Identification of Cald1 as a novel regulator of Linggui Zhugan decoction for improving insulin resistance in vivo and in vitro.

OBJECTIVE: To identify Cald1 as a novel regulator of Linggui Zhugan decoction for improving insulin resistance in vivo and in vitro. METHODS: Sprague-Dawley rats were randomly assigned to 3 groups that were received a normal rat chow diet, high-fat diet (HFD), and an HFD plus LGZGD, respectively. The homeostatic model assessment (HOMA)-insulin resistance (IR) index was used to determine IR. Gene microarray methodology was used to identify differentially expressed genes (DEGs) in the three groups of rats. The DEGs associated with IR were confirmed by quantitative real-time polymerase chain reaction. Additionally, Mouse 3T3-L1 pre-adipocytes were differentiated into mature 3T3-L1 adipocytes, which were then treated with tumor necrosis factor (TNF)-α to induce cellular IR. Lipid accumulations were identified by Oil Red O staining. Glucose uptake was assessed using the 3 H-2-DG test. RESULTS: In this study, we found Cald1 was further screened to validate its biological function in 3T3-L1 adipocytes induced to develop IR. In vitro experiments showed that insulin-stimulated 3H2-DG uptake by IR 3T3-L1 adipocytes was increased after LGZGD intervention, which was associated with a down-regulation of Cald1 expression. CONCLUSION: LGZGD ameliorates HFD-induced IR in rats and TNF-α induced IR in adipocytes by down-regulating Cald1 expression.

Systematic and practical solvent system selection strategy based on the nonrandom two-liquid segment activity coefficient model for real-life counter-current chromatography separation.

Solvent system selection is the first step toward a successful counter-current chromatography (CCC) separation. This paper introduces a systematic and practical solvent system selection strategy based on the nonrandom two-liquid segment activity coefficient (NRTL-SAC) model, which is efficient in predicting the solute partition coefficient. Firstly, the application of the NRTL-SAC method was extended to the ethyl acetate/n-butanol/water and chloroform/methanol/water solvent system families. Moreover, the versatility and predictive capability of the NRTL-SAC method were investigated. The results indicate that the solute molecular parameters identified from hexane/ethyl acetate/methanol/water solvent system family are capable of predicting a large number of partition coefficients in several other different solvent system families. The NRTL-SAC strategy was further validated by successfully separating five components from Salvia plebeian R.Br. We therefore propose that NRTL-SAC is a promising high throughput method for rapid solvent system selection and highly adaptable to screen suitable solvent system for real-life CCC separation.

Enhancement of secondary metabolites from Bacillus Licheniformis XY-52 on immune response and expression of some immune-related genes in common carp, Cyprinus carpio.

This study was undertaken to isolate active secondary metabolites from immunostimulatory Bacillus Licheniformis XY-52 and evaluate their activities at 0%, 0.5%, 1.0%, and 2.0% doses supplementation with feed on immune response in common carp at weeks 1, 2, and 3. By applying chromatography techniques and successive recrystallization, two purified metabolites were obtained and identified by spectral data (mass spectrometry and nuclear magnetic resonance) as: Cyclo-(Phe-Tyr) and Cyclo-(Phe-Gly). The results revealed that humoral innate immune parameters (lysozyme activity, phagocytic activity and bactericidal activity) were significantly (P < 0.05) increased after feeding on the two active compounds-supplemented diet. Furthermore, administration of the two active compounds significantly (P < 0.05) up regulated IL-1ß, Type 1 IFN, IFN g2b, IL10 and TNF-α gene expression. Heat shock protein 70 (HSP70) gene expression was significantly (P < 0.05) lower as compared to control group at the end of trial. Common carp fed with the two compounds had higher survival rates (69.3%) compared to the controls (32.0%) after challenged with the pathogen, Aeromonas hydrophila. The present study indicates that the two isolated active compounds could positively influence immune response and enhance disease resistance of common carp against A. hydrophila infection.

Separation of nine compounds from Salvia plebeia R.Br. using two-step high-speed counter-current chromatography with different elution modes.

Nine compounds were successfully separated from Salvia plebeia R.Br. using two-step high-speed counter-current chromatography with three elution modes. Elution-extrusion counter-current chromatography was applied in the first step, while classical counter-current chromatography and recycling counter-current chromatography were used in the second step. Three solvent systems, n-hexane/ethyl acetate/ethanol/water (4:6.5:3:7, v/v), methyl tert-butyl ether/ethyl acetate/n-butanol/methanol/water (6:4:1:2:8, v/v) and n-hexane/ethyl acetate/methanol/water (5:5.5:5:5, v/v) were screened and optimized for the two-step separation. The separation yielded nine compounds, including caffeic acid (1), 6-hydroxyluteuolin-7-glucoside (2), 5,7,3',4'-tetrahydroxy-6-methoxyflavanone-7-glucoside (3), nepitrin (4), rosmarinic acid (5), homoplantaginin (6), nepetin (7), hispidulin (8), and 5,6,7,4'-tertrahydroxyflavone (9). To the best of our knowledge, 5,7,3',4'-tetrahydroxy-6-methoxyflavanone-7-glucoside and 5,6,7,4'-tertrahydroxyflavone have been separated from Salvia plebeia R.Br. for the first time. The purities and structures of these compounds were identified by high-performance liquid chromatography, electrospray ionization mass spectrometry, (1)H and (13)C NMR spectroscopy. This study demonstrates that high-speed counter-current chromatography is a useful and flexible tool for the separation of components from a complex sample.

A strategy that iteratively retains informative variables for selecting optimal variable subset in multivariate calibration.

Nowadays, with a high dimensionality of dataset, it faces a great challenge in the creation of effective methods which can select an optimal variables subset. In this study, a strategy that considers the possible interaction effect among variables through random combinations was proposed, called iteratively retaining informative variables (IRIV). Moreover, the variables are classified into four categories as strongly informative, weakly informative, uninformative and interfering variables. On this basis, IRIV retains both the strongly and weakly informative variables in every iterative round until no uninformative and interfering variables exist. Three datasets were employed to investigate the performance of IRIV coupled with partial least squares (PLS). The results show that IRIV is a good alternative for variable selection strategy when compared with three outstanding and frequently used variable selection methods such as genetic algorithm-PLS, Monte Carlo uninformative variable elimination by PLS (MC-UVE-PLS) and competitive adaptive reweighted sampling (CARS). The MATLAB source code of IRIV can be freely downloaded for academy research at the website: http://code.google.com/p/multivariate-calibration/downloads/list.

Nonlinear alignment of chromatograms by means of moving window fast Fourier transfrom cross-correlation.

The preprocessing of chromatograms is essential to modern chromatography for further qualitative and quantitative analysis, especially when chromatographic instruments are used for herb products analysis involving large number of samples. To accurately compare and analyze the obtained chromatograms, it is necessary to preprocess, especially align retention time shifts. Here moving window fast Fourier transform (FFT) cross-correlation is introduced to perform nonlinear alignment of high-throughput chromatograms. Since elution characteristics of chromatograms will produce local similarity in retention time shifts, moving window procedure seems to be a better substitute of segmentation steps. The retention time shifts can be calculated and accelerated by FFT cross-correlation. The artifacts can be detected and eliminated from the retention time shifts profile since the continuity of moving window procedure. The proposed method is demonstrated in comparison with recursive alignment by FFT on chromatographic datasets from herb products analysis. It is shown that the proposed method can address nonlinear retention time shift problem in chromatograms with the simple moving window procedure, which will not introduce segments size optimization problem. In additional, the parameters are intuitive and easy to adjust, which makes it off-the-shelf toolbox for alignment of chromatograms.

Multiscale peak alignment for chromatographic datasets.

Chromatography has been extensively applied in many fields, such as metabolomics and quality control of herbal medicines. Preprocessing, especially peak alignment, is a time-consuming task prior to the extraction of useful information from the datasets by chemometrics and statistics. To accurately and rapidly align shift peaks among one-dimensional chromatograms, multiscale peak alignment (MSPA) is presented in this research. Peaks of each chromatogram were detected based on continuous wavelet transform (CWT) and aligned against a reference chromatogram from large to small scale gradually, and the aligning procedure is accelerated by fast Fourier transform cross correlation. The presented method was compared with two widely used alignment methods on chromatographic dataset, which demonstrates that MSPA can preserve the shapes of peaks and has an excellent speed during alignment. Furthermore, MSPA method is robust and not sensitive to noise and baseline. MSPA was implemented and is available at http://code.google.com/p/mspa.

Progallin A isolated from the acetic ether part of the leaves of Phyllanthus emblica L. induces apoptosis of human hepatocellular carcinoma BEL-7404 cells by up-regulation of Bax expression and down-regulation of Bcl-2 expression.

AIM OF THE STUDY: This study was aimed to investigate the effects of Progallin A isolated from the acetic ether part of the leaves of Phyllanthus emblica L. on apoptosis in human hepatocellular carcinoma BEL-7404 cells and its possible mechanisms, and to measure the immune toxicity of Progallin A in vitro. MATERIALS AND METHODS: Progallin A was isolated from the acetic ether part of the leaves of Phyllanthus emblica L. with column chromatography. The proliferation of spleen lymphocytes and the viability of BEL-7404 cells were assessed with MTT assay. Inverted microscope, light microscope and fluorescence microscope were utilized to observe the morphological changes of BEL-7404 cells respectively. AnnexinV/PI double labeling and TUNEL assay were used to detect early apoptosis and DNA fragmentations of BEL-7404 cells respectively. In addition, cell cycle distribution was analyzed by using flow cytometry (FCM). Bax and Bcl-2 protein levels were determined by immunofluorescence staining and western blot respectively. RESULTS: The results showed that Progallin A had low immune toxicity and the proliferation of BEL-7404 cells was strongly inhibited by Progallin A in a time- and dose-dependent manner and that the characteristics of apoptosis of BEL-7404 cells were observed. The results also showed that apoptosis rates and the number of apoptotic cells significantly increased with prolongation of the action time. The results of flow cytometry (FCM) indicated that Progallin A induced significant G1/M and G2/M arrest of BEL-7404 cells. Immunofluorescence staining and western blot showed that the expression of Bax was found to be noticeably up-regulated and the expression of Bcl-2 was down-regulated significantly. CONCLUSIONS: These results indicate that Progallin A has low immune toxicity in vitro and induces apoptosis of human hepatocellular carcinoma BEL-7404 cells which is related to the G1/M and G2/M arrest, and it exerts its apoptotic effect by up-regulation of Bax expression and down-regulation of Bcl-2 expression.

Supercritical CO2 extraction of emodin and physcion from Polygonum cuspidatum and subsequent isolation by semipreparative chromatography.

Emodin and physcion are abundant anthraquinone compounds found in the traditional Chinese medicinal herb Polygonum cuspidatum Sied. et Zucc. In this paper, emodin and physcion were successfully extracted with supercritical CO2 plus ethanol modifier after the extraction conditions were optimized with uniform design-sequential optimization. Results showed that the ethanol modifier concentration was the main factor for the effective extraction of the emodin. The optimal extraction condition was obtained: 20 MPa, 30 degrees C, and 95% ethanol, at which the yields of emodin and physcion were 0.616 and 0.178 g/100 g, respectively. The yield obtained by supercritical fluid extraction (SFE) was a little lower than that obtained by sonication extraction (SE). The crude extract obtained by SFE was further isolated and purified by semipreparative chromatography with the mobile phase composed of methanol-water (90:1, v/v). Emodin and physcion were obtained with purity 98.6 and 99.1%, respectively, when determined by HPLC, and identification was performed by retention time and UV spectra of the standards. The result suggested that SFE is an alternative and promising method for extraction of the two compounds from P. cuspidatum owing to its environment-friendly properties and fewer coextracts.

Tentative fingerprint-efficacy study of Houttuynia cordata injection in quality control of traditional Chinese medicine.

To establish potent fingerprint for quality control of traditional Chinese medicine, Houttuynia cordata (Saururaceae) injection (HCI), the attempt on fingerprint-efficacy was developed in this study. HCI from ten different factories were determined by gas chromatography-mass spectrum (GC-MS) and classified by hierarchical clustering. The anti-inflammatory effect of HCI was characterized with the rat pleurisy model induced by carrageenin and the mice ear edema model by xylene. The results showed that anti-inflammatory effect of the injections from most of factories on the two models was significant. There was corresponding relationship between the fingerprint of HCI and efficacy to certain extent. The main common constitutes in injection from the factories that possess anti-inflammatory activity were analysed with GC-MS and identified using the NIST Mass Spectral Database. This common pattern of HCI based on the efficacy was helpful for the purpose of quality control.

[Profile-effect on quality control of Houttuynia cordata injection].

AIM: To find corresponding relationship between the fingerprint of Houttuynia cordata injections from different factories and their effects. METHODS: Houttuynia cordata injections from six different factories were determined by gas chromatography (GC) and gas chromatography-mass spectra (GC-MS), and GC fingerprints were classified by hierarchical clustering. The anti-inflammatory activity of Houttuynia cordata injections was characterized through the rat pleurisy model induced by carrageenin and the mice ear edema model by dimethylbenzene. RESULTS: The anti-inflammatory effect of the injections from the first class factories on the two model was significant, while those from the second class not. GC-MS analysis result indicated that main effect compounds in Houttuynia cordata injections are methyl n-nonyl ketone, decanoylacetaldehyde, lauryl aldehyde, capryl aldehyde, beta-pinene, beta-linalool, 1-nonanol, 4-terpineol, alpha-terpineol, bornyl acetate, n-decanoic acid and acetic acid, geraniol ester etc. CONCLUSION: There is corresponding relationship between the fingerprint of Houttuynia cordata injections and effect to a certain extent.