new compound named frachromone B, compounds 2 and 3 are reported for the first time from family Dryopteridaceae.
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Objective: To study the chemical constituents from the whole plants of Dryopteris fragrans. Methods: The chemical constituents were isolated by various column chromatographic methods. The structures of the compounds were elucidated on the basis of physiochemical properties and spectral analysis. Results: Four compounds named 2-ethyl-5, 7-dihydroxy-1'-O-β-Dglucopyranosyl chromone (1), vitexin (2), isoquercitrin (3), and divarin-3-O-β-glucopyranoside (3-hydroxy-5-propylphenyl-O-β-Dglucopyranoside, 4) were isolated and identified. Conclusion: Compound 1 is a new compound named as frachromone A, compound 2 is obtained from this plant for the first time, and compound 4 is reported for the first time from the plants of family Dryopteridaceae.
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Objective To determine the genetic diversity of germplasm resource in Dryopteris fragrans by amplified fragment length polymorphism(AFLP) technique.Methods Forty-six samples from six populations of D.fragrans were analyzed by AFLP DNA markers,and the genetic diversity was evaluated by NTSYS-PC 2.10 and PopGen32.Results The percentage of polymorphic bands(PPB) reached to 76.67%,observed number of alleles(Na) was 1.766 7, effective number of alleles(Ne) was 1.504 6,Nei′s gene diversity index(H) was 0.292 1,Shannon information index(I) was 0.431 6,and genetic differentiation index(Gst) was 0.412 5.Conclusion Genetic diversity is high,gene diversity in populations(Hs) is 0.170 5,and gene diversity among populations(Dst) is 0.119 8.Because of environmental specificity,its resource on the spot should be protected.
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Objective To study the chemical components from Dryopteris fragrans and their biological activities.Methods By means of HPLC and other column chromatography,the chemical components were isolated from D.fragrans and their chemical structures were identified by spectral and chemical tests in accordance with their physicochemical characteristics.Furthermore,the cytotoxicity of the chemical components for Bel-7402,HCT-8,and A-549 cells was measured by MTT method.Results Four compounds,geniposide(Ⅰ),3S,5R,6R,7E,9S-megastigman-7-ene-3,5,6,9-tetrol-3-O-?-D-glucopyranoside(Ⅱ),(6S,9R)-3-oxo-?-ionol-9-O-?-D-glucopyranoside(Ⅲ),and fragranoside a(Ⅳ) were iso-lated and identified.Conclusion Compound Ⅳ is a new compound.And all the other three compounds Ⅰ-Ⅲ are found in plants of Dryopteris Adanson for the first time.The results of pharmacological experiments indicate that compounds Ⅰ and Ⅲ show strong cytotoxicity.