Isolation, characterization and modulatory potentials of β-stigmasterol, ergosterol and xylopic acid from Anchomanes difformis on mitochondrial permeability transition pore in vitro / 中草药·英文版

OBJECTIVE@#Secondary metabolites and polyphenolic compounds from medicinal plants have been demonstrated to have multiple biological functions with promising research and development prospects. This study examined the effect of β-stigmasterol (with ergosterol) and xylopic acid isolated from Anchomanes difformis on liver mitochondrial permeability transition pore (mPTP).@*METHODS@#The compounds were isolated by vacuum liquid chromatography. Mitochondrial swelling was assessed as changes in absorbance under succinate-energized conditions.@*RESULTS@#1H and 13C NMR spectroscopic elucidation of the isolates affirmed the presence of β-stigmasterol with ergosterol (1:0.3) and xylopic acid. The isolates reversed the increase in lipid peroxidation and inhibited the opening of mitochondrial permeability transition pores caused by calcium and glucose. Pharmacological inhibition of mPTP offers a promising therapeutic target for the treatment of mitochondrial-associated disorders.@*CONCLUSION@#Reduction in the activity of calcium ATPase and the expression of Caspase-3 and -9 were observed, suggesting that they could play a role in protecting physicochemical properties of membrane bilayers from free radical-induced severe cellular damage and be useful in the management of diseases where much apoptosis occurs.

Stigmasterol protects human brain microvessel endothelial cells against ischemia-reperfusion injury through suppressing EPHA2 phosphorylation / 中国天然药物

Stigmasterol is a plant sterol with anti-apoptotic, anti-oxidative and anti-inflammatory effect through multiple mechanisms. In this study, we further assessed whether it exerts protective effect on human brain microvessel endothelial cells (HBMECs) against ischemia-reperfusion injury and explored the underlying mechanisms. HBMECs were used to establish an in vitro oxygen and glucose deprivation/reperfusion (OGD/R) model, while a middle cerebral artery occlusion (MCAO) model of rats were constructed. The interaction between stigmasterol and EPHA2 was detected by surface plasmon resonance (SPR) and cellular thermal shift assay (CETSA). The results showed that 10 μmol·L-1 stigmasterol significantly protected cell viability, alleviated the loss of tight junction proteins and attenuated the blood-brain barrier (BBB) damage induced by OGD/R in thein vitro model. Subsequent molecular docking showed that stigmasterol might interact with EPHA2 at multiple sites, including T692, a critical gatekeep residue of this receptor. Exogenous ephrin-A1 (an EPHA2 ligand) exacerbated OGD/R-induced EPHA2 phosphorylation at S897, facilitated ZO-1/claudin-5 loss, and promoted BBB leakage in vitro, which were significantly attenuated after stigmasterol treatment. The rat MCAO model confirmed these protective effects in vivo. In summary, these findings suggest that stigmasterol protects HBMECs against ischemia-reperfusion injury by maintaining cell viability, reducing the loss of tight junction proteins, and attenuating the BBB damage. These protective effects are at least meditated by its interaction with EPHA2 and inhibitory effect on EPHA2 phosphorylation.

Isolamento do estigmasterol do extrato hexânico das folhas de Sphagneticola trilobata Pruski (Asteraceae) / Isolation of stigmaterol from hexanic extract of the leaves of Sphagneticola trilobata Pruski (Asteraceae)

O interesse de pesquisadores e da população geral é bastante evidente, e tem crescido bastante na atualidade, no uso de plantas medicinais. A espécie de interesse deste trabalho, a Sphagneticola trilobata Pruski é uma planta herbácea pertencente à família Asteraceae. Alguns relatos têm mostrado a presença de diversos constituintes químicos nesta planta, principalmente diterpenos. O presente trabalho realizou o estudo fitoquímico de uma fração do extrato hexânico, devido ao baixo rendimento da fração diclorometânica, através da utilização de diversos métodos cromatográficos, na qual isolouse o fitoesterol estigmasterol. Este composto foi confirmado por meio de dados espectroscópicos e de literaturas, e o isolamento de compostos desta classe de substâncias indicam uma nova vertente para o uso medicinal desta planta como um futuro hipocolesterolêmico. Para isto, estudos serão direcionados para o isolamento de fitoesteróis, com posterior realização de ensaios biológicos (AU)

The interest of researchers and the general population is quite evident, and has grown considerably today, in the use of medicinal plants. The species of interest in this work, Sphagneticola trilobata Pruski is a herbaceous plant, belonging to Asteraceae family. Some reports have shown the presence of several chemical constituents in this plant, mainly diterpenes. The present work carried out the phytochemical study of a fraction of the hexane extract, due to the low yield of the dichloromethane fraction, by several chromatographic methods, in which the phytosterol estigmasterol was isolated. This compound has been confirmed by means of spectroscopic and literature data, and the isolation of compounds of this class of substances indicates a new strand for the medicinal use of this plant as a hypocholesterolemic future. For this, studies will be directed to the isolation of phytosterols, with subsequent biological tests (AU)

Feasibility of Xiaochaihu Decoction on fever induced by coronavirus disease 2019 (COVID-19) based on network pharmacology / 中草药

Objective: The aim of this article was to study the potential antivirus and fever reducing mechanisms of Xiaochaihu Decoction (XCHD) on novel coronavirus pneumonia based on network pharmacology and molecular docking method. Methods: Firstly, the potential targets and pathways of XCHD on fever were analyzed using network pharmacology. Compounds and potential targets in XCHD were screened using TCMSP and PharmMapper databases. The targets in fever reducing were identified from OMMI and Genecards databases. The protein-protein interaction network was established by String database to analyze key targets. The gene oncology (GO) analysis and Kyoto encyclopedia of genes and genomes (KEGG) analysis of key targets were also conducted to generate the relative pathways based on DAVID and KOBAS 3.0 databases, respectively. The compound-target-pathway network was established using Cytoscape 3.2.7. In addition, we used molecular docking method to identify the crucial compounds with higher connectivity on SARS-CoV-2 and the angiotensin-converting enzyme 2 (ACE2). ACE2 has been identified as the key target of SARS-CoV-2 entering cells. The possible binding sites of compounds on SARS-CoV-2 and ACE2 were predicted. Results: Network pharmacology analysis indicated that 165 active compounds and 168 relative targets were selected. A total of 7006 targets related to fever were identified. In addition, 141 potential targets of XCH on fever were identified. Totally, 292 GO terms of XCHD on fever and 30 pathways were identified using GO and KEGG analysis. Furthermore, molecular docking indicated that main active compounds in XCHD exhibited higher affinity with both SARS-CoV-2 and ACE2. Beta-sitosterol, stigmasterol, 3’-hydroxy-4’-O-methylglabridin were top three candidates with highest affinity. Conclusion: In summary, our study identified the potential mechanisms of XCHD on fever. Besides, Beta-sitosterol, stigmasterol, 3’-hydroxy-4’-O-methylglabridin could be the key compounds to exert anti-viral effects against SARS-CoV-2. Our prediction also provided the research fields to further study the mechanisms of XCH on SARS-CoV-2 infection in future.

Exploring active compounds of Jinhua Qinggan Granules for prevention of COVID-19 based on network pharmacology and molecular docking / 中草药

Objective: To explore the effective chemical constituents of Jinhua Qinggan Granules for treatment of coronavirus disease 2019 (COVID-19). Methods: The compounds and action targets of eleven herbal medicines in Jinhua Qinggan Granules were collected via TCMSP. The genes corresponding to the targets were queried by the UniProt database, then the “herbal medicine-compound-target” network was established by Cytoscape software. The gene ontology (GO) function enrichment analysis and KEGG pathway enrichment analysis were performed by DAVID to predict their mechanism. Molecular docking was used to analyze the binding force of the core effective compounds in the “herbal medicine-compound-target” network with SARS-CoV-2 3CL hydrolase and angiotensin converting enzyme II (ACE2). Results: The “herbal medicine-compound-target” network contained 154 compounds and 276 targets, and the key targets involved PTGS2, HSP90AB1, HSP90AA1, PTGS1, NCOA2, etc. GO function enrichment analysis revealed 278 items, including ATP binding, transcription factor activation and regulation of apoptosis process, etc. KEGG pathway enrichment screened 127 signaling pathways, including TNF, PI3K/Akt and HIF-1 signaling pathways related to lung injury protection. The results of molecular docking showed that formononetin, stigmasterol, beta-sitosterol, anhydroicaritin and other key compounds have a certain degree of affinity with SARS-CoV-2 3CL hydrolase and ACE2. Conclusion: The effective compounds in Jinhua Qinggan Granules regulate multiple signaling pathways via binding ACE2 and acting on targets such as PTGS2, HSP90AB1, HSP90AA1, PTGS1, NCOA2 for the prevention of COVID-19.

Passiflora mucronata leaves extracts obtained from different methodologies: a phytochemical study based on cytotoxic and apoptosis activities of triterpenes and phytosterols constituents

Cancer is one of the most prevalent diseases worldwide and the natural products could be a source of bioactive compounds. Passiflora mucronata (PM) belongs to a very known vegetal genus, although, there are no studies about cytotoxic activity or isolated compounds. Different extracts from PM were obtained by liquid-liquid partition (P), Soxhlet (Sox) and supercritical fluid (SFE1-5) extraction techniques, being compared concerning their yields, chemical profile and cytotoxicity. The Sox extracts showed the highest yields (6.03%: hexane; 2.51%: dichloromethane) followed by SFE (from 4.34 to 1.63%) and partitions (1.06 and 2.26%). The hexane partition (HP) showed the best cytotoxic activity against K562 cell line (IC50 = 18.72 µg.mL-1). From HP, the following compounds were identified and analysed its cytotoxic activities: β-amyrin (IC50 = 3.92 µg.mL-1), β-sitosterol (IC50 = 3.37 µg.mL-1), stigmasterol (IC50 = 3.31 µg.mL-1) and oleanolic acid. Stigmasterol induced about 75% of K562 total apoptosis. The compounds were tested against MA-104 cell line and the selective index (SI) attributed (SI >10 for all compounds). This indicates good selectivity to K562 cell line at the expense of MA-104. This is the first time, identifying those compounds to PM .

A new triterpenoid and a new steroid from roots of Vladimiria souliei / 中草药

Objective: To study the chemical compositions from the roots of Vladimiria souliei. Methods: The ethanol extract of the roots of V. souliei was isolated by silica gel, Sephadex LH-20, MCI, HPLC, and C18 reverse-phase column chromatographies. Their physical and chemical structures were elucidated by analysis of NMR and MS spectroscopic data, and the absolute configuration of compound 1 was confirmed by Cu-Kα X-ray crystallographic analysis. Results: Three compounds were isolated from the roots of V. souliei, and named as 25-hydroxy-uvaol (1), vlasoudiol (2), and stigmasterol (3), respectively. Conclusion: Compounds 1 and 2 are identified as new compounds, and compound 3 is isolated from the roots of V. souliei for the first time.

Efficacy network and mechanism prediction of Chaihu Longgu Muli Decoction in treatment of essential hypertension / 中草药

Objective: To study the efficacy network and potential mechanisms of effects of Chaihu Longgu Muli Decoction in the treatment of essential hypertension by using network pharmacology methods. Methods: TCMSP, TCMID, and Stitch were used to obtain the components and their corresponding targets. PubMed, CTD, TTD, OMIM, and DrugBank were used to search disease targets of essential hypertension. The common targets between components and disease targets were screened and builded the “compound - target” efficacy network and the protein-protein interaction network by STRING and Cytoscape. The key components and core targets of Chaihu Longgu Muli Decoction in the treatment of hypertension were screened in these networks. Finally, relevant software was applied to GO analysis and pathway analysis of core targets to predict potential mechanisms. Results: A total of 137 components of Chaihu Longgu Muli Decoction and 168 targets of essential hypertension were screened. According to the analysis, quercetin, β-sitosterol, kaempferol, and stigmasterol were found as the four key components and 12 core targets such as IL-6, AKT1, and MAPK8 were found involving the Chaihu Longgu Muli Decoction-induced treatment of essential hypertension. The result of GO analysis and KEGG pathway analysis showed that the mechanisms of Chaihu Longgu Muli Decoction for treating essential hypertension were related to pathways such as activation of AP-1 family transcription factors, interleukin-10 signaling, interleukin-4 signaling, interleukin-13 signaling, the intrinsic pathway of apoptosis, and MAPK targeting/by MAP kinase-mediated nuclear events. Conclusion: The mechanisms of the effect of Chaihu Longgu Muli Decoction in the treatment of essential hypertension were through the above-mentioned “multi-components-multi-targets-multi-pathways”. This study provides a foundation for further investigation of the effective compound and specific mechanisms of Chaihu Longgu Muli Decoction in the treatment of essential hypertension.

Triterpenes and sterols from Hoya diversifolia Blume.

Chemical investigation of the dichloromethane extracts of Hoya diversifolia Blume led to the isolation of β-amyrin cinnamate (1), squalene (2), β-sitosterol (3), a mixture of β-amyrin (4a), α-amyrin (4b) and lupeol (4c) in a 4:2:1 ratio and saturated hydrocarbons from the leaves; and 2, taraxerol (5), lupeol cinnamate (6), and a mixture of 3 and stigmasterol (7) in a 2:1 ratio from the stems. The structures of 1-7 were identified by comparison of their NMR data with those reported in the literature.

Chemical constituents of Pilea sinofasciata / 中草药

Objective: To study the chemical constituents of Pilea sinofasciata. Methods: Column chromatography, such as silica gel, Sephadex LH-20, and preparative HPLC, was used to isolate and purify the compounds. Spectroscopic methods like MS, 1H-NMR, and 13C-NMR, and the physical constants were used to elucidate their structures. Results: Nineteen compounds were isolated from 95% ethanol extracts of P. sinofasciata, including α-tocopherol (1), stigmasterol (2), epihernandulcin (3), hernandulcin (4), benzoic acid (5), ethyl linolenate (6), ethyl hexadecanoate (7), α-amyrin (8), palmitic acid (9), behenic acid (10), adenosine (11), indole-3-carboxylic acid (12), protocatechuate (13), gallic acid (14), betulinic acid (15), oleanolic acid (16), potassium nitrate (17), diosmetin 7-O-β-D-glucopyranoside (18), and 3-O-β-D-xylopyranosyl (1→2)-β-D-glucopyranosyl-28-O-β-D-glucopyranosyl oleanolic acid (19). Conclusion: All of the compounds are isolated from the plant for the first time.

Chemical Constituents of Hoya buotii Kloppenb.

Chemical investigation of the dichloromethane extracts of Hoya buotii Kloppenb. afforded taraxerone (1), taraxerol (2), a mixture of β-sitosterol (3a) and stigmasterol (3b) in about 2:1 ratio, and a mixture of α-amyrin cinnamate (4a) and β-amyrin cinnamate (4b) in about 1:2 ratio from the stems; 1, 2, and 3a from the roots; a mixture of 4a and 4b in about 3:2 ratio from the flowers; and 3a, squalene (5) and saturated hydrocarbons from the leaves. The structures of 1-5 were identified by comparison of their NMR data with those reported in the literature.

Cytotoxic Triterpenes and Sterols from Pipturus arborescens (Link) C.B. Rob.

The triterpenes, squalene (1), friedelin (2) and a mixture of ursolic acid (3a) and oleanolic acid (3b) in a 2:3 ratio, and a mixture of β-sitosterol (4a) and stigmasterol (4b) in a 2:1 ratio, obtained from the dichloromethane extract of Pipturus arborescens (Link) C.B. Rob., were evaluated for their anti-proliferative activities against three human cancer cell lines, breast (MCF-7) and colon (HT-29 and HCT-116), and a normal cell line, human dermal fibroblast- neonatal (HDFn) using the in vitro PrestoBlue® cell viability assay. The HCT-116 cell line was most susceptible to the compounds and mixtures tested. Triterpene 1 was most cytotoxic against HCT-116 and MCF-7 with IC50 values of 4.21 and 5.92 μg/mL, respectively. Triterpene 2 and the mixture of 3a and 3b were highly anti-proliferative against HCT-116 cells (IC50 of 1.22 and 1.66 μg/mL, respectively) and moderately inhibitory against MCF-7 cells (IC50 of 16.51 and 23.97 μg/mL, respectively). The mixture of 4a and 4b exhibited high cytotoxicity against HCT-116 cells (IC50 of 1.14 μg/mL). Compounds 1-4b showed the least activity against HT-29 cells (IC50 of 11.97 to 52.52 μg/mL). Cytotoxic effect was not observed against HDFn cells (>100 μg/mL). Comparing the effects of 1-4b on the two colon cancer cell lines, the IC50 values of 1-4b against HCT-116 were lower than those of HT-29.

Chemical Constituents of Ixora philippinensis Merr.

Chemical investigations of the dichloromethane extracts of Ixora philippinensis Merr., a plant endemic to the Philippines, led to the isolation of syringaresinol (1), pinoresinol (2), isoscopoletin (3), squalene (4), β-sitosterol (5a), and stigmasterol (5b) from the stems; and 4, 5a, 5b, lupeol (6), and lutein (7) from the leaves. The structures of 1 and 3 were elucidated by extensive 1D and 2D NMR spectroscopy, while those of 2 and 4-7 were identified by comparison of their NMR data with literature data.

Chemical constituents of Cycas aenigma.

Chemical investigation of Cycas aenigma, a plant endemic to the Philippines, led to the isolation of a rare neolignan, 2-[2-hydroxy-5-(3-hydroxypropyl)-3-methoxyphenyl]-1-(4-hydroxy-3-methoxyphenyl)propane-1,3-diol (1), pinoresinol (2), and fatty alcohols (3) from the leaflets; and triglycerols (4), and a mixture of β-sitosterol (5a) and stigmasterol (5b) from the petiole and rachis. The structure of 1 was elucidated by extensive 1D and 2D NMR spectroscopy, while those of 2-5b were identified by comparison of their 1H and/or 13C NMR data with literature data.

Simultaneous Estimation of Stigmasterol and Withaferin A in Union Total Herbal Formulation Using Validated HPTLC Method.

Union Total is herbal formulation made in the form of capsule which contains two standardized plant extracts Cissus quadrangularis (CQ) and Withania somnifera (WS). The present work describes development and validation of High Performance Thin Layer Chromatographic method for simultaneous analysis of Stigmasterol (STG) in Cissus quadrangularis (CQ) and Withaferin A (WFA) in Withania somnifera (WS). Stigmasterol and Withaferin A were identified on silica G60 F254 HPTLC plates by post derivatization technique and robustness study was performed by applying a central composite design (CCD) with k factor having 2k factorial runs, 2k axial experiments and five center points. In HPTLC good separation was obtained with chloroform: methanol: toluene: formic acid (6.5: 0.5: 3: 0.25 v/v/v/v) as mobile phase and anisaldehyde sulphuric acid as a derivatizing reagent at detection wavelength 530 nm. Linearity was obtained in the concentration range of 100-200 ng/band for WFA and 200-700 ng/band for STG and the % recoveries were found in the range of 100.06 % to 100.46 % for WFA and 99.97 % to 100.94 % for STG respectively. HPTLC method was found to be sensitive, precise, accurate and reproducible, which would be of use in quality control of these tablets.

Chemical Constituents of Cycas sancti-lasallei.

Chemical investigation of the dichloromethane extracts of Cycas sancti-lasallei, a plant endemic to the Philippines, led to the isolation of squalene (1), β-sitosterol (2a), stigmasterol (2b), and triglycerides (3) from the sarcotesta; 2a, 2b, 3,and phytyl fatty acid esters (4) from the endotesta; 2a, 2b, 3,and β-sitosteryl fatty acid esters (5) from the sclerotesta; and 3 and 5 from the bark. The structures of 1-5 were identified by comparison of their 1H NMR and/or 13C NMR data with those reported in the literature.

Terpenoids and Sterols from Hoya multiflora Blume.

Chemical investigation of the dichloromethane extracts of Hoya multiflora Blume led to the isolation of lupeol (1a), α-amyrin (1b), β-amyrin (1c), lupeol acetate (2a), α-amyrin acetate (2b), and β-amyrin acetate (2c) from the stems; and 1b, bauerenol (3), squalene (4), lutein (5), β-sitosterol (6a), and stigmasterol (6b)from the leaves. The structures of 1-6 were identified by comparison of their 1H and/or13C NMR data with those reported in the literature.

Effect of Ipomea carnea Jacq. flowers on hematological changes in toluene diisocyanate-induced inflammation in Wistar rats / 中国天然药物

AIM@#To investigate the active chloroform fraction of the ethanol extract of Ipomoea carnea flowers on hematological changes in toluene diisocyanate-induced inflammation in Wistar rats.@*METHOD@#Except for the control group, all of the rats were sensitized with intranasal application of 5 μL of 10% toluene diisocyanate (TDI) for 7 days. One week after second sensitization, all of the rats were provoked with 5 μL of 5% TDI to induce airway hypersensitivity. After the last challenge, blood and bronchoalvelor lavage (BAL) fluid were collected and subjected to total and differential leucocytes count. Flash chromatography was performed on the most active chloroform fraction to isolate an individual component.@*RESULTS@#Treatment with the ethanolic extract and its chloroform fraction at an oral dose of 200 mg·kg⁻¹ showed a significant decrease in circulating neutrophil and eosinophil in blood and BAL as compared with standard dexamethasone (DEXA). The structure of the compound obtained from chloroform fraction of Ipomea carnea was elucidated as stigmast-5, 22-dien-3β-ol on the basis of spectral data analysis.@*CONCLUSION@#The chloroform fraction was found to be more effective to suppress airway hyper reactivity symptoms, and decreased count of both total and differential inflammatory cells.

Chemical constituents from Orobanche cernua / 中草药

Objective: To study the chemical constituents from Orobanche cernua in Orobanchaceae. Methods: The chemical constituents were separated and purified by macroporous resin, silica gel, Sephadex LH-20, and MCI column chromatographies. Their structures were determined by physicochemical properties and spectral data. Results: Twelve compounds were isolated from 70% ethanol extract of O. cernua. Among them, eight phenylpropanoid glycosides were identified as acteoside (1), campneoside II (2), crenatoside (3), campneoside I (4), isocrenatoside (5), isoacteoside (6), leucosceptoside A (7), and cistanoside F (8), three liganans were identified as (+)-syringaresinol-4'-O-β-D-glucopyranoside (9), (+)-pinoresinol-4'-O-β-D-glucopyranoside (10), and isoeucommin A (11), and one steroide was stigmasterol-3-O-β-D-glucoside (12). Conclusion: Compounds 7 and 12 are isolated from the family Orobanchaceae for the first time; Compounds 4, 7, and 9-12 are isolated from the plants of Orobanche L. firstly, and compounds 2, 4, and 6-12 are found from O. cernua for the first time.

Chemical constituents from leaves of Acanthopanax evodiaefolius / 中草药

Objective: To study the chemical constituents from the leaves of Acanthopanax evodiaefolius. Methods: The compounds were isolated and purified by silica gel and Sephadex LH-20 column chromatography. Their structures were identified on the basis of their physicochemical properties and spectroscopic data. Results: Thirteen compounds were isolated from the methanol extract of A. evodiaefolius and their structures were identified as heptadecanoic acid (1), heptacosanol (2), trans-cinnamic acid (3), ursolic acid (4), oleanolic acid (5), betulinic acid (6), corosolic acid (7), apigenin (8), naringenin (9), maltol (10), syringic acid (11), stigmasterol (12), and daucosterol (13), respectively. Conclusion: Compounds 1-3 and 7-10 are obtained from the plans in genus Acanthopanax Miq. for the first time and all the compounds are obtained from A. evodiaefolius for the first time.