RESUMO
Acacetin, a flavonoid compound, possesses a wide range of pharmacological effects, including antimicrobial, immune regulation, and anticancer effects. Some key steps in its biosynthetic pathway were largely unknown in flowering plants. Here, we present the first haplotype-resolved genome of Chrysanthemum indicum, whose dried flowers contain abundant flavonoids and have been utilized as traditional Chinese medicine. Various phylogenetic analyses revealed almost equal proportion of three tree topologies among three Chrysanthemum species (C. indicum, C. nankingense, and C. lavandulifolium), indicating that frequent gene flow among Chrysanthemum species or incomplete lineage sorting due to rapid speciation might contribute to conflict topologies. The expanded gene families in C. indicum were associated with oxidative functions. Through comprehensive candidate gene screening, we identified five flavonoid O-methyltransferase (FOMT) candidates, which were highly expressed in flowers and whose expressional levels were significantly correlated with the content of acacetin. Further experiments validated two FOMTs (CI02A009970 and CI03A006662) were capable of catalyzing the conversion of apigenin into acacetin, and these two genes are possibly responsible acacetin accumulation in disc florets and young leaves, respectively. Furthermore, combined analyses of ancestral chromosome reconstruction and phylogenetic trees revealed the distinct evolutionary fates of the two validated FOMT genes. Our study provides new insights into the biosynthetic pathway of flavonoid compounds in the Asteraceae family and offers a model for tracing the origin and evolutionary routes of single genes. These findings will facilitate in vitro biosynthetic production of flavonoid compounds through cellular and metabolic engineering and expedite molecular breeding of C. indicum cultivars.
Assuntos
Chrysanthemum , Evolução Molecular , Flavonas , Genoma de Planta , Filogenia , Proteínas de Plantas , Chrysanthemum/genética , Chrysanthemum/metabolismo , Chrysanthemum/enzimologia , Flavonas/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Genoma de Planta/genética , Haplótipos , Diploide , Flavonoides/metabolismo , Flavonoides/biossíntese , Flores/genética , Flores/enzimologia , Flores/metabolismo , Metiltransferases/genética , Metiltransferases/metabolismoRESUMO
BACKGROUND: Isatis tinctoria Linnaeus and Isatis indigotica Fortune are very inconsistent in their morphological characteristics, but the Flora of China treats them as the same species. In this work, a new technology that differs from conventional barcodes is developed to prove that they are different species and to clarify their classification. RESULTS AND METHODS: I. indigotica was indistinguishable from I. tinctoria when using ITS2. CPGAVAS2 was used to construct the chloroplast genomes. MAFFT and DnaSP were used to calculate nucleotide polymorphism, the chloroplast genomes of the two have high diversity in the rpl32 ~ trnL-UAG short region. When using this region as a mini barcode, it was found that there are obvious differences in the base numbers of I. tinctoria and different ploidy I. indigotica were found, but diploid and tetraploid I. indigotica had the same number of bases. Moreover, the reconstruction of the maximum likelihood (ML) tree, utilizing the mini-barcode, demonstrated that I. tinctoria and both diploid and tetraploid I. indigotica are located on distinct branches. The genome size of tetraploid I. indigotica was approximately 643.773 MB, the heterozygosity rate was approximately 0.98%, and the repeat sequence content was approximately 90.43%. This species has a highly heterozygous, extremely repetitive genome. CONCLUSION: A new method was established to differentiate between I. indigotica and I. tinctoria. Furthermore, this approach provides a reference and basis for the directional breeding of Isatis.
Assuntos
Genoma de Cloroplastos , Isatis , Isatis/genética , Tetraploidia , Melhoramento Vegetal , ChinaRESUMO
This study explored the correlation between color and chemical components of Chrysanthemi Indici Flos(CIF), aiming at providing a reference for its procurement, evaluation, and breeding. Colorimeter and ultra-performance liquid chromatograph(UPLC) were used to determine the color(lightness-shade chromaticity value L~*, red-green chromaticity value a~*, yellow-blue chromati-city value b~*) and chemical components(cynaroside, linarin, luteolin, apigenin, and chlorogenic acid) of 84 CIF germplasms, respectively. Diversity analysis, correlation analysis, regression analysis, and cluster analysis were performed. The results showed that the color and chemical components of CIF were diversified. Chlorogenic acid was in significantly positive correlation with L~* and b~* and significantly negative correlation with a~*. Cynaroside and grey relational grade γ_i of chemical components were in significantly po-sitive correlation with b~* and L~*, respectively, whereas linarin, luteolin, and apigenin had no significant correlation with L~*, a~*, or b~*. The 84 CIF germplasms were clustered into 4 clades. In addition, germplasms in clade ⠢ had higher γ_i and total color value(E~*_(ab)) than those in other clades, with the best quality and color, and a germplasm with the highest quality, bright yellow color, and highest content of linarin was screened out in this clade. Thus, CIF with bright yellow color had high content of cymaroside and chlorogenic acid and thereby high quality. In summary, the color can be used to quickly predict the quality of CIF. Our results provided data for the evaluation of CIF quality by color and a reference for its procurement and breeding.
Assuntos
Chrysanthemum , Apigenina/análise , Ácido Clorogênico/análise , Cromatografia Líquida de Alta Pressão/métodos , Chrysanthemum/química , Luteolina/análise , Melhoramento VegetalRESUMO
Chrysanthemi Indici Flos(CIF), the capitulum of Chrysanthemum indicum, is widely used in proprietary Chinese medicine and daily chemical products. At present, CIF is mainly produced from wild resources and rarely cultivated. This study aims to reveal the correlations between linarin content in CIF and climatic factors in different habitats, and provide a theoretical basis for suitable zoning and rational production of medicinal materials. The content of linarin in CIF was determined by HPLC. Grey relational analysis and Pearson correlation analysis were carried out for linarin content with climatic factors. The results showed that the content of linarin in CIF was significantly different among different habitats. The grey relational degrees of climatic factors with linarin content was in an order of average annual precipitation>annual average sunshine hours>annual average temperature>longitude>annual frost-free period>latitude>altitude. Longitude, annual average temperature and average annual precipitation had significantly positive correlations with the content of linarin in CIF, whereas latitude and altitude showed negative correlations with it. The annual frost-free period and annual average sunshine hours had no significant correlation with the content of linarin in CIF. The content of linarin in CIF varied significantly in different habitats. High longitude, low latitude, low altitude, high annual average temperature and high annual average precipitation could be used as indicators for the habitats of high-quality Ch. indicum. This study provides a reference for selecting suitable producing areas of Ch. indicum and establishing artificial cultivation system.
Assuntos
Chrysanthemum , Cromatografia Líquida de Alta Pressão , Ecossistema , GlicosídeosRESUMO
To provide theoretical basis for the formation of Chrysanthemum indicum resource and quality,the differences in biology,ecology,yield and quality of Ch. indicum population in the wild tending,the cultivated and the wild were analyzed and compared in this study. The results showed that in the aspect of biology,there was no significant difference between the condition of wild tending and cultivated Ch. indicum at the height of plant,both were significantly higher than wild Ch. indicum. In the aspect of ecology,in the condition of wild tending is less likely to occur water logging than cultivated Ch. indicum. In the condition of wild tending is less likely to occur plant diseases and insect pests than Ch. indicum in the condition of wild. In the aspect of production,the average output of the sample of the condition of wild tending was significantly higher than that the condition of artificial cultivation and wild quadrat. There was no significant difference between the three kinds of condition on the traits of capitulum dichasium,the content of buddleoside of the wild tending Ch. indicum was significantly higher than that of 0. 80% required by the Chinese Pharmacopoeia( 2015 edition). Therefore,the artificial supplement and the appropriate artificial management of wild tending is a mode of production to increase the population density of Ch. indicum and obtain high quality and high yield medicinal materials.
Assuntos
Chrysanthemum , ÁguaRESUMO
The Cd stress of Chrysanthemum indicum was treated by different concentrations of Cd Cl2 solution in the culture substrate. The content of Cd in different parts of Ch. indicum and the content of buddleoside and the total flavonoids in Ch. indicum were determined. The absorption characteristics of Cd elements in Ch. indicum were analyzed. And the influence of Cd elements on the quality of the herbs. The results showed that the application of soil Cd in the range of 0-100 mg·kg~(-1) had no significant effect on the biomass of Ch. indicum,and the root-shoot ratio showed a decreasing-increasing-decreasing trend. The content of Cd in different parts of Ch. indicum was significantly different,and the content of aboveground part was higher than that of underground part. The enrichment factors of Cd elements in different parts of Ch. indicum are different. The enrichment coefficient of aboveground parts is larger than that of underground parts. The whole parts and plants show an increase first and then decrease,and the overall enrichment factor is greater than1. The transfer coefficient of the aerial part/underground part of Ch. indicum showed a decreasing-increasing-decreasing-increasing trend with the increase of the amount of Cd applied in the soil,and the transfer coefficient was higher than 1. The contents of buddleoside and total flavonoids in Ch. indicum after Cd stress treatment were lower than the control,and the overall performance was lower and then increased,but it was still significantly lower than the control,indicating that Cd pollution directly led to the decrease of chemical quality of Ch. indicum.
Assuntos
Chrysanthemum , Cádmio , Flavonoides , Solo , Poluentes do SoloRESUMO
BACKGROUND: Chrysanthemum indicum L., an important ancestral species of the flowering plant chrysanthemum, can be used as medicine and for functional food development. Due to the lack of hereditary information for this species and the difficulty of germplasm identification, we herein provide new genetic insight from the perspective of intraspecific transcriptome comparison and present single sequence repeat (SSR) molecular marker recognition technology. RESULTS: Through the study of a diploid germplasm (DIWNT) and a tetraploid germplasm (DIWT), the following outcome were obtained. (1) A significant difference in Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) annotations for specific homologous genes was observed using the OrthoMCL method for the identification of homologous gene families between the two cytotypes. Ka/Ks analysis of common, single-copy homologous family members also revealed a greater difference among genes that experienced positive selection than among those experiencing positive selection. (2) Of more practical value, 2575 SSR markers were predicted and partly verified. We used TaxonGap as a visual tool to inspect genotype uniqueness and screen for high-performance molecular loci; we recommend four primers of 65 randomly selected primers with a combined identification success rate of 88.6% as priorities for further development of DNA fingerprinting of C. indicum germplasm. CONCLUSIONS: The SSR technology based on next-generation sequencing was proved to be successful in the identification of C. indicum germplasms. And the information on the intraspecfic genetic divergence generated by transcriptome comparison deepened the understanding of this complex species' nature.
Assuntos
Chrysanthemum/genética , Repetições de Microssatélites/genética , Transcriptoma , Análise por Conglomerados , Perfilação da Expressão Gênica , Genes de Plantas , Marcadores Genéticos , Variação Genética , Anotação de Sequência Molecular , Ploidias , RNA de Plantas/química , RNA de Plantas/genética , RNA de Plantas/metabolismo , Análise de Sequência de DNA , Especificidade da EspécieRESUMO
Three new triterpenoid saponins, namely asprellinoids A-C (1-3), featuring a sulfate substitution in sugar moiety, were isolated from the stems of Ilex asprella (Hook. et Arn.) Champ. ex Benth. Their structures were elucidated by the spectroscopic data analyses including HR-ESI-MS, IR, and NMR spectra, and chemical method.
Assuntos
Ilex/química , Caules de Planta/química , Saponinas/química , Triterpenos/química , Espectroscopia de Ressonância Magnética , Estrutura MolecularRESUMO
ICP-MS was used to determine the contents of five heavy metals such as lead (Pb), cadmium (Cd), mercury (Hg), arsenic (As) and copper (Cu) in Chrysanthemum indicum and soil samples. Pearson correlation and path analysis were used to analyze the effects of latitude and longitude and altitude on the content of heavy metal elements in Ch. indicum. To explore the relationship between geographical location of heavy metal content in Ch. indicum and the content of heavy metal elements in soil. The results showed that the Ch. indicum had high enrichment of Cd and had a synergistic effect on the absorption of five heavy metal elements. Elevation directly affected the accumulation of Pb and Cd in Ch. indicum and was a positive effect. The direct positive effect of latitude acts on the accumulation of Hg and Cu in Ch. indicum. The effect of longitude on the Cd content was the negative effect of longitude. The geographical location had different effects on the accumulation of heavy metal elements in Ch. indicum. There was a certain interaction between direct and indirect effects. Ch. indicum had different absorption and enrichment characteristics for different heavy metal species. When controlling the heavy metal content and medicinal material quality of Ch. indicum, we must fully consider the influence of geographical location on the quality of medicinal materials, select suitable geographic regions to carry out acquisitions, and choose a good ecological environment. Regional establishment of Chinese herbal medicine planting bases, or adopting certain agricultural measures to reduce the absorption of heavy metal elements in Ch. indicum herbs, while conducting research on chemical composition, strengthening the research on the effective forms of heavy metals, and paying attention to the safety of medicinal herbs.
Assuntos
Chrysanthemum , China , Metais Pesados , Solo , Poluentes do SoloRESUMO
Phytochemical investigation on Ilex asprella stems by using various chromatographic techniques led to the isolation of 18 phenolic constituents. Based on spectroscopic data analyses and/or comparison of the spectroscopic data with those in literature, these constituents were identified, including two lignans (1, 2), five phenylpropanes (3-7), six chlorogenic analogues (8-13), and five benzoic analogues (14-18). Among them, compounds 3-7, 9, 11, 13, 14, 17, and 18 were isolated from genus Ilex for the first time, and 2, 8, 10, 15, and 16 were isolated from this species for the first time. The in vitro anti-inflammatory assay results showed that compounds 8, 9, 11, 13, and 15 possessed moderate inhibition on the NO production in RAW264.7 cells with IC50 values of 51.1-85.8 µmol·L⻹. The present study brought preliminary reference for the clarification of therapeutic ingredients of I. asprella with anti-inflammatory efficacy and its quality evaluation.
Assuntos
Anti-Inflamatórios/farmacologia , Ilex/química , Fenóis/química , Compostos Fitoquímicos/farmacologia , Animais , Anti-Inflamatórios/isolamento & purificação , Camundongos , Óxido Nítrico/metabolismo , Compostos Fitoquímicos/isolamento & purificação , Caules de Planta/química , Células RAW 264.7RESUMO
Phytochemical investigation on the stems of Ilex asprella by using various chromatographic techniques led to the isolation of 13 compounds. By spectroscopic analyses and comparisons the spectral data with those in literatures, these compounds were identified as salicifoneoliganol(1), rel-(7R,8S)-3,3',5-trimethoxy-4',7-epoxy-8,5'-neolignan-4,9,9'-triol 9-ß-D-glucopyranoside(2),(+)-cycloolivil(3),(+)-syringaresinol-4'-O-ß-D-monoglucoside(4), liriodendrin(5), caffeic acid (6), 3,4-dihydroxy-5-methoxybenzaldehyde(7), benzene-1,2,4-triol(8), 3,4,5-trimethoxyphenyl-1-O-ß-D-apiofuranosyl(1â³â6')-glucopyranoside(9), aeculetin(10), cryptochlorogenic acid ethyl ester(11), chlorogenic acid ethyl ester(12), and rel-5-(3S,8S-dihydroxy-1R,5S-dimethyl-7-oxa-6-oxobicyclo [3,2,1]oct-8-yl)-3-methyl-2Z,4E-pentadienoic acid(13). Among them, compounds 7, 8, 11, and 13 were isolated from genus Ilex for the first time, and 1-3, 9, 10, and 12 were isolated from this speciesfor the first time. The anti-inflammatory assay results of these compounds showed that compounds 1 and 9 showed moderate inhibitory effect against NO production in RAW 267. 4 cells with IC50 values of 35.7 and 50.6 µmolâ¢L⻹, in vitro respectively, whereas compound 10 showed weak inhibition(IC50 value 98.7 µmolâ¢L⻹).
Assuntos
Ilex/química , Compostos Fitoquímicos/farmacologia , Caules de Planta/química , Animais , Anti-Inflamatórios/isolamento & purificação , Anti-Inflamatórios/farmacologia , Camundongos , Estrutura Molecular , Óxido Nítrico/metabolismo , Compostos Fitoquímicos/isolamento & purificação , Células RAW 264.7RESUMO
Objective: To provide the experimental evidence for expansion of medicinal parts of Zanthoxylum nitidum by comparing the effects of anti-gastritis,gastric mucosal protection and gastrointestinal movement promotion of its root and stem. Methods: The pharmacological effects between root and stem of Zanthoxylum nitidum were compared by observing the anti-gastritis effect on rats with chronic superficial gastritis induced by iodoacetamide, evaluating the gastric mucosal protective effect on rats' gastric ulcer induced by stress, indometacin and pylorus ligation test, and investigating gastrointestinal movement promotion effect on mice gastric evacuation and intestinal propelling. Results: Both root and stem of Zanthoxylum nitidum showed effects of relieving the inflammation symptoms of rats' gastric mucosa induced by iodoacetamide, gastric ulcer respectively induced by stress, and presenting a strong inhibition of free acid and pepsin activity in gastric juice. Furthermore stem parts of Zanthoxylum nitidum in promoting gastrointestinal motility even showed better efficacy than root. Conclusion: Stem of Zanthoxylum nitidum has similar effects of anti-gastritis, gastric mucosal protection and gastrointestinal movement promotion with root of Zanthoxylum nitidum.
Assuntos
Raízes de Plantas , Zanthoxylum , Animais , Suco Gástrico , Mucosa Gástrica , Gastrite , Inflamação , Camundongos , Caules de Planta , Ratos , Úlcera GástricaRESUMO
Phytochemical investigation on the roots of Zanthoxylum nitidum led to the isolation of four new unsaturated alkylamides, zanthoxylumamides A-D. Their structures were established via the detailed spectroscopic analyses.
Assuntos
Medicamentos de Ervas Chinesas/isolamento & purificação , Alcamidas Poli-Insaturadas/isolamento & purificação , Zanthoxylum/química , Medicamentos de Ervas Chinesas/química , Estrutura Molecular , Ressonância Magnética Nuclear Biomolecular , Raízes de Plantas/química , Alcamidas Poli-Insaturadas/químicaRESUMO
OBJECTIVE: To provide the scientific evidence for expansion of medicinal parts of Zanthoxylum nitidum by comparing the effects of anti-contusion injury, analgesia and anti-inflammation of its root and stem. METHODS: The pharmacological effects between root and stem of Zanthoxylum nitidum were compared by observing the anti-injury effect in rats with injury struck by hammer. The analgesic effect in mice was evaluated by writhing test and hot plate test, and the anti-inflammatory effect on paw edema induced by carrageenan and granuloma induced by cotton pellet were investigated in rats. RESULTS: Both root and stem of Zanthoxylum nitidum relieved the exterior and histological symptoms of rats' injury legs struck by hammer, decreased the numbers of mice's writhing, enhanced pain threshold of mice on heat plate, inhibited the edema of rats induced by carrageenan, and suppressed the granuloma of rats induced by cotton pellet. CONCLUSION: Stem of Zanthoxylum nitidum has similar effects of anti-contusion injury, analgesia and anti-inflammation with root of Zanthoxylum nitidum.
Assuntos
Analgésicos/farmacologia , Anti-Inflamatórios/farmacologia , Contusões/tratamento farmacológico , Extratos Vegetais/farmacologia , Zanthoxylum/química , Animais , Carragenina , Edema/induzido quimicamente , Edema/tratamento farmacológico , Inflamação/tratamento farmacológico , Camundongos , Dor/tratamento farmacológico , Limiar da Dor , Raízes de Plantas/química , Caules de Planta/química , RatosRESUMO
Chrysanthemi Indici Flos (CIF) is the inflorescence of Chrysanthemum indicum L., which exists in various shades of yellow and has pharmacologically active constituents. It is widely used for medicinal purposes in China, Japan, and South Korea to treat inflammatory diseases. Its external color is usually used to judge its internal quality in trade; however, the correlation between its color and chemical constituents is unknown. Here, we simultaneously determined five phenylpropanoids (neochlorogenic acid, chlorogenic acid, and isochlorogenic acids A, B, and C) and four flavonoids (linarin, luteolin, apigenin, and acacetin) of 70 CIF germplasms using a newly established UPLC method; furthermore, we measured their color parameters (L*, a*, and b*) using a spectrophotometer. Our results showed considerable variations in the bioactive constituent contents and color parameters of CIF. The content of the five phenylpropanoids and the relative correlation degree γi of the nine constituents were positively correlated with color parameters, which could be rapidly predicted based on L* and/or b*. Moreover, we screened out a high-quality germplasm with a high linarin content and bright colors using the hierarchical clustering method. Our results provide comprehensive insight into CIF's quality evaluation process, particularly the methods for procuring high-quality medicinal materials and breeding by color.
Assuntos
Chrysanthemum , Medicamentos de Ervas Chinesas , Melhoramento Vegetal , Flavonoides/análise , Luteolina , Medicamentos de Ervas Chinesas/análise , Apigenina , Flores/química , Cromatografia Líquida de Alta Pressão/métodosRESUMO
There are two source plants for the traditional Chinese medicine Murrayae Folium et Cacumen (MFC) in Chinese Pharmacopoeia, i.e. Murraya exotica L. and M. paniculata (L.) Jack. Herein, a chemical comparison of M. exotica and M. paniculata by high performance liquid chromatography (HPLC) fingerprint analysis coupled with chemometrics and network pharmacology was performed. The main peaks in the fingerprints were identified by liquid chromatography coupled with ion trap/time-of-flight mass spectrometry (LC-IT-TOF-MS) and authenticated by references. The chemometrics results showed that the HPLC fingerprints of these two species were clearly divided into two categories using hierarchical cluster analysis (HCA) and principal component analysis (PCA), and a total of 13 significantly differentiated markers were screened out by orthogonal partial least squares-discriminant analysis (OPLS-DA). However, the following network pharmacology analysis showed that these discriminated markers were found to act via many common targets and metabolic pathways, indicating the possibly similar pharmacological effects and mechanisms for M. exotica and M. paniculata. The above results provide valuable evidence for the equivalent use of these two plants in clinical settings. Moreover, the chromatographic fingerprint analysis coupled with chemometrics and network pharmacology supplies an efficient approach for the comparative analysis of multi-source TCMs like MFC.
Assuntos
Murraya , Cromatografia Líquida de Alta Pressão , Cromatografia Líquida , Espectrometria de Massas , Análise de Componente PrincipalRESUMO
Flos Chrysanthemi Indici (FCI), the flower of Chrysanthemum indicum L., is a common functional food and a well-known traditional Chinese medicine (TCM) for the treatment of inflammatory diseases. Previous studies have revealed that FCI has anti-inflammatory activity, but little is known about its anti-inflammatory chemical profile. In this study, the potential anti-inflammatory constituents of FCI were investigated by ultra-high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS) combined with the network pharmacology approach, and further confirmed on a LPS activated RAW264.7 macrophage model. As a result, a total of forty-two compounds, including thirty-two flavonoids, nine phenolic acids and one sesquiterpene, were identified. Among them, fourteen compounds including eight flavonoids (11, 17, 24, 28, 32, 39, 41 and 42) and six caffeoylquinic acids (3, 4, 5, 13, 15 and 20) were recognized as potential key anti-inflammatory constituents of FCI through network pharmacology analysis, because they accounted for 92% of the relative peak area in the UPLC-Q-TOF/MS chromatogram and acted on 87 of 97 the inflammatory targets of FCI. However, only 16 targets were shared between the flavonoids and caffeoylquinic acids, indicative of both acting on more different targets. Further the anti-inflammatory effects of the fourteen constituents were validated with the decreased levels of NO, TNF-α, IL-6 and PGE2 in RAW264.7 macrophage cells treated with LPS. Our results indicated that both flavonoids and caffeoylquinic acids were responsible for the anti-inflammatory effect of FCI through synergetic actions on multi-targets. Moreover, 3,5-dicaffeoylquinic acid (15), luteolin (24) and linarin (28) were the most important active constituents of FCI and could be selected as chemical markers for quality control of FCI. Overall, the findings not only explore the anti-inflammatory chemical constituents of FCI, but also provide novel insights into the effective constituents and mechanism of TCMs.
Assuntos
Anti-Inflamatórios/farmacologia , Cromatografia Líquida de Alta Pressão , Chrysanthemum/química , Medicamentos de Ervas Chinesas/farmacologia , Flores/química , Espectrometria de Massas em Tandem , Animais , Anti-Inflamatórios/química , Sobrevivência Celular , Medicamentos de Ervas Chinesas/química , Flavonoides/análise , Glicosídeos , Lipopolissacarídeos/efeitos adversos , Luteolina/análise , Medicina Tradicional Chinesa/métodos , Camundongos , Ácido Quínico/análogos & derivados , Células RAW 264.7RESUMO
Encouraged by the in vivo anti-inflammatory effect of aqueous extract of Ilex asprella stems, a further phytochemical investigation on I. asprella stems oriented by the in vitro NO production inhibition in RAW264.7 cells was conducted, which led to the isolation of eight new phenolic constituents, namely asprenols A-H (1-8), together with 12 known ones (9-20). The structures of the new compounds were established by extensive spectroscopic data analyses of HR-ESI-MS, IR, UV, and 1D and 2D NMR, and the absolute configurations were determined by comparison of experimental and calculated ECD analyses. All isolated were evaluated for the inhibition against NO production in RAW 264.7 cells, and several compounds showed moderate inhibitory effect.
Assuntos
Anti-Inflamatórios/farmacologia , Ilex/química , Fenóis/farmacologia , Caules de Planta/química , Animais , Anti-Inflamatórios/isolamento & purificação , China , Camundongos , Estrutura Molecular , Óxido Nítrico , Fenóis/isolamento & purificação , Células RAW 264.7RESUMO
The root of Ilex asprella is a commonly used herb in Southern China, and also constitutes the main raw material of Canton herbal tea. I. asprella is readily contaminated by mildew because of rich nutrients. Aspergillus versicolor producing sterigmatocystin is one of the most common molds that contaminate foodstuffs and medicinal herbs. Previous study on the evaluation of fungal contamination on samples of I. asprella revealed that A. versicolor was the dominant contaminant. In this study, experiments based on response surface methodology combined with central composite design were carried out to determine the optimal storage conditions of I. asprella to minimize the contamination of sterigmatocystin. The herb, manually innoculated with A. versicolor, was stored under different temperatures (20â»40 °C) and humidity (80â»95%) for seven days. The effects of temperature and humidity were evaluated using total saponin, polysaccharide and the sterigmatocystin levels as quality indexes. The results showed that A. versicolor grew quickly and produced large amounts of sterigmatocystin on I. asprella, at humidity ranging from 85% to 90% and temperatures above 26 °C. Meanwhile, total saponin and polysaccharide amounts were reduced significantly. These findings suggested that I. asprella samples should be stored in an environment with humidity and temperature below 85% and 26 °C, respectively, to reduce A. versicolor growth and sterigmatocystin production.
Assuntos
Aspergillus/crescimento & desenvolvimento , Aspergillus/metabolismo , Contaminação de Medicamentos/prevenção & controle , Ilex/química , Ilex/microbiologia , Esterigmatocistina/análise , Armazenamento de Medicamentos , Umidade , Raízes de Plantas/química , Raízes de Plantas/microbiologia , Plantas Medicinais , Polissacarídeos/análise , Saponinas/análise , Esterigmatocistina/metabolismo , TemperaturaRESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Ilex asprella (Hook. et Arn.) Champ. ex Benth. (IA) is a representative medicinal plant from the South of the Five Ridges of China. Its roots (RIA) and stems (SIA) have been traditionally used for the inflammation-related diseases, such as acute and chronic pharyngitis, cough, and sore throats. AIM OF THE STUDY: To evaluate the in vivo anti-inflammatory effects of IA extracts to provide evidence for its traditional use and to enhance the knowledge of the medicinal properties of IA. MATERIALS AND METHODS: Models of xylene-induced ear edema in mice and carrageenan-induced paw edema in rats were used for the pharmacological evaluations. The mice were randomly divided into eight groups (nâ¯=â¯10 per group): a model group, a positive control group [dexamethasone (Dex), 10â¯mg/kg, intragastrically (i.g.)], RIA aqueous extract groups with three dosages (30, 15, and 7.5â¯mg/kg, i.g.), and SIA aqueous extract groups with three dosages (60, 30, and 15â¯mg/kg, i.g.). The rats were randomly divided into eight groups (nâ¯=â¯6 per group): a model group, a positive control group [acetylsalicylic acid (ASA), 300â¯mg/kg, i.g.], RIA groups with three dosages (80, 40, and 20â¯mg/kg, i.g.) and SIA aqueous extract groups with three dosages (160, 80, and 40â¯mg/kg, i.g.). Histological examinations of the ear and paw tissues were observed by hematoxylin-eosin (HE) staining, and neutrophil elastase levels were assessed in ear tissues by immunohistochemical analysis. Serum levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and interleukin-1ß (IL-1ß) were measured by ELISAs, and expression levels of TNF-α, IL-6, and IL-1ß in rat paw tissues were measured by RT-PCR. The signal transduction proteins p65, IκBα, Janus kinase 2 (JAK2), signal transducer and activator of transcription 3 (STAT3), p38 mitogen-activated protein kinases (p38), extracellular regulated protein kinases (ERK1/2), and c-Jun N-terminal kinase (JNK) in the rat paw tissues were investigated by western blot analyses. RIA and SIA were characterized by HPLC and LC-MS analyses, and the components were confirmed by comparison with isolated compounds. RESULTS: Intragastric administration with RIA (30â¯mg/kg) and SIA (60, 30â¯mg/kg) significantly mitigated ear edema in mice. RIA administration at 80 and 40â¯mg/kg reduced paw edema in rats 2â3â¯h after injection. SIA administration with 160â¯mg/kg inhibited paw edema in rats after the injection of carrageenan for 1â4â¯h, and SIA administration at 80â¯mg/kg inhibited paw edema after the injection of carrageenan for 2â4â¯h. Meanwhile, RIA (80, 40â¯mg/kg) and SIA (160, 80â¯mg/kg) reduced inflammatory cell infiltration in the ear and paw tissues and infiltration of neutrophil leukocytes in rat paw tissues. RIA (80, 40, and 20â¯mg/kg) and SIA (160, 80, and 40â¯mg/kg) notably inhibited the increases of TNF-α, IL-6 and IL-1ß in the serum and mRNA expression in the rat paw tissues. RIA (80, 40â¯mg/kg) and SIA (160, 80â¯mg/kg) reduced the p-p65/p-IκBα, p-JAK2/p-STAT3, and p-p38/p-ERK1/2/p-JNK levels in the pathological tissues of the animals. Phenolic acids and triterpenoids likely contributed to the anti-inflammatory activity. CONCLUSIONS: Both RIA and SIA aqueous extracts showed anti-inflammatory effects in vivo in a dose-independent manner (20â80 and 40â160â¯mg/kg, respectively). The underlying mechanisms are mediated by inhibiting the pro-inflammatory cytokines TNF-α, IL-6, and IL-1ß via regulation of the NF-κB, JAK2/STAT3, and MAPK signaling pathways. The present results provided pharmacological evidence that stems are alternative medicinal parts of IA but function at different doses. Additionally, this study supports the use of IA as an anti-inflammatory herbal medicine.