Therapeutic effect and mechanism of Anemarrhenae Rhizoma on Alzheimer's disease based on multi-platform metabolomics analyses.

Anemarrhenae Rhizoma (AR) has multiple pharmacological activities to prevent and treat Alzheimer's disease (AD). However, the effect and its molecular mechanism are not elucidated clear. This study aims to evaluate AR's therapeutic effect and mechanism on AD model rats induced by D-galactose and AlCl3 with serum metabolomics. Behavior study, histopathological observations, and biochemical analyses were applied in the AD model assessment. Gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-mass spectrometry (LC-QTOF/MS) were combined with multivariate statistical analysis to identify potential biomarkers of AD and evaluate the therapeutic effect of AR on AD from the perspective of metabolomics. A total of 49 biomarkers associated with the AD model were identified by metabolomics, and pathway analysis was performed to obtain the metabolic pathways closely related to the model. With the pre-treatment of AR, 32 metabolites in the serum of AD model rats were significantly affected by AR compared with the AD model group. The regulated metabolites affected by AR were involved in the pathway of arginine biosynthesis, arginine and proline metabolism, ether lipid metabolism, glutathione metabolism, primary bile acid biosynthesis, and steroid biosynthesis. These multi-platform metabolomics analyses were in accord with the results of behavior study, histopathological observations, and biochemical analyses. This study explored the therapeutic mechanism of AR based on multi-platform metabolomics analyses and provided a scientific basis for the application of AR in the prevention and treatment of AD.

The unique carrier mobility of monolayer Janus MoSSe nanoribbons: a first-principles study.

Charge-carrier mobility is a determining factor of the transport properties of semiconductor materials and is strongly related to the optoelectronic performance of nanoscale devices. Here, we investigate the electronic properties and charge carrier mobility of monolayer Janus MoSSe nanoribbons by means of first-principles simulations coupled with deformation potential theory. These simulations indicate that zigzag nanoribbons are metallic. Conversely, armchair nanoribbons are semiconducting and show oscillations in the calculated band gap as a function of edge-width according to the 3p < 3p + 1 < 3p + 2 rule, with p being the integer number of repeat units along the non-periodic direction of the nanoribbon. Although the charge-carrier mobility of armchair nanoribbons oscillates with the edge-width, its magnitude is comparable to its two-dimensional sheet counterpart. A robust room-temperature carrier mobility is calculated for 3.5 nm armchair nanoribbons with values ranging from 50 cm2 V-1 s-1 to 250 cm2 V-1 s-1 for electrons (e) and holes (h), respectively. A comparison of these values with the results for periodic flat sheet (e: 73.8 cm2 V-1 s-1; h: 157.2 cm2 V-1 s-1) reveals enhanced (suppressed) hole (electron) mobility in the Janus MoSSe nanoribbons. This is in contrast to what was previously found for MoS2 nanoribbons, namely larger mobility for electrons in comparison with holes. These differences are rationalized on the basis of the different structures, edge electronic states and deformation potentials present in the MoSSe nanoribbons. The present results provide the guidelines for the structural and electronic engineering of MoSSe nanoribbon edges towards tailored electron transport properties.

Cardioprotective mechanisms of salvianic acid A sodium in rats with myocardial infarction based on proteome and transcriptome analysis.

Ischemic heart diseases (IHDs) cause great morbidity and mortality worldwide, necessitating effective treatment. Salvianic acid A sodium (SAAS) is an active compound derived from the well-known herbal medicine Danshen, which has been widely used for clinical treatment of cardiovascular diseases in China. This study aimed to confirm the cardioprotective effects of SAAS in rats with myocardial infarction and to investigate the underlying molecular mechanisms based on proteome and transcriptome profiling of myocardial tissue. The results showed that SAAS effectively protected against myocardial injury and improved cardiac function. The differentially expressed proteins and genes included important structural molecules, receptors, transcription factors, and cofactors. Functional enrichment analysis indicated that SAAS participated in the regulation of actin cytoskeleton, phagosome, focal adhesion, tight junction, apoptosis, MAPK signaling, and Wnt signaling pathways, which are closely related to cardiovascular diseases. SAAS may exert its cardioprotective effect by targeting multiple pathways at both the proteome and transcriptome levels. This study has provided not only new insights into the pathogenesis of myocardial infarction but also a road map of the cardioprotective molecular mechanisms of SAAS, which may provide pharmacological evidence to aid in its clinical application.

UPLC-QTOF MS-Based Serum Metabolomic Profiling Analysis Reveals the Molecular Perturbations Underlying Uremic Pruritus.

As one of the most troublesome complications in patients with chronic renal disease, the etiology of uremic pruritus remains unknown, and the current therapeutic approaches are limited and unsatisfactory. To identify potential biomarkers for improving diagnosis and treatment and obtain a better understanding of the pathogenesis of uremic pruritus, we compared serum metabolome profiles of severe uremic pruritus (HUP) patients with mild uremic pruritus (LUP) patients using ultraperformance liquid chromatography-quadruple time-of-flight mass spectrometry (UPLC-QTOF MS). Partial least squares discriminant analysis (PLS-DA) showed that the metabolic profiles of HUP patients are distinguishable from those of LUP patients. Combining multivariate with univariate analysis, 22 significantly different metabolites between HUP and LUP patients were identified. Nine of the 22 metabolites in combination were characterized by a maximum area-under-receiver operating characteristic curve (AUC = 0.899) with a sensitivity of 85.1% and a specificity of 83.0% distinguishing HUP and LUP. Our results indicate that serum metabolome profiling might serve as a promising approach for the diagnosis of uremic pruritus and that the identified biomarkers may improve the understanding of pathophysiology of this disorder. Because the 9 metabolites were phospholipids, uremic toxins, and steroids, further studies may reveal their possible role in the pathogenesis of uremic pruritus.

Comprehensive two-dimensional APTES-decorated MCF7-cell membrane chromatographic system for characterizing potential anti-breast-cancer components from Yuanhu-Baizhi herbal medicine pair.

Rhizoma corydalis and Radix Angelicae Dahurica (Yuanhu-Baizhi) herbal medicine pair has been used for thousands of years and has been reported to be potentially active in recent cancer therapy. But the exact active components or fractions remain unclear. In this study, a new comprehensive two-dimensional (2D) 3-aminopropyltriethoxysilane (APTES)-decorated MCF7-cell membrane chromatography (CMC)/capcell-C18 column/time-of-flight mass spectrometry system was established for screening potential active components and clarifying the active fraction of Yuanhu-Baizhi pair. APTES was modified on the surface of silica, which can provide an amino group to covalently link cell membrane fragments with the help of glutaraldehyde in order to improve the stability and column life span of the MCF7 CMC column. The comprehensive 2D MCF7-CMC system showed good separation and identification abilities. Our screen results showed that the retention components are mainly from the alkaloids in Yuanhu (12 compounds) and the coumarins (10 compounds) in Baizhi, revealing the active fractions of Yuanhu-Baizhi herbal medicine pair. Oxoglaucine, protopine, berberine, osthole, isopimpinellin and palmitic acid were selected as typical components to test the effects on cell proliferation and their IC50 were calculated as 38.17 µM, 29.45 µM, 45.42 µM, 132.7 µM, 156.8 µM and 90.5 µM respectively. Cell apoptosis assay showed that the drug efficacy was obtained mainly through inducing cell apoptosis. Furthermore, a synergistic assay results demonstrated that oxoglaucine (representative of alkaloids from Yuanhu) and isopimpinellin (representative of coumarins from Baizhi) showed significant synergistic efficacy with GFT, indicating that these components may act on other membrane receptors. The proposed 2D CMC system could also be equipped with other cells for further applications. Besides, the follow-up in-vitro experimental strategy using cell proliferation assay, cell apoptosis assay and synergistic assay proved to be a practical way to confirm the active fractions of herbal medicine.

Target Identification of Kinase Inhibitor Alisertib (MLN8237) by Using DNA-Programmed Affinity Labeling.

Accurate identification of the molecular targets of bioactive small molecules is a highly important yet challenging task in biomedical research. Previously, a method named DPAL (DNA-programmed affinity labeling) for labeling and identifying the cellular targets of small molecules and nucleic acids was developed. Herein, DPAL is applied for the target identification of Alisertib (MLN8237), which is a highly specific aurora kinase A (AKA) inhibitor and a drug candidate being tested in clinical trials for cancer treatment. Apart from the well-established target of AKA, several potential new targets of MLN8237 were identified. Among them, p38 mitogen-activated protein kinase (p38) and laminin receptor (LAMR) were validated to be implicated in the anticancer activities of MLN8237. Interestingly, these new targets were not identified with non-DNA-based affinity probes. This work may facilitate an understanding of the molecular basis of the efficacy and side effects of MLN8237 as a clinical drug candidate. On the other hand, this work has also demonstrated that the method of DPAL could be a useful tool for target identification of bioactive small molecules.

Identification of a ligand for tumor necrosis factor receptor from Chinese herbs by combination of surface plasmon resonance biosensor and UPLC-MS.

Identification of bioactive compounds directly from complex herbal extracts is a key issue in the study of Chinese herbs. The present study describes the establishment and application of a sensitive, efficient, and convenient method based on surface plasmon resonance (SPR) biosensors for screening active ingredients targeting tumor necrosis factor receptor type 1 (TNF-R1) from Chinese herbs. Concentration-adjusted herbal extracts were subjected to SPR binding assay, and a remarkable response signal was observed in Rheum officinale extract. Then, the TNF-R1-bound ingredients were recovered, enriched, and analyzed by UPLC-QTOF/MS. As a result, physcion-8-O-ß-D-monoglucoside (PMG) was identified as a bioactive compound, and the affinity constant of PMG to TNF-R1 was determined by SPR affinity analysis (K D = 376 nM). Pharmacological assays revealed that PMG inhibited TNF-α-induced cytotoxicity and apoptosis in L929 cells via TNF-R1. Although PMG was a trace component in the chemical constituents of the R. officinale extract, it had considerable anti-inflammatory activities. It was found for the first time that PMG was a ligand for TNF receptor from herbal medicines. The proposed SPR-based screening method may prove to be an effective solution to analyzing bioactive components of Chinese herbs and other complex drug systems. Graphical abstract Scheme of the method based on SPR biosensor for screening and recovering active ingredients from complex herbal extracts and UPLC-MS for identifying them. Scheme of the method based on SPR biosensor for screening and recovering active ingredients from complex herbal extracts and UPLC-MS for identifying them.

LC-MS/MS analysis and pharmacokinetic study on five bioactive constituents of Tanreqing injection in rats.

Tanreqing injection (TRQ), a well-known traditional Chinese medicine formula, is commonly used to treat respiratory diseases. In the present study, a rapid, selective, and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated to simultaneously determinate the plasma contents of 5 major constituents of TRQ, including chlorogenic acid (CHA), caffeic acid (CFA), baicalin (BA), ursodeoxycholic acid (UDCA) and chenodeoxycholic acid (CDCA) in rats after intravenous administration of TRQ. Chromatographic separation was performed on an Agilent Zorbax SB-C18 column (3.5 µm, 100 mm × 2.1 mm), with acetonitrile and 0.1% aqueous formic acid as mobile phase at a flow rate of 0.3 mL·min-1. The calibration curves were linear over the ranges of 27.0-13 333.0 ng·mL-1 for CFA, 30.0-14 933.0 ng·mL-1 for CHA, 50.0-50 333.0 ng·mL-1 for BA, 550.0-55 000.0 ng·mL-1 for UDCA, and 480.0-48 000.0 ng·mL-1 for CDCA, respectively. Intra- and inter-day precisions (relative standard deviations, RSDs) were from 3.11% to 14.08%. The extraction recoveries were greater than 71% and accuracy (relative recovery) was from 89% to 137% for all analytes, except endogenous bile acids. This validated method was successfully applied to the first pharmacokinetic study of CFA, CHA, BA, UDCA and CDCA in rat plasma after intravenous administration of TRQ.

Detection of structurally similar adulterants in botanical dietary supplements by thin-layer chromatography and surface enhanced Raman spectroscopy combined with two-dimensional correlation spectroscopy.

Thin-layer chromatography (TLC) coupled with surface enhanced Raman spectroscopy (SERS) has been widely used for the study of various complex systems, especially for the detection of adulterants in botanical dietary supplements (BDS). However, this method is not sufficient to distinguish structurally similar adulterants in BDS since the analogs have highly similar chromatographic and/or spectroscopic behaviors. Taking into account the fact that higher cost and more time will be required for comprehensive chromatographic separation, more efforts with respect to spectroscopy are now focused on analyzing the overlapped SERS peaks. In this paper, the combination of a TLC-SERS method with two-dimensional correlation spectroscopy (2DCOS), with duration of exposure to laser as the perturbation, is applied to solve this problem. Besides the usual advantages of the TLC-SERS method, such as its simplicity, rapidness, and sensitivity, more advantages are presented here, such as enhanced selectivity and good reproducibility, which are obtained by 2DCOS. Two chemicals with similar structures are successfully differentiated from the complex BDS matrices. The study provides a more accurate qualitative screening method for detection of BDS with adulterants, and offers a new universal approach for the analysis of highly overlapped SERS peaks.

Bioactive compound reveals a novel function for ribosomal protein S5 in hepatic stellate cell activation and hepatic fibrosis.

UNLABELLED: Liver fibrosis and its endstage, cirrhosis, represent a major public health problem worldwide. Activation of hepatic stellate cells (HSCs) is a central event in hepatic fibrosis. However, the proteins that control HSC activation are incompletely understood. Here we show that (6aS, 10S, 11aR, 11bR, 11cS)-10-methylamino-dodecahydro-3a, 7a-diaza-benzo [de]anthracene-8-thione (MASM) exhibits potent inhibitory activity against liver fibrosis in vitro and in vivo associated with the reduction of Akt phosphorylation. Furthermore, ribosomal protein S5 (RPS5) was identified as a direct target of MASM, which stabilized RPS5 in cultured HSCs and in the liver of experimental animals after dimethylnitrosamine (DMN) or bile duct ligation (BDL). Functional studies revealed that RPS5 could prevent HSC activation. RPS5 overexpression in HSCs resulted in Akt dephosphorylation at both Ser473 and Thr308, and led to subsequent dephosphorylation of GSK3ß or P70S6K. Progression of DMN- and BDL-induced hepatic fibrosis was aggravated by Rps5 knockdown and alleviated by RPS5 overexpression, which correlated with the modulation of Akt phosphorylation and HSC number in the fibrotic livers. Moreover, RPS5 was substantially reduced in the transdifferentiated HSCs, experimental fibrotic livers, and human cirrhosis samples. CONCLUSION: These results demonstrate that RPS5 is implicated in hepatic fibrogenesis and may represent a promising target for potential therapeutic intervention in liver fibrotic diseases.

UPLC-Q-TOF/MS based metabolomic profiling of serum and urine of hyperlipidemic rats induced by high fat diet.

Hyperlipidemia is considered to be a high lipid level in blood, can induce metabolic disorders and dysfunctions of the body, and results in some severe complications. Therefore, hunting for some metabolite markers and clarifying the metabolic pathways in vivo will be an important strategy in the treatment and prevention of hyperlipidemia. In this study, a rat model of hyperlipidemia was constructed according to histopathological data and biochemical parameters, and the metabolites of serum and urine were analyzed by UPLC-Q-TOF/MS. Combining pattern recognition and statistical analysis, 19 candidate biomarkers were screened and identified. These changed metabolites indicated that during the development and progression of hyperlipidemia, energy metabolism, lipid metabolism, amino acid metabolism and nucleotide metabolism were mainly disturbed, which are reported to be closely related to diabetes, cardiovascular diseases, etc. This study demonstrated that a UPLC-Q-TOF/MS based metabolomic approach is useful to profile the alternation of endogenous metabolites of hyperlipidemia.

High-performance liquid chromatography-electrospray ionization time-of-flight mass spectrometry analysis of Radix Saposhnikoviae for metabolomic research.

In this study, metabolite profiling of Radix Saposhnikoviae from different geographical locations was performed using high-performance liquid chromatography-electrospray ionization time-of-flight mass spectrometry (HPLC-ESI-TOFMS) and multivariate statistical analysis technique. Principle component analysis (PCA) of the data shows that these samples could be roughly separated into three groups: Guan Fangfeng, Kou Fangfeng and Chuan Fangfeng. The potential chemical markers were discovered through the loading plot of PCA. Based on accurate mass measurements and subsequent fragment ions of TOFMS after in-source collision induced dissociation, as well as matching of empirical molecular formulae with those of published components in the in-house chemical library, 10 potential markers, such as 4'-O-glucosyl-5-O-methylvisamminol, cimifugin, prim-O-glucosylcimifugin and 3'-O-angeloylhammaudol, were tentatively identified and partially verified by the available reference standards. The results of this study indicate that it is an effective and novel approach to identify traditional Chinese medicine (TCM) from different sources, and that performing quantity determination of corresponding marker compounds could optimize the quality control of TCM.

Using cell membrane chromatography and HPLC-TOF/MS method for in vivo study of active components from roots of Aconitum carmichaeli.

An offline two-dimensional system combining a rat cardiac muscle cell membrane chromatography time-of-flight mass spectrometry (CMC-TOF/MS) with a high Performance liquid chromatography time-of-flight mass spectrometry (HPLC-TOF/MS) was established for investigating the parent components and metabolites in rat urine samples after administration of the roots of Aconitum carmichaeli. On the basis ofthe analysis of the first dimension, retention components of the urine sample were collected into 30 fractions (one fraction per minute). Then offline analysis of the second dimension was carried out. 34 compounds including 24 parent alkaloids and 10 potential metabolites were identified from the dosed rat urine, and then binding affinities of different compounds on cell membranes were compared and influences of some functional groups on activity were estimated with the semi-quantification and curve fitting method. As a result, binding affinities decreased along with the process of deacylation, debenzoylation and demethylation, which may be related to the alleviation of toxicity in the procedure of herb processing or metabolism. Moreover, some minor components in rat urine (Songorine, 14-benzoylneoline, Deoxyaconitine, etc.) exerted relatively strong affinity on cell membranes are worth exploring. The results delivered by the System suggest that the CMC can be applied to in vivo study.

Comparative analysis of essential oils found in Rhizomes Curcumae and Radix Curcumae by gas chromatography-mass spectrometry.

A comparison of the volatile compounds in Rhizomes Curcumae (Ezhu) and Radix Curcumae (Yujin) was undertaken using gas chromatography-mass spectrometry (GC-MS). Ultrasonic extraction and GC-MS methods were developed for the simultaneous determination of five sesquiterpenes, namely, α-pinene, ß-elemene, curcumol, germacrone and curdione, in Ezhu and Yunjin. Good linearity (r>0.999) and high inter-day precision were observed over the investigated concentration ranges. The validated method was successfully used for the simultaneous determination of five sesquiterpenes in Ezhu and Yujin. The quantitative method can be effectively used to evaluate and monitor the quality of Chinese curcuma in clinical use.

Liquid chromatography coupled with time-of-flight and ion trap mass spectrometry for qualitative analysis of herbal medicines.

With the expansion of herbal medicine (HM) market, the issue on how to apply up-to-date analytical tools on qualitative analysis of HMs to assure their quality, safety and efficacy has been arousing great attention. Due to its inherent characteristics of accurate mass measurements and multiple stages analysis, the integrated strategy of liquid chromatography (LC) coupled with time-of-flight mass spectrometry (TOF-MS) and ion trap mass spectrometry (IT-MS) is well-suited to be performed as qualitative analysis tool in this field. The purpose of this review is to provide an overview on the potential of this integrated strategy, including the review of general features of LC-IT-MS and LC-TOF-MS, the advantages of their combination, the common procedures for structure elucidation, the potential of LC-hybrid-IT-TOF/MS and also the summary and discussion of the applications of the integrated strategy for HM qualitative analysis (2006-2011). The advantages and future developments of LC coupled with IT and TOF-MS are highlighted.

[Fast determination of volatile components in rhizoma curcuma by headspace gas chromatography-mass spectrometry].

OBJECTIVE: To analyze the chemical constituents of essential oil extracted from Rhizoma Curcumae. METHODS: Headspace-GC/MS was employed to prepare essential oil from rhizome curcuma, with TR-5 quartz capillary column, extraction temperature 90 degrees C, hold 30 min, injection volume 1.5 mL. CONCLUSION: A rapid, simple and reliable method using HS-GC/MS was developed to analyze 79 volatile components, which can be an alternative for quality control for TCMs such as Rhizome Curcuma.

[Local straight line screening method for the detection of Chinese proprietary medicines containing undeclared prescription drugs].

The present paper constructs a new approach named local straight-line screening (LSLS) to detect Chinese proprietary medicines (CPM) containing undeclared prescription drugs (UPD). Different from traditional methods used in analysis of multi-component spectrum, LSLS is proposed according to the characteristics of original infrared spectra of the UPD and suspected CPM, without any pattern recognition or concentration model establishment. Spectrum-subtraction leads to the variance in local straight line, which serves as a key in discrimination of whether suspected CPD is adulterated or not. Sibutramine hydrochloride, fenfluramine hydrochloride, sildenafil citrate and lovastatin were used as reference substances of UPD to analyze 16 suspected CPM samples. The results show that LSLS can obtain an accurate quantitative and qualitative analysis of suspected CPM. It is possible for the method to be potentially used in the preliminary screening of CPM containing possible UPD.

Characterization of constituents in Stellera chamaejasme L. by rapid-resolution liquid chromatography-diode array detection and electrospray ionization time-of-flight mass spectrometry.

A reliable and rapid method based on rapid-resolution liquid chromatography-diode array detection (RRLC-DAD) and electrospray ionization time-of-flight mass spectrometry (ESI-TOF/MS) has been developed for the isolation and characterization of multiple constituents in the root of Stellera chamaejasme L., which was extracted by sonication with methanol in an optimized procedure. Separation of the multiple constituents was achieved on an Agilent Zorbax XDB-C18 (50x3.0 mm i.d.; 1.8 microm) column using a gradient elution at a flow rate of 0.4 mL/min. The detection wavelength was 210 nm. Mass spectra were acquired in both positive and negative modes. A formula database of the known chemical constituents in the root of Stellera chamaejasme L. was established by an Agilent software. Twenty-two obvious peaks appeared in the total ion chromatogram and nine of them were characterized by TOF/MS. The RRLC-DAD and ESI-TOF/MS method with ultrasonic extraction would be useful for rapid and effective characterization of chemical constituents in the root of Stellera chamaejasme L.

[Discrimination of adulterated traditional Chinese medicines by infrared spectroscopy-two dimensional correlation analysis].

A new method was established, based on infrared spectroscopy two dimensional (2D) correlation analysis, for the discriminative analysis of adulteration in traditional Chinese medicines (TCM). Fenfluramine hydrochloride (FH) and sibutramine hydrochloride (SH) were taken as examples of synthetic drugs (adulterant), and the correlative peaks of their synchronous 2D correlation spectra were found. Then the characteristics of the synchronous 2D correlation spectrum of the suspected TCM were compared with those of FH. Since the correlative peaks in the synchronous spectrum of the suspected TCM coincide well with those of FH, a positive conclusion could be drawn after further investigation of the asynchronous spectra of TCM, which could provide the information about the source of correlative peaks. On the contrary, the dissimilarity of the synchronous spectra of SH and TCM directly implies that the suspected TCM is not adulterated with SH. The method can be used for a correct discrimination on whether the TCM is adulterated with the synthetic drugs, it does not rely on sample separation, and provides a new simple and cost-effective alternative to test the adulteration of TCM.

[Determination of three constituents in Radix Astragali by HPLC-MS].

AIM: To determine calycosin-7-O-beta-D-glucoside, astragaloside IV and formononetin in Radix Astragali and other relative samples by HPLC-MS. METHODS: HPLC was carried out with Agilent 1100LC/MSD, equipped with Agilent Zorbax SB C18 column (250 mm x 4.6 mm ID, 5 microm) and mass spectrum detector. The mobile phase (CH3CN-H2O) was eluted in gradient mode. RESULTS: The calibration curves of calycosin-7-O-beta-D-glucoside, astragaloside IV and formononetin were linear in the range of 0.03 - 1.21 microg x mL(-1), 0.35 - 13.86 microg x mL(-1) and 0.38 - 15.22 microg x mL(-1), respectively. These recoveries of samples were from 95% to 105% with RSD less than 1.5%. CONCLUSION: The method was employed to analyse 25 samples of Radix Astragali and other relative samples, including Radix Astragali slice, Radix Astragali Preparata, Hedysarum polybotrys Hand. -Mazz, Astragalus ernestii Comb. The contents of three constituents vary greatly because of the species, place of collection and season of harvesting. This method could apply to evaluate the quality of Radix Astragali and it is simple, sensitive and reliable.