Mechanism of action of formononetin in alleviating allergic asthma through DRP1-NLRP3 signaling pathway / 中国药理学通报

Aim To investigate the mechanism by which formononetin (FN) inhibits mitochondrial dynamic-related protein 1 (DRP1) -NLRP3 axis via intervening the generation of ROS to reduce allergic airway inflammation. Methods In order to establish allergic asthma mouse model, 50 BALB/c mice aged 8 weeks were divided into the control group, model group, FN treatment group and dexamethasone group after ovalbumin (OVA) induction. Airway inflammation and collagen deposition were detected by HampE and Masson staining. Th2 cytokines and superoxide dismutase (SOD), catalase (CAT), malondialdehyde (MDA), and IgE levels in bronchoalveolar lavage fluid (BALF) were measured by ELISA, ROS in BEAS-2B cells was assessed by DCFH-DA staining, DRP1 expression in lung tissue and BEAS-2B cells was detected by immunohistochemistry and immunofluorescence, and the DRP1-NLRP3 pathway was analyzed by immunoblotting. Results FN treatment could effectively ameliorate the symptoms of asthmatic mouse model, including reducing eosinophil accumulation, airway collagen deposition, decreasing Th2 cytokine and IgE levels, reducing ROS and MDA production, increasing SOD and CAT activities, and regulating DRP1-NLRP3 pathway-related protein expression, thereby relieving inflammation. Conclusion FN ameliorates airway inflammation in asthma by regulating DRP1-NLRP3 pathway.

Formononetin regulates dilated cardiomyopathy-mediated heart failure in rats via HSP90/AKT cardiomyocyte apoptosis and mechanism / 西安交通大学学报(医学版)

【Objective】 To investigate the effects of formononetin (FMN) on cardiomyocyte apoptosis and HSP90/AKT in rats with dilated cardiomyopathy-mediated heart failure. 【Methods】 Echocardiography, ELISA, histological staining, and TUNEL staining were used to observe the protective effect of different doses of FMN on dilated cardiomyopathy-mediated heart failure in rats and the apoptosis of cardiomyocytes. The potential targets of formononetin on dilated cardiomyopathy-mediated heart failure were obtained from TCMSP, DisGeNet, GeneCards, and other databases, the key targets were obtained according to the protein-protein interaction (PPI) network, and the key targets were verified by molecular docking. Western blotting was used to further verify the regulatory role of key targets in the treatment of dilated cardiomyopathy-mediated heart failure with formononetin. 【Results】 Formononetin could reduce the levels of LVIDS, LVIDD, NT-pro BNP, cTn-T, CK, CK-MB, and LDH in rats with dilated cardiomyopathy-mediated heart failure, increase the levels of EF and FS, and reduce the apoptosis of cardiomyocytes. FMN had a strong binding effect on 10 key targets (AKT1, HSP90AA1, CASP3, MAPK1, MMP9, SRC, ALB, HRAS, IGF1, and EGFR) screened by network pharmacology, with HSP90AA1 and AKT1 having the strongest binding effect. Formononetin decreased the expression of HSP90, AKT and downstream CASP3 protein, but increased the expression of p-AKT in myocardial tissue. 【Conclusion】 Formononetin may inhibit the expression of HSP90, promote phosphorylation of AKT to p-AKT, and inhibit the expression of CASP3, thereby reducing the apoptosis of cardiomyocytes and improving myocardial tissue damage, so as to achieve the purpose of treating dilated cardiomyopathy-mediated heart failure.

Inhibitory effects of formononetin on lipopolysaccharide-induced apoptosis and inflammatory response in alveolar epithelial cells / 中国药房

OBJECTIVE To investigate the inhibitory effects of formononetin on lipopolysaccharide （LPS）-induced apoptosis and inflammatory response in alveolar epithelial cells through phosphoinositide 3-kinase （PI3K）/protein kinase B （Akt） signaling pathway. METHODS Human lung cancer alveolar basal epithelial cells A549 were cultured in vitro and divided into control group （no intervention）， model group （1 μg/mL LPS）， different concentrations of formononetin groups （1 μg/mL LPS+6.25， 12.5， 25， 50 μmol/L formononetin）. The levels of inflammatory factors （interleukin-8， tumor necrosis factor-α） and cell viability were detected in each group. Another A549 cells were divided into control group， model group （1 μg/mL LPS）， LPS+25 group （1 μg/mL LPS+25 μmol/L formononetin）， inhibitor group （1 μg/mL LPS+20 μmol/L LY294002）， formononetin+inhibitor group （1 μg/mL LPS+25 μmol/L formononetin+20 μmol/L LY294002） and formononetin+activator group （1 μg/mL LPS+25 μmol/L formononetin+ 10 μmol/L SC79）. The secretion levels and mRNA expressions of inflammatory factors， cell apoptosis， and expressions of the key proteins of PI3K/Akt signaling pathway were detected in each group. RESULTS Compared with model group， the levels of inflammatory factors were decreased significantly after the intervention of 25 μmol/L of formononetin， and the cell viability was increased significantly （P＜0.05）. Compared with the control group， the secretion levels and mRNA expressions of inflammatory factors， apoptotic rate， and relative expressions of phosphorylated Akt and phosphorylated PI3K of the model group were increased significantly （P＜0.05）. Compared with the model group， the above indexes of the LPS+25 group and the inhibitor group were decreased significantly （P＜0.05）. Compared with the LPS+25 group， the above indicators of formononetin+inhibitor group were further decreased， while those of formononetin+activator group were increased significantly （P＜0.05）. CONCLUSIONS Formononetin can inhibit LPS-induced epithelial cell apoptosis and improve inflammatory response， and the mechanism may be related to the inhibition of the PI3K/Akt signaling pathway.

The effect and mechanism of formononetin on alleviating no-reflow after myocardial ischemia and reperfusion by up-regulating the PI3K/Akt/eNOS signal pathway activated by GPER / 药学学报

To investigate the cardioprotective effect of formononetin (FMN) on no-reflow (NR) after myocardial ischemia-reperfusion and its molecular mechanism based on integrated pharmacology and experimental verification, firstly, human breast cancer MCF-7 cells and myocardial NR rats were used to confirm the estrogenic activity and the effect of alleviating NR of FMN, respectively. Male SD rats were divided into Sham, NR, FMN (20 mg·kg-1) and sodium nitroprusside (SNP, 5.0 mg·kg-1) groups, which were administered once a day for one week, the experiment was approved by the Ethics Committee of Tianjin University of Traditional Chinese Medicine (TCM-LAEC2019095). The pharmacological analysis and in vivo study of NR rats were integrated to reveal the mechanism of FMN improving NR. The results showed that FMN had estrogenic effect and reduced NR by improving cardiac structure and function, reducing NR, ischemic myocardial area and pathological injury of cardiomyocytes. Integrated pharmacology predicts that the mechanism of FMN improving NR is mainly related to phosphatidyinositol-3-kinase-protein kinase B (PI3K-Akt) signal pathway. Phytoestrogens play a role in cardiovascular protection mainly by activating G protein-coupled estrogen receptor (GPER). GPER is also an important regulator in the upstream of PI3K-Akt signaling pathway. This study found that FMN can significantly activate GPER, p-PI3K, p-Akt and phospho-endothelial nitric oxide synthase (p-eNOS). It has good binding ability with GPER and eNOS protein. In this study, through the integration of pharmacology and experimental evaluation, it is revealed that FMN activates PI3K/Akt/eNOS signal pathway by activating GPER, thus significantly improving NR.

Research progress on Astragali Radix and prediction of its quality markers (Q-markers) / 中国中药杂志

Astragali Radix, a medicinal herb for invigorating Qi, has anti-aging, anti-tumor, immunoregulatory, blood sugar-and lipid-lowering, anti-fibrosis, anti-radiation and other pharmacological effects. This article reviewed the studies about the chemical components and pharmacological effects of Astragali Radix. According to the theory of quality markers(Q-markers) of Chinese medicinal materials, we predicted the Q-markers of Astragali Radix from traditional efficacy, chemical component validity, measurability, plant phylogeny, and pharmacokinetis. The results showed that total polysaccharides, flavonoids(e.g., calycosin-7-O-β-D-glucoside, formononetin, calycosin, quercetin, and ononin), and saponins(e.g., astragalosides Ⅱ, Ⅲ, and Ⅳ) can be taken as the main Q-markers. This review lays a foundation for regulating the quality research and standard establishment of Astragali Radix, and benefits the control and quality supervision of the production process of Astragali Radix and its related products.

Determination of formononetin and harmful elements of heavy metals in Jinji Pill / 国际中医中药杂志

Objective:To use the high performance liquid chromatography method to determine the content of formononetin in Jinji Pills and by using atomic absorption spectrophotometry,method to determine the harmful elements of heavy metal in Jinji Pills in orer to provide the scientific foundation for improving its quality standards and safety evaluation. Methods:Use Waters XBridge? C18 column (4.6 mm × 250 mm, 5 μm), set mobile phase at acetonitrile-1% phosphoric acid solution (27:73), flow rate 1.0 ml/min, column temperature 30 ℃, detection wavelength 249 nm, column temperature 30 ℃; Lead (Pb) and cadmium (Cd) was detected by graphite furnace method; arsenic (As) was detected by cold steam series graphite furnace method; copper (Cu) was detected by flame method; mercury (Hg) was detected by cold steam method.Results:The formononetin had a good linear relationship between 0.02-2.01 μg, the recovery rate was 98.5%, RSD was 1.53%. Lead (Pb) recovery rate was 103.6%, cadmium (Cd) recovery rate was 95.7%, arsenic (As) recovery rate was 92.4%, mercury (Hg) recovery rate was 104.9%, copper (Cu) recovery rate was 112.5%. Conclusion:This method is of accuracy, specificity, high sensitivity and good reproducibility, which could provide strong evidence for quality improvement and safety use of Jinji pill.

Research Progress on Anti-tumor Mechanism of Formononetin / 中国实验方剂学杂志

Formononetin is a kind of plant isoflavones extracted from medicinal herbs such as Trifolium pratense，Astragalus membranaceus and Spatholobi Caulis have shown that formononetin has strong anti-tumor biological activity，and can be used as an anti-tumor drug in the treatment of various malignant tumors. Many studies so far have shown that formononetin can inhibit cell proliferation，induce cell apoptosis，inhibit cell migration and invasion，and induce cell cycle arrest on tumors through a variety of molecular mechanisms and pathways. These antitumor activities can be observed in cells of various tumors such as breast cancer，colorectal cancer，prostate cancer，bladder cancer and lung cancer in trials and animal models. Examples of these effects include triggering the generation of reactive oxygen species （ROS），regulating phosphatidylinositol-3-kinase/protein kinase B/mammalian target of rapamycin（PI3K/Akt/mTOR） and Mitogen-activated protein kinases（MAPK） signaling pathways，inhibiting the activation of tyrosine kinase（JAK1 and JAK2 ）and nonreceptor tyrosine kinase（c-Src），and regulating cytokeratin 19（CK19），matrix metalloproteinases（MMP），microRNA-21（miR-21），lamin A/C antibody（Lamin A/C），expression of Cyclin D1 and Cyclin E1. In addition， the anti-tumor effects of formononetin derivatives were reviewed in this paper. By modifying the chemical structure of formononetin，many related derivatives have been obtained. Experimental results have shown that some derivatives of formononetin have stronger anti-tumor activity and lower cytotoxicity，but the related molecular mechanism of action still needs to be explored further in-depth. In conclusion，formononetin and its derivatives may become potential anti-tumor drugs.

Therapeutic effect and mechanism of sodium formononetin-3′-sulphonate on collagen-induced rheumatoid arthritis / 中国药科大学学报

@#To investigate the therapeutic effect and mechanism of sodium formononetin-3′-sulphonate (SFS) on collagen-induced rheumatoid arthritis in mice, C57 mice were induced with chicken type II collagen to establish a model of rheumatoid arthritis (collagen-induced arthritis, CIA), and were injected intraperitoneally with different doses of SFS (50,100,200 mg/kg). Body weight, food intake and foot swelling of all groups were observed during the experiment.After the treatment, TNF-α, IL-6, and IL-10 in the serum were detected with the CBA kit; NF-κB p65, p-NF-κB p65 (p-p65), TIPE2, PCNP and IκB-α in spleen tissue were determined by Western blot; the organ index, pathological changes of ankle joint cartilage tissue and the positive expression of NF-κB p65 in ankle joint tissue were also observed.The results showed that, compared with the model group, the body weight and food intake of mice in the treatment group increased, while the degree of foot swelling decreased; the expression levels of inflammatory factors TNF-α and IL-6 in serum decreased, while the expression of anti-inflammatory factor IL-10 increased; the expression levels of NF-κB p65, p-p65 and PCNP in spleen tissue decreased, while the expression of TIPE2 and IκB-α protein increased; the index of spleen and thymus of the CIA mice in the treatment group, the infiltration of inflammatory cells in the ankle joint, the destruction of synovial tissue and cartilage, and the positive expression of NF-κB p65 decreased.Among them, the high-dose group of SFS showed a better therapeutic effect.It is suggested that SFS has a therapeutic effect on CIA mice, and the mechanism may be achieved by regulating the NF-κB p65 signaling pathway and inhibiting the expression of inflammatory factors.

Formononetin alleviates diabetic cardiomyopathy by inhibiting oxidative stress and upregulating SIRT1 in rats

Objective: To evaluate the effect of formononetin on type 2 diabetic cardiomyopathy. Methods: Diabetes was induced by feeding high-fat diet for 2 weeks and administration of 35 mg/kg of streptozotocin in rats. Formononetin was administered at 10, 20 and 40 mg/kg for 16 weeks once a day. Plasma glucose, lipid parameters, and cardiac markers in blood samples were measured. Body weight and relative heart weight were recorded. Hemodynamic parameters, oxidative stress parameters and silence information regulator 1 (SIRT1) expression in cardiac tissue were estimated. Histopathological changes in cardiac tissue were also observed. Results: Formononetin significantly reduced the levels of glucose, triglycerides, cholesterol, low density lipoprotein, creatine kinase-MB, lactate dehydrogenase and aspartate aminotransferase. In addition, formononetin significantly improved hemodynamic parameters, alleviated oxidative stress and increased SIRT1 expression. Conclusions: The study indicates that formononetin can improve hyperglycemia and hyperlipemia, reduce oxidative stress and increase SIRT1 expression. It can be a potential therapeutic agent for diabetic cardiomyopathy.

The effect of four compounds of astragalus isoflavones on the differentiation of PC 12 cells / 药学实践杂志

Objective To investigate the effects of calycosin, formononetin, calycosin-7-glucoside and ononin on PC 12 cells differentiation. Methods PC 12 cells were cultured and treated with different concentrations of nerve growth factor (NGF), calycosin, formononetin, calycosin-7-glucoside and ononin for 5 days, once a day, 3 times in a row. The neurite outgrowth of PC 12 cells was observed and the expression of β III-tubulin were measured by immunofluorescence. Results Compared with the vehicle group, neurite outgrowth and the expression of β III tubulin in PC 12 cells had not promoted by calycosin, formononetin, calycosin-7-glucoside and ononin (0.01-10.00 μmol/L). Conclusion PC 12 cells differentiation could not be induced by calycosin, formononetin, calycosin-7-glucoside and ononin.

Network pharmacological analysis and mechanism prediction of Xiaochaihu Decoction in treatment of COVID-19 with syndrome of pathogenic heat lingering in lung and obstructive cardinalate / 中草药

Objective: To study the efficacy network and potential mechanism of Xiaochaihu Decoction (XCHD) in the treatment of coronavirus disease 2019 (COVID-19) with syndrome of pathogenic heat lingering in the lung and obstructive cardinalat, and analyze the active ingredients of XCHD with anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) efficacy. Methods: The correspondence between COVID-19 and XCHD was analyzed by literature consulting. Based on network pharmacology, Cytoscape 3.6.0 and other software were then used to construct XCHD efficacy network of “Chinese medicine prescription-active ingredient-key target” for pneumonia and immune regulation, in order to confirm anti-SARS-CoV-2 active ingredients in the prescription. Some softwares were used to analyze XCHD for COVID-19 treatment in multiple aspects. Results: A total of 48 active ingredients with potential anti-SARS-CoV-2 effect in herbs were collected; 140 active ingredients in XCHD for pneumonia treatment and immune regulation were analyzed, of which 12 ingredients had direct anti-SARS-CoV-2 activity including baicalein, formononetin, quercetin, etc. The active ingredients in XCHD exerted efficacy for pneumonia treatment and immunoregulation through 95 key targets such as IL-6, NOS2, and ESR1, involving multiple pathways such as the TNF signaling pathway, IL-17 signaling pathway, and influenza A. Analysis of gene co-expression and PPI interaction analysis found that ACE2 only co-expressed with NOS2 in the above targets, and also interacted with only five targets in the PPI interaction network. It is speculated that the ACE2 target only plays an important role when SARS-CoV-2 invaded the human body, and had little effect in the treatment of pneumonia after viral infection. Conclusion: The active ingredients in XCHD play a role in treating COVID-19 by inhibiting SARS-CoV-2 activity, blocking the SARS-CoV-2 invasion pathway, inhibiting cytokine storm, and regulating immunity. It is worth noting that drugs designed for the ACE2 target can block virus invasion, but may not be effective for diseases such as alveolar inflammation, Therefore, this study also provides a multi-target and multi-directional space for XCHD for early COVID-19 treatment. In addition, when XCHD is used in the early treatment of COVID-19, we should pay attention to the precise use of drugs based on syndrome accurate identification, one is to avoid adverse reactions, the other is to avoid cytokine damage caused by re-crown disease.

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.

Chemical composition identification and mechanism study of Tenghuang Jiangu Capsule based on UHPLC-Q-Orbitrap HRMS and network pharmacology / 中草药

Objective: To systematically study the chemical components of Tenghuang Jiangu Capsule, explore the main mechanism of action, and provide some evidences for the research of its pharmacodynamic substances. Methods: In this study, UHPLC-Q-Orbitrap HRMS was used to comprehensively analyze the main chemical components of Tenghuang Jiangu Capsule. According to the MS/MS spectrometry information of compounds, the chemical information of this herbal formula can be quickly and accurately identified by comparison with standards or references. Next, the BAT-MAN-TCM database was used to predict the targets of the identified chemical components. The KEGG pathway annotation analysis and GO enrichment analysis were further carried out through the DAVID database to screen out the main pharmacodynamic substances of Tenghuang Jiangu Capsule and explore the potential mechanisms. Results: A total of 34 chemical components were identified in Tenghuang Jiangu Capsule. The "component-target" network analysis indicated that the major components including kaempferol, isoflavoues aglycone, ursolic acid, formononetin, and stigmasterol might act on some key targets such as Bcl-2, BAX, AKt, PPARG, PTGS1, PTGS2, TNF, IL6, F7, IL1B, etc. The results indicated that osteoclast differentiation, NF-κB, PI3K-Akt, renal cell, and platelet activation might be the main action pathways of exerting the therapeutic effect of bone protection, nourishing kidney, promoting blood circulation and relieving pain of Tenghuang Jiangu Capsule. Conclusion: In this study, UHPLC-Q-Orbitrap HRMS combined with network pharmacology was used to preliminarily clarify the chemical composition and reveal potential mechanism of Tenghuang Jiangu Capsule. The results provided scientific theoretical basis for screening the effective ingredients and further clarifying the mechanism of action of Tenghuang Jiangu Capsule.

Establishment of HPLC fingerprints of Qiwei Tangmaishu tablet and simultaneous determination of six constituents / 国际药学研究杂志

Objective: To establish HPLC fingerprints of Qiwei Tangmaishu tablet and simultaneously determine the contents of the six constituents, acteoside, martynoside, calycosin 7-O-β- D-glucopyranoside, schisandrin, formononetin and deoxyschizan- drin. Methods: The analysis of 50% methanol extract of this drug was performed on a 30℃ thermostatic Waters Symmetry C18 column, with the mobile phase comprising of acetonitrile(A)-0.2% formic acid solution(B)flowing at 1.0 ml/min in a gradient elution manner. The detection wavelength was set at 330 nm for acteoside and martynoside, 254 nm for calycosin 7-O-β-D-glucopyranoside, schisandrin, formononetin and deoxyschizandrin. Results: There were thirteen common peaks in the fingerprints of ten batches of samples with the similarities more than 0.9. Acteoside, martynoside, calycosin 7-O-β-D-glucopyranoside, schisandrin, formononetin and deoxyschizandrin showed good linear relationship within the ranges of 1.47-36.75, 0.66-16.50, 0.89-22.25, 2.58-64.50, 1.91-47.75 and 0.77-19.25 μg/ml(r≥0.9993), and their average recoveries were 97.57%, 99.22%, 96.69%, 100.01%, 98.79% and 96.77%, with the RSD of 0.79%, 1.54%, 0.61%, 0.64%, 0.83% and 0.36%, respectively. Conclusion: The established method is easily operable and repeatable, which could be used for quality control of Qiwei Tangmaishu tablet.

Formononetin alleviates diabetic cardiomyopathy by inhibiting oxidative stress and upregulating SIRT1 in rats

Objective: To evaluate the effect of formononetin on type 2 diabetic cardiomyopathy.Methods: Diabetes was induced by feeding high-fat diet for 2 weeks and administration of 35 mg/kg of streptozotocin in rats. Formononetin was administered at 10, 20 and 40 mg/kg for 16 weeks once a day. Plasma glucose, lipid parameters, and cardiac markers in blood samples were measured. Body weight and relative heart weight were recorded. Hemodynamic parameters, oxidative stress parameters and silence information regulator 1 (SIRT1) expression in cardiac tissue were estimated. Histopathological changes in cardiac tissue were also observed. Results: Formononetin significantly reduced the levels of glucose, triglycerides, cholesterol, low density lipoprotein, creatine kinase-MB, lactate dehydrogenase and aspartate aminotransferase. In addition, formononetin significantly improved hemodynamic parameters, alleviated oxidative stress and increased SIRT1 expression. Conclusions: The study indicates that formononetin can improve hyperglycemia and hyperlipemia, reduce oxidative stress and increase SIRT1 expression. It can be a potential therapeutic agent for diabetic cardiomyopathy.

Simultaneous determination of six kinds of components in Buyang-Huanwu decoction by UHPLC-MS/MS / 国际中医中药杂志

Objective To develop the UHPLC-MS/MS method for the determination of amygdalin, paeoniflorin, ferulic acid, calycosin glucosidase, quercetin and formononetin in Buyang-Huanwu decoction. Methods Isocratic elution was carried out with mobile phase consisting of methanol- 2 mM ammonium formate. The separation was performed on Agilent ZORBAX SB-C18 maintained at 35 ℃. The flow rate was 200 μl/min, and the injection volume was 2 μl. The mass spectrometer was operated in the positive and negative ionization electrospray (ESI) mode using multiple monitoring (MRM) for analysis of six components. The mass spectrometric conditions were that ion source temperature 400 ℃, dry gas flow 500 L/h, atomization gas flow rate 75.8 Kpa, spray voltage 4000 V, dry gas temperature 400 ℃. Results The amygdalin, paeoniflorin, ferulic acid, calycosin glucosidase, quercetin and formononetin were all analyzed exactly, and the linear ranges were 0.5-32, 0.2-12.8, 0.1-6.4, 0.8-51.2, 0.4-25.6, 0.08-5.12 ng, respectively. The r were 0.9921, 0.9945, 0.9928, 0.9958, 0.9947, 0.9966, respectively. The recoveries of six analytes ranged from 99.21％ to 101.44％ and the relative standard deviations were all below 2.05％. Conclusions A sensitive, accuracy and suitable UHPLC-MS/MS method has been developed, and the method could be applied for the determination of amygdalin, paeoniflorin, ferulic acid, calycosin glucosidase, quercetin and formononetin in Buyang-Huanwu decoction.

Simultaneous determination of seven index components in Naoxintong capsule by HPLC wavelength switching method / 中国药科大学学报

@#An HPLC-DAD wavelength switching method(240 nm, 280 nm, 316 nm, 403 nm)was developed for simultaneous determination of seven index components: hydroxysafflor yellow A, paeoniflorin, ferulic acid, salvianolic acid B, kaempferol, formononetin and tanshinone IIA in Naoxintong capsule. The qualities of different batches of Naoxintong capsules were evaluated by statistical analysis. Seven index components in 20 batches of Naoxintong capsules were simultaneously determined by HPLC wavelength switching method with Capcell PAK C18 MG II column(250 mm × 4. 6 mm, 5. 0 μm). The mobile phase consisted of methanol-acetonitrile(25 ∶75, A)-0. 1% formic acid aqueous solution(B)with a gradient elution program and a flow rate of 1. 0 mL/min, and the column temperature was 30 °C. The results were analyzed by statistical analysis to evaluate the differences in the quality of Naoxintong capsules. Results showed that the seven active components were well separated and showed good linearity hydroxysafflor yellow A(403 nm)2. 30- 11. 50 mg/L(r=0. 999 2), paeoniflorin(240 nm)8. 81- 44. 05 mg/L(r=0. 999 6), ferulic acid(316 nm)1. 22- 6. 10 mg/L(r=0. 999 6), salvianolic acid B(280 nm)11. 61- 58. 05 mg/L(r=0. 999 4), kaempferol(403 nm)1. 16-5. 80 mg/L(r=0. 999 4), formononetin(240 nm)0. 12- 0. 60 mg/L(r=0. 999 5)and tanshinone IIA(280 nm)2. 28- 11. 40 mg/L(r=0. 999 5). The precision was good and RSD was less than 2. 0%, The repeatability was good and RSD was less than 2. 0%. The stability was good in 24 h. The average recoveries were between 97. 35%- 101. 02% and RSD was less than 2. 0%. The contents of target components in Naoxintong capsules, hydroxysafflor yellow A was 0. 213- 0. 369 mg/g, paeoniflorin was 1. 535- 3. 217 mg/g, ferulic acid was 0. 153- 0. 236 mg/g, salvianolic acid B was 2. 563- 3. 271 mg/g, kaempferol was 0. 103- 0. 181 mg/g, formononetin was 0. 022- 0. 028 mg/g, and tanshinone IIA was 0. 466- 0. 698 mg/g. HPLC wavelength change and gradient elution method was established for simultaneous determination of seven index components in Naoxintong capsule. The method is accurate, sensitive, reliable, and repeatable, and can be used for the quality control of Naoxintong capsule.

UPLC-Q-TOF-MS/MS Analysis of Brain and Blood Absorption Components of Buyang Huanwu Tang After Intragastric Administration / 中国实验方剂学杂志

Objective:To analyze and identify the brain and blood absorption components of rats after intragastric administration of Buyang Huanwu Tang(BYHWT). Method:The brain tissue,plasma of normal rats and the cerebral ischemia-reperfusion rats were analyzed by UPLC-Q-TOF-MS/MS.The prototype components in BYHWT were identified according to retention time,accurate relative molecular weight,primary and secondary mass spectrometry data. Result:After the administration of BYHWT,five compounds were found to enter the normal brain tissue through the blood-brain barrier and identified as calycosin-7-glucoside,albiflorin,formononetin-7-O-β-D-glucoside-6″-O-acetyl,safflower yellow A and astragaloside A;two compounds penetrated the blood-brain barrier and entered modeling brain tissue,and they were identified as calycosin-7-glucoside and formononetin-7-O-β-D-glucoside-6″-O-acetyl;seven compounds entered normal plasma and were identified as calycosin-7-glucoside,albiflorin,hydroxysafflor yellow A,et al;three compounds entered model plasma and identified as calycosin-7-O-β-D-glucoside-6″-O-acetyl,6″-O-acetyl-(6αR,11αR)-9,10-dimetho-xypterocarpan-3-O-β-D-glucoside and formononetin-7-O-β-D-glucoside-6″-O-acetyl. Conclusion:BYHWT has different pharmacological material basis in normal and cerebral ischemia-reperfusion rats.

Effect of main active components combination of Astragalus and Angelica on hematopoietic function in mice with bone marrow suppression / 中国药理学通报

Aim To explore the effect of the main active components combination of Astragalus promoting hematopoiesis on mouse model of bone marrow hematopoietic suppression. Methods The contents of five main active components such as ferulic acid, astragalo-side IV, formononetin, calycosin and calycosin glycoside were determined after the extract of Astragalus combined with Angelica at 1 : 1 ratio was prepared by water extraction, thus ascertaining the combination dosage. Mice were randomly divided into blank control, model, active component alone, five active components' combination and Astragalus-Angelica combination groups. The blank control group was given solvent for 7 days; the model group was given the intragastric administration as above and received intraperitoneal injection of cyclophosphamide on day 3 for 3 days; each drug group was treated intragastrically and established the model as above. The peripheral blood, the content of serum hematopoietic growth factor and the colony forming of hematopoietic progenitor cells were measured. Results The active components' combination and Astragalus-Angelica combination could significantly increase the numbers of peripheral blood leukocytes, e-rythrocytes, platelets, the content of hemoglobin and the area of bone marrow hematopoietic tissue (P < 0. 05 or 0. 01 ) five active components could promote the recovery of erythrocytes and hemoglobin; ferulic acid could promote the recovery of leukocyte number and bone marrow hematopoietic tissue area; ferulic acid and astragaloside IV could promote the recovery of platelet. The contents of granulocyte-macrophage colony-stimulating factor ( GM-CSF ), thrombopoietin (TPO), erythropoietin ( EPO) in serum significantly increased in Astragalus-Angelica combination and the active components combination groups ( P < 0. 05 or 0. 01); five components could increase TPO content, ferulic acid and calycosin could increase the contents of GM-CSF and EPO, calycosin glycoside could increase EPO. Astragalus-Angelica combination and the active components combination could significantly increase the colony numbers of granulocyte-monocyte colony fo-ming unit ( CFU-GM ) , megakaryocyte colony forming unit (CFU-MK), erythroid colony forming unit (CFU-E) , burst forming unit-erythroid (BFU-E) ; except for astragaloside IV, the others could increase the number of CFU-GM, five active components could all increase the numbers of CFU-MK, CFU-E, but only ferulic acid could increase the number of BFU-E. Conclusions The main active substances that the combination of Astragalus and Angelica enriches the blood are the above five active components, which exert the synergistic effect through acting on different links of hematopoiesis such as increasing the content of hematopoietic growth factor in blood, promoting the proliferation of hematopoietic progenitor cells,and so on.

Optimization of Extraction Technology of Flavonoids in Glycyrrhiza uralensis by Single Factor Experiment Combined with Box-Behnken Design-response Surface Methodology / 中国药房

OBJECTIVE： To optimize the extraction technology of the flavonoids from Glycyrrhiza uralensis. METHODS： Using total contents of four flavonoids， liquiritinapioside， glycyrrhizin， isoliquiritin apioside and formononetin as indexes， types and volume fractions of extraction solvents （water， ethanol）， volume of addition and extraction time as factors， based on single factor experiment， Box-Behnken design-response surface method was used to optimize the extraction technology of flavonoids from G. uralensis. Validation test was also conducted. RESULTS： The optimal extraction technology was 50 mL 50％ ethanol as extraction solvent， 0.200 g G. uralensis， ultrasonic extraction for 50 min. In validation test， the extraction amounts of liquiritinapioside， glycyrrhizin， isoliquiritin apioside and formononetin were 10.733 0， 27.784 9， 3.441 9， 0.429 1 mg/g， respectively （all RSDs＜3.0％， n＝3）. The average total extraction amount of four flavonoids was obtained was 42.388 9 mg/g， the relative error of which to predicted value （42.173 2 mg/g） was 0.52％ （n＝3）. CONCLUSIONS： The optimized extraction technology is simple， rapid and stable， and can be used for the extraction of flavonoids from G. uralensis.