Qualitative and quantitative study of constituents in Lysionoti Herba based on UHPLC-Q-Exactive Orbitrap HRMS and HPLC-UV / 中国中药杂志

Ultra-high performance liquid chromatography-quadrupole-Exactive Orbitrap high resolution mass spectrometry(UHPLC-Q-Exactive Orbitrap HRMS) was employed to systematically analyze the chemical constituents in Lysionoti Herba, and high perfor-mance liquid chromatography-ultraviolet(HPLC-UV) to determine the content of main compounds. A Synergi~(TM) Hydro-RP 100 Å colu-mn(2 mm×100 mm, 2.5 μm) was used for gradient elution with acetonitrile-0.1% aqueous formic acid as the mobile phase at a flow rate of 0.2 mL·min~(-1) and a column temperature of 40 ℃. MS and MS/MS were conducted with electrospray ionization(ESI) in both positive and negative modes. The chemical components in Lysionoti Herba were identified by comparison with the retention time and mass spectra of reference compounds and the relevant mass spectral data reported in MS databases and relevant literature. Furthermore, the content of five constituents(neochlorogenic acid, chlorogenic acid, forsythoside B, acteoside, and nevadensin) in different Lysiono-ti Herba samples was simultaneously determined by HPLC-UV at the wavelength of 330 nm. A total of 84 compounds were identified in Lysionoti Herba, including 27 flavonoids, 20 phenylethanoid glycosides, 5 amino acids, 18 organic acids, 1 alkaloid, 6 nucleosides, and 7 others. The content of neochlorogenic acid, chlorogenic acid, forsythoside B, acteoside, and nevadensin showed good linear relationship(r>0.999) with the peak area within certain concentration ranges, which were 3.22-102.90, 12.84-410.82, 31.63-1 012.01, 25.00-800.11, and 4.08-130.51 μg·mL~(-1), respectively. The instrument precision, method repeatability, and solution stability all met requirement, and the average recovery rate was 97.31%-100.2%, with RSD ranging from 0.95% to 2.4%. The content of the five components varied among different Lysionoti Herba samples collected from different regions of Guizhou, and the average content of forsythoside B was the highest. The established qualitative method can rapidly and efficiently identify the chemical components of Lysionoti Herba, and the developed HPLC-UV method can simultaneously determine the content of five components in a simple, ra-pid, and accurate manner, providing a scientific basis for the quality evaluation of Lysionoti Herba.

Effective Components of Periploca forrestii Against Rheumatoid Arthritis by Targeting TNF-α / 中国实验方剂学杂志

ObjectiveTo analyze the effective components of Periploca forrestii against rheumatoid arthritis（RA）by targeting tumor necrosis factor （TNF）-α based on network pharmacology and experimental verification. MethodThe preliminary research of the research group found that the alcohol extracts of P. forrestii （CDLF and CQAF） had significant anti-RA activities，and 10 monomers with such activities were identified. The anti-RA activities of active monomers，CDLF， and CQAF were compared by the enzyme-linked immunosorbent assay （ELISA）with interleukin（IL）-6，nitric oxide （NO），IL-1β， and prostaglandin E2（PGE2）as indicators. Network pharmacology was employed to analyze the possible molecular mechanism of P. forrestii against RA. The targeting ability of P. forrestii chemical monomers to TNF-α was verified by TNF-α molecular docking，surface plasmon resonance （SPR）， and TNF-α-induced L929 injury model. ResultELISA showed that the anti-RA activities of CDLF and CQAF were significantly stronger than those of identified 10 active monomers. Network pharmacology analysis showed that the core targets of P. forrestii against RA were signal transducer and activator of transcription protein 3 （STAT3），TNF， and IL-6. Gene Ontology（GO） analysis revealed collagen catabolism，inflammatory response，positive regulation of nuclear factor kappa-B（NF-κB） transcription factor activity，and positive regulation of B cell proliferation. Kyoto Encyclopedia of Genes and Genomes （EKGG） pathway enrichment analysis demonstrated TNF signaling pathway，phosphoinositide 3-kinase（PI3K）/protein kinase B（Akt） signaling pathway，NF-κB signaling pathway，Toll-like receptor signaling pathway，mitogen-activated protein kinase（MAPK） signaling pathway， etc. Verification experiments by TNF-α molecular docking，SPR， and TNF-α-induced L929 injury model found that CDLF and CQAF had good binding activities and could manifestly antagonize TNF-α. However， the active components separated and identified from CDLF and CQAF did not show the same anti-TNF-α activity. ConclusionThe CDLF and CQAF of P. forrestii may treat RA by targeting TNF-α. The experiments found that the isolated chemical components had weaker binding activity to TNF-α than CDLF and CQAF. Meanwhile，the research group isolated chemical components with a minimum mass fraction of 0.25 ng·g-1 from P. forrestii， which suggested that the active components generated by binding to TNF-α with anti-RA activities were presumedly trace components .

Proteomics Analysis of Mechanism of Shenxiong Glucose Injection in Antagonizing H2O2-Induced Oxidative Damage in H9c2 Cells / 中国实验方剂学杂志

ObjectiveTo explore the mechanism of Shenxiong glucose injection （SGI） in inhibiting hydrogen peroxide （H2O2）-induced oxidative damage in H9c2 cells by tandem mass tags （TMT）-labeled quantitative proteomics. MethodH9c2 cells cultured in vitro were exposed to H2O2 for inducing oxidative damage. The cell viability was measured by cell proliferation and cytotoxicity assay （MTS）， followed by peptide fractionation by high performance liquid chromatography （HPLC） and protein expression detection in H9c2 cells by ultrahigh performance liquid chromatography-mass spectrometry. MaxQuant （v1.5.2.8） was utilized for data retrieval， and the high-resolution mass spectrometry was conducted to screen out differentially expressed proteins， which were then subjected to gene ontology （GO） and Kyoto encyclopedia of genes and genomes （KEGG） enrichment analysis. The protein expression levels of perilipin 2 （Plin2） and tropomyosin 1 （Tpm1） in cells were measured by Western blot. ResultThe spectral analysis yielded 48 608 specific peptide fragments and 5 903 quantifiable proteins. Compared with the model group，the SGI group exhibited 82 differentially expressed proteins，of which 22 were up-regulated and 60 were down-regulated. GO analysis results showed that the differentially expressed proteins were mainly involved in biological processes such as programmed cell death regulation，regulation of cell proliferation，cardiovascular system development， and cell migration. As revealed by KEGG analysis， these proteins were mainly related to peroxisome proliferator-activated receptor （PPAR），focal adhesion，phosphatidylinositol 3-kinase/protein kinase B（PI3K/Akt），and Ras-related protein 1 （Rap1） pathways. Western blot results demonstrated that compared with the model group，SGI significantly increased the Plin2 protein expression and decreased the Tpm1 protein expression （P<0.01），consistent with the proteomics results. ConclusionSGI may inhibit cell apoptosis and antagonize H2O2-induced cell oxidative damage by regulating PPAR，focal adhesion，PI3K/Akt and Rap1 pathways，which should be further verified by subsequent experiments.

Intestinal absorption characteristics of Polygonum orientale extract in normal and isoproterenol-induced myocardial ischemia model rats via everted intestinal sac models / 中国中药杂志

The present study is to investigate the absorption characteristics of the main components in Polygonum orientale extract in normal and isoproterenol-induced myocardial ischemia model rats with everted intestinal sac models. Intestinal sac fluid samples were collected in different part of intestine(duodenum, jejunum, ileum, colon) at different time after administration of different concentration of P. orientale extract(5.0,10.0, 20.0 mg·mL~(-1)). An UPLC-TQD method was employed for the determination of six components including orientin, isoorientin, vitexin, protocatechuic acid, kaempferol-3-O-β-D-glucoside and quercitrin in the intestinal sac samples. The absorption rate and cumulative absorption were calculated to analyze the intestinal absorption characteristics of six components in normal and myocardial ischemia model rats. The P-glycoprotein(P-gp) inhibitor was applied to investigate influence of intestinal absorption of six components in P. orientale extract. The results showed that the main absorption sites were concentrated on the duodenum at low concentration, while they were the colon at the medium concentration and the ileum at high concentration in control groups. In the condition of myocardial ischemia model, the main absorption sites focus on the ileum and jejunum at low concentration; the main absorption sites were in the ileum at the medium concentration and main absorption sites were the duodenum and ileum at high concentration. Compared with the normal group, the absorption rate and cumulative absorption of the six components significantly decreased in the model group. P-gp inhibitor markedly increased the absorption rate and cumulative absorption of six components in the model group, inferring that the 6 components may be the substrates of P-gp, and the mechanism needs further study. In this study, it is revealed that the six components of P. orientale extract can be absorbed into the intestinal sac, and it is an effective method to assess the intestinal absorption characteristics of P. orientale extract through everted intestinal sac model, providing data support for the clinical application and further development of P. orientale.

Effect of Aidi Injection on Expression of Cytochrome P450 Enzyme in DEN-induced Hepatocellular Carcinoma Rats / 中国实验方剂学杂志

Objective:To study the effect of Aidi injection （AD） on the expression of cytochrome P450 isoenzyme 1A2，2E1，3A2，2C11（CYP1A2，2E1，3A2，2C11）mRNA and protein in rats with N-nitrosodiethylamine （DEN） chemically induced primary hepatocellular carcinoma（HCC）. Method:Three healthy SD male rats were randomly selected as the blank group， and the remaining rats were treated with DEN intermittently induced primary hepatocellular carcinoma rat model. After success of the model， the rats were randomly divided into model group and AD group， with 3 rats in each group. The rats in the blank group and model group were intraperitoneally injected with 10 mL·kg^-1 saline， while those in the AD group were intraperitoneally injected with 10 mL·kg^-1AD once a day， a total of 14 d intervention. Real-time quantitative polymerase chain reaction（Real-time PCR） and Western blot were used to detect the mRNA and protein expressions of CYP1A2， CYP2E1， CYP3A2 and CYP2C11， respectively. Result:Real-time PCR results showed that after 14 days of drug administration， compared with the blank group， the mRNA expressions of CYP1A2， CYP2E1， CYP3A2 and CYP2C11 were all down-regulated in para-cancerous tissue （PCT） and cancerous tissue （CT） in model group， and there were significant differences （<italic>P</italic><0.05，<italic>P<</italic>0.01）. Compared with the model group， the mRNA expressions of the four subtype enzyme were significantly down-regulated in PCT in the AD group（<italic>P</italic><0.05，<italic>P<</italic>0.01）， while the mRNA expressions of the four subtype enzyme were significantly up-regulated in CT （<italic>P</italic><0.05）， and the expression was down-regulated overall. Western blot results showed that compared with the blank group， the protein expressions of CYP1A2， CYP2E1， CYP3A2 and CYP2C11 in CT of the model group were significantly down-regulated （<italic>P</italic><0.01）， and the protein expressions of CYP3A2 and CYP2C11 were significantly down-regulated in PCT （<italic>P</italic><0.01）. Compared with the model group， the protein expressions of CYP1A2， CYP2E1， CYP3A2 and CYP2C11 in CT and PCT were down-regulated in the AD group， but the differences were not statistically significant. Conclusion:AD can down-regulate the mRNA and protein expressions of CYP1A2， CYP2E1， CYP3A2 and CYP2C11 in rat liver tissues. In clinical use of AD， attention should be paid to drug interactions that may be caused by CYP450 enzyme inhibition.

Pharmacokinetic study of Polygonum orientale extract in H9c2 cells by UPLC-MS/MS / 中国中药杂志

A detection method of ultra-performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS) was established to detect concentrations of isoorientin, orientin, quercetin, vitexin and kaempferol-3-O-β-D-glucoside in H9 c2 cells and applied to the pharmacokinetic study of Polygonum orientale extract in the cells. H9 c2 cells were treated with 100 μg·mL~(-1) P. orientale extract and then they and the corresponding nuclei, mitochondria and Golgi bodies were collected at the set time. After protein precipitation, UPLC-MS/MS was used to determine concentrations of isoorientin, orientin, quercetin, vitexin and kaempferol-3-O-β-D-glucoside in the whole cells and subcellular structures. Also, related pharmacokinetic parameters were calculated. The results showed that the peak time was 8 h for all these components. Orientin, vitexin, quercetin and isoorientin have high affinities to nuclei and mitochondria, while the affinity of kaempferol-3-O-β-D-glucoside is higher with mitochondria compared to nuclei. It is suggested that these chemical components of P. orientale may mainly act on nuclei or mitochondria to exert pharmacological effects of protecting cardiomyocytes.

Simultaneous determination of four components of Shuganning Injection in rat plasma by UPLC-MS/MS and its application to a pharmacokinetic study / 中国中药杂志

This study aims to reveal the pharmacokinetics of Shuganning Injection in normal rats. In this experiment,ultra-high performance liquid chromatography-electrospray-tandem mass spectrometry( UPLC-ESI-MS/MS) was used to establish an analytical method for simultaneous determination of chlorogenic acid,gardenioside,oroxylin A and baicalin in rat plasma. Then,the non-compartmental model( NCA) in Phoenix WinN onL in 6. 4 software was used to fit pharmacokinetic parameters. The methodological validation showed that the linear relationship of the components in rat plasma samples were good( r>0. 995). The recovery rate and matrix effect of plasma samples with low,middle and high concentration were 79. 14%-101. 4%. The intra-day and inter-day precision,accuracy and stability meet the requirements of biological sample analysis. The half-life( t1/2) of chlorogenic acid,gardenioside,oroxylin A did not change significantly and the area under blood concentration-time curve( AUC0-t) is proportional to the dose,which suggested that three components showed a linear kinetic characteristics,but baicalin showed nonlinear kinetic characteristics. Moreover,the retention time of each component in rats was short. The established UPLC-MS/MS quantitative analysis method is rapid,sensitive and accurate,which can be used for the determination of chlorogenic acid,gardenioside,oroxylin A and baicalin in rat plasma and pharmacokinetic study of Shuganning Injection.

Absorptive characteristic of Bletilla striata extract in Caco-2 cell monolayer / 中国中药杂志

To investigate the active fraction from Bletilla striata in Caco-2 cell monolayer,so as to explore its absorption mechanism of oral administration preliminarily.Active fraction from B.striata in Caco-2 cell monolayer was analyzed by UPLC-Q-TOF and detected by UPLC-MS/MS,and the effects of different concentrations,pH and P-glycoprotein inhibitors on Caco-2 cells Monolayer were investigated.Six compounds were isolated from the active fraction of B.striata in Caco-2 cell monolayer by UPLC-Q-TOF,and identified as B6,B12,B14,B17,B19 and B23,with concentration dependence.Within the 0-180 min,the uptake of B12 and B14 had a time dependence,while B6,B17,B19 and B23 tended to saturate after 60 min.All of the components had a good absorption in an acidic environment.B6 had a good absorption at pH 6.0,while the other components B12,B14,B17,B19 and B23 had a good absorption at pH4.0.The absorption of the 6 main components of B.striata were not be affected by P-glycoprotein inhibitors(verapamil/cyclosporin A).Compared with the control group,there was no difference in the absorption of B6 and B12,and the absorption of B14,B17,B19 and B23 increased,but with no significant difference.The absorption characteristic of B.striata extract across the Caco-2 cell monolayer is probably passive diffusion,and the absorption process was not affected by P-glycoprotein.

Shenxiong Glucose Injection inhibits H_2O_2-induced apoptosis of H9c2 cells by activating PI3K/AKT pathway / 中国中药杂志

The aim of this paper was to explore the mechanism of Shenxiong Glucose Injection antagonizing apoptosis of H9 c2 cells induced by H_2O_2. H9 c2 cells were pretreated with 1. 7%,3. 4% and 6. 8% Shenxiong Glucose Injection,and then H_2O_2 was introduced to induce apoptosis in vitro. Cell viability was detected by MTS assay,morphological changes of apoptosis were observed by AO/EB fluorescence staining,apoptosis rate was detected by Annexin/PI method,cell expression profile was detected by gene chip technology,the mRNA of PIK3 CA,Bcl-2,Bax,caspase-3 and GAPDH were detected by qRT-PCR,the protein expression levels of PIK3 CA,AKT,P-AKT,Bcl-2,Bax and caspase-3 were detected by Western blot,and the contents of LDH and MDA were detected by kit. The results showed that Shenxiong Glucose Injection of different concentrations significantly increased the viability of H9 c2 cells treated with H_2O_2( P<0. 01),and reversed H_2O_2-induced apoptosis( P< 0. 01). The microarray experiments showed that 138 genes were altered in H9 c2 cells after treatment with Shenxiong Glucose Injection. The differential expression fold of PIK3 CA associated with PI3 K/AKT pathway was 3. 59. The results of qRT-PCR and Western blot showed that Shenxiong Glucose Injection could down-regulate the mRNA and protein expression levels of caspase-3( P<0. 01),up-regulate the mRNA and protein expression level of PIK3 CA and Bcl-2( P<0. 01),and up-regulate the phosphorylation levels of AKT( P<0. 01) in H_2O_2-treated H9 c2 cells. The protective effect of Shenxiong Glucose Injection on H_2O_2 cells injury was significantly inhibited by LY294002,a PI3 K/AKT pathway inhibitor. The results suggested that Shenxiong Glucose Injection may inhibit H_2O_2-induced H9 c2 cells apoptosis by regulating PI3 K/AKT signaling pathway.

Effect of Intestinal Flora in Rats on Metabolism and Transformation of Effective Components in Polygonum orientale Inflorescences / 中国实验方剂学杂志

Objective:To observe the metabolic characteristics of effective components from Polygonum orientale inflorescences in intestinal flora of rats. Method:The incubating samples of effective components from P. orientale inflorescences in rat intestinal flora in vitro were detected by UPLC-ESI-Q-TOF-MS/MS, the mobile phase was consisted of 0.1%formic acid solution-0.1%formic acid acetonitrile solution and eluted in gradient mode at a flow rate of 0.3 mL·min-1.The mass spectral analysis was detected with electrospray ionization under positive ion mode and negative ion mode.The metabolites and possible biotransformation pathways of effective components form P. orientale inflorescences in rat intestinal flora in vitro was analyzed by Metabolite ToolsTM, mass defect filtration(MDF) and other metabolite analysis techniques and combined with the accurate relative molecular weight of the compounds, the fragment ion information and the literature data. Result:Eighteen metabolites were detected after incubation of effective components from P. orientale inflorescences in rat intestinal flora.The main biotransformation pathways were reduction, oxidation, hydrolysis in Ⅰ phase reaction and methylation in Ⅱ phase reaction. Conclusion:The effective components of P. orientale inflorescences can be transformed into a variety of metabolites under the action of intestinal flora in rats.It is suggested that whether the metabolites are bioactive components should be considered when P. orientale inflorescences is used as medicine.

Kinetic Analysis of Enzymatic Reaction of Active Components from Xinshao Formula in Liver Microsomes of Sham-Operated and Cerebral Ischemia-reperfusion Injury Rats / 中国实验方剂学杂志

Objective: To investigate and compare enzymatic kinetics of scutellarin,apigenin-7-O-glucronide and paeoniflorin from Xinshao fomula in liver microsomes of sham-operated rats and middle cerebral artery occlusion(MCAO) rats with focal cerebral ischemia-reperfusion injury. Method: Xinshao fomula were incubated respectively with liver microsomes of sham-operated rats and MCAO rats,UPLC-MS and substrate elimination method was employed,Michaelis constant(Km),maximum velocity of enzymatic reaction(Vmax) and intrinsic clearance(CLint) of these three components from Xinshao fomula in liver microsomes of sham-operated rats and MCAO rats were calculated,these parameters between different groups were evaluated by statistical analysis. Result: The Km values of scutellarin,apigenin-7-O-glucronide and paeoniflorin in liver microsomes of MCAO rats were (0.798±0.031),(0.213±0.017),(0.499±0.029) μmol·L-1,which were quite different to these in liver microsomes of sham-operated rats.Compared with the sham-operated group,Vmax and CLint values of scutellarin and paeoniflorin in liver microsomes of MCAO rats were significantly reduced(PPVmax of apigenin-7-O-glucronide in liver microsomes of MCAO rat was also significantly reduced(PConclusion: Metabolic rates of these three active components from Xinshao fomula in liver microsomes of MCAO rats with focal cerebral ischemia-reperfusion injury decrease with low elimination rate.

Absorption of Inula cappa extract based on everted intestinal sac method / 中国中药杂志

To investigate the absorptive characteristics of Inula cappa extract based on the rat everted intestinal sac method . Nine representative ingredients in I. cappa extract were selected as the study objects. An UPLC-MS/MS method was established to determine and detect their cumulative absorption amount for expounding the absorptive characteristics of ingredients in different intestinal sections. According to the results， the transport mechanism of 8 compounds showed passive diffusion by the reverted gut sac method. And scopolin was actively transported in the intestine. The best absorption site of chlorogenic acid was duodenum. The best absorption site of cryptochlorogenic acid， 1，3--dicaffeoylquinic acid， luteolin-7-glucoside and 3，4--dicaffeoylquinic acid were jejunum. The best absorption site of neochlorogenic acid， scopolin， 4，5--dicaffeoylquinic acid and 3，5--dicaffeoylquinic acid was ileum. The absorption of all the compounds was affected by pH and bile. All of the nine ingredients in I. cappa extract could be absorbed in intestines， but with differences in the absorption rate， the best absorptive site and mechanism， indicating that the intestinal absorption of I. cappa extract was selective.

Metabolism of Inula cappa extract by rat intestinal bacteria in vitro / 中国中药杂志

To investigate the metabolism of major components in Inula cappa by rat intestinal bacteria in vitro. I. cappa extract was incubated for 24 h with rat intestinal bacteria under anaerobic environment. After the samples were precipitated by n-butanol, UPLC-Q-TOF-MS/MS was applied for the qualitative analysis of the metabolites, combined with data software such as Metabolite Tools, Data Analysis and so on. The potential metabolites in rat intestinal bacteria were analyzed by comparing the total ion current of the test samples and blank samples and analyzing the quasi-molecular ion and fragment ion of all chromatograms. The results injected that fourteen metabolites were detected in rat intestinal flora. Various types of metabolic reactions happen to caffeoylquinic acid in intestinal flora, including isomerization, hydrolyzation, there were also methylation, hydrogenation and acetylation of caffeic acid. At the same time, a methylate of dicaffeoylquinic acid was also detected. Presumably, caffeoylquinic acids were gradually transformed into more hydrophobic metabolites with smaller molecular mass, which were better absorbed by the intestinal tract.

Isolation and characterization of two new 2-isobutylmalates from Bletilla striata / 中国天然药物

The present study isolated 17 compounds from the tubers of Bletilla striata (Orchidaceae), using various chromatographic techniques. Their structures were identified based on their physical-chemical properties and spectroscopic analyses. Among them, two new 2-isobutylmalates, named bletimalates A (1) and B (2), together with other fifteen known compounds (3-17), were isolated and identified. Additionally, compounds 3, 4, and 8 were isolated from this plant for the first time.

Serum pharmacochemistry of stem bark of Eucommia ulmoides / 中草药

Objective: To investigate the bioactive constituents in the stem bark of Eucommia ulmoides and study on their serum pharmacchemistry. Methods: The UHPLC-Q-TOF-MS was used to analyze the ingredients in the serum samples in rats, and the chromatogram was compared among the peaks of extracts in the stem bark of E. ulmoides, rat serum with drug and blank serum sample was used to determine them after the administration of extracts by comparing the finger-print. Results: Seven compounds absorbed into blood were be detected, and had the same retention time with those in the finger-print of extracts, after repeated experiments, which were original constituents of the extracts. The extracts were identified by comparing the retention times with standard substance, five compounds were obtained and identified as geniposidic acid, protocatechuic acid, geniposide, pinoresinol diglucoside, and pinoresinol monoglucoside. Through review of the literature, two compounds may be 1-hydroxypinoresinol glucoside, and eucommiol. Conclusion: The compounds are absorbed into the blood are the effective constituents, and the research provides a scientific fundament for material basis of effectiveness of E. ulmoides.

Structures and hepatocytotoxicity of co-occurring substances in oleanolic acid tablets

Tablets of oleanolic acid [OA] have been approved by SFDA in China as an adjuvant therapy for acute and chronic hepatitis. Co-occurring substances present in the tablets of OA and their hepatocytotoxicity have not yet been reported. In the current investigation, the crude OA drug was separated by repeated column chromatography. The structures of the isolated compounds were characterized by spectral analysis and the cytotoxicity of each compound was evaluated in vitro against the human normal liver cell L02 at concentrations from 0.125 to 1000 micromol/L using the MTT method. As a result, OA and its 11 co-occurring trace compounds including one new triterpenoid, 3-O- [4-oxopentanoyl]- olean-12- en-28-oic acid, were isolated and structurally characterized. Cytotoxicity tests indicated that these compounds were all non-toxic at concentrations up to 50micromol/L. Clear structure-activity relationship [SAR] was also observed. The results suggested that OA tablets of similar origin might not cause obvious cytotoxicity to the normal liver cell. The work may facilitate further SAR studies of OA-type triterpenoids.

Quality evaluation of bletillae rhizoma based on hemostatic biopotency / 中国中药杂志

This dissertation is to determine the biopotency of hemostat which processed in different places by establishing a bioassay method of Bletillae Rhizoma based on the thrombin time. Contrast test is the main methodology. Specifically, the reference substance of Bletillae Rhizoma is determined by comparing with the control substance of vitamin K1 using thrombin time, which is calibrated the Bletillae Rhizoma. The hemostatic biopotency is calculated by using the method of "parallel line assay method based on quantitative responses" (3.3) from different processed products. It indicates that there is a strong linear correlation between Bletillae Rhizoma and control drugs (Y = 66.332-23.913X, R2 = 0.995 3). The hemostatic biopotency of Bletillae Rhizoma from different processed products ranged between 821.93-1 187.53 U x g(-1) shown in the paper, and all of them can meet the requirements of the test. The methodology has an appropriate instrument precision (RSD 3.8%), intermediate precision (RSD 4.6%), repeatability (RSD 3.2%) and stability (RSD 3.7%). Therefore, it can be turned out that the methodology which established in the dissertation is good at determinating the hemostatic biopotency of Bletillae Rhizoma and it is reliable, simple and repeatable.

Simultaneous determination of nine chemical markers of bletillae rhizoma by ultra performance liquid chromatography / 中国中药杂志

A UPLC method has been developed in the current investigation for simultaneous determination of nine chemical markers of Bletilla striata, 4-hydroxymethylphenyl beta-D-glucoside, blestroside, dactylorhin A, militarine, dihydrophenanthrene 5, gymnoside V, dihydrophenanthrene 1, benzylphenanthrene 3 and gymnosides IX. Separation was performed at 45 degrees C on an ACQUITY UPLC BEH C18 column (2.1 mm x 150 mm, 1.7 microm) with a gradient solvent system of acetonitrile-water as the mobile phase. The flow rate was 0.3 mL x min(-1), the detection wavelength was 280 nm. The results showed that the nine chemical markers could be well resolved and that in the selected linear range, all calibration curves of the nine chemical markers showed good linearity (r > or = 0.999 3). The recoveries (n = 6) were in the range of 98.15% - 102.2% and RSDs were between 2.1% - 3.6%. The data suggested that the developed UPLC-UV method had good reproducibility, robustness, and accuracy, which was suitable for the quality control of Bletilla striata. Applications of the method showed that the nine chemical markers had higher contents in the wild B. striata than in the cultivated ones.

Study on fingerprint-pharmacology correlation of protective effect of Polygonum orientale on myocardial cell oxidative injury induced by H202 / 中国中药杂志

<p><b>OBJECTIVE</b>To study the protective effect of flowers, branch with leaves and main stem of Polygonum orientale on H9c2 myocardial cells oxidative injury induced by H202 and discuss its efficacious substance foundation of anti-myocardial ischemia.</p><p><b>METHOD</b>UPLC fingerprint was established for flowers, branch with leaves and main stem of P. orientale, and the chromatographic peak was identified. Meanwhile, the protective effect of composites of flowers, branch with leaves and main stem of P. orientale in different proportions on myocardial cells oxidative injury was observed (with MTT method as indicators of concentration screening and cell livability) and MDA were detected. The correlation analysis was made between activity information of composites and chemical information of UPLC fingerprint to infer efficacious substance foundation.</p><p><b>RESULT</b>Branch with leaves and flowers from Polygonum orientale showed a protective effect on H9c2 myocardial cells oxidative injury. According to the study on spectrum-activity relationship, peak 3-5, 11-14, 18-19, 21-25 showed a positive correlation with their antioxidant activities.</p><p><b>CONCLUSION</b>This study determined activity of Polygonum orientale on anti-oxidative injury, laying an experimental foundation for in-depth study and development of P. orientale herbs.</p>

Effect of scutellarin on expressions of nicotinic acetylcholine receptor protein and mRNA in the brains of dementia rats / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To observe the effect of Scutellarin (Scu) on expressions of nicotinic acetylcholine receptor (nAChR) subunit protein and mRNA in dementia rats, and to study its possible mechanism on dementia.</p><p><b>METHODS</b>Forty-two Wistar rats were randomly divided into 5 groups, i.e., the normal control group (n=6), the sham-operative group (n=6), the memory deficit model group, the Scu treatment group (n=10), and the positive drug (piracetam) control group (n=10). The dementia rat model was established by bilateral ventricle injection with beta-amyloid peptide (Abeta)(25-35) and abdominal cavity injection with D-galactose. Rats in the Scu treatment group or the piracetam control group were treated with Scu or piracetam by gastrogavage. The learning and memory ability of rats were detected by Morris water maze test, nAChR alpha4, alpha7, and beta2 subunits at protein and mRNA levels were detected by Western blot and Real-time PCR respectively.</p><p><b>RESULTS</b>Compared with the normal control group and the sham-operative group, the learning and memory ability decreased in rats of the model group (P<0.05). nAChR alpha4 and alpha7 subunit protein expressions were obviously lowered (P<0.05), but changes of beta2 were not obvious. No obvious change of mRNA expressions in all three nAChR subunits was seen (P>0.05). After treatment of Scu, the learning and memory ability was greatly improved, nAChRs alpha4 and alpha7 subunit protein expressions increased in rats with dementia (all P<0.05). No obvious change of mRNA expressions in all three nAChR subunits was seen (P>0.05). No obvious difference of each index was shown between the Scu treatment group and the positive drug (piracetam) control group.</p><p><b>CONCLUSIONS</b>Scutellarin could improve the learning and memory ability of dementia rats. Its mechanism might be associated with its up-regulation of nAChR expressions.</p>