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We performed an extensively targeting metabolomic detecting using ultra-performance liquid chromatography-mass spectrometry (UPLC-MS/MS) to compare the secondary metabolites in Dang shen [Codonopsis pilosula (Franch.) Nannf.] from Shanxi and Gansu provinces. The findings showed that 161 secondary metabolites in 6 groups (phenolic acids, flavonoids, lignans and coumarins, alkaloids, terpenoids, others) were found from Dang shen in Changzhi city of Shanxi province and Dingxi city of Gansu province. There were 98 secondary metabolites which is differed significantly. In comparison to Dingxi city, 33 different secondary metabolites of Dang shen in Changzhi city had a greater relative content, whereas relative content of 65 different metabolites in Dingxi city was higher. Metabolic pathway enrichment analysis revealed that phenolic acids and flavonoids were significantly different in the secondary metabolites of Dang shen from different producing places. This may be one of the reasons for the difference in the quality of Dang shen in Shanxi and Gansu provinces. This work compared and analyzed the secondary metabolites of Dang shen from Dingxi city in Gansu province and Changzhi city in Shanxi province for the first time, which lays the foundation for further study on the quality of Dang shen.
ABSTRACT
Lu Dangshen, a traditional authentic medicinal material of Codonopsis Radix is mainly produced in Shangdang (Changzhi) area of Shanxi Province. Baitiao Dangshen is mainly produced in Gansu Province. Codonopsis Radix contains many kinds of components such as phenylpropanoids, polyalkynes, alkaloids, terpenes, fatty acids, flavonoids, and so on. At present, the effect of producing areas on its chemical compositions has not been systematically studied. This study analyzed the differences of metabolites among Codonopsis pilosula from different producing areas by UPLC-HRMS. PCA, OPLS-DA coupled with Thermo mzcloud online and local databases were used to compare the overall differences of metabolites among Codonopsis pilosula from different producing areas, and the chemical constituents were identified to further screen and find out the different metabolites and analyze the metabolic pathways by information retrieval in HMDB, PubChem, Chemspider and KEGG databases. The results showed that 72 differential metabolites were identified in this study. There were 15 kinds of up-regulated and 57 kinds of down-regulated metabolites of Lu Dangshen compared with Baitiao Dangshen. The top 30 metabolic pathways were analyzed by KEGG enrichment, and the most important metabolic pathways were phenylpropanoid biosynthesis, which was demonstrated that phenylpropanoid biosynthesis pathway and related intermediate metabolites could be used as the characteristics of distinguishing Lu Dangshen from different habitats of Codonopsis pilosula. The present study provided a basis for analyzing the influence of producing areas on the chemical components of Codonopsis pilosula and reasonably evaluating the quality of Codonopsis Radix, and also provided a new idea for expounding the authenticity of Lu Dangshen.
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Objective: To study the chemical constituents of Codonopsis pilosula. Methods: The chemical constituents were isolated and purified by AB-8 macroporous adsorption resin, silica gel, and Sephadex LH-20 column chromatography. The structures of the isolated compounds were identified on the basis of spectroscopic methods including 1H-NMR, 13C-NMR, CD, and MS spectra. Results: Seven compounds were identified as (-)-(8R, 9R, 2E, 6E, 10E)-tetradeca-2, 6, 10-triene-4-yne-8, 14-diol-9-β-D-glucopyranoside (1), phenyl-β-D-glucoside (2), (E)-isoconiferin (3), coniferin (4), syringin (5), apigenin (6), and adenosine (7). Conclusion: Compound 1 is a new compound, named lobetyolin A, and compounds 2 and 3 are all isolated from this plant for the first time.
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Objective: To study the correlation between monosaccharide compositions and their in vitro cytotoxic activities. Methods: Twenty-six Codonopsis pilosula polysaccharides (CPPs) were extracted by water extraction and alcohol precipitation method from C. pilosula collected from 26 different habitats. Total carbohydrate contents of the polysaccharides were determined by the phenol-sulfuric acid colorimetric method. The methods of m-hydroxydiphenyl and GC were all applied to the determination of the galacturonic acid contents in polysaccharides, and the compositional analysis was performed using the aldononitrile acetate method while the fructose contents was analyzed by the trimethylsilyl ether method. In vitro cytotoxic activities against human hepatocellular carcinoma HepG2 cells of the 26 batches of CPPs were evaluated by MTT. Then, hierarchical clustering analysis (HCA) was used to study the classification of 26 batches of C. pilosula and partial least squares (PLS) was used to investigate the correlation between the monosaccharide compositions and in vitro cytotoxic activities of the polysaccharides. Results: The results showed that all the 26 batches of CPPs possessed the cytotoxic activities against HepG2 cells. The 8th CPPs sample, which was extracted from the C. pilosula collected in Wen county (Gansu province, China), had the highest growth inhibitory rate on HepG2 cells and the inhibitory rate was 36.36%. In addition, the monosaccharide contents of the 26 batches of CPPs were different from each other. The results of HCA showed that the C. pilosula could not be classified according to the monosaccharide compositions. The PLS results indicated that galacturonic acid, arabinose, rhamnose, galactose, and fructose were correlated positively with the cytotoxic activities, while mannose, xylose, and glucose were correlated negatively with the cytotoxic activities. Conclusion: The in vitro cytotoxic activities of CPPs are highly correlative to their monosaccharide compositions, and the CPPs containing much galacturonic acid have the obviously cytotoxic activities.
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Objective: To explore the effect of different treatment on Codonopsis pilosula seed germination and seedling growth, and then find out the best treatment method. Methods: C. pilosula was pretreated by different methods, culture was carried out by thermostatic cultivation, and C. pilosula seed germination and seedling growth (30 d) were observed, then the data were analyzed with statistic. Results: Temperature and soil moisture had the significant impact on C. pilosula seed germination and seedling growth, the optimum temperature was 20-30℃, optimum humidity was 45%-75%. C. pilosula seed germination and seedling growth could be effectively promoted by potash treatment, chemicals at suitable concentration, and growth regulators. However, the growth regulators could change the seedling morphological characteristics, C. pilosula seed germination and seedling growth fast with potash treatment, plant was normal and has less disease. Conclusion: Potash treatment is beneficial to seed germination and seedling growth, so as to effectively improve the seedlings yield and quality. Potash treatment could be widely used in C. pilosula field seedling production.
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Objective: To rapidly and largely identify the functional simple sequence repeat (SSR) in transcriptome dataset of Codonopsis pilosula based on bioinformatics analysis. Methods: MicroSAtellite (MISA) sofeware was used to analyze the distributing frequency of high-flux transcriptome SSR and the basic characteristics of repeat motifs. Primers were designed using Primer3, and SSRFinder was used to check SSR and screen the SSR primers. Results: SSR loci (7 327) were obtained from 45 511 unigenes, distributed in 6 017 unigenes (12.22%). There are 415 kinds of repeat motifs existing in C. pilosula transcriptome. On average, SSRs occurred every 4 520 bp in length. Dinucleotid repeats predominated with an occurrence frequency of 58.67%, and AG/CT was the most frequent one among all the repeat types. A total of 4 329 primer pairs were designed for marker development. Conclusion: The large number of SSR genetic markers developed in the present study should contribute greatly to research into genetic diversity and germplasm characterization in C. pilosula.
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Objective To study the process of the root development of Codonopsis pilosula as well as theaccumulatinglocationsofthemainmedicinalcompositionandstoragenutritionintheroot. MethodsThe development process of the roots was studied by paraffin section methods. Storage nutrition was loca-lized by PAS reaction and Sudan Ⅲ reaction. The medicinal composition of C. pilosula, terpene and steroid were stained with mix reagent of 5% vanillin-acetic acid and perchloric acid. Results The development process of the root is shown as four stages: promeristem, primary meristem, primary structure, and secondary growth. Abundant laticifers exist in mature root and the prime laticifers could be found in phloem as early as the first vessel of primary xylem appears. Then, the laticifers exist in phloem at all time. Starch granule accumulated in mature root as reserved substance. Terpene and steroid accumulated in parenchyma and laticifer. Conclusion Parenchyma and laticifer in the roots of C. pilosula are the main parts of the roots to store medicinal composition and storage nutrition.