[Mechanism of Cordyceps militaris against non-small cell lung cancer: based on serum metabolomics].

This study investigated the potential mechanism of Cordyceps militaris(CM) against non-small cell lung cancer(NSCLC) based on serum untargeted metabolomics. Specifically, Balb/c nude mice were used to generate the human lung cancer A549 xenograft mouse model. The tumor volume, tumor weight, and tumor inhibition rate in mice in the model, cisplatin, Cordyceps(low-, medium-, and high-dose), and CM(low-, medium-, and high-dose) groups were compared to evaluate the influence of CM on lung cancer. Gas chromatography-mass spectrometry(GC-MS) was used for the analysis of mouse serum, SIMCA 13.0 for the compa-rison of metabolic profiles, and MetaboAnalyst 5.0 for the analysis of metabolic pathways. According to the pharmacodynamic data, the tumor volume and tumor weight of mice in high-dose CM group and cisplatin group decreased as compared with those in the model group(P<0.05 or P<0.01). The results of serum metabolomics showed that the metabolic profiles of the model group were significantly different from those of the high-dose CM group, and the content of endogenous metabolites was adjusted to different degrees. A total of 42 differential metabolites and 7 differential metabolic pathways were identified. In conclusion, CM could significantly inhibit the tumor growth of lung cancer xenograft mice. The mechanism is the likelihood that it influences the aminoacyl-tRNA biosynthesis, the metabolism of D-glutamine and D-glutamate, metabolism of alanine, aspartate, and glutamate, metabolism of glyoxylate and dicarboxylic acid, biosynthesis of phenylalanine, tyrosine, and tryptophan, arginine biosynthesis as well as nitrogen metabolism. This study elucidated the underlying mechanism of CM against NSCLC from the point of metabolites. The results would lay a foundation for the anticancer research and clinical application of CM.

A target lipidomics approach to investigate the acute inflammatory irritation induced by indolealkylamines from Chansu water fraction in rats.

Chansu has demonstrated adverse reactions in clinical settings, which is associated with its toxicity and limits its clinical applications. But there are methodological limitations for drug safety evaluation. In the current study, ultra-high performance liquid chromatography, lipidomic profiling, and molecular docking were used to systemically assess Chansu-induced acute inflammatory irritation and further identify the underlying drug targets. Compared with the EtOAc extract, Chansu water fraction containing indolealkylamines caused acute inflammatory irritation in rats, including acute pain (spontaneous raising foot reaction), and inflammation (paw edema). At the molecular level, lipids analysis revealed significantly higher levels of pro-inflammatory mediators of the COX and LOX pathways. However, anti-inflammatory mediators from the CYP 450, ALA, and DHA pathways markedly decreased after exposure to Chansu water fraction. Moreover, four indolealkylamines from Chansu showed a high theoretical affinity to a known irritation target, 5-HT2AR. These results suggest that Chansu-induced inflammatory irritation is related to the distinct dysregulation of inflammatory lipids, and peripheral 5-HT2AR is a potential target for irritation therapy. The strategy used in this study can be a crucial approach in the safety evaluation of natural medicinal substances.

[Structural Chinese medicine: new research field on pharmacodynamic substance basis of traditional Chinese medicine].

The research on the pharmacodynamic substance basis of traditional Chinese medicine(TCM) is a key scientific issue for the inheritance and development of TCM. At present, a large number of remarkable achievements have been made in the field of chemical components in Chinese medicine, however, another important aspect, namely the physical structure and mode of action of the multi-component assembly of TCM, has not been clearly understood and deeply studied. From the bottleneck of restricting material ba-sic research, we objectively analyzed the common cause of the existing problems. Based on the new discoveries and advances of active substances from TCM emerging in recent years, we extracted and summarized the concept of structural Chinese medicine, elaborated the basic ideas, main features and research modes, hoping to provide theoretical and practical references for the study on the pharmacodynamic substance basis and other research fields of TCM.

[Quality evaluation of fried Glycyrrhizae Radix et Rhizoma pieces by HPLC fingerprint and multicomponent quantitative analysis].

To establish the HPLC fingerprint and multi-component determination method of fried Glycyrrhizae Radix et Rhizoma pieces. HPLC analysis was performed on Thermo Acclaim ~(TM)120 C_(18) column(4.6 mm×250 mm, 5 µm). Acetonitrile-0.1% phosphoric acid aqueous solution was taken as the mobile phase for gradient elution. The flow rate was 1 mL·min~(-1),the column temperature was maintained at 30 ℃, and the detection wavelength was 237 nm and 360 nm. The similarity of 15 batches of fried Glycyrrhizae Radix et Rhizoma pieces was higher than 0.849, and 17 common peaks were identified. Liquiritin, isoliquiritin apioside, isoliquiritin, liquiritigenin, isoliquiritigenin and glycyrrhizic acid were identified; among them, the mass fractions of Liquiritin, isoliquiritin apioside, isoliquiritin, liquiritigenin, glycyrrhizic acid were were 0.519%-3.058%, 0.227%-0.389%, 0.070%-0.439%, 0.038%-0.173%, 1.381%-4.252%, respectively. According to the cluster analysis, the 15 batches of decoction pieces were classified into three categories; principal component analysis screened out four principal components, with the cumulative variance contribution rate of 86.630%, indicating that the principal components contained most information of original data. Partial least squares discriminant ana-lysis marked 6 differential components in the decoction pieces. The established fingerprint and multicomponent determination are stable and reliable, and can provide a reference for the quality control of Radix Glycyrrhizae Radix et Rhizomae and fried Glycyrrhizae Radix et Rhizoma pieces.

Challenges and strategies in progress of drug delivery system for traditional Chinese medicine Salviae Miltiorrhizae Radix et Rhizoma (Danshen).

Traditional Chinese medicines (TCMs), with a history of thousands of years, are widely used clinically with effective treatment. However, the drug delivery systems (DDSs) for TCMs remains major challenges due to the characteristics of multi-components including alkaloids, flavones, anthraquinones, glycosides, proteins, volatile oils and other types. Therefore, the novel preparations and technology of modern pharmaceutics is introduced to improve TCM therapeutic effects due to instability and low bioavailability of active ingredients. Salviae Miltiorrhizae Radix et Rhizoma, the radix and rhizomes of Salvia miltiorrhiza Bunge (Danshen in Chinese), is a well known Chinese herbal medicine for protecting the cardiovascular system, with active ingredients mainly including lipophilic tanshinones and hydrophilic salvianolic acids. In this review, this drug is taken as an example to present challenges and strategies in progress of DDSs for TCMs. This review would also summary the characteristics of active ingredients in it including physicochemical properties and pharmacological effects. The purpose of this review is to provide inspirations and ideas for the DDSs designed from TCMs by summarizing the advances on DDSs for both single- and multi-component from Danshen.

Elaboration in type, primary structure, and bioactivity of polysaccharides derived from mollusks.

Over the past decades, numerous Mollusca species have received more attention in development and utilization as valuable bio-resources. Many efforts have been focused on investigating mollusk polysaccharides because of their rich content, ease of extraction, diversified sorts, specific structure, various biofunctions and potent activities. To date, many mollusks, especially species of gastropods, bivalves, or cephalopods, have been reported containing polysaccharide compounds in tissues with abundant amount, and most of polysaccharides are obtainable through combining techniques of extraction, separation and purification. The polysaccharides isolated from mollusks appeared with various structural and physicochemical characteristics, ranged from neutral polysaccharides and sulfated polysaccharides, to GAGs series (including Hep/HS, CS/DS, HA and similarities), even to heterogeneous glycan with high molecular weight. This review article provides comprehensive knowledge of recent researches on type classification, tissue origins and possible biofunctions of various polysaccharides from mollusks. The highlights were placed in structure variation including molecular weight, sulfation pattern, linkages and monomer compositions for repeating unit, and primary molecular construction of the mollusks polysaccharides. In addition, this article covers general information on exhibition of mollusks polysaccharide extracts or preparations in the various bioactivities, such as anticoagulant, antiatherogenic, antioxidant, immunomodulatory, antivirus and antitumor activities, which would reveal their possible potentials in medical application. Furthermore, the article presents a brief overview on several challenges and future scope in this field.

Pudilan xiaoyan oral liquid alleviates LPS-induced respiratory injury through decreasing nitroxidative stress and blocking TLR4 activation along with NF-ΚB phosphorylation in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Pudilan xiaoyan oral liquid (PDL), collected in Chinese Pharmacopoeia, has been used clinically for treating inflammatory diseases such as upper respiratory tract infection diseases. However, its potential anti-inflammation and the mechanism are still unclear. MATERIALS AND METHODS: lipopolysaccharide (LPS) was used to induce respiratory inflammation of mice by intratracheal administration. UPLC/MS was performed for components analysis of PDL. Enzyme-linked immune sorbent assay (ELISA) was conducted for determining interleukin-6(IL-6), interleukin-1ß(IL-1ß) and tumor necrosis factor-α(TNF-α) in serum and supernatant of tracheal tissue while Nitric oxide assay kit for nitric oxide (NO) content. Hematoxylin-Eosin (HE) staining was applied to evaluate pathological lesions. Western blotting analysis (WB) and Immunohistochemistry(IHC) were employed for the determination of Toll-like receptors 4(TLR4), TNF-α, IL-6, inducible nitric oxide synthase(iNOS) and nuclear factor-kappa B p65 (NF-κB p65) protein expressions. RESULTS: Seven major compounds of PDL were analyzed simultaneously. The treatment of PDL could attenuate LPS-induced histopathological damage of tracheal tissues, followed by reducing pro-inflammation mediators including TNF-α and IL-6 in serum and supernatant of tracheal tissue. LPS-induced nitroxidative stress including NO content and iNOS expression was inhibited significantly by PDL. Furthermore, PDL also down-regulated NF-kB p65 phosphorylation and TLR4 expressions. CONCLUSION: The results indicated that the PDL had a protective effect on LPS-induced respiratory inflammation injury in mice. Our findings for the first time provide experimental evidence for the application of PDL on respiratory inflammation injury in clinical practice.

A selective HPLC-MS/MS method for quantification of SND-117 in rat plasma and its application to a pharmacokinetic study.

Rheumatoid arthritis (RA), a chronic systemic inflammatory disorder affects many adults. Sinomenine, a natural product, has been clinically available for the treatment of RA in China. SND-117, a sinomenine derivative with much more potent activity, might serve as a candidate for anti-arthritis. The aim of the present study was to develop a sensitive and rapid high performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) method for quantification of SND-117 in rat plasma and to understand its absolute bioavailability. The HPLC-MS/MS method was developed and fully validated for determination of SND-117 in rat plasma, and the pharmacokinetic differences were investigated after different administration routes. The pharmacokinetics parameters were calculated by non-compartment model with DAS 3.0 software. After the oral or intravenous administration of different doses of SND-117, the time to peak is 1.5h, half-life time is 8-10h. The absolute oral bioavailability of SND-117 in rats was 9.60%. The results showed that SND-117 in rats was quickly absorbed, slowly eliminate, and the kinetics were linear. This method was suitable for pharmacokinetic studies of SNA-117 in rats.

Components of Goutengsan in Rat Plasma by Microdialysis Sampling and Its Protection on Aß1-42-Induced PC12 Cells Injury.

Goutengsan, a Chinese herbal formula, potential protection on Alzheimer's disease (AD) has been less reported. In current study, we investigated the protection of Goutengsan on Aß1-42-induced pheochromocytoma-derived cells (PC12). Furthermore, the components from Goutengsan in rat plasma were identified by microdialysis (MD) for in vivo sampling. Meanwhile, the protection of components identified was also verified. At last, we found that Goutengsan has a potential protective effect on Aß1-42-induced PC12 cells via reducing cells damage and increasing cells vitality as well as six components (pachymic acid, liquiritin, rhynchophylline, isorhynchophylline, corynoxeine, and isocorynoxeine) which may be effective components. This study helps to understand the treatment of Goutengsan for AD and would facilitate the clinical and further studies for this formula.

Application of Traditional Chinese Medical Herbs in Prevention and Treatment of Respiratory Syncytial Virus.

Respiratory syncytial virus (RSV) is a common viral pathogen of the lower respiratory tract, which, in the absence of effective management, causes millions of cases of severe illness per year. Many of these infections develop into fatal pneumonia. In a review of English and Chinese medical literature, recent traditional Chinese medical herb- (TCMH-) based progress in the area of prevention and treatment was identified, and the potential anti-RSV compounds, herbs, and formulas were explored. Traditional Chinese medical herbs have a positive effect on inhibiting viral attachment, inhibiting viral internalization, syncytial formation, alleviation of airway inflammation, and stimulation of interferon secretion and immune system; however, the anti-RSV mechanisms of TCMHs are complicated, which should be further investigated.

Preparation and Physicochemical and Pharmacokinetic Characterization of Ginkgo Lactone Nanosuspensions for Antiplatelet Aggregation.

The aim of this study was to investigate the potential of nanosuspensions (NSs) in improving the dissolution and absorption of poorly water-soluble ginkgo lactones (GLs), including ginkgolide A, ginkgolide B, and ginkgolide C. Liquid GL-NSs were prepared by a combined bottom-up and top-down approach with response surface methodology design, followed by freeze-drying solidification. Physicochemical characterization of the prepared freeze-dried GL-NSs was performed by photon correlation spectroscopy, scanning electron microscopy, powder X-ray diffraction, and differential scanning calorimetry. In vitro dissolution and in vivo bioavailability of ginkgolide A, ginkgolide B, and ginkgolide C in freeze-dried GL-NSs were evaluated with GLs coarse powder as control. Their inhibitory effects on platelet aggregation were also comparatively analyzed. GLs existed in an amorphous state in the prepared freeze-dried GL-NSs. The particle size, polydispersity index, zeta potential, and redispersibility index of freeze-dried GL-NSs were around 286 nm, 0.26, -25.19 mV, and 112%, respectively. The particle size reduction resulted in much more rapid and complete dissolution of ginkgolides from GL-NSs than coarse powder. Comparison with GLs coarse powder, freeze-dried GL-NSs showed a significant decreased Tmax, 2-fold higher peak concentration, and 2-fold higher area under plasma concentrations curve for 3 ginkgolides and exhibited significantly higher antiplatelet aggregation effect.

[Tissue Distribution of Alkaloids from Coptidis Rhizoma in Rats Determined by UPLC-MS].

Objective: To develop an UPLC-MS method for simultaneous determination of alkaloids from Coptidis Rhizoma in rat tissues, and to study the tissue distribution of alkaloids from Coptidis Rhizoma in rats. Methods: The samples were extracted with ethyl acetate, and analyzed by UPLC-MS with acetonitrile-0. 2% formic acid solution in a gradient elution mobile phase, the flow rate was 0. 2m L/min. Tetrahydropalmatine was used as an internal standard. The mass spectrometer was operated in selected reaction monitoring( SIM) mode with positive electrospray ionization, the transition were m/z 191. 904 /118. 973( noroxyhydrastinine), m/z 335. 877 /308. 072( 8-ocoptisine),m/z 351. 94 /294. 554( palmatine chloride),m/z 335. 94 /262. 112( epiberberine), m/z 337. 94 /322. 422( columbamine), m/z 319. 904 /292. 037( coptisine), m/z 355. 977 /192. 036( tetrahydropalmatine),m/z 335. 94 /320. 036( berberine hydrochloride),m/z 351. 94 /321. 995( oxyberberin), m/z 337. 94 /322. 949( jatrorrhizine respectively). Results: Excellent linearity was observed in all alkaloids in their linear range( r & 0. 9901). The RSD of precision of the developed method was less than 15%,and the accuracy and stability were less than ± 15%,the extraction recovery was 72. 1% ~ 82. 9% with RSD less than 15%. Coptisine,epiberberine,berberine,jatrorrhizine,columbamine,palmatine were widely distributed in rat tissues. Noroxyhydrastinine,8-ocoptisine,oxyberberin could only be determined in liver and heart or kidney. Conclusion: The established method is simple and accurate. Satisfactory results are obtained with applying this method to the tissue distribution study of alkaloids from Coptidis Rhizoma.

A metabolomics approach to studying the effects of Jinxin oral liquid on RSV-infected mice using UPLC/LTQ-Orbitrap mass spectrometry.

ETHNOPHARMACOLOGICAL RELEVANCE: Jinxin oral liquid (JOL) is a traditional Chinese medicine (TCM) formula modified from ma-xing-shi-gan-tang, an ancient formula widely used in the treatment of respiratory diseases such as bronchitis, pneumonia, and asthma. In our previous studies, JOL was shown to safely and effectively treat viral pneumonia, especially that involving respiratory syncytial virus (RSV). AIM OF THE STUDY: To investigate the mechanism of the effect of JOL in RSV infected mice, using a metabolomics approach based on ultra-performance liquid chromatography coupled with linear ion trap quadrupole-Orbitrap mass spectrometry (UPLC/LTQ-Orbitrap-MS). MATERIALS AND METHODS: BALB/c mice were divided into four groups, the control group (saline inoculation/no treatment), RSV group (RSV inoculation/saline treatment), RSV+JOL group (RSV inoculation/JOL treatment), and RSV+Riba group (RSV inoculation/ribavirin treatment). Plasma and lung tissue samples were collected 7 days after the inoculation/treatment protocols, and UPLC/LTQ-Orbitrap-MS method based on metabolomics was developed. Principal component analysis (PCA) and orthogonal partial least squares-discriminant analysis (OPLS-DA) were utilized to identify biomarkers potentially associated with the anti-RSV activity of JOL. RESULTS: JOL was associated with reduced inflammatory responses in RSV-infected lung tissue. The combination of PCA and OPLS-DA revealed deviations in 11 biomarkers in plasma, and 16 biomarkers in lung tissue induced by RSV that were corrected with JOL treatment. These biomarkers were primarily components of metabolic pathways involving glycerophosphocholines, sphingolipids, and glycerolipids. JOL was able to restore the abnormal levels of these biomarkers detected in the plasma and lung tissue of RSV-infected mice to approximately normal levels. CONCLUSIONS: This study suggested that JOL can treat RSV pneumonia effectively, partially by ameliorating the associated disturbances to lipid metabolism. The results provided insight into the anti-RSV mechanism of JOL, and also demonstrated that metabolomics is a valuable tool for investigating the efficacy of TCM treatment for RSV pneumonia, and the associated biomarkers involved.

Effects of Gancao on pharmacokinetic profiles of platycodin D and deapio-platycodin D in Jiegeng.

ETHNOPHARMACOLOGICAL RELEVANCE: Jiegeng (Radix Platycodi), the dried root of Platycodon grandiflorum A. DC (Campanulaceae), has been used to treat cough, sore throat, bronchitis, and bronchial asthma for thousands of years. It is commonly prescribed with Gancao (Radix et Rhizoma Glycyrrhizae) as a herbal combination in traditional Chinese medicine (TCM) to produce synergistic effects. AIM OF THE STUDY: To elucidate the herbaceous compatibility of Jiegeng and Gancao, we investigated the comparative pharmacokinetics, intestinal absorption, and microbial metabolism of platycodin D (PD) and deapio-platycodin D (DPD), the platycodins contained in Jiegeng. MATERIALS AND METHODS: In the comparative pharmacokinetic study, the concentrations of PD and DPD in Jiegeng extract (JE) and the Jiegeng-Gancao herb pair (JGHP) were determined in rat plasma using liquid chromatography-tandem mass spectrometry (LC-MS/MS). In addition, the main pharmacokinetic parameters were calculated using data analysis software (DAS). Furthermore, in vitro studies using Caco-2 cells and fecal lysates were performed to contradistinguish the intestinal absorption and microbial metabolism of PD and DPD in JE from those in JGHP. RESULTS: The peak concentration (Cmax) and area under the plasma concentration curve (AUC) of PD in rats orally administrated JGHP significantly increased compared to that in rats treated with JE. In addition, the time to reach peak concentration (Tmax) and half-life (t1/2) of PD and DPD in combination with JGHP were all prolonged compared with those of JE. There was no significant difference in the absorption of PD between JE and JGHP in Caco-2 cells. However, the hydrolysis of both PD and DPD in JGHP were weaker than that in JE after a 2-h incubation in fecal lysate which might be responsible for the different pharmacokinetic profiles of the platycodins in JE and JGHP. CONCLUSION: In this study, we discovered that Gancao might influence the pharmacokinetic profiles of PD and DPD in Jiegeng. Furthermore, the difference in profiles may be attributable to the inequable microbial metabolism rather than intestinal absorption of the platycodins in JE and JGHP. The results of this study elucidated the pharmacokinetic compatibility and rationale for the use of JGHP.

[HPLC specific chromatogram spectrum-effect relationship for Shuanghuanglian on MDCK cell injury induced by influenza A virus (H1N1)].

To establish HPLC specific chromatogram and its correlation with the protection effect of Shuanghuanglian on MDCK (Madin-Darby canine kidney) cell injury induced by influenza A virus( H1N1). Nine recipes of Shuanghuanglian based on the official prescription were prepared according to orthogonal test for HPLC analysis and MDCK cells protection experiment separately (cytopathic effect (CPE) method was used for observing the virus infectivity and MTT staining results were used as the determining indexes for drug concentration selection and analyzing cell viability). The results suggested that all the other Shuang-Huang-Lian recipes except recipe1 demonstrate protecting effect on MDCK cell injury induced by influenza A virus (P < 0.01, P < 0.001). Stepwise regression analysis was used for analyzing the relationships between HPLC fingerprint and the protecting effect of Shuanghuanglian on influenza A virus induced MDCK cell injury. Peak 2, 3, 6, 8 and 12 were found to be strongly related with anti-influenza A virus efficacy. Stepwise regression analysis of recipes data and efficacy data showed that Lonicerae Japonicae Flos and Forsythiae Fructus were positively associated with the protecting effect of cells injury. From HPLC fingerprints, we found that peak 2, 3, 12 were from Lonicerae Japonicae Flos and peak 6, 8 were from Forsythiae Fructus. Four peaks were identified through comparing the retention time between the standard and Shuanghuanglian recipes, and they were chlorogenicacid, cryptochlorogenic acid, forsythoside B and 3,4-dicaffeoylquinic acid respectively. Caffeic acid derivatives in Lonicerae Japonicae Flos and Forsythiae Fructus were found to be greatly correlated with anti-influenza A virus efficacy and maybe the substance basis of Shuanghuanglian.

[Comparison of total phenol of magnolia solid dispersion prepared by different methods].

To compare the difference of total phenol of magnolia solid dispersion prepared by different methods. Hot melt extrusion, solvent evaporation method, and fusion-cooling method were used to prepare total phenol of Magnolia accessory solid dispersion, Plastone S-630 and HPC. The drug dispersion state in the prepared solid dispersion was evaluated with DSC and X-ray diffraction; FT-IR method was used to analyze the possible connections between drug and accessories. Finally, accelerated stability-in vivo dissolution test was use to compare the stability differences between these three processes. The results of DSC and X-ray diffraction showed that all of the drug in solid dispersion processed by three processes can exist in amorphous form; FT-IR results also could not distinguish the difference between the three processes; accelerated stability-in vivo dissolution test showed the stability of solid dispersion prepared by HPC was better than Plastone S-630, and the same kinds of materials solid dispersion prepared by hot melt extrusion showed a better stability than the other two processes.

[Active ingredients and its pharmacokinetic behavior and anti-inflammatory effects of ginseng with different steamed times].

HPLC analysis was performed to study the changes in chemical composition of ginseng extracts prepared from high quality ginseng with 0, 2, 4, 8 h of steamed times. An UFLC-MS/MS multiple-reaction monitoring (MRM) quantitative analysis was made to investigate the pharmacokinetic behavior differences of ginsenosides in mice ig administered of ginseng extracts with different steamed times in the negative ion mode, with Digoxin as the internal standard substance. The mice were injected with LPS to establish inflammation model after ig administration of ginseng for a week and the contents of tumor necrosis factor-α (TNF-α) and interleukin-1ß (IL-1ß) in mice plasma were detected by ELISA, in order to study on anti-inflammatory effects of ginseng with different steamed times. It was determined that levels of TNF-α and IL-1ß were significantly decreased in inflammation model group ig administered of ginseng extracts with 8h of steamed time. The results showed that the chemical components in ginseng changed after steaming and the components into the blood changed, correspondingly. Ginseng with steamed 8 h contributes to anti-inflammatory effects. These results provided an experimental basis for revealing the active substance basis and dose-effect relationship of ginseng on anti-inflammatory effect.

Simultaneous determination of ten alkaloids of crude and wine-processed Rhizoma Coptidis aqueous extracts in rat plasma by UHPLC-ESI-MS/MS and its application to a comparative pharmacokinetic study.

Rhizoma coptidis (R.C.), a widely used traditional Chinese medicine, has been used for centuries in the treatment of hypertension, inflammation, dysentery and liver diseases, etc. Wine-processing is a specialized technology by sautéing crude herbal medicine using Chinese rice wine. This paper was designed to establish a simultaneous quantitative method of ten alkaloids (berberine, coptisine, palmatine, jatrorrhizine, epiberberine, magnoflorine, columbamine, noroxyhydrastinine, oxyberberine and 8-oxocoptisine) in rat plasma. Furthermore, the pharmacokinetics of those alkaloids after administration of crude and wine-processed R.C. aqueous extracts was compared. As a result, a ultra high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UHPLC-ESI-MS/MS) method was developed and validated for the first time. Chromatographic separation was achieved on a C18 column using gradient elution with the mobile phase consisting of acetonitrile and water (containing 0.2% formic acid) at a flow rate of 0.2 ml/min. The validated method showed good linearity over a wide concentration range (r>0.99), and lower limits of quantification less than 5.46 ng/ml for the each analyte. The intra- and inter-day assay variability was below 9.9% and 10.5% for all analytes, respectively. The extraction recovery of those alkaloids and I.S. ranged from 65.3% to 90.7%. The validated method has been successfully applied to pharmacokinetic comparison after administration of crude and wine-processed R.C. aqueous extracts. Pharmacokinetic comparative study showed that Cmax of coptisine and 8-oxocoptisine and AUC0-t of coptisine, palmatine and 8-oxocoptisine were increased significantly (p<0.05) after wine-processing, while other compounds didn't show significant difference, which suggested that wine-processing exerted limited effects on the absorption of alkaloids. These results might be helpful for R.C.' clinical reasonable application and further studies on its wine-processing mechanism.

Protection of Tong-Sai-Mai Decoction against Apoptosis Induced by H2O2 in PC12 Cells: Mechanisms via Bcl-2-Mitochondria-ROS-INOS Pathway.

Tong-Sai-Mai decoction (TSM) is a Chinese materia medica polyherbal formulation that has been applied in treating brain ischemia for hundreds of years. Because it could repress the oxidative stress in in vivo studies, now we focus on the in vitro studies to investigate the mechanism by targeting the oxidative stress dependent signaling. The relation between the neurogenesis and the reactive oxygen species (ROS) production remains largely unexamined. PC12 cells are excitable cell types widely used as in vitro model for neuronal cells. Most marker genes that are related to neurotoxicity, apoptosis, and cell cycles are expressed at high levels in these cells. The aim of the present study is to explore the cytoprotection of TSM against hydrogen peroxide- (H2O2-) induced apoptosis and the molecular mechanisms underlying PC12 cells. Our findings revealed that TSM cotreatment with H2O2 restores the expression of bcl-2, inducible nitric oxide synthase (INOS), and mitochondria membrane potential. Meanwhile, it reduces intracellular [Ca(2+)] concentration, lactate dehydrogenase (LDH) release, and the expression of caspase-3 and bax. The results of the present study suggested that the cytoprotective effects of the TSM might be mediated, at least in part, by the bcl-2-mitochondria-ROS-INOS pathway. Due to its nontoxic characteristics, TSM could be further developed to treat the neurodegenerative diseases which are closely associated with the oxidative stress.

[Absorption and transportation of calycosin in Astragali Radix by using Caco-2 monolayer model].

Flavonoids are a class of important active ingredients in traditional Chinese medicine, pharmacological activity and in vivo process is the focus of research in recent years. Calycosin is the main active ingredients of flavonoids in Astragali Radix, recent studies indicate that it has many kinds of pharmacological activity, but the absorption and transport characteristics in vivo is unclear. The experiment using Caco-2 cell model, with apigenin as internal standard substance, using the method for the determination of drug concentration by HPLC, were studied at different concentrations and absorption transport characteristics of respectively adding different types of protein inhibitors. Data were analyzed by Q test, the results show that low, middle, high concentration of P(app)(BL-AP)/ P(app)(AP-BL) = 1.38 < 1.5, respectively adding different types of protein inhibitors, compared with the control group of P(app)(BL-AP)/ P(app)(AP-BL), there were no significant differences. Calycosin absorption may mainly passive transport, also involved in active transport mechanism, the transport may not be affected by the P-protein, MRP2 protein, SGLT protein.