Study on extraction technology of Sophora flavescens-Phellodendron chinense drug pair / 中国药房

OBJECTIVE To study the extraction technology of Sophora flavescens-Phellodendron chinense drug pair and provide a reference for the development of new drugs for the treatment of anorectal diseases. METHODS Using the contents of total alkaloids of S. flavescens （matrine+oxymatrine）， berberine hydrochloride and total flavonoid， and extract yield as evaluation indicators， analytic hierarchy process-entropy weight method was used to calculate the weight coefficient of each indicator， and was combined with Box-Behnken design-response surface method to study the extraction technology of S. flavescens-P. chinense drug pair and verify it. RESULTS The optimal extraction technology of S. flavescens-P. chinense drug pair was immersed in 12-fold amount of 58% ethanol for 30 minutes and extracted twice， each time for 120 minutes. The relative error between the verification experimental results and the predicted value was 1.88%. CONCLUSIONS The obtained extraction technology is stable and feasible and can provide reference for the application of S. flavescens-P. chinense drug pair and development of new drugs.

Discovery, isolation and structural identification of alkaloid glycosides in six traditional Chinese medicine such as Coptis chinensis / 中国中药杂志

Alkaloids are important active ingredients occurring in many traditional Chinese medicines, and alkaloid glycosides are one of their existence forms. The introduction of saccharide units improves the water solubility of alkaloid glycosides thus presenting better biological activity.Because of the low content in plants, alkaloid glycosides have been not comprehensively studied. In this study, ultrahigh performance liquid chromatography-quadrupole time of flight-tandem mass spectrometry(UPLC-QTOF-MS/MS) was employed to identify and analyze the alkaloid glycosides in Coptis chinensis, Phellodendron chinense, Menispermum dauricum, Sinomenium acutum, Tinospora sagittata and Stephania tetrandra. The results showed that except Tinospora sagittata, the other five herbal medicines contained alkaloid glycosides. Furthermore, the alkaloid glycosides in each herbal medicine were identified based on UV absorption spectra, quasimolecular ion peaks in MS, fragment ions information in the MS/MS, and previous literature reports. A total of 42 alkaloid glycosides were identified. More alkaloid glycosides were identified in C. chinensis and Menispermum dauricum, and eleven in C. chinensis were potential new compounds. Furthermore, the alkaloid glycosides in the water extract of C. chinensis were coarsely se-parated by macroporous adsorption resin, purified by column chromatography with D151 cation exchange resin, ODS and MCI, combined with semi-preparative high performance liquid chromatography. Two new alkaloid glycosides were obtained, and their structures were identified by mass spectrometry and NMR data as(S)-7-hydroxy-1-(p-hydroxybenzyl)-2,2-N,N-dimethyl-1,2,3,4-tetrahydroisoquinoline-6-O-β-D-glucopyranoside and(S)-N-methyltetrahydropalmatubine-9-O-β-D-glucopyranoside, respectively. This study is of great significance for enriching the information about the chemical composition and the in-depth development of C. chinensis. Meanwhile, it can provide a reference for rapid identification and isolation of alkaloid glycosides from other Chinese herbal medicines.

Preliminary study on the material basis for expectorant and cough relief effects of the fruits of Phellodendron chinense var. glabriusulum Schneid / 中国药房

OBJECTIVE To explore the material basis and mechanism of expectorant and cough relief effects of the fruits of Phellodendron chinense var. glabriusulum Schneid. METHODS The expectorant and cough relief effects of volatile oil and water decoction of the fruits of P. chinense var. glabriusulum Schneid. were studied by ammonia water cough induction and drug expectorant model mice experiments； GC-MS and UPLC-MS technologies were used to identify its volatile oils and non-volatile components of the fruits of P. chinense var. glabriusulum Schneid. The active ingredients， core targets and pathways of expectoration and cough relief were analyzed by network pharmacology. RESULTS The volatile oil （0.8， 0.2 g/kg， calculated by volatile oil） and water decoction （12， 3 g/kg， calculated by crude drug） of the fruits of P. chinense var. glabriusulum Schneid. both had obvious expectorant and cough relief effects， and showed obvious dose-dependent relationship. A total of 38 volatile oil components were identified from the medicinal herbs， and the relative percentage contents of 8 components were greater than 1%， such as α -pinene， myrcene， β -caryophyllene， germanene D， isospathulenol； a total of 69 non-volatile oil components were identified， mainly including phenolic compounds， alkaloids， and flavonoids. The active ingredients screened from the identified components included 13 compounds such as α-pinene， myrcene， chlorogenic acid， luteolin， berberine. There were a total of 55 intersection targets with diseases， and the core targets were tumor necrosis factor （TNF）， epidermal growth factor receptor （EGFR）， vascular endothelial growth factor A （VEGFA）， serine/threonine kinase proteins （AKT1） and Toll-like receptor 4 （TLR4）. The molecular docking results showed that the active ingredients and the core targets had good binding ability. GO functional analysis found that the targets were significantly enriched in biological processes such as the reaction affecting lipopolysaccharides， the positive regulation of peptidyl serine phosphorylation， and the positive regulation of the biosynthesis process of nitric oxide. KEGG pathway enrichment analysis found that the targets were significantly enriched in the signaling pathways such as cancer， non-small cell lung cancer， proteoglycans in cancer. CONCLUSIONS Fruits of P. chinense var. glabriusulum Schneid. have obvious expectorant and cough relief effects， and its material basis may be α-pinene， myrcene， chlorogenic acid， luteolin， berberine， etc.， and mainly act on TNF， EGFR， VEGFA， AKT1， TLR4 and its significantly enriched signaling pathway.

Globally Ecological Suitability Analysis of Phellodendron chinense and Phellodendron amurense / 中国实验方剂学杂志

Objective: Phellodendri Cortex, one of the "three wood medicine materials", is a Chinese traditional medicinal material and also a national second-class protected plant in China. Its is considered as excellent trees for the Natural Forest Conservation Program and the Grain-to-Green Program because of its high economic value and ecological value. The Phellodendron Cortex is divided into Phellodendron chinense and P. amurense according to species and origins. The global potential suitable areas predicted by Global Geographic Information System for Medicinal Plant (GMPGIS) can provide data for us to decide which specie can be selected in different areas. Method: Sample ecological information was collected from global genuine areas, main producing areas and wild distribution areas, and a total of 364 sampling sites of P. chinense and 247 sampling sites of P. amurense were used by GMPGIS to analyze the suitable growth areas in the world. Result: A clear geographical line existed between P. chinense and P. amurense. P. chinense was mainly distributed in tropical monsoon climate and had the most suitable areas in Asia, Europe, North America, South America and Oceania, including 65 countries and regions such as China, the United States, France, Brazil, Japan, Italy and New Zealand. P. amurense was mainly distributed in temperate monsoon climate and had the most suitable areas in Asia, Europe, and North America, including 30 countries and regions such as the United States, China, Russia and Canada.. Conclusion: The results of GMPGIS can provide scientific data for selecting correct species and cultivation areas for Phellodendris Cortex in future.

Sensitive and Rapid Detection of Traditional Chinese Herbs by Loop-Mediated Isothermal Amplification Method and Real-Time Fluorescence Quantitative PCR

ABSTRACT Adulterant herbal materials are threats to import and export trade and consumer safety. In this study, we established a simple and rapid examination system for the detection of Phellodendron chinense Schneid. Two detection methods, real-time fluorescence quantitative PCR (real-time PCR) and loop-mediated isothermal amplification (LAMP), were developed for traditional Chinese medicine detection, and their specificity and sensitivity were compared. The DNA of P. chinense was extracted and its special periods amplified with designed primers. Real-time PCR and LAMP experiments were conducted to test the specificity of primers in contrast to other similar species. The template concentration was diluted from 101 ng/µL to 10-5 ng/µL in order to contrast sensitivity between real-time PCR and LAMP. Real-time PCR and Lamp method has shown specificity because P. chinense was positive as opposed to other negative similar species. The Lamp method could detect a limited DNA concentration of 10-4ng/µL in 60 minutes with same sensitivity to real-time PCR. The results indicate that real-time PCR and LAMP are sensitive, accurate and specific in detection of P. chinense. However, LAMP is more convenient and cast less time. What's more, expensive equipments are not necessary for LAMP detector. For a better detection, we suggest an establishment of a real-time PCR and LAMP method for TCM market supervision which depends on DNA barcode sequences and LAMP.

Accumulation of main alkaloids composition in growth process of different parts of Phellodendron chinense / 中草药

Objective: To analyze the content variation of phellodendrine and berberine from Phellodendron chinense in different years, different seasons, and different parts, and to provide the basis for increasing the planting benefit of Chinese herbal medicine. Methods: High performance liquid chromatography (HPLC) analysis method was used to determine the contents. Results: The contents of phellodendrine and berberine from different parts of P. chinense showed as the following order: root barks > barks > roots. The contents of the two kinds of alkaloids changed each year. In first three years, the content change trend was obvious, and the content in barks was close to 5% in the third year. The young P. chinense had development value. After the third year, the contents of the two kinds of alkaloids changed slowly. The contents of phellodendrine and berberine in October were higher. Conclusion: The metabolic accumulation rules of berberine and phellodendrine are summarized, in order to provide the basis for the sustainable utilization of P. chinensis resources.

Effect of Processing Method on the Content of 3 Kinds of Alkaloids in Phellodendron chinense / 中国药房

OBJECTIVE:To study the influence of processing method on the content of 3 kinds of alkaloids in Phellodendron chinense. METHODS:HPLC was used to determine the content of 3 kinds of alkaloids in P. chinense,such as stir-frying with wine,salt-roast process and carbonization. The separation was performed on Zorbax SB-C18(250 mm?4.6 mm,5 ?m) column. The mobile phase consisted of acetonitrile-0.1% phosphoric acid (48 ∶ 52,0.1 g SDS per 100 mL) with flow rate of 1.0 mL?min-1. Detection wavelength was set at 265 nm and column temperature was set at room temperature. Injection volume was 10 ?L. RESULTS:Except for special case,the content of berberine hydrochloride,pamatine hydrochoride and jateorizine hydrochoride were all decreased after processing,especially carbonization.CONCLUSION:The 3 kinds of alkaloids can be also used as index of quality control of processed P. chinense. The method is simple,rapid and accurate for the determination of 3 kinds of alkaloids in P. chinense.

Studies on biopharmacy of ultra-fine particles of Atractylodes lancea, Phellodendron chinense and a pill composed of the two drugs Ⅰ. Characterization by scanning electron microscopy / 中草药

To characterize the ultra-fine particles of Atractylodes lancea (Thunb.) DC., Phellodendron chinense Schneid and the ERMIAO PILL*, a compounded preparation of the above two drugs, for the purpose to observe a deeper view on their physico-chemical properties. Methods　By comparing their particle size, porosity and specific surface area with light and scanning electron microscope. Results　The size of the ultra-fine particles were more uniform in size, 90% of which were under 20 μm, their specific area were increased by 60% ～ 190% and bulk density were around 0.42 g/cm3, and the great majority of the plant cells were broken as compared with the conventional coarse powder. Conclusion　Both A. lancea and P. chinense together with their combined preparation became smaller in particle size, more uniformly distributed with increased specific surface area and broken cell wall.

Study on extracting purified active components of Picria fel-terrae, Scutellaria baicalensis and Phellodendron chinense with macroporous resins / 中草药

Object To study the application of macroporous resins to the purified active components of Picria fel-terrae Lour., Scutellaria baicalensis Georgi, and Phellodendron chinense Schneid. Methods With the purified active components of Chinese materia medica (CMM) as a standard, we selected the suitable macroporous resins and studied the optimum technological parameters of the adsorption and elution. Results The suitable three types of macroporous resins which were used to the purified active components of CMM were HPD500 for P. fel-terrae., HPD300 for S. baicalensis, and HPD100 for P. chinense. In the absorption course, when the ratios between the medicinal materials amount and the volume of macroporous resins were 1.5 for P. fel-terrae, 0.5 for S. baicalensis and 1 for P. chinense, the purified active components of CMM appeared leaking, and were no more absorbed when the ratios were 11 for P. fel-terrae, 2 for S. baicalensis and 4.75 for P. chinense. In the elution course, the optimum alcohol concentrations were 50% for P. fel-terrae, 30% for S. baicalensis and 50% for P. chinense. Conclusion It is obviously different to refine the active components of CMM, while using the diverse types of macroporous resins.

Studies on biopharmacy of ultra-fine particles of Atractylodes lancea, Phellodendron chinense and a pill composed of the two drugs Ⅰ. Characterization by scanning electron microscopy / 中草药

Object To characterize the ultra fine particles of Atractylodes lancea (Thunb ) DC., Phellodendron chinense Schneid and the ERMIAO PILL *, a compounded preparation of the above two drugs, for the purpose to observe a deeper view on their physico chemical properties Methods By comparing their particle size, porosity and specific surface area with light and scanning electron microscope Results The size of the ultra fine particles were more uniform in size, 90% of which were under 20 ?m, their specific area were increased by 60% ～ 190% and bulk density were around 0 42 g/cm 3, and the great majority of the plant cells were broken as compared with the conventional coarse powder Conclusion Both A. lancea and P chinense together with their combined preparation became smaller in particle size, more uniformly distributed with increased specific surface area and broken cell wall

Reversed-phase ion pair high performance liquid chromatography in determination of berberine and palmatine in Phellodendron chinense Schneid.and its granules / 第二军医大学学报

Objective:To determine berberine and palmatine in Phellodendron chinense Schneid.and its granules.Methods: The reversed-phase ion pair high performance liquid chromatography(RP-HPLC) was used and the validation of the method was tested.The chromatography condition was with Lichrospher C18 column(4.6 mm?250 mm,5 ?m), mobile phase was ACN∶25 mmol/L NaH 2PO 4∶25 mmol/L SDS(2∶1∶1),flow speed was 1.0 ml/min,detection wavelength was 345 nm,and temperature of column was 25℃,Phellodendron chinense Schneid.and its granules were extracted with methanol solution of hydrochloric acid.Results: The theoretical plate number of berberine and palmatine were 14 906 and 14 847,the resolution were 2.33 and 2.86,the tailing factor were 1.09 and 1.06,respectively; all the parameters were suitable for determination.The calibration curves were linear in the range of 40-500 ng,Y=698 278X-3 846,r=1.000(berberine) and 20-250 ng, Y= 536 632X- 7 738, r=0.999 9,r=0.999 4(palmatine).The intra-day and inter-day precision(RSD) at low,middle and high injection amount was all less than 2.5%(berberine) and 1.5%(palmatine).The stability(RSD) was 0.66%(berberine) and 0.70% (palmatine) in 48 h.The recurrence(RSD,n=5) was 0.11%(berberine) and 0.12%(palmatine).The limits of detection was 2.0 ng(berberine) and 1.0 ng(palmatine).The recoveries were 100.4% (RSD=0.12%,n=3) for berberine and 99.80%(RSD=0.22%,n=3) for palmatine.The contents of berberine and palmatine in 3 batch of Phellodendron chinense Schneid.and 5 batch of its granules were determined.Conclusion: Our method can be used for determination of berberine and palmatine in Phellodendron chinense Schneid.and its granules, which is simple and reliable.