Effectss of hyperoside on traumatic brain injury rats by regulating RhoA/ROCK signaling pathway / 中国临床药理学与治疗学

AIM: To investigate the effects of hyperoside on traumatic brain injury (TBI) rats by regulating the Ras homolog gene family, member A (RhoA)/Rho-associated coiled coil-forming kinase (ROCK) signal pathway. METHODS: The TBI rat model was established by modified Feeney free fall hit method, and was randomly divided into model group, low-dose hyperoside (60 mg / kg) group, high-dose hyperoside (120 mg / kg) group, high-dose hyperoside (120 mg / kg) + no load group, and high-dose hyperoside (120 mg / kg) + RhoA overexpression group, with 10 rats in each group, another 10 healthy rats were set as sham operation group, after hyperoside and plasmid were grouped, the nerve injury was detected by modified neurological deficit score (mNSS) and dark avoidance test; Evans blue (EB) quantitative method was used to detect the permeability of blood brain barrier in rats; ultrastructural damage of blood-brain barrier was observed by transmission electron microscopy; the levels of tumor necrosis factor- α (TNF- α), interleukin-8 (IL-8), superoxide dismutase (SOD) and malondialdehyde (MDA) in serum and brain tissue of rats were measured with the kit; and the expression of RhoA/ROCK pathway related proteins in rat brain was detected by Western blot. RESULTS: Compared with the sham operation group, the blood brain barrier structure of the model group rats was damaged, the step-through latency and SOD level decreased obviously (P0.05). CONCLUSION: Hyperoside can inhibit neuroinflammation and oxidative stress in TBI rats by down-regulating RhoA / ROCK signal pathway, thereby reducing the damage of blood brain barrier and repairing its neural function.

Hyperoside protects mouse spermatocytes GC-2 cells from oxidative damage by activating the Keap1/Nrf2/HO-1 pathway / 南方医科大学学报

OBJECTIVE@#To study the protective effect of hyperoside (Hyp) against ydrogen peroxide (H2O2)- induced oxidative damage in mouse spermatocytes GC-2 cells and explore the role of the Keap1/Nrf2/HO-1 pathway in this protective mechanism.@*METHODS@#GC-2 cells were treated with 2.5 mmol/L azaacetylcysteine (NAC), 50, 100, and 200 μmol/L hyperoside, or the culture medium for 48 h before exposure to H2O2 (150 μmol/L) for 2 h. CCK-8 assay was used to detect the changes in cell viability, and cell apoptosis was analyzed using flow cytometry. Enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of superoxide dismutase (SOD), glutathione peroxidase (GSH-PX), catalase (CAT) activity and malondialdehyde (MDA) in the culture medium. Western blotting and RT-qPCR were used to detect the protein and mRNA expression levels of nuclear factor erythroid 2-related factor2 (Nrf2), Kelch-like ECH-associated protein 1 (Keap1), and heme oxygenase-1 (HO-1); the nuclear translocation of Nrf2 was detected using immunofluorescence assay.@*RESULTS@#Exposure to H2O2 significantly lowered the proliferation rate, reduced the activities of SOD, GSH and CAT, and obviously increased MDA content, cell apoptosis rate, and the expressions of Keap1 and Nrf2 mRNA and Keap1 protein in GC-2 cells (P < 0.05 or 0.01). Treatment of the cells prior to H2O2 exposure with either NAC or 200 μmol/L hyperoside significantly increased the cell proliferation rate, enhanced the activities of SOD, GSH-PX and CAT, and lowered MDA content and cell apoptosis rate (P < 0.05). Treatment with 200 μmol/L hyperoside significantly decreased the mRNA and protein expressions of Keap1 and increased the expressions of HO-1 mRNA and the protein expressions of Nrf2 and HO-1 (P < 0.05 or 0.01). Hyperoside also caused obvious nuclear translocation of Nrf2 in the cells (P < 0.05).@*CONCLUSION@#Hyperoside protects GC-2 cells against H2O2- induced oxidative damage possibly by activation of the Keap1/Nrf2/HO-1 signaling pathway.

Study on the preparation of hyperoside mixed nanomicelles and its intestinal absorption / 中国药房

OBJECTIVE To prepare hyperoside mixed nanomicelles （Hyp-F127/TPGS） and optimize its preparation technology，and to investigate its intestinal absorption characteristics. METHODS Hyp-F127/TPGS was prepared by thin film dispersion method. Based on single factor test and Plackett-Burman design ，combined with Box-Behnken response surface method ， the preparation process was optimized and validated using entrapped efficiency （EE）and drug loading （DL）as evaluation indexes ， F127-TPGS mass ratio ，hydration time and the amount of Hyp as factors. The appearance and microscopic morphology of Hyp-F127/TPGS obtained by the optimal technology were observed ，and the particle size ，polydispersity index （PDI）and Zeta potential were also determined. The critical micelle concentration （CMC）of blank micelle （F127/TPGS），in vitro release behavior and preliminary stability of Hyp-F 127/TPGS were investigated ，and absorption characteristics of Hyp-F 127/TPGS were investigated by in situ unidirectional intestinal perfusion model. RESULTS The optimal preparation technology of Hyp-F 127/TPGS included F127-TPGS mass ratio of 2∶1，hydration time of 2 h，and Hyp amount of 9 mg. Results of three validation tests showed that the EE of Hyp-F 127/TPGS was （87.20±0.99）％，and the DL was （5.02±1.20）％，deviations from predicted values were 0.92％ and 2.39％. The micelles prepared by optimal technology were yellow ，clear and transparent solution ，with good Tyndall effect ；under transmission electron microscope ，they were spherical ，complete and evenly distributed ；the particle size was （15.02±0.16）nm， the PDI was 0.092±0.031，and the Zeta potential was （－6.67±1.47）mV. The CMC of F 127/TPGS was 21 μg/mL，Hyp-F127/ TPGS was stable after 4 weeks of storage at 4 ℃，and the cumulative release rates of Hyp-F 127/TPGS and Hyp control were （66.30±2.93）％（96 h）and（99.24±0.27）％（60 h），respectively. Hyp-F 127/TPGS and Hyp reference were absorbed in each intestinal segment ，and the main absorption sites were jejunum and duodenum respectively ；drug absorption rate constant andapparent absorption coefficient of the former were significantly higher than those of the latter （P＜0.05 or P＜0.01）. E-mail：zhangyuhangxz@163.com CONCLUSIONS The optimized preparation technology of Hyp-F127/TPGS is stable and feasible ；prepared Hyp-F 127/ TPGS shows a sustained -release effect ，which promotes the intestinal absorption of H yp to a certain extent.

Content determination of five flavonoids in Tibetan medicine Rhododendron anthopogonoides by quantitative analysis of multi-components by single marker (QAMS) / 中国中药杂志

To establish a quantitative analysis of multi-components by single marker(QAMS) method for five flavonoids in Rhododendron anthopogonoides and verify its feasibility and applicability in the medicinal materials of R. anthopogonoides. With hyperoside as the internal reference, relative correction factors(RCF) of rutin, quercetin, quercitrin and kaempferol were established by high-performance liquid chromatography(HPLC) analysis. RCFs were used to calculate the content of each component, system durability and relative retention time. Simultaneously, QAMS and external standard method(ESM) were used to determine the content of five flavonoids in 12 batches of R. anthopogonoides from different origins. The results were statistically analyzed to verify the accuracy and feasibility. The fingerprints and cluster analysis data of R. anthopogonoides analyzed and discussed differences among the batches. According to the results, the RCFs of rutin, quercetin, quercetin and kaempferol in R. anthopogonoides were 1.242 6, 0.990 5, 0.535 0, and 0.781 3, respectively. The RCFs represented a good reproducibility under different experimental conditions. Besides, there was no significant difference between QAMS and ESM. Besides, the fingerprint and cluster analysis data showed the consistency between the classification and with the origin distribution of the herbs. In conclusion, the QAMS method shows a good stability and accuracy in the quality control of R. anthopogonoides.

Study on potential mechanism of hyperoside on improving ischemia/reperfusion injury based on network pharmacology / 中国药理学与毒理学杂志

OBJECTIVE To predict the potential targets of hyperoside (Hyp) on improving ischemia/reperfusion injury by network pharmacology, and explore its possible mechanism combined with related literature. METHODS The action targets of Hyp and ischemia/reperfusion injury were obtained by TCMSP, Swiss Target Prediction, Pharm Mapper, Simi?larity ensemble approach, Online Mendelian Inheritance in Man, DisGENT and database. The common targets of drugs and diseases were screened by Omishare and STRING database respectively, and the protein-protein interaction (PPI) network map was constructed. Then the interaction network between Hyp and disease targets was constructed by Cyto?scape software and topological cross-linking analysis was carried out. Then the interaction network between Hyp and disease targets was constructed and cross-linked analysis was carried out by using Cytoscape software. The gene ontol?ogy (GO) of the core target was analyzed by David database, and then the related pathways of the core target were enriched by KEGG database. RESULTS A total of 54 GO enrichment processes were obtained by GO enrichment anal?ysis of 44 common genes, including 38 biological processes (BP), 15 cell composition (CC) processes, and 1 molecular functional (MF) process. 43 items were obtained by signal pathway enrichment analysis in KEGG database. CONCLU?SION It is suggested that the mechanism of Hyp may be related to PI3K-Akt, RAP1, RAS, VEGF and other signal trans?duction pathways. The above results laid a theoretical foundation for the study of the mechanism and clinical application of the treatment of ischemia/reperfusion injury.

Hyperoside ameliorates the injury of heart and thoracic aorta in mice with myocardial infarction by regulating autophagy pathway / 中国临床药理学与治疗学

AIM: To explore the protection of Hyperoside (Hyp) on heart and thoracic aorta in mice with myocardial infarction (MI) as well as its potential mechanism. METHODS: The MI model was generated by a ligation on the left anterior descending coronary artery. The mice were then randomly divided into sham group (saline, 0.1 mg/10 g), model group (saline, 0.1 mg/10 g), Hyp-low, moderate and high concentration groups (Hyp, 9, 18 and 36 mg/kg), Fosinopril group (Fosinopril, 15 mg/kg), and Hyp-high concentration + 3-MA group (Hyp, 36 mg/kg; 3-MA, 30 mg/kg). The mice were treated with Hyp and Fosinopril for two weeks, and then the changes of heart weight versus body weight (HW/BW), electrocardiogram (ECG) remodeling, cardiac function, oxidative stress level in serum, thoracic aorta remodeling and endothelial function were investigated. RESULTS: In the model group, the HW/BW was elevated (P<0.01). The width of QRS in ECG was elevated, as a company with the reduction of height of QRS (P<0.01). The echocardiography assay showed that the cardiac cavity was enlarged (P<0.01). The oxidative stress level in serum was enhanced (P<0.01). The thoracic aorta remodeling and endothelial dysfunction became more serious (P<0.01). After being treated with different concentrations of Hyp and fosinopril for two weeks, the changes above were reserved (P<0.05, P<0.01). Co-treatment with 3-MA, an autophagy inhibitor, suppressed the protective effects of Hyp on impaired hearts and vessels (P<0.05, P<0.01). CONCLUSION: Hyp has protective prospects on injured heart and thoracic aorta remodeling, as well as endothelial dysfunction in MI mice; 3-MA, an autophagy inhibitor, can reverse the cardiovascular protection of Hyp. The mechanism may be explained that Hyp can elevate autophagy level in heart, which further weaken oxidative injury in MI mice.

Evaluation on content difference of multi-index components between Yao medicine Young Leonurus heterophyllus and Leonurus japonicus / 中草药

Objective: To study the content difference of multi-index components between Yao medicine Young Leonurus heterophyllus (YLH) and Leonurus japonicus, and evaluate the quality with the content. Methods: A high performance liquid chromatography (HPLC) method was established for the simultaneous content determination of five chemical constituents of chlorogenic acid, leonurine hydrochloride, rutin, hyperin and isoquercitrin in Yao medicine YLH and Leonurus japonicus. SPSS 25.0 statistical software was used to conduct paired samples t-test and one-way ANOVA to infer the content differences among the samples. Principal component analysis (PCA) was used to evaluate the quality of medicinal materials. Results: T-test results of paired samples of Yao medicine YLH and Leonurus japonicus showed that there was significant difference in rutin content among the five chemical constituents, with no significant difference in other constituents. The results of one-way ANOVA analysis showed that there were significant differences in five chemical constituents of Yao medicine YLH and Leonurus japonicus from different habitats (P < 0.01). The results of principal component analysis showed that the higher scores of Leonurus japonicus from different habitats were from Nanning and Yulin, with the lowest in Shangsi County. Conclusion: There is a significant difference between the content of Yao medicine YLH and Leonurus japonicus. The quality of medicinal materials from Nanning and Yulin is better. The results provide data support for the comparison of the content between two kinds of Leonurus heterophyllus and Leonurus japonicus from different habitats.

Quality control of Prunella vulgaris based on HPLC fingerprints combined with chemical pattern recognition and content determination / 中草药

Objective: The quantitative method for the quality makers screened by a chemical pattern recognition method combined with the HPLC fingerprint was established, so as to provide reference for scientific and comprehensive quality evaluation of Prunella vulgaris. Methods: Fingerprints of 30 batches of P. vulgaris were established by HPLC. Similarity evaluation was performed by using Similarity Evaluation System for Fingerprint Chromatogram of TCM (2004A) to confirm the common peak. Principal component analysis and orthogonal partial least squares discriminant analysis were used to screen out the components that caused the quality differences in the batches. Quantitative method of the screened quality makers was established, and its content in 61 batches of samples was determined. Results: The HPLC fingerprints of P. vulgaris were obtained, and a total of 28 common peaks were marked. The similarity of 30 batches of samples was higher than 0.970, which indicated that the overall quality of P. vulgaris was relatively stable. Caffeic acid (F5), hyperoside (F9), isoquercitrin (F10), salviaflaside (F11), and rosmarinic acid (F12) were recognized as the quality makers using principal component analysis and orthogonal partial least squares discriminant analysis. Five markers, which showed good linear relationship, were used as indicators for content determination. The average recovery was 95.0%-105.0%, with the RSD value less than 3%. Conclusion: The analysis method established was scientific, accurate, and reliable. A more perfect, reasonable, and effective method for quality evaluation of P. vulgaris was constructed using a fingerprint combined with chemical pattern recognition technique.

Fingerprint and molecular identification of rbcL sequence of Mongolian medicine Althaeae Roseae Flos by HPLC / 中草药

Objective: To establish the fingerprint of Althaeae Roseae Flos by HPLC and the molecular identification method of DNA barcode of rbcL sequence. Methods: The fingerprint establishment of Althaeae Roseae Flos was performed on Welchrom Column C18 (300 mm × 4.6 mm, 5 μm) with acetonitrile - 0.1% formic acid solution as mobile phase for gradient elution, with flow rate of 1.0 mL/min, column temperature of 35 ℃, detection wavelength of 365 nm, injection volume of 10 μL. DNA barcode molecular identification method was used for PCR amplification and determination of rbcL sequence. Results: The fingerprints of 11 samples were established, 21 common peaks were obtained, their similarities were calculated, and four components (hyperoside, quercetin, apigenin and kaempferide) were determined. The total length of rbcL sequence of 11 samples was measured, and the G + C content was 44.10%-44.40% and genetic distance (K2P) was 0.001 4. There were 10 ectopic points and the similarity was 99.00%. Conclusion: The two methods are stable and reliable, which can provide basis for the identification and quality control of A. rosea.

Optimization of pH response and mitochondrial targeting bifunctional hyperoside liposomes by central composite design response surface methodology and its in vitro evaluation / 中草药

Objective: To optimize preparation of mitochondrial targeting hyperoside liposomes (DLD/Hyp-Lip), and study its stability in fetal bovine serum, in vitro release behavior and mitochondrial targeting. Methods: DLD/Hyp-lip was prepared by film dispersion method. Single factor experiment was carried out with entrapment efficiency and drug loading as indexes to investigate the effects of the ratio of phospholipids to hyperoside (Hyp) and DSPE-PEG (distearoyl phosphoethanolamine-polyethylene glycol) to DLD on DLD/Hyp-Lip. The formulation of DLD/Hyp-Lip was further optimized by central composite design response surface methodology. The appearance, size and potential of liposomes were observed by transmission electron microscope and particle size analyzer. The stability and drug release rate of liposomes in fetal bovine serum were evaluated by serum stability test and in vitro drug release test. The drug delivery system was evaluated by mitochondrial targeting. Results: The optimal formula of DLD/ Hyp-Lip was as follows: the ratio of total phospholipids to hyperoside was 12.50:1, the ratio of total phospholipids to cholesterol was 6.00:1, and the dosage ratio of DSPE-PEG to DLD was 3:5, the encapsulation efficiency was (95.57 ± 0.56) %, the drug loading was (8.55 ± 0.57) %. The prepared liposomes had good appearance, the particle size of the lip was (124.9 ± 3.4) nm, and the potential was (-6.2 ± 1.9) mV. It was stable in fetal bovine serum and accumulated in vitro release medium for 24 h. Mitochondrial targeting experiments showed that DLD/Hyp-Lip could promote the accumulation of drugs in the mitochondria. Conclusion: This method is simple and convenient, and can accurately and effectively optimize the preparation process of DLD/Hyp-Lip. The prepared DLD/Hyp-Lip has high encapsulation efficiency, small particle size, uniform distribution and good sustained-release effect, which lays the foundation for further in vivo research of DLD/Hyp-Lip. DLD/Hyp-Lip with hyperoside has good mitochondrial targeting of liver cancer cells and is a potentially efficient mitochondrial targeted drug delivery system for liver cancer cells.

HPLC characteristic chromatograms and quantitative analysis of Kunxian Capsules / 中草药

Objective: In order to provide a scientific basis for the quality control of Kunxian Capsules (KC), HPLC characteristics chromatogram combined with multi-components determination was established. Methods: The analysis was performed on Agilent Zorbax SB-C18 column (250 mm × 4.6 mm, 5 μm), using acetonitrile and 0.1% phosphoric acid solution as the mobile phase at a flow rate of 0.8 mL/min for gradient elution, the column temperature was 33 ℃, and the detection wavelength was 270 nm. The fingerprints of 15 batches of KC were established and evaluated by the similarity evaluation system of TCM (2012A version), hierarchical cluster analysis and discriminant analysis of partial least squares. Furthermore, the content of hyperoside, epimedin A, epimedin B, epimedin C, icariin and baohuoside Ⅰ were determined. Results: The HPLC fingerprint with 21 common peaks of KC was established, and the similarities of samples were over 0.9. The linearity relationships separated with hyperoside, epimedin A, epimedin B, epimedin C, icariin and baohuoside Ⅰ were good, and the contents of the above-mentioned components in 15 batches of preparations were 2.817-7.527, 7.287-9.103, 8.730-18.675, 33.377-70.371, 35.297-50.291 and 4.059-9.079 mg/g, respectively. Conclusion: The combination methods of HPLC characteristic chromatograms and simultaneous determinations of multiple components are rapid, simple and reproducible, which can provide methodological reference for the quality control of KC.

Optimization of the Extraction Technology of Senecio scandens by Orthogonal Design Combined with Informa- tion Entropy Weighting Method / 中国药房

OBJECTIVE：To optimize the extraction technology of Senecio scandens ，and to provide reference for the further development and utilization of the medicinal material. METHODS ：The method of reflux extraction with 80 ℃ water bath was used to extract S. scandens . HPLC method was adopted to determine the contents of chlorogenic acid and hyperoside. The determination was performed on Diamonsil C 18 with mobile phase consisted of 0.2％ glacial acetic acid water solution-methanol （82∶18，V/V）at the flow rate of 1.0 mL/min；the column temperature was set at 35 ℃；the detection wavelength was 327 nm，and the sample size was 5 μL. UV-Vis spectrophotometry was used to determine the contents of total flavonoids（by rutin ）and total alkaloids （by matrine）and the detection wavelengths were set at 509 nm and 208 nm，respectively. Based on single factor tests ，using ethanol volume fraction （A，％），solvent folds （B，mL/g），extraction time （C，h），extraction times （D，times）as factors ，using the contents of chlorogenic acid ，hyperoside，total flavonoids and total alkaloids as indexes ，and t he weight coefficients of above indicator components were calculated based on information entropy weighting method so as to calculate their comprehensive scores ， then L 9（34）orthogonal design was used to further optimize the extraction technology. The validation tests were also performed. RESULTS：The optimal extraction technology of S. scandens included that 8-fold 50％ ethanol，extracting for 3 times，lasting for 1.5 h each time. Results of 3 times of validation tests showed that RSDs of the contents of chlorogenic acid ，hyperoside，total flavonoids and total alkaloids were all lower than 1.5％（n＝3）. CONCLUSIONS ：The optimized technology is reproducible ， stable and feasible ，and can be used for the extraction of S. scandens .

Chemical constituents of underground parts of Astragalus tatsienensis var. incanus / 中草药

Objective To study the chemical constituents of the underground parts of Astragalus tatsienensis var. incanus. Methods The compounds were separated and purified by RP18, Sephadex LH-20 column chromatography, as well as preparative liquid chromatography. The structures of the isolated compounds were determined by means of modern spectroscopic analysis and physicochemical properties. Results Four compounds were isolated from the methanol extract of the underground part of A. tatsienensis var. incanus, and identified as 3β-O-α-L-rhamnopyranosyl (1→2)-β-D-galactopyranosyl (1→2)-β-D-glucuronopyranosyl- 22β-hydroxy-11-oxo-olean-12-ene (1), hyperoside (2), wighteone (3), and 4-hydroxybenzoic acid (4). Conclusion Compounds 2-4 are isolated from this plant for the first time, and compound 1 is a new triterpene glycosides compound, named astratatincoside A.

Optimization of extraction of total flavonoids from leaves of Psidium guajava by orthogonal test based on information entropy weighting / 中草药

Objective: To optimize the extraction of total flavonoids from leaves of Psidium guajava. Methods: L9(34) orthogonal test was used to screen the optimal extraction technology of total flavonoids from leaves of P. guajava with ethanol concentration, solid-liquid ratio, extraction time, and extraction times as factors. The weight coefficient of the four evaluation indicators, including the yield of total flavonoids, hyperoside, quercetin-3-O-β-D-pyranarabinoside, and quercetin-3-O-α-L-furanarabinoside was calculated by information entropy weighting so as to calculate comprehensive score. We obtained the optimal technology by orthogonal analysis based on the comprehensive score. Results: The optimal extraction technology was that using 8-fold 70% ethanol water to extract 2 h for three times. The mean comprehensive score of the three batches was 0.142 1 and the RSD was 2.37%. Conclusion: The optimal extraction technology of total flavonoids from leaves of P. guajava was stable and feasible with high yield.

Enhancement effects of hyperoside on killing activity of human NK cells against pancreatic cancer PANC1 cells / 中华胰腺病杂志

Objective To investigate the effect of hyperoside on proliferation and killing activity of NK cells against pancreatic cancer PANC1 cells in vitro,and explore its potential mechanism.Methods Peripheral blood mononuclear cells of healthy donors were isolated,NK cells were induced with medium contained with IL-2 and different concentrations of hyperoside (0.3,1.6,8,40 and 200 μg/ml) for 12 days.Cell viability was observed by trypan blue staining.Phenotype and perforin,granzyme B expression of NK cells were detected by flow cytometry.Killing activity of NK cells against PANC1 cells were analyzed with lactate dehydrogenase (LDH) releasing method.Results The proportion of NK cells in control group and experimental group treated with different concentration of hyperoside both reached about 80％,respectively.The proliferation of CDs-CD56 + NK cells treated by hyperoside at 0.3,1.6 and 8 μg/ml was (93.76 ±8.77),(106.67 ± 12.35) and (118.50 ± 11.51) times,respectively,which were significantly higher than (73.70 ± 9.43) times of the control group.The expressions of perforin in NK cells treated with hyperoside at 1.6,8 and 40 μg/ml were significantly higher than those of the control group [(82.34 ± 2.90) ％,(89.15 ±3.54) ％,(81.78 ± 2.81)％ vs (72.93 ± 2.06)％].The expressions of granzyme B in NK cells treated with hyperoside at 1.6 and 8 μg/ml were significantly higher than those of the control group [(87.30 ± 1.70) ％,(92.16 ±3.05)％ vs (82.35 ±2.73)％].The killing activity of NK cells against PANC1 cells treated by hyperoside at 1.6 and 8 μg/ml was significantly higher than those of the control group [(63.18 ± 3.77)％,(65.34 ± 4.97) ％ vs (52.16 ± 5.48) ％].The differences were statistically significant (all P ＜ 0.05).Conclusions Hyperoside could promote the proliferation of NK cells at certain concentrations and maybe enhance the killing effect against pancreatic cancer PANC1 cells through up-regulating the expression of perforin and granzyme B in NK cells.

Effects of hyperoside on the myocardical activities of ATPases and expressions of Cx43 and Kir2.1 in arrhythmia rats induced by ischemia-reperfusion / 中成药

AIM To investigate the effects of hyperoside,an anti-arrhythmic agent capable of reducing myocardial infarct size,on arrhythmic rats induced by ischemia-reperfusion (I/R) and the corresponding mechanism.METHODS Male SD rats were randomly assigned to sham operation group,model group and hyperoside group (50 mg/kg,n =15).The I/R model was reconstructed by the ligation of left anterior descending coronary artery for 30 min ischemia.Rats in the hyperoside group were injected with 50 mg/kg hyperoside intraperitoneally 10 min before ischemia.Heart rate,mean arterial pressure (MAP) and heart rate systolic blood pressure product (RPP) at time points of 10 min before ischemia (T0),30 min after ischemia (T1),30 min (T2),60 min (T3),120 min (T4) after reperfusion were recorded.ELISA method was used to determine serum CK-MB and cTnI,spectrophotometry to measure Na+-K+-ATPase and Ca2+-Mg2+-ATPase levels,HE staining to observe myocardial tissue changes,immunohistochemistry to investigate Cx43 protein,and Western blot to detect Kir2.1 protein expression.RESULTS At T1,T2,T3 and T4,the model group demonstrated significantly lower HR,MAP and RPP than those in the sham operation group (P ＜ 0.05),whereas the hyperoside group had higher HR,MAP and RPP than the model group.Both hyperoside group and the model group shared significantly higher arrhythmia score,levels of CK-MB and cTnI than the sham operation group (P ＜ 0.05) while their lower activities of Na +-K +-ATPase and Ca2+-Mg2+-ATPase,protein expressions of Cx43 and Kir2.1 than the sham operation group were noticed as well.But the hyperoside group displayed its lower arrhythmia score,levels of CK-MB and cTnI,and yet higher activities of Na +-K +-ATPase and Ca2 +-Mg2 +-ATPase,protein expressions of Cx43 and Kir2.1 than the model group (P ＜0.05).CONCLUSION Hyperoside in improving ventricular arrhythmia of I/R rats may contribute to the activity restoration of Na +-K +-ATPase and Ca2 +-Mg2 +-ATPase,and the up-regulation of Cx43 and Kir2.1 protein expressions.

Determination of seven effective constituents in crude and processed Cuscuta australis by quantitative analysis of multi-components with single marker / 中草药

Objective To establish a quantitative analysis of multi-components with single marker (QAMS) for the simultaneous determination of chlorogenic acid, cryptochlorogenin acid, caffeic acid, hyperoside, isoquercitrin, astragalin, kaempferol in crude and processed Cuscuta australis, which is proved to be a scientific and feasible method in the quality analysis in C. australis. Methods Six relative correction factors (RCFs) of chlorogenic acid, cryptochlorogenin acid, caffeic acid, isoquercitrin, astragalin, kaempferol was established in the HPLC method with the hyperoside as the internal standard (IS), which was to calculate the mass fraction of each. The mass fraction of seven effective constituents in crude and processed C. australis was calculated by the external standard method (ESM) at the same time. Compared with the content results determined by the ESM and QAMS, the feasibility and accuracy of QAMS method were verified. Results The relative correction factor (RCF) was perfect. The detection calculated by QAMS was consistent with the results by ESM. Conclusion The method with a single marker, using the hyperoside as IS, is accurate and feasible for the quantitative analysis of six other effective constituents in C. australis.

Chemical constituents from Simiao Yongan Decoction / 中草药

Objective: To study the chemical constituents of Simiao Yongan Decoction. Methods: The silica gel, ODS, Sephadex LH-20, and AB-8 macroporous adsorption resin column chromatography were used to isolate and purify the chemical constituents from Simiao Yongan Decoction. The structures of the constituents were identified on the basis of physiochemical properties, NMR, MS, and other data. Results: Twenty-two compounds were isolated and identified as 5(S)-5-carboxystrictosidine (1), harpagoside (2), geniposide (3), glycyrrhetinic acid (4), glycyrrhizic acid (5), hyperoside (6), liquiritin (7), isoliquiritoside (8), liquiritigenin (9), isoliquiritigenin (10), luteolin (11), quercetin (12), 2-(3-hydroxy-4-methoxyphenyl)ethyl O-α-arabinopyranosyl-(1→6)-O-α- rhamnopyranosyl-(1→3)-O-β-glucopyranoside (13), angroside C (14), acteoside (15), cinnamic acid (16), ferulic acid (17), (E)-aldosecologanin (18), protocatechuic acid (19), stigmasterol (20), hentriacontanol (21), and daucosterol (22). Conclusion: Compounds 1-3, 5, 6, 8, 11, 13-15, 18-21 are isolated from Simiao Yongan Decoction for the first time.

Spectrum-effect relationship of anti-inflammatory effect of different part from Polygonum capitatum / 中草药

Objective To study the spectrum effect relationship between the water and ethanol extracts of Polygonum capitatum and its anti-inflammatory activity. Methods The inflammatory model of mice RAW264.7 cells induced by lipopolysaccharide (LPS) was established, and the release of inflammatory factors was detected by NO and TNF-α kit. The relationship of chemical constituents of P. capitatum was established by using the partial least square method to associate the fingerprint data established by UPLC-MS method and the pharmacodynamics index of different extracts of P. capitatum. Results Different extracts of P. capitatum had no cytotoxicity in the range of concentration less than 250 mg/L, and all of them had anti-inflammatory effect, which had a dose-effect relationship. By using partial least square method, the correlation degree between TNF-α and chromatographic peak was compared. The results showed that the correlation among the peaks 24, 17, 22, 23, 20 and the anti-inflammatory effect was positive, and the anti-inflammatory effect was negatively correlated with the peaks 11, 1, 7, 15, 5, 3. From the analysis of the five peaks, the results showed that all the four peaks were flavonoids except for the tannic acid at peak 17. Conclusion Different extracts of P. capitatum inhibited the inflammatory response of RAW264.7 cells. The flavonoids in P. capitatum had significant anti-inflammatory activity. The results showed that quercetin, tannic acid, and hyperoside had great contribution to the anti-inflammatory effect of P. capitatum.

Determination of ten flavones in Kunxian Capsules with quantitative analysis of multi-components by single marker / 中草药

Objective To establish quantitative analysis of multi-components by single marker (QAMS) method for simultaneous determining the content of ten flavonoids in Kunxian Capsules (KC), and evaluate the adaptation and application of QAMS method in the quality control of KC. Methods The relative factor (fs/i) of epimedin A, epimedin C, epimedin B, icariin, luteolin, quercetin, nobiletin, kaempferol and baohuoside I were established by HPLC method with hyperoside as internal standard, which were used to calculate the content of ten flavonoids in the samples of KC. Meanwhile, external standard method (ESM) was used to calculate the content of ten flavonoids. The difference between QAMS and ESM were analyzed to evaluate the accuracy of QAMS. T-test was used to compare the content of ten flavonoids in KC. Results The fs/i of epimedin A, epimedin C, epimedin B, icariin, luteolin, quercetin, nobiletin, kaempferol, and baohuoside I were 0.756 5, 2.199 4, 1.232 7, 1.008 5, 0.635 7, 0.576 0, 0.487 5, 0.545 6, 0.675 8. The content determination results of ten batches of KC samples were calculated by the method of QAMS and ESM, with no significant difference in RSD < 2.0%. Conclusion The fs/i established in the QAMS method with hyperoside as the internal reference substance is accurate and feasible. The QAMS method can be used for the quality evaluation of KC.