Comparison of Wild and Cultivated Bupleurum chinense Based on Traditional Quality Evaluation / 中国实验方剂学杂志

ObjectiveIn order to understand the quality differences between wild and cultivated Bupleurum chinense（BC）， modern analytical techniques were used to systematically compare the quality of wild and cultivated BC in terms of appearance characteristics， primary and secondary metabolites. MethodSamples of wild and cultivated BC were collected from the main production areas of Shanxi， Shaanxi and Hebei， and images of BC were collected and their length and diameter were measured using vernier caliper to compare and analyze the characteristics of the two. Referring to the method under extract of CP in the 2020 edition of Chinese Pharmacopoeia， the extract contents of the two species were determined. The cellulose， hemicellulose and lignin compositions of both were determined using fiber analyzer. Quantitative determination of representative saikosaponins， flavonoids and saccharides in BC by ultra performance liquid chromatography（UPLC）， headspace gas chromatography-mass spectrometry（HS-GC-MS） was used to determine the types and relative contents of volatile components， and UPLC-quadrupole-time-of-flight mass spectrometry（UPLC-Q-TOF-MS） coupled with multivariate statistical analysis was used to screen and identify the differential compounds between wild and cultivated BC. ResultThere were significant differences in the appearance characteristics between wild and cultivated BC， the wild BC had a large root head， twisted and thick axial root， rough epidermis， and often had a stem base and lateral root with dark color and strong odor. However， the cultivated BC has long and straight taproots， delicate epidermis， few lateral roots， light root color and light smell. In terms of primary and secondary metabolites， the contents of alcohol-soluble extract and lignin of wild BC was significantly higher than those of cultivated BC， while the contents of water soluble extract and quercitrin was higher than those of cultivated BC， but the difference was not significant. The contents of cellulose， five saikosaponins， rutin， narcissoside and isorhamnetin-3-O-glucoside in cultivated BC were significantly higher than those of wild BC， and the total water-soluble polysaccharides， sucrose， hemicellulose and starch of cultivated BC were higher than those of wild BC， but the difference was not significant. The results of HS-GC-MS identification showed that a total of 67 volatile components were identified in wild and cultivated BC， 59 in wild BC and 51 in cultivated BC， with a total of 43 compounds in both， and the screening based on variable importance in the projection（VIP） value>1 revealed that the differential components were mainly concentrated in the aromatic and fatty acid compounds. The results of UPLC-Q-TOF-MS-based non-targeted metabolomics combined with multivariate statistical analysis showed that the two were significantly different in saikosaponins and the differential compounds had higher response values in cultivated BC. ConclusionThere are significant differences in the appearance， primary and secondary metabolite contents between wild and cultivated BC. At present， the quality evaluation system of cultivated BC is not perfect， and this study provides theoretical references for updating and revising the quality evaluation standard of cultivated BC and guiding the production of high-quality BC.

Study on the intestinal absorption characteristics of saikosaponins / 中国药房

OBJECTIVE To explore the intestinal absorption characteristics of saikosaponins. METHODS Based on everted intestinal sac model， using accumulative absorption amount （Q） and absorption rate constant （Ka） as indexes， UHPLC-MS/MS technique as a method， the absorption of saikosaponin A， B2， C， D and F from total saponins of Bupleurum chinense （8 g/mL， by crude drug） in the duodenum， jejunum and ileum was detected. RESULTS The correlation coefficients （r） of the regression equations for the absorption of saikosaponins A， B2， C and F in the duodenum， jejunum and ileum were all higher than 0.95， while the r of saikosaponin D in the above intestinal segments was lower than 0.95； compared with the absorption of the same composition in the duodenum， the Q and Ka of saikosaponin A and C circulating in jejunum and ileum for 120 min， as well as the Q and Ka of saikosaponin F circulating in the ileum for 120 min were significantly decreased （P＜0.05）. CONCLUSIONS Saikosaponin A and the other 4 saikosaponins are all absorbed in the duodenum， jejunum and ileum； among them， saikosaponin A， B2， C and F are linearly absorbed， which conforms to the zero-order absorption characteristics， but saikosaponin D shows non- linear absorption.

Correlations between appearance traits and internal quality of Bupleurum scorzonerifolium roots based on theory of "quality evaluation through morphological identification" / 中国中药杂志

It is generally believed that high-quality Bupleurum scorzonerifolium roots possess specific morphological characteristics, being red, robust, and long with strong odor. However, the scientific connotation of these characteristics has not been elucidated. According to the theory of "quality evaluation through morphological identification", we studied the correlations between appearance traits(the RGB value of root surface, root length, root diameter, dry weight, and ratio of phloem to xylem) and content of main chemical components(volatile oils, total saponins, total flavonoids, total polysaccharides, and seven saikosaponins) of B. scorzonerifolium roots. Epson Scanner and ImageJ were used to scan the root samples and measure the appearance traits. Ultraviolet spectrophotometry and HPLC were employed to determine the content of chemical components. The correlation, regression, and cluster analyses were performed to study the correlations between the appearance traits and the content of chemical components. The results showed that the content of volatile oils and saikosaponins were significantly correlated with RGB value, root length, and root diameter, indicating that within a certain range, the roots being redder, longer, and thicker had higher content of volatile oils and saikosaponins. According to the appearance traits and chemical component content, the 14 samples from different producing areas were classified into four grades, and the differences in morphological traits and chemical component content were consistent among different grades. The findings in this study demonstrate that appearance traits(RGB value, root length, and root diameter) can be used to evaluate the quality of B. scorzonerifolium roots. Meanwhile, this study lays a foundation for establishing an objective quality evaluation method for B. scorzonerifolium roots.

Rapid determination of microbial contamination of Bupleurum chinense DC. decoction slices by ATP bioluminescence technology combined with statistical analysis methods / 药学学报

This study focuses on the microbial quality control of the Chinese herbal decoction pieces. In view of the shortcomings of traditional culture methods such as slow detection speed and inability to detect unculturable microorganisms, a new method based on ATP bioluminescence technology combined with statistical analysis methods was established to rapidly predict and quantitatively detect the total aerobic microbial count (TAMC) and total yeast and mold count (TYMC) contaminated Bupleurum chinense DC. decoction pieces. Based on the optimized ATP bioluminesence detection system, accurate detection of pure bacterial solution of Escherichia coli, Bacillus subtilis and Staphylococcus aureus can be achieved, with detection limits of 47.86, 89.13 and 1 862.09 CFU·mL-1, respectively. The detection time was 6.5 h, and the detection cost was as low as 2 yuan/time. The upper and lower warning limits of TAMC were determined by the misjudgment rates of 10% and 20%, respectively. And the warning limit of TYMC was determined by the misjudgment rate of 20%. The proposed crossing method could quickly predict the amount of microbial contamination in Bupleurum chinense DC. decoction pieces. The constructed partial least squares regression (PLSR) model could accurately quantify the quantity of microbial contamination in Bupleurum chinense DC. decoction pieces. The optimal PLSR prediction model for TAMC had a correction coefficient (R2) of 0.826, a root mean square error of correction set (RMSEE) of 0.468 and a root mean square error of cross-validation set (RMSECV) of 0.465. The R2, RMSEE and RMSECV in the prediction model of TYMC were 0.778, 0.543 and 0.541, respectively. The aim of this study is to establish a kind of rapid detection method and prediction models for the microbial limit of traditional Chinese medicine and Chinese herbal decoction pieces, and to provide a more convenient and sensitive detection technology for the microbial quality process control of traditional Chinese medicine products.

Analysis of codon usage patterns in Bupleurum falcatum chloroplast genome / 中草药·英文版

OBJECTIVE@#In order to distinguish the traditional Chinese medicine Bupleurum falcatum and its adulterants effectively and develop a better understanding of the factors affecting synonymous codon usage, codon usage patterns of chloroplast genome, we determine the complete chloroplast (cp) genome of B. falcatum and clarify the main factors that influence codon usage patterns of 78 genes in B. falcatum chloroplast genome.@*METHODS@#The total genomic DNA of fresh leaves from a single individual of B. falcatum was extracted with EASYspin plus Total DNA Isolation Kit and 2 μg genome DNA was sequenced using Illumina Hiseq 2500 Sequencing Platform. The cp genome of B. falcatum was reconstructed with MITObim v1.8 and annotated in the program CPGAVAS2 with default parameters. Python script and Codon W were used to calculate the codon usage bias parameters.@*RESULTS@#The full length of B. falcatum cp genome was 155 851 bp, 132 different genes were annotated in this cp genome containing 80 protein-coding genes, 30 tRNA genes, and four rRNA genes. The codon usage models tended to use A/T-ending codons. The neutrality plot, ENC plot, PR2-Bias plot and correspondence analysis showed that both compositional constraint under selection and mutation could affect the codon usage models in B. falcatum cp genome. Furthermore, three optimal codons were identified and most of these three optimal codons ended with G/U.@*CONCLUSION@#The cp genome of B. falcatum has been characterized and the codon usage bias in B. falcatum cp genome is influenced by natural selection, mutation pressure and nucleotide composition. The results will provide much more barcode information for species discrimination and lay a foundation for future research on codon optimization of exogenous genes, genetic engineering and molecular evolution in B. falcatum.

Exploration of the Mechanism about Couplet Medicine of Bupleurum falcatum-Scutellaria baicalensis Intervening in Podocyte Lesion Based on Network Pharmacology / 中国药房

OBJECTIVE：To predict the pot ential target and mechanism of Xintong Changluo Method carrier-Compund Shensu Ⅱ couplet medicine of Bupleurum falcatum-Scutellaria baicalensis intervening in podocyte lesion ，and to provide reference for the development of sequential clinical and basic research of Xintong Changluo Method in the prevention and treatment of podocyte lesion. METHODS ：Based on TCMSP database ，chemical components and target protein of B. falcatum and S. baicalensis were retrieved，and Cytoscape 3.2.1 software was used to draw a “TCM-component-target”network. The targets related to podocyte lesion were searched from OMIM database ，DrugBank database and Digsee online text ，and the intersection genes of above targets and“B. falcatum -S. baicalensis ”target were obtained by Venny 2.1.0 online mapping tool. The protein-protein interaction （PPI） network was constructed by STRING database ，and the core targets were obtained by topology analysis of the network by using CytoNCA plug-in Cytospace 3.2.1 software. With the help of DAVID database ，the fu nction of Gene Ontology （GO）was annotated and KEGG pathway was enriched ；and the enrichment results were visualized through OmicShare Tools online mapping platform. RESULTS ：Based on retrieval results of TCMSP database，44 active components were obtained ，involving 13 com of B. falcatum and 32 of S. baicalensis ； stigmasterol is common component of B. falcatum and S. baicalensis Quercetin，kaempferol and wogonin were the compounds withmain potential targets. T he target proteins wi th high node degree were prostaglandin endoperoxide synthase 2（PTGS2），PTGS1， nuclear receptor coactivator 2 and heat shock protein 90α，which were associated with 37，30，25 and 25 active components respectively. Twenty genes were obtained from the interaction between “B. falcatum -S. baicalensis ”and podocyte lesion related targets，including PTGS2，VEGFA，MMP9，TNF and IL6. PPI network diagram of the above intersection genes contained 20 nodes and 110 lines，with MMP9，VEGFA，IL6 and other genes at the core. The results of GO analysis showed that a total of 154 biological information items were obtained （P＜0.05），including 139 biological process items ，8 cell composition items and 7 molecular function items. Among them ，biological processes mainly involved in the positive regulation of NO biosynthesis process ， inflammatory response ，immune response. Cell composition mainly involved in extracellular space ，extracellular region ，external side of plasma membrane ，etc.，and molecular function mainly involved in protein binding ，cytokine activity ，growth factor activity，etc. At the same time ，47 KEGG pathways were obtained （P＜0.05），mainly including cytokine-cytokine receptor interaction，rheumatoid arthritis ，malaria，cancer signaling pathways. CONCLUSIONS ：The active components of Compund Shensu Ⅱ couplet medicine of “B. falcatum -S. baicalensis ”may act on MMP9，VEGFA，IL6，TNF and other targets through cytokine-cytokine receptor interaction ，rheumatoid arthritis ，malaria，cancer signal pathway ，so as to play its intervention effect on podocyte lesion.

Complete plastid genomes of Bupleurum chinense DC. and B. boissieuanum H. Wolff, with comparative and phylogenetic analyses of medicinal Bupleurum species / 药学学报

italic>Bupleurum L. (Apiaceae) is an economically important genus, in which many species are of medicinal value. In this study, the complete plastid genomes (plastomes) of B. chinense DC. and B. boissieuanum H. Wolff were sequenced and their characteristics were investigated. Comparative and phylogenetic analyses were conducted with other published Bupleurum plastomes. The complete plastomes of B. chinense and B. boissieuanum were 155 458 and 155 800 bp in length, and both exhibited the typical quadripartite circular structure consisting of a large single copy region (LSC, 85 343 and 85 804 bp), a small single copy region (SSC, 17 495 and 17 410 bp), and a pair of inverted repeat regions (IRa/b, 26 310 and 26 293 bp), respectively. A total of 129 genes, including 84 protein-coding genes, 37 transfer RNA (tRNA) genes, and eight ribosomal RNA (rRNA) genes were identified from each of the two plastomes. Repeat sequences detected were similar in types and distribution patterns, but the numbers were slightly different. Comparative analyses revealed that the Bupleurum plastomes were highly conserved in length, structure, the guanine and cytosine (GC) content, and gene content and order, both intraspecifically and interspecifically, and no obvious expansion or contraction of the inverted repeat regions occurred. Sequence variation was lower within the same species than among different species, noncoding sequences (including intergenic regions and introns) showed a higher divergence than the protein-coding sequences, and sequences in the LSC and SSC regions were more divergent than those in the IR regions. In addition, 11 sequences with higher nucleotide diversity among species were detected in the LSC and SSC regions. All studied Bupleurum species were inferred forming a monophyletic group with a 100% bootstrap value. Bupleurum chinense and B. boissieuanum were phylogenetically closest to B. commelynoideum and B. falcatum, separately, with all three B. chinense accessions clustered into a distinct clade. These results provide genetic information for further species identification, phylogenetic resolution, and will assist in exploration and utilization of medicinal Bupleurum species.

Effects of Bupleurum chinense on the diversity of intestinal flora in depressed mice / 中国中药杂志

Bupleuri Radix, serving as the sovereign medicinal in many antidepressant compound preparations, has been proved effective in treating depression in mice, but its effect on the intestinal flora remains unclear. The present study aimed to investigate the effects of Bupleurum chinense(one of the original materials of Bupleuri Radix) on the behaviors and the diversity of intestinal flora of depressed mice. A depression mouse model was induced by repeated social defeat stress. Specifically, C57 BL/6 J male mice were exposed to the attack from the CD-1 mice. Then, C57 BL/6 J male mice were divided into a depression group and a B. chinense group, with normal saline and B. chinense administered(ig) respectively. Sucrose preference test and tail suspension test were conducted during and after the experiment respectively, to analyze the effects of B. chinense on the behaviors of the depressed mice. The feces were collected after the experiment. The V3-V4 16 S rDNA regions of intestinal flora of mice in each group were sequenced by Ion S5 TMXL for the analysis of the number of operational taxonomic units(OTUs), richness, alpha and beta diversity indexes, and differential phyla and genera. The results indicated that B. chinense could decrease depressive-like behaviors of mice, increase sucrose preference, and shorten the time of immobility in tail suspension test. After B. chinense intervention, the relative abundance of Firmicutes was significantly decreased, while that of Bacteroidetes was increased at the phylum level. At the genus level, the relative abundance of Lactobacillus and Lachnoclostridium decreased(P<0.05), while that of Bacteroides, Alistopes, etc. was elevated(P<0.05). The findings demonstrate that B. chinense can regulate the intestinal flora and improve the depressive-like behaviors of mice with depression.

Molecular Identification of Bupleurum chinense Seeds Based on DNA Barcoding Technology / 中国实验方剂学杂志

Objective:To establish a molecular identification method for Bupleurum chinense seeds based on ribosomal DNA internal transcribed spacer (ITS) sequence, ensuring the species authenticity of the cultivated seeds of B. chinense. Method:A total of 59 seeds samples of B. chinense and its main cultivated species, marketed B. chinense were collected. The effect of different sampling amounts and different water bath conditions on DNA extraction quality of the seeds was investigated, a DNA extraction method for seeds of Bupleurum was established. Their ITS sequences were obtained by polymerase chain reaction (PCR) and bidirectional sequencing. In addition, 34 ITS sequences of main cultivated Bupleurum species, such as B. chinense, B. scorzonerifolium, B. falcatum and B. smithii, were downloaded from GenBank to enrich identification database of B. chinense seeds. The neighbor-joining (NJ) dendrogram were constructed by MEGA-X 10.0.5 software to investigate the the species identification ability of ITS sequences for B. chinense seeds. And DNA barcoding identification of marketed B. chinense seeds was conducted based on BLAST method and NJ dendrogram method. Result:In total, 59 ITS sequences were obtained. ITS sequences of B. chinense could be divided into six haplotypes, including seven variable sites. The NJ dendrogram indicated that all the haplotypes of B. chinense could form independent branches, which could be distinguished from other cultivated species of Bupleurum in the collected samples, and possessed the ability to identify species of B. chinense seeds. Based on ITS sequence barcoding identification, 3 of the 19 marketed B. chinense seeds were B. falcatum with a counterfeit rate of 15.8%. Conclusion:DNA barcoding technology based on ITS sequence can accurately and reliably identify B. chinense seeds and its adulterants, providing reference for the standardization construction of Chinese medicinal materials seeds.

Identification of Bupleurum Seeds by ITS2 Genetic Barcoding and Seed Morphology / 中国实验方剂学杂志

Objective:To accurately identify Bupleurum seeds by traditional morphological identification method combined with DNA barcoding technique. Method:A total of 41 seed samples on the market were collected and 75 ribosomal DNA internal transcribed spacer 2 (ITS2) sequences of 15 varieties were downloaded from the GenBank database as experimental materials. The seeds were measured and observed by stereomicroscope and vernier caliper, and their 1 000-grain weights were calculated. Genomic DNA was extracted from the seeds and used as a template, and ITS2 sequences were amplified using polymerase chain reaction (PCR) and bidirectional sequencing. Species identification was conducted based on BLAST method, neighbor-joining (NJ) phylogenetic tree method, Kimura two-parameter model (K2P) genetic distance method, and secondary structure of ITS2 sequence. Result:There were slight differences in the length, width, cross-section, and 1 000-grain weight among Bupleurum seeds from different origins. The ITS2 sequences of B. chinense seeds had 2 intraspecific variable sites and 3 haplotypes, the maximum intraspecific genetic distance (0.009) was far smaller than the minimum interspecific genetic distance (0.032). B. chinense and B. scorzonerifolium in the NJ phylogenetic tree were clustered into independent branches with good monophyletic property. The secondary structure of ITS2 sequences could make up for the shortcomings of NJ tree in identifying variants. The collected 41 seeds included 30 B. chinense seeds, 3 B. scorzonerifolium seeds, 5 B. falcatum seeds, 2 B. marginatum var. stenophyllum seeds, and 1 B. smithii var. parvifolium seeds. Conclusion:The B. chinense seeds on the market have problems of diverse sources and chaotic origins. Based on the combination of ITS2 gentic barcoding and seed morphological identification, the Bupleurum seeds can be accurately identified, which provides scientific bases for establishing the quality standard of Bupleurum seeds, standardizing the cultivation of B. chinense, and solving the quality problems of B. chinense from the source, and provides a reference for the accurate identification of other medicinal plant seeds or seed medicinal materials.

Quality Difference Characteristics of Bupleurum Formulae Granules Based on the Analysis of Dual-Wavelength Fingerprint Profiles and Chemometric Methods / 中国药学杂志

OBJECTIVE: To distinguishing the Bupleurum formulae granules from different origins and promoting the consistency of quality of them,with the analysis of fingerprint profiles and chemometric methods. METHODS: In the study, a method of dual-wavelength (210 & 254 nm) HPLC fingerprint of Bupleurum formulae granules was developed. Based on the fingerprint method, the quality differences of the samples of Bupleurum formulae granules were analyzed by similarity analysis, hierarchical cluster analysis, principal component analysis and partial least squares discriminant analysis. RESULTS: The fingerprints of 12 batchs of Bupleurum formulae granules are established. The results indicated that the fingerprints of Bupleurum formulae granules from different origins could be differentiated well and the characteristic peaks could be marked. CONCLUSION: The 210 nm fingerprints of Bupleurum formulae granule have the most characteristic differences; It is recommended that the dual-wavelength fingerprints could be combined for analysis and application. Saikosaponin a, saikosaponin b2 and saikosaponin b1 are the key components of the quality difference, which should be focused on in the quality evaluation and control of Bupleurum formula granules.

Identification of different Bupleurum varieties based on carbohydrate-specific chromatograms / 药学学报

Characterization of the polysaccharides and monosaccharides of Bupleurum chinense was undertaken to identify differences in the Bupleurum chinense's sugar profiles, so as to provide a basis for the identification of different varieties. High performance liquid chromatography (HPLC) was used to generate chromatograms of the total polysaccharides of Bupleurum using an Evaporation Light Detector (ELSD), and a monosaccharide chromatogram was generated using a UV-detector (UV) following polysaccharide derivatization. The data were analyzed using SIMCA software and SPSS software to distinguish different varieties of Bupleurum. The results show that the yield of polysaccharides from Bupleurum falcatum is the highest, while the yield of polysaccharides from Bupleurum chinense is the lowest. The polysaccharide spectrum shows that the molecular weights of the polysaccharides in different Bupleurum differ, and their percentages of the total peak area are also different. The four Bupleurum polysaccharides are composed of mannose, glucuronic acid, rhamnose, galacturonic acid, glucose, galactose, and arabinose, but differ in length. The ratio of glucose to arabinose in Bupleurum chinense, Bupleurum scorzonerifolium, Bupleurum falcatum and Bupleurum marginatum var. stenophyllum is: 3.0-4.0, 5.5-7.0, 12.0-17.0, 9.0-12.0. In this study, a sugar profile technique was developed to provide a new method for the identification of different varieties of Bupleurum.

Identification of Bupleurum marginatum var. Wall. ex DC stenophyllum (Wolff) Shan et Li and its analogues based on ITS2 barcode / 中草药

Objective: DNA barcoding technology, a molecular identification method, is applied to distinguish Bupleurum marginatum var. stenophyllum from its analogues in order to ensure the quality and clinical curative effect. Methods: In this study, the internal transcribed spacer 2 (ITS2) regions of 50 samples were amplified by PCR and sequenced bi-directionally. Obtained sequences were assembled using CodonCode Aligner. The genetic distances were computed by MEGA 6.0 in accordance with the kimura 2-parameter (K2P) model and the phylogenetic tree was constructed by Neighbor-joining (NJ) method. Moreover, the secondary structure of ITS2 was predicted using ITS2 database websites. Results: The intra-specific genetic distances were smaller than inter-specific ones in ITS2 regions of B. marginatum var. stenophyllum. The NJ tree and secondary structure results could distinctly differentiate B. marginatum var. stenophyllum and its analogues. Conclusion: ITS2 sequence can scientifically and reliably identify the authenticity of B. marginatum var. stenophyllum and could provide more new and reliable techniques to ensure clinical safety of this traditional Chinese medicine.

Effects of soil factors on saikosaponin content of Bupleurum chinense in different habitats / 中草药

Objective: Soil is an important factor affecting the formation and accumulation of active ingredients in Chinese medicinal herbs. Taking 29 sample wild and cultivated Bupleurum chinense of 11 areas as study materails, using mathematical statistical analysis methods to explore the relationship between saikosaponin accumulation and soil factors in order to improve the quality of B. chinense by soil ecological regulation. Methods: HPLC analysis of the contents of saikosaponins a, c, d, e, and f from different habitats; pH, organic matter, conductivity, soil water, total nitrogen, alkaline nitrogen, available phosphorus, available potassium, effective calcium, effective magnesium, effective iron, effective copper, and effective manganese were determined by conventional soil physicochemical property assay methods, cluster analysis method was used to analyze the content of saikosaponin from different habitats, analysis of the relationship between soil factors and the content of saikosaponin by Pearson correlation analysis, analysis of soil factors using principal component analysis. Results: Determination of saikosaponin showed that higher content of saikosaponin in Henan province (habitat 5, habitat 6), total content of saikosaponin a and saikosaponin d in habitat10 was 5.16 times the national standard.Cluster analysis of B. chinense from different origins, according to the content of saikosaponin, the B. chinense from 29 habitats were grouped into three types. Pearson correlation analysis of saikosaponin content and soil factors that the organic matter in the soil was significantly positively correlated with saponin a and total saponins (P < 0.05), there was a significant positive correlation between available zinc and available iron and each saikosaponin (P < 0.05) properly improve the organic matter, effective zinc, effective ironcontent in the soil can promote the accumulation of saikosaponin.Analysis of the principal components of soil from different habitats, the higher score was habitat 9, habitat 10, habitat 11, habitat 14, habitat 19, habitat 28. This is basically consistent with the results of cluster analysis of saikosaponin content. Based on the analysis of the main soil indicators of B. chinense from different habitats that in a certain range, the higher organic and effective zinc content, the more favorable were the accumulation of saikosaponins. This was basically consistent with the results of Pearson correlation analysis. Conclusion: These results indicated enhancement of organic matter, effective zinc, in the soil can improve the saikosaponin content in cultivated B. chinense.

Prescription regularity of traditional Chinese medicine for treating alcoholic liver disease based on traditional Chinese medicine inheritance support system / 中草药

Objective: To summarize the prescription rules of traditional Chinese medicine (TCM) for treating alcoholic liver disease (ALD) based on TCM inheritance support system (V2.50). Methods: The literatures about TCM prescriptions for treating ALD were collected from CNKI, Wanfang database, and VIP database. The TCM inheritance platform system was used to analyze the prescription rules of TCM in the treatment of alcoholic liver disease. Results: Statistics showed that the majority of prescriptions were used to treat alcoholic fatty liver, alcoholic hepatitis, and alcoholic cirrhosis. Through "frequency statistics" analysis, 107 prescriptions were found involving 149 flavors of TCM, with a cumulative frequency of 1 195 times. Twenty-three Chinese medicines with a frequency of ≥ 15 times were used, and the cumulative frequency was 737 times (62%). The most frequently used medicines were blood-activating and stasis-removing drugs, water-diffusing and damp-permeating drugs, tonics, heat-clearing drugs, antialcoholic poisons and qi-regulating drugs. The commonly used doses of Salvia miltiorrhiza, Poria cocos, Bupleurum chinense, Glycyrrhiza uralensis, Atractylodes macrocephala, Alismatis Rhizoma, and Curcumae Radix in the top 10 medicines ranked in the frequency of medication accorded with the prescribed doses in the Pharmacopoeia of the People's Republic of China (2015 edition), while Crataegi Fructus, Artemisiae Scopariae Herba, and Puerariae Lobatae Radix exceeded the prescribed doses. In the frequency analysis of drug pairs, the combination of S.miltiorrhiza and B. chinense was the most widely used. According to the association rules of drug combination, the correlation between Curcumae Radix and S. miltiorrhiza was the strongest, that was, the probability of S. miltiorrhiza appearing with the emergence of Curcumae Radix was 88%. From the network display chart, it was indicated that S. miltiorrhiza and P. cocos were the main herbs for treatment. Through unsupervised entropy hierarchical clustering algorithm, 14 core combinations for new clustering were extracted, and seven new prescriptions can be obtained by further clustering. Conclusion: The basic principles of TCM treatment of ALD include promoting blood circulation and removing blood stasis, removing dampness, tonifying, detoxifying alcohol, and promoting qi, and with "protecting spleen and stomach function" as its purpose, which accords with the theoretical basis of traditional Chinese medicine in treating alcoholic liver disease. Core combinations and new prescriptions provide references for clinical drug use and new drug research and development, but new prescriptions must be further evaluated with the combination of traditional Chinese medicine theory and clinical practice.

Study on basis of liver toxicity of decoction Bupleurum chinense based on integrated model of spectrum toxicity relationship and liver toxicity network / 中草药

Objective: To study the HPLC fingerprint of Bupleurum chinense and the relationship between the spectrum and the toxicity in vitro, construct the network of the liver toxicity induced by B. chinense, and integrate the spectrum and the network toxicology to predict the material basis of the liver toxicity induced by B. chinense. Methods: The fingerprints of 10 batches of decoction of B. chinense were established by HPLC, the contents of ALT and AST in L02 cells were determined by decoction of B. chinense, and the toxic components of liver were preliminarily determined by the method of grey correlation analysis. Combined with network toxicology, the candidate components of hepatotoxicity were predicted. Integrated analysis on the components of liver toxicity induced by decoction of B. chinense. Results: The established fingerprint of B. chinense was calibrated with 29 common peaks. The results of grey correlation analysis showed that common peaks 13, 12, 8, 26, 10, 14, 27 and 11 had a high correlation degree with the ALT content of L02 cells. The common peaks of 12, 13, 26, 8, 10, 14, 27 and 11 had a high correlation degree with the AST content of L02 cells. According to the network toxicology, 17 components of B. chinense, stigmasterol, baicalin, etc were speculated to be the hepatotoxicity components of B. chinense. The integrated analysis initially determined that the hepatotoxicity components of B. chinense were 13, 12, 8, 26, 10, 14, 27 and 11 peaks in total, and stigmasterol, baicalin, saikosaponin D, etc. Conclusion: The hepatotoxicity of B. chinense is the result of the interaction of various components. This study can provide data support for the further research on the material basis of B. chinense hepatotoxicity.

Correlation between content of saikosaponin and expression of key enzyme genes in different parts of Bupleurum chinense / 中草药

Objective: To select reference genes suitable for the expression analysis of Bupleurum chinense, and analyze the relationship between the content of saikosaponin and the gene expression of key enzymes in different tissues of B. chinense. Methods The roots, stems, leaves and fruits of B. chinense were used as test materials, and five commonly used internal reference genes of Actin, α-tubublin, β-tubulin, Cyclophilin and EF-1α were selected as candidates by real-time quantitative PCR. Based on the selected internal reference gences, tissue expression pattern of ACAT, FPS, HMGR, IPPI, PMD, PMK, SE, SS, β-AS, UGT1, UGT3, UGT6, UGT8, UGT10, P450-7 and P450-12 genes in B. chinense was analyzed. The content of saikosaponin a, saikosaponin c and saikosaponin d were determined by HPLC, and correlation analysis was performed by SPSS. Results:The EF-1α gene with the best stability in the five candidate genes (EF-1α, Cyclophilin, Actin, β-tubulin, α-tubublin) was selected as the internal reference gene. The expression levels of 16 key enzymes in the roots of B. chinense were measured. The results showed that ACAT, PMK, IPPI, SS, SE, UGT1, UGT3, UGT6, and UGT8 were the highest in the aboveground parts, the levels of HMGR, β-AS, P450-7 and P450-12 were higher in the roots than those in the aboveground parts, but PMD, FPS and UGT10 were not significantly different in the tissues. The content of saponins in the root was much higher than that in the aerial parts (stem, leaf and fruit) by HPLC. The results of correlation analysis showed that 16 key enzyme genes in the upstream ACAT, HMGR, PMD, SE and so on were significantly correlated with downstream P450-7, P450-12, UGT3, UGT6 and UGT8 (P < 0.05). It showed that the key enzyme genes were closely related to each other and regulated the synthesis of saikosaponin in common. The correlation analysis between the 16 key enzyme genes and the content of saikosaponin showed: HMGR, P450-7, p450-12 and the total of three saponins were significantly positively correlated (P < 0.01), and β-AS was significantly correlated with the total content of three saponins (P < 0.05), and HMGR, P450-7, P450-12, and β-AS were significantly correlated with the monomer saponins a, c, d (P < 0.05). These four genes jointly regulated the synthesis of saikosaponin and had an important effect on the accumulation of saponin. Conclusion:The HMGR, β-AS, P450-7 and P450-12 genes in the saikosaponin synthesis pathway have a consistent distribution in saikosaponin synthesis and play an important role in the regulation of saponin synthesis.

Analysis of tissues difference of flavonoids and key enzyme genes expression in Bupleurum chinense and B. scorzonerifolium / 中草药

Objective To investigate the relationship between the content of flavonoids and the key enzyme genes expression in different tissues of Bupleurum chinense and B. scorzonerifolium. Methods The roots, stems, leaves, and fruits of B. chinense and B. scorzonerifolium were used as test materials, determination of flavonoids (rutin, quercetin, kaempferol, and isorhamnetin) in different tissues by HPLC, determination of total flavonoids by UV spectrophotometry, the tissues expression of key enzyme genes (IFS, F3H, and DFR) in flavonoids synthesis was determined by real-time quantitative PCR, correlation analysis was performed with SPSS. Results The content of flavonoids in the aerial parts (stems, leaves, and fruits) of B. chinense and B. scorzonerifolium was significantly higher than that in roots, the content of flavonoids was mainly rutin, and the content of rutin in the leaves of B. chinense leaves was up to 106.961 mg/g; The distribution of total flavonoids in B. chinense and B. scorzonerifolium was obviously different, the content was from high to low: leaves ≥ fruit > stem > root; The expression of B. chinense IFS, F3H, and DFR gene in the aerial parts was much higher than that in roots, IFS gene was significantly positive correlated with rutin (P < 0.05), F3H gene was significantly positive correlated with DFR gene (P < 0.05), but the expression of IFS, F3H, and DFR gene in each tissues of B. scorzonerifolium was at lower level. Conclusion The content of flavonoids in different parts of B. chinense and B. scorzonerifolium was consistent with the expression of flavonoids synthesis key enzyme genes, the differential expression of key enzyme genes regulates the synthesis and accumulation of flavonoids in different tissues.

Discussion on Quality Control Methods for Chinese Patent Medicines Containing Bupleurum / 中国药学杂志

There are many problems in the use of Bupleurum medicinal materials such as confusion in variety, unclearness in origin and quality problems, e.g., adulteration of authentic Bupleurum and mixing of non-medicinal parts. Therefore, the quality of Chinese patent medicines containing Bupleurum chinense is variable, and it is necessary to establish targeted quality control methods. Based on the quality control methods of Bupleurum and its Chinese patent medicines, combined with the example of Xiaochaihu granules, this paper discussed the quality standard improvement and supplementary test methods of Bupleurum and its Chinese patent medicines, aiming to provide reference for the quality control, quality supervision and standardized production of Chinese patent medicines containing Bupleurum and promote the improvement of its intrinsic quality.

Academic thought and clinical experience of professor Chen Qiaolin in diagnosis and treatment of acute pancreatitis / 中国中西医结合急救杂志

Acute pancreatitis (AP) is an acute inflammatory and cell-damaging process of the pancreas. In this paper, using the AP onset location and its characteristics combined with review of the ancient and modern literatures and summary of the clinical practices, the professor Chen Qiaolin's academic ideas and clinical experiences in treating AP are introduced and demonstrated. Dr. Chen believes that the development of AP is a dynamic process, the key point is to grasp the Shaoyang syndrome complicated with Yangming, the internal administration of the self-modified Dachaihu decoction is the basic therapy accompanied by applying paste-like Traditional Chinese Medicine TCM attached on the acupoints and acupuncture for comprehensive treatment of AP, the therapeutic result is confirmed and its summary is as follows.