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Objective: As traditional techniques for microscopic identification of Chinese medicines currently lack objective and high-quality reference images, here we developed a systemic procedure to be used in microscopic identification of Chinese medicines, which would lead to more objective, effective and accurate identification process. Methods: Spatholobi Caulis (Jixueteng in Chinese) was used as the specimen in the development of such procedure. Jixueteng samples were microscopically examined in bright- and dark-field microscopy. Microscopic images were obtained by regular, EDF, and image stitching techniques. Results: The microscopic images of the characteristics in pulverized Jixueteng were captured, thanks to EDF imaging and image stitching techniques which allowed the detailed and full sighting of each characteristic to be obtained simultaneously. Different layers in anatomical transverse section, including cork, phelloderm, cortex, phloem, cambium, xylem and pith, were distinctively observed. Moreover, by comparing images of bright- and dark-field microscopy, birefringent and non- birefringent components could readily be distinguished. Conclusion: With application of the developed procedure, high-definition, panoramic and microscopic images were acquired, which could be used as the reference images for microscopic identification of Chinese medicines.
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OBJECTIVE： To study the mechanism of couplet medicine of Tripterygium hypoglaucum-Spatholobus suberectus in the treatment of rheumatoid arthritis （RA）. METHODS： The RA targets were retrieved and obtained by therapeutic target database （TTD）， DrugBank and DisGeNET databases， and the protein protein interaction （PPI） network was constructed to screen its key targets. Using oral bioavailability（OB）≥30％， drug like（DL）≥0.18 and drug half-life （HL） ≥4 h as index， active components were obtained from couplet medicine of T. hypoglaucum-S. suberectus by using TCM systematic pharmacological analysis platform （TCMSP） and TCM integrated database （TCMID）， and the targets were predicted. The active component-target network of couplet medicine of T. hypoglaucum-S. suberectus was constructed. Systems Dock Web Site online platform and Genomics platform were used to screen the active component and common targets of RA of couplet medicine of T. hypoglaucum-S. suberectus； KEGG signaling pathways of common targets were analyzed by using Cluego plug-in unit of Cytoscape 3.2.1 software. RESULTS： Totally 1 956 RA targets were retrieved， involving 11 key targets [such as IL-6， TNF， VEGFA]. The couplet medicine contained 30 active components （including luteolin， erythroxanthin， β-sitosterol and triptolide） and 229 targets. There were 37 common targets for couplet medicine of T. hypoglaucum-S. suberectus and RA （including MMP2， TNF， VEGFA）. KEGG signaling way involved cell apoptosis， IL-17 signaling pathway， Th17 cell differentiation pathway and TNF signaling pathway. CONCLUSIONS： The couplet medicine of T. hypoglaucum-S. suberectus may play a role in the treatment of RA by acting on cell apoptosis， IL-17 signaling pathway and other signaling pathways through MMP2， TNF， and VEGFA target. The results of this study can provide a reference for further study on the mechanism of the effects of couplet medicine of T. hypoglaucum-S. suberectus on RA.
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Objective To establish an HPLC method for the fingerprint analysis of Spatholobus suberectus, so as to provide evidence for the quality control of it. Methods Separation was performed on a ZORBAX SB-C18 column (150 mm × 4.6 mm, 5 μm) and the mobile phase was acetonitrile-0.2% phosphoric acid with gradient elution. The flow rate was 1 m/min and the wavelength was 260 nm. The column temperature was 30 ℃. The cluster analysis (CA) and principal component analysis (PCA) were applied to studying the HPLC fingerprint and chemical pattern recognition. Results The HPLC fingerprint of S. suberectus has been established with 35 common peaks, and six of common peaks were identified. The values of similarity evaluation mostly agreed with the result of CA and PCA. Conclusion The application of HPLC combined with CA and PCA could quickly and objectively evaluate the quality of S. suberectus from different regions.
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In this study,DNA molecular identification technology and chemical fingerprint method were adopted to evaluate the quality system of precise powder decoction pieces (PPDP) of S.suberectus dunn (SSD).ITS2 sequence was taken as DNA barcode to indentify SSD.Different specifications of PPDP were prepared,their dry extract contents were quantified in contrast with that of original slices.Three batches of SSD original slices were gleaned and the content uniformity,fingerprint and similarity evaluation before and after the mixing and pulverization were valued by HPLC-DAD.As a result,ITS2 successfully and accurately identified the SSD in this study.The extract rate of PPDP was 15.5％,1.11 times as much as the original slices.RSD of inter-assay dissolution of cepicatechin from the original slices was 11.0％,which was reduced to 1.0％ after mixing and preparing into PPDP.The relative peak area of the 14 common peaks identified by fingerpringts were larger,while the RSD values significantly decreased.It was concluded that the PPDP of SSD improved the extraction efficiency and uniformity of the original slices,featuring quite prospective in more reasonable and scientific clinical use.
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OBJECTIVE:To develop and improve the quality standard for Kanggu zengsheng tablet. METHODS:TLC was used for the qualitative identification of Drynaria fortunei,Epimedii Folium and Spatholobus suberectus;HPLC was used for the contents determination of icariin and acteoside:the column was Diamonsil C18 with mobile phase of acetonitrile-0.1% formic acid solution (gradient elution) at a flow rate of 1.0 mL/min;detection wavelength was 270 nm for icariin and 334 nm for acteoside, Cdumn temperature was 25 ℃,and the injection volume was 10 μL. RESULTS:The TLC spots of D. fortunei,Epimedii Folium and S. suberectus were clear and well separated,negative control without interference. The linear range was 0.0188-1.88 μg for ac-teoside(r=0.9999)and 0.107-2.14 μg for icariin(r=0.9999);RSDs of precision,stability and reproducibility tests were lower than 2.0%;recoveries were 100.2%-105.0%(RSD=1.6%,n=9) and 96.2%-99.5%(RSD=1.4%,n=9). CONCLUSIONS:The improved standard can more effectively control the quality of Kanggu zengsheng tablet.
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To study the anti-radiation effect and mechanism of ethanol extracts from Spatholobus suberectus and its active component catechin, ICR mice were exposed to 6Gy irradiation and randomly divided into normal group, model group, positive control group (amifostine, 43.6 mg•kg⁻¹, iv 30 min before irradiation), SSD group (10, 20, 40 g•kg⁻¹) and catechin group (50, 100, 200 mg•kg⁻¹). The mice were administered the appropriate drugs once a day after irradiation for 28 consecutive days. Blood samples were collected from the tail end and the number of peripheral blood cells was counted before irradiation and on day 1, 3, 7, 14, 21 and 28 using a microcell counter. Changes of thymus and spleen index of mice on day 7 were observed. The serum SOD, GSH-Px activity and MDA level were detected by the colorimetric method. The colony forming ability of bone marrow hematopoietic progenitor cells on day 7 was detected by semi solid culture method. The HE staining was adopted to observe the pathological changes. The apoptosis of bone marrow cells was detected by flow cytometry. The expression of cleaved caspase-3 and Bax of bone marrow cells were measured separately by western-blotting and immunohistochemistry method. SSD and catechin can both significantly revert the irradiated-induced decline in hematological parameters (RBC, WBC, PLT, Hb), improve thymus and spleen index, significantly enhance serum SOD and GSH-Px activity and decrease the MDA level. The proliferation and differentiation of hematopoietic progenitor cells in bone marrow were promoted, the apoptosis of bone marrow cells was significantly up-regulated and the expression of cleaved caspase-3 and Bax was significantly reduced in SSD and catechin group. SSD and catechin have significant anti-radiation effect and its mechanism may be related to hematopoietic promoting, antioxidant and anti-apoptotic effects.
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Objective The dynamic accumulation trend of secondary metabolite in cultivated Spatholobus suberectus Dunn at different growth time was studied to provide scientific basis for confirming its reasonable harvesting time. Methods Ethanol-soluble extractives were measured by the method recorded in Chinese Pharmacopoeia ( A) Appendix XA ( 2010 edition). The content of total flavonoids was tested by ultraviolet spectrophotometer. The contents of protocatechuic acid, catechin, and epicatechin in Spatholobus suberectus Dunn were determined by high performance liquid chromatography ( HPLC). Results The contents of ethanol-soluble extractives, total flavonoids and the major components such as protocatechui cacid, catechin and epicatechin in Spatholobus suberectus Dunn showed increasing trend with the prolongation of growth time. The changes of contents of alcohol-soluble extract, total flavonoids and epicatechin in samples collecting at different months in one year showed a parabolic trend, and the contents reached the peak values from July to September. Catechin content kept increasing continually. The content of ethanol -soluble extractives of Spatholobus suberectus Dunn aged 4-6 years met the requirements of the current edition of Chinese Pharmacopoeia when harvested from July to September. Conclusion It is preliminarily recommended that Spatholobus suberectus Dunn cultivated for more than 7 years is suitable for harvesting, but the optimal harvest period and the harvest month still need to be further studied.
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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.
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This study was aimed to establish an identification method for Spatholobus suberectus and its adulterants by near-infrared spectroscopy (NIRS). Near-infrared diffuse reflection spectroscopy (NIRDRS) spectra of different S. suberectus and its adulterants were acquired by using OPUS INDENT analysis software. NIRDRS spectra clustering analysis model and identification model were established and verified. The results showed that S. suberectus from dif-ferent regions and its adulterants were identified successfully by clustering analysis model and identification model. It was concluded that Spatholobus suberectus and its adulterants can be identified rapidly and non-destructively by NIRS.
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Objective To determine the content of protocatechuic acid in Spatholobus suberectus Dunn.Methods HPLC method was developed for the determination of protocatechuic acid in Spatholobus suberectus Dunn.The separation was performed on C18 column with acetonitrile-water-orthophosphoric acid(8 ∶92 ∶0.1) as a mobile phase with the flow rate being 1.0 mL/min.The detection wavelength was set at 260 nm.Results A satisfactory separation between protocatechuic acid and impurity was obtained.The calibration curve was linear in the range of 0.020～2.008 ug,r=0.999 9.The average recovery was 98 %,and RSD was 0.1 %(n=5).Conclusion The method is accurate and can be used for the detemrmination of protocatechuic acid in Spatholobus suberectus Dunn.
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Aim To study effects of nine compounds extracted from Spatholobus suberectus Dunn (SSD) on proliferation of hematopoietic progenitor cell (HPC) in marrow-depressed mice. Methods Serum pharmacology experiment was used to observe the influence of nine compounds on growth of CFU-E、BFU-E、CFU-GM、CFU-Meg in marrow-depressed mice. Results Compared with the control, all compounds except pyromucic acid and ononin could significantly stimulate the growth of CFU-GM (P