The mechanism of Belamcanda chinensis in the treatment of glioma based on network pharmacology and molecular simulation / 西安交通大学学报(医学版)

【Objective】 To explore the potential molecular biological mechanism of Belamcanda chinensis in the treatment of glioma based on network pharmacology, molecular docking technology and in vitro cell experiments. 【Methods】 ① The active components, targets of Belamcanda chinensis and targets of glioma were obtained by database search. String database was used to analyze protein-protein interaction relationship, R project was used to analyze gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway, Cytoscape software was used to build "compound-target-disease" network and PPI network, and AutoDock software was used to verify molecular docking. ② Western blotting, qRT-PCT and apoptosis assay were used to verify the enrichment results of network pharmacology targets and protein pathway. 【Results】 ① We screened out 32 types of active components, 484 types of targets and 464 types of glioma targets, and obtained 62 kinds of therapeutic targets after mapping. We obtained 12 kinds of key pharmacodynamic molecules such as Isoiridogermanal, Iridobelamal A and Rhamnazinand and other key pharmacodynamic molecules, as well as AKT1, STAT3, HRAS and other core targets by network topology analysis. Enrichment analysis results demonstrated that they were mainly involved in biological processes such as peptide serine phosphorylation, protein kinase B signal transduction, peptide serine modification, and pathways including PI3K/AKT signal pathway and Rap1 signal pathway. The results of molecular docking verified the good binding activity of the key pharmacodynamic molecules with the core targets. ② The results of Western blotting showed that the protein expressions of VEGF and MMP9 of Belamcanda chinensis extracts in 8 mg/mL and 16 mg/mL groups were significantly lower than those in the blank control group (P<0.01 or P<0.001). Compared with the blank control group, the early apoptosis rate of Belamcanda chinensis extracts at 8 mg/mL and 16 mg/mL were significantly decreased (P<0.001 or P<0.000 1). qRT-PCR results showed that the mRNA expression levels of VEGF and MMP9 in Belamcanda chinensis extracts at 8 mg/mL and 16 mg/mL were significantly decreased (P<0.001 or P<0.0001). 【Conclusion】 The treatment of glioma with Belamcanda chinensis is the result of multi-component, multi-target and multi-channel interactions. The results of cell experiments confirmed that Belamcanda chinensis extracts can affect the expressions of related target proteins of PI3K/AKT signal pathway and VEGF and MMP9, which verified the results of network pharmacology. The results provide a theoretical basis for the clinical application of Belamcanda chinensis and studies on glioma.

Hypolipidemic properties of the extracts of Belamcanda chinensis leaves (BCLE) in KK-A y mice

Abstract The extract of Belamcanda chinensis leaves (BCLE) is a traditional Chinese herbal medicine for the treatment of diabetes-related hyperlipidemia in Hainan province, South China. In this study, the lipid-decreasing effects of BCLE on obese diabetes were investigated on KK-Ay mice. The component F2 ameliorated lipid disorder, as indicated by decreased levels of body weight, liver index, levels of TC, TG and LDL-c in the serum and liver. The enhancement effect of F2 on liver SOD and the inhibitory effect of F2 on MDA demonstrated that F2 exhibited significant antioxidant activity on liver injury. F2 also prevented vacuolar degeneration and reduced pathological tissue injury in liver. In addition, the component F1 decreased the levels of TG, LDL-c and MDA in the liver. These findings suggest that F2 may have therapeutic potential in the prevention and therapy of hyperlipemia and liver disease associated with obesity-related diabetes.

Identification and biological characteristics of southern blight causing root rot on three medicine plants of Iridaceae in Dabie Mountains / 中国中药杂志

In order to identify the species and biological characteristics of the pathogen of southern blight from three kinds of Chinese medicine of Iridaceae(Belamcanda chinensis, Iris tectorum and I. japonica) in Dabie Mountains, the isolation, identification, pathogenicity and biological characteristics of the pathogens were studied according to Koch's postulates. In addition, 9 chemical fungicides, 3 botanical fungicides and 5 microbial fungicides were used to evaluate their inhibition to the isolates in vitro. The results showed that all the strains(SG-Q, YW-Q, and HDH-Q) isolated and purified from the diseased plants of B. chinensis, I. tectorum and I. japonica, respectively, were identified as Sclerotium rolfsii through morphological observation and sequence aligement of 18 S rDNA, rDNA-ITS and TEF. Field observations showed that the intensity of the disease incidence of three Iridaceae plants was B. chinensis>I. japonica> I. tectorum, and the pathogenicity of the strains was SG-Q>YW-Q>HDH-Q. For biological characteristics, SG-Q strain was suitable for growth under the 12 h light/12 h dark cycle, with the optimal growth temperature of 30 ℃ and pH of 5. Among the 9 tested chemical fungicides, 29% lime sulphure and 10% flusilazole had stronger inhibitory effect on mycelia growth of SG-Q. For 3 botanical fungicides, 1% osthol, 20% eugenol and 0.5% berberine could effectively inhibt the mycelial growth of SG-Q and cause the morphological variation of the pathogen. For 5 microbial fungicides, Trichoderma harzianum and Bacillus subtilis had better inhibition on the mycelium growth of SG-Q.

Analysis on Seed Quality Grading Standard of Belamcanda chinensis / 中国实验方剂学杂志

Objective:To study the quality inspection standard of seeds of <italic>Belamcanda chinensis</italic> and establish the quality grading standard of seeds of <italic>B. chinensis</italic>. Method:The purity of seeds of <italic>B. chinensis</italic> from different producing areas was analyzed，and the 1 000-grain weight，water content，viability and germination rate of seeds of <italic>B. chinensis</italic> with different diameters were determined after diameter classification. K-means was used for cluster analysis to preliminarily formulate the quality grading standard of seeds of <italic>B. chinensis</italic>. Result:There were obvious regional differences in the size of seeds of <italic>B. chinensis</italic>. The particle size of seeds of <italic>B. chinensis</italic> of Xiaochang，Dawu and Huanggang Academy of Agricultural Sciences in Hubei Province was larger and mostly between 4.5 mm to 5.5 mm. The seeds of<italic> B. chinensis </italic>from Tuanfeng，Qujing，Yunnan，and Anguo，Hebei province had smaller particle sizes than those from the other three producing areas，which were mostly between 3.5 mm to 5.0 mm. Arils in seeds of <italic>B. chinensis</italic> were removed by 10% NaClO to effectively break seed dormancy and significantly improve the seed germination rate. On this basis，the seed diameter was taken as the main grading index，and the seed moisture content，viability，germination rate and purity were taken as important reference indexes. The quality of seeds of <italic>B. chinensis</italic> was preliminarily divided into three grades，grade Ⅰ：seed diameter≥5.5 mm，moisture content≤10%，viability≥94%，germination rate≥60%，and cleanliness≥93%；grade Ⅱ：seed diameter≥4.5 mm，moisture content≤10%，viability≥90%，germination rate≥55%，and cleanliness≥85%；and grade Ⅲ：seed diameter≥3.5 mm，moisture content≤10%，viability≥84%，germination rate≥45%，cleanliness≥80%. Conclusion:In this study，the quality grading standard of seeds of <italic>B. chinensis</italic> was preliminarily established to provide reference for the quality evaluation of seeds of <italic>B. chinensis</italic> and the breeding of improved varieties. In addition，the maturity and the storage time of seeds of <italic>B. chinensis</italic> have a greater impact on the quality of seeds，so it is recommended to select fully mature（dark black） seeds and new seeds for production.

Content Determination of 10 Isoflavones in Belamcanda chinensis by UPLC / 中国药房

OBJECTIVE： To establish a method for simultaneous determination of 10 isoflavones in Belamcanda chinensis， and to evaluate the differences of active ingredient content of B. chinensis from different areas. METHODS： UPLC method was adopted. The determination was performed on Waters ACQUITY UPLC BEH C18 column with mobile phase consisted of 0.5 ％ methyl-β-cyclodextrin and 0.1％ phosphate as water phase， acetonitrile as organic phase （gradient elution） at the flow rate of 0.2 mL/min. The column temperature was set at 35 ℃， and the detection wavelength was set at 265 nm. The sample size was 2 μL， and analysis time was 20 min. The contents of 10 isoflavones in 26 samples from 8 provinces， including tectoridin， iristectorin A， iristectorin B， iridin， tectorigenin， iristectorigenin B， iristectorigenin A， irigenin， irisflorentin， dichotomitin， were determined. RESULTS： The linear ranges of tectoridin， iristectorin A， iristectorin B， iridin， tectorigenin， iristectorigenin B， iristectorigenin A， irigenin， irisflorentin， dichotomitin were 8.569 5-342.78， 0.643-25.72， 1.119 8-44.79， 2.187 8-87.51， 0.770 3-30.81， 0.421 3- 16.85， 0.288 5-11.54， 1.795 3-71.81， 0.560 8-22.43， 0.086-3.44 μg/mL（all r≥0.999 6）. The limits of quantitation were 0.015， 0.102， 0.096， 0.013， 0.036， 0.088， 0.102， 0.019， 0.067， 0.092 μg/mL. RSDs of precision， stability（24 h）and reproducibility tests were lower than 2.00％ （n＝6）. The recoveries ranged 95.30％-103.30％ （all RSD≤2.33％， n＝6）. Among 26 samples of B. chinensis， the content of tectoridin was the highest （3.66％-57.79％）， and the content of dichotomitin was the lowest （0.09％- 0.59％）， the contents of irisflorentin were 0.29-2.80 mg/g. The contents of isoflavones in B. chinensis from different areas were different greatly.  CONCLUSIONS： The established method is sensitive， with short analysis time and good repeatability， and can be used to determine the content of 10 isoflavones and evaluate the content difference of each component.

Genetic diversity of endophytic fungi strains Fusarium proliferatum isolated from Belamcanda chinensis using ISSR and SRAP markers / 中草药

Objective: In order to understand the genetic diversity of endophytic fungi strain Fusarium proliferatum isolated from Belamcanda chinensis. Methods: A total of 52 ISSR primers and 90 SRAP primers were used to detect the genetic diversity among 17 F. proliferatum strains. Results: The results indicated that 27 ISSR primers and 38 SRAP primers were screened out for the genetic diversity analysis. 178 bands were amplified from 27 ISSR primers, among which 131 (63%) allelic variations were detected. However, 357 bands were amplified by 38 SRAP primers, among which 323 (91%) allelic variations were detected. The value of allelic polymorphism information content (PIC) of ISSR primers ranged from 0.19 to 0.91, with the average of 0.70 per primer. The value of PIC of SRAP primers ranged from 0.00 to 0.93, with the average of 0.72 per primer. The value of Nei's genetic similarity (GS) indexes of 17 strains based on ISSR, SRAP and ISSR + SRAP genetic locus varied from 0.73-0.99, 0.72-0.95 and 0.73-0.95, and with the average of 0.84, 0.85 and 0.85, separately. Cluster analysis showed that the 17 strains in this study could be clustered into three groups, three strains from the roots were clustered together, and F. proliferatum strains isolated from stems and leaves were gathered in other two groups. Cluster analysis revealed that genetic similarity of 17 strains were high, this suggested that the 17 strains had a near relationship, in accordance with the traditional morphology identification. Conclusion: The results show that the ISSR and SRAP technology is more efficient than traditional morphology identification. It is also found that ISSR and SRAP markers could more really reflect the genetic diversity of endophytic fungi strain F. proliferatum from B. chinensis, which can provide the basis for the application of molecular biotechnology in endophytic fungi of F. proliferatum from B. chinensis.

Determination of six active ingredients in different parts of Belamcanda chinensis and Iris tectorum and their anti-inflammatory activity / 中国中药杂志

In order to explore the anti-inflammatory activity and active ingredient basis from the leaves of the Belamcanda chinensis and Iris tectorum, we established an HPLC method for simultaneous determination of six anti-inflammatory active ingredient contents in the root of the B. chinensis and I. tectorum as well as their leaves with different dry methods, and the anti-inflammatory effects of the extract were studied by the mouse ear swelling experiment. The HPLC analysis was performed on an Agilent WondaSil© C₁₈-WR(4.6 mm×250 mm，5 μm)，with isocratic elution of acetonitrile-0.1% ortho-phosphoric acid solution at a flow rate of 1. 0 mL·min⁻¹ and the detection was carried out at 265 nm. The chemical compositions of the B. chinensis and I. tectorum are similar but the contents of them are obviously different. Both rhizome and leaf extract of B. chinensis and I. tectorum had inhibitory effects on inflamed mice induced by dimethylbenzene and had anti-inflammatory effects by animal experiment, which could lay the material and active foundation for the development of the non-medicinal parts of the B. chinensis and I. tectorum.

Textual Research on Traditional Effect Verification of Belamcanda chinensis and Its Pharmacological Experimental Verification / 世界科学技术-中医药现代化

Through reviewing ancient and modern literatures,the effect evolution and disease treatment changes of Belamcanda chinensis were understood.The pharmacological experiments were used to verify its main effects.The combination of featured advantages of traditional Chinese medicine (TCM) and modern science and technology contributed to the promotion of TCM modernization.It had important significance for the development of effective components,selection of disease types in the treatment for research and development of new TCM drugs.The indication of Belamcanda chinensis was verified from textual research.The treatment of disease types by Belamcanda chinensis was verified from medical books.The treatment of disease types by Belamcanda chinensis compound was analyzed based on the Pujifang database management system.Main indications of Belamcanda chinensis were summarized.Modern pharmacological studies on anti-inflammatory mechanism of main components of Belamcanda chinensis were combined to screen animal models and investigation indexes for the preliminary verification of the efficacy of Belamcanda chinensis.The comprehensive application of classical herbal medicine books and prescription database analysis results showed that removing phlegm and relieving sore throat were the efficacy of Belamcanda chinensis,which was an important medicine in the treatment of pharyngitis and sore throat.In the modern research,serum of experimental group,IL-4 in throat tissues,as well as IgE and LTC4 level in serum and lung tissues were significantly reduced compared to the model group (P＜0.05).It was concluded that the treatment effect of Belamcanda chinensis extract to chronic pharyngitis may be through the decreasing of IgE level in serum and lung tissues,inhibiting IL-4 expression in serum and throat tissues,and the LTC4 expression in serum.

Studies on chemical constituents of Belamcanda chinensis / 第二军医大学学报

Objective: To investigate the chemical constituents of 80% ETOH extract from rhizome of Belamcanda chinensis (L.) DC. Methods: The constituents were isolated and purified by chromatography on silica gel, Sephadex LH-20 and HPLC; the structures of the compounds were determined by MS and NMR spectral analysis. Results: Ten constituents were identified from rhizome of Belamcanda chinensis (L.) DC, including tectoridin (I), acetovanillone (II), 4-hydroxy-acetophenone (III), β-sitosterol (IV), β-daucosterol (V), 5, 7, 4′-trihydroxyl-3′, 5′-dimethoxyflavone (VI), luteolin (VII), apigenin (VIII), 5,7,4′-trihydroxyflavanones (IX), and isorhamnetin (X). Conclusion: Compounds VI, VI, VIII, and IX have been isolated from rhizome of Belamcanda chinensis for the first time.

Quantitative Determination of Mangiferin in Rhizoma Belamcandae and Its Substitute of Iris L / 中草药

Mangiferin, one of the active constituents of Rhizoma Belamcandae, in samples of Be-lamcanda chinensis (L.) DC. or its substitute was determined quantitatively by RP-HPLC. The 11 sam-ples collected from different localities for analysis were: 7 rhizomes of wildly grown or cultivated B. chi-nensis, 1 of its leaf and stem, and 3 substitutes (a wildly grown and another commercially available Iristectorum Maxim. and a I. dichotoma Pall. ). Results of the analysis showed that the contents of mangiferinin Rhizoma Belamcandae were significantly higher than that of its substitutes I. tectorum and I. di-chotoma. There were also certain significant differences between samples from different localities (P<0.05), but with no statistically significant difference between the rhizome or leaf and stem, neither be-tween cultivated and wildly grown samples, (P>0.05). The method was proved to be quick, simple andreproducible, and may provide a reliable basis for the quality control and evaluation of B. chinensis.

Rhizomatous induction of Belamcanda chinensis in vitro / 中草药

Objective To optimize the medium for rhizomatous induction of Belamcanda chinensis in vitro. Methods By plant tissue culture technology, the effects of various carbon source, NAA, and active carbon at different concentration on the rhizomatous formation of B. chinensis in vitro were studied. Results MS+6-BA 2.0 mg/L+NAA 0.5 mg/L+6% white sugar was the optimal medium for the rhiz-omatous formation of B. chinensis in vitro. Active carbon should not be added to the medium. The germination rate of rhizomatous in vitro was 61.03% on the MS + BA 2.0 mg/L+3% white sugar. Conclusion Sugar concentration is the main factor of the influence on the rhizomatous formation of B. chinensis in vitro.

Study on chemical constituents of Belamcanda chinensis (Ⅱ) / 中草药

ObjectTo investigate the chemical constituents in the rhizoma of Belamcanda chinensis (L.) DC. MethodsThe chemical constituents were isolated and purified by solvent extraction together with various chromatographic techniques. The structures were elucidated on the basis of chemical evidence and spectral data. ResultsThree compounds were isolated from the EtoAC extracts of the rhizome of B. chinensis which were isorhamnetin (Ⅹ), hispidulin (Ⅺ), dichotomitin ( ⅩⅡ); four compounds were isolated from n-BuOH extracts, which were iridin ( ⅩⅢ), tectoridin (ⅩⅣ), daucosterol ( ⅩⅤ), vittadinoside or stigmasterol-3-O-glucoside ( ⅩⅥ). ConclusionCompound Ⅺ is isolated from this medicinal plant for the first time.