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Depression is a common complex psychiatric disorder but its pathophysiological mechanism is not yet fully understood. Metabonomics by GC-MS and multivariate statistical analysis were used to select potential biomarkers associated with CUMS (chronic unpredictable mild stress) depression. The dynamic metabolic changes in rat serum were investigated to find potential disease biomarkers and to investigate the pathology of depression induced by the CUMS depression model. The changes in behavior and serum metabolic profiles were investigated during a three-week CUMS exposure. Serum samples were collected on days 0, 6, 9, 12, 15 and 21, and the serum metabolic profiling was carried out using GC-MS, followed by multivariate analysis. The potential biomarkers were screened from metabolites by principal component analysis and correlation analysis. The peak area of potential biomarkers was used to identify changes in depression in rats and describe their dynamics. Exposure to CUMS for three weeks caused depression-like behavior in rats, as indicated by significant decreases in weight gain, sucrose consumption, ambulation number and rearing numbers. Six potential biomarkers in serum, including glycine (Gly), glutamic acid (Glu), fructose, citric acid, glucose and hexadecanoic acid, were subjected to screening by metabonomics and multivariate statistical analysis. It was found that fructose, glucose and Gly were increased in the model group, while hexadecanoic acid, Glu and citric acid were reduced in the model group. According to the results of principal component analysis and correlation analysis, the correlation coefficient between the behavior scores and potential biomarkers in serum were all more than 0.9. This result suggests that the progression of depression may be associated with perturbation of glycometabolism, amino acid metabolism and energy metabolism. Gly, Glu, fructose, citric acid, glucose and hexadecanoic acid appear to be suitable quantitative diagnostic biomarkers for depression. The representative and unique nature of these biomarkers needs to be verified by pharmacological experiments, including molecular pharmacology investigations of enzymes or genes.

The major therapeutic strategy used to treat exacerbated cystic fibrosis (CF) is antibiotic treatment. As this approach easily generates antibiotic-resistant strains of opportunistic bacteria, optimized antibiotic therapies are required to effectively control chronic and recurrent bacterial infections in CF patients. A promising future for the proper use of antibiotics is the management of lung microbiota. However, the impact of antibiotic treatments on CF microbiota and vice versa is not fully understood. This study analyzed 718 sputum samples from 18 previous studies to identify differences between CF and uninfected lung microbiota and to evaluate the effects of antibiotic treatments on exacerbated CF microbiota. A reference-based OTU (operational taxonomic unit) picking method was used to combine analyses of data generated using different protocols and platforms. Findings show that CF microbiota had greater richness and lower diversity in the community structure than uninfected control (NIC) microbiota. Specifically, CF microbiota showed higher levels of opportunistic bacteria and dramatically lower levels of commensal bacteria. Antibiotic treatment affected exacerbated CF microbiota notably but only transiently during the treatment period. Limited decrease of the dominant opportunistic bacteria and a dramatic decrease of commensal bacteria were observed during the antibiotic treatment for CF exacerbation. Simultaneously, low abundance opportunistic bacteria were thriving after the antibiotic treatment. The inefficiency of the current antibiotic treatment against major opportunistic bacteria and the detrimental effects on commensal bacteria indicate that the current empiric antibiotic treatment on CF exacerbation should be reevaluated and optimized.

Plant metabolomics combined with GC-MS was used to investigate metabolic fingerprinting of Tussilago farfara at different growth stages.Dried Samples were extracted by two-phase solvent system to obtain polar and nonpolar parts, which were subjected to GC-MS analysis. Metabolites were identified by NIST data base search and comparison with the authentic standards. The data were introduced into SIMCA-P 11.0 software package for multivariate analysis after pretreatment.Fifty-four metabolites were identified, including 35 polar metabolites and 19 nonpolar compounds. The score plot for PCA showed clear separation of the different development stages of flower buds of T. farfara, showing a trend of gradual change. Samples of October, November, December were in close proximity on the plot, indicating that the metabolome of these three periods was similar, samples from September (early development) and March (after flowering) were far away, showing big chemical differences. Content comparison results of some representative metabolites reveals that, the content of proline, lysine and linoleic acid increased gradually to the highest in the medium term, but sharply decreased to the lowest after flowering; the content of malic acid and citric acid were the lowest in the medium term; sucrose content decreased gradually, and then reached the lowest level after blooming.It is obvious that metabolites of the early development and flowering stage were quite different with those of the traditional harvest time, suggesting that they can not be used as traditional medicine. This study will provide a research basis for harvest time determination and bioactive compounds of T. farfara.

The present study was aimed to discriminate the anti-depression of Xiao-yao-san series prescriptions on forced swimming test (FST) rat model via metabonomic. Xiao-yao-san (XY), Dan-zhi-xiao-yao-san (DZ) and Chai-hu-shu-gan-san (CH) formula were extracted with aqueous solution, and the final concentration was 46.3, 48.9 and 7.9 g/kg (the total amount of crude drug), respectively. The saline, venlafaxin and Xiao-yao-san series extracts were orally treated to the rat groups for 2 weeks. The forced swimming test was used to evaluate the action of antidepressant and the plasma was collected simultaneously. Plasma samples were prepared and measured using NMR, and the data were processed with SIMCA-P software. Variable importance in score plot and loading plot were used to screen relative biomarkers. Significant difference was observed in the FST and control group, which was an illustration that the pharmacological model was duplicated successfully. The effect of Xiao-yao-san was similar to that of positive drug venlafaxin, while the effect of Dan-zhi-xiao-yao-san and Chai-hu-shu-gan-san is not as good as Xiao-yao-san. Compared with the control group, significant changes were observed in lactate, thrronine, Glu, choline, glucose in plasma of venlafaxin and Xiao-yao-san groups. The 1H NMR analysis suggested that Xiao-yao-san prescription was of significant anti-depression effect which is better than that of Dan-zhi-xiao-yao-san and Chai-hu-shu-gan-san.

Ethnopharmacological relevance Xiaoyaosan (XYS), a well-known formula for relieving depression, was originated from the book of “Taiping Huimin Heji Jufang” in Song Dynasty (960–1127 AD), composed of Radix Bupleuri , Radix Angelicae Sinensis , Radix Paeoniae Alba , Rhizoma Atractylodis Macrocephalae , Poria , Herba Menthae , Rhizoma Zingiberis Recens and Radix Glycyrrhizae with dose proportion of 6:6:6:6:6:3:2:2. It is commonly used for the treatment of depression-related syndromes in China. In the formula, Radix Bupleuri usually serves as the principal drug, Radix Angelicae Sinensis and Radix Paeoniae Alba serve as the ministerial drugs, Rhizoma Atractylodis Macrocephalae , Poria , Herba Menthae and Rhizoma Zingiberis Recens serve as adjunctive drugs, Radix Glycyrrhizae serves as messenger drug, they coordinate with each other and enhance the effect of the formula. In our previous experiments, the antidepressant effect of XYS was revealed. However, the antidepressant part (or component) of this prescription was still obscure. Materials and methods An experimental despair animal model: the mice tail suspension test (TST) was used to evaluate the antidepressant activity of XYS and its fractions. GC–MS method was developed to identify the volatile components and determine 4 major volatile components in active fraction. Results In the TST test, the effect of a low polar fraction (XY-EA) was superior to other fractions of XYS. 13 volatile compounds in the XY-EA were identified on the basis of standards, isolation and structural determination in our laboratory, NIST 05 database and literature data. The content of 4 major volatile compounds in XY-EA which is 6.703%. Conclusions The petroleum ether fraction (XY-EA) appears to be the active fraction of XYS. 4 major components Z -ligustilide, palmitic acid, atractylenolide I, and atractylenolide II may be the antidepressant active compounds.

The flower bud of Tussilago farfara L. is widely used for the treatment of coughs, bronchitis and asthmatic disorders in traditional Chinese medicine. In Europe, the plant has been used as herbal remedies for virtually the same applications, but the leaves are preferred over flowers.To systematically evaluate the chemical profiles of Tusssilago farfara leaves and flowers along with the identification of the polar and non-polar metabolites.Metabolic profiling carried out by means of ¹H-NMR spectroscopy and multivariate data analysis was applied to crude extracts from flowers and leaves. Metabolites were identified directly from the crude extracts through one-dimensional and two-dimensional NMR spectra.A broad range of metabolites were detected without any chromatographic separation. Principal component analysis (PCA) and partial least squares-discriminant analysis (PLS-DA) of ¹H-NMR data provided a clear separation between the samples. The corresponding loadings plot indicated that higher levels of phenylpropanoids, amino acids, organic acids and fatty acids, as well as lower levels of sugars, terpenoids and sterols were present in the leaves, as compared with flowers. For the flowers, more phenylpropanoids were present in fully open flowers, while more sugars and fatty acids were present in flower buds.NMR spectra (one- and two-dimensional) are useful for identifying metabolites, especially for the overlapped signals. The NMR-based metabolomics approach has great potential for chemical comparison study of the metabolome of herbal drugs.

Ethnopharmacological relevance Flower bud of Tussilago farfara L. is widely used for the treatment of cough, bronchitis and asthmatic disorders in the Traditional Chinese Medicine. However, due to the increasing demands, adulteration with rachis is frequently encountered in the marketplace. No report demonstrated the chemical and pharmacological differences between flower bud and rachis before. Materials and methods The water extracts were orally administrated to mice. Ammonia induced mice coughing model was used to evaluate the antitussive activity. The expectorant activity was evaluated by volume of phenol red in mice's tracheas. Metabolites were identified directly from the crude extracts through 1D- and 2D-NMR spectra. A metabolic profiling carried out by 1H NMR spectroscopy and multivariate data analysis was applied to crude extracts from flower bud and rachis. Results Flower bud significantly lengthened the latent period of cough, decreased cough frequency caused by ammonia and enhanced tracheal phenol red output in expectorant evaluation. Principal component analysis (PCA) yielded good separation between flower bud and rachis, and corresponding loading plot showed that the phenolic compounds, organic acid, sugar, amino acid, terpene and sterol contributed to the discrimination. Conclusions These findings provide pharmacological and chemical evidence that only flower bud can be used as the antitussive and expectorant herbal drug. The high concentration of chlorogenic acid, 3,5-dicaffeoylquinic acid, rutin in flower buds may be related with the antitussive and expectorant effects of Flos Farfara. To guarantee the clinical effect, rachis should be picked out before use.

Chronic unpredictable mild stress (CUMS) is a well-validated model of depression. In this study, a urinary metabonomics method based on the NMR spectrometry was used to study the metabolic perturbation in CUMS-induced rat depression model. With pattern recognition analysis, a clear separation of CUMS rats and healthy controls was achieved, and nine endogenous metabolites contributing to the separation were identified. CUMS-treated rats were characterized by the increase of glycine, pyruvate, glutamine, and asparagines, as well as the decrease of 2-oxoglutarate, dimethylglycine, citrate, succinate, and acetate. The urinary biochemical changes related to the metabolic disturbance in CUMS induced depression, and the possible correlations with live qi stagnation in traditional Chinese medicine are discussed. The work shows that CUMS is a reliable model for studying depression, and the noninvasive urinary metabolomic method is a valuable tool to investigate the biochemical pertubations in depression as an early diagnostic means.

a b s t r a c t Ethnopharmacology: Xiaoyaosan (XYS), a famous Chinese prescription, composed of Radix Bupleuri (Bupleurum chinense DC.), Radix Angelicae Sinensis (Angelica sinensis (Oliv.) Diels), Radix Paeoniae Alba (Paeonia lactiflora Pall.), Rhizoma Atractylodis Macrocephalae (Atractylodes macrocephala Koidz.), Poria (Poria cocos (Schw.) Wolf), Radix Glycyrrhizae (Glycyrrhiza uralensis Fisch.), Herba Menthae (Mentha haplocalyx Briq.), and Rhizoma Zingiberis Recens (Zingiber officinale Rosc.), has been widely used in the clinic for treating mental disorders. Behavior and biochemical analyses indicate XYS has obvious anti-depression activity. However, there is no report on the effects of XYS using a metabolomics approach. Aim of the study: Depression is a prevalent complex psychiatric disorder and its pathophysiological mech- anism is not yet well understood. This paper was designed to study metabonomic characters of the depression induced by chronic unpredictable mild stress (CUMS) and the therapeutic effects of XYS, classic traditional Chinese medicine (TCM) in treating the depression. Material and methods: A plasma metabonomics method based on gas chromatography/mass spectrom- etry (GC/MS) was developed. Principal component analysis (PCA) was utilized to classify and reveal the differences between the model group and control group. In turns, the concentration of these differences was analyzed with t-test to determine whether XYS was possible to influence the metabolic pattern induced by CUMS. Results: The significant difference in metabolic profiling was observed from model group compared with drug-dose group by using the PCA, indicating the recovery effect of XYS on CUMS rats. Some signif- icantly changed metabolites like glycine, glucose and hexadecanoic acid have been identified. These biochemical changes are related to the disturbance in amino acid metabolism, energy metabolism and gly- cometabolism, which are helpful to further understand the CUMS and the therapeutic mechanism of XYS. Conclusions: Metabonomic approach is helpful to further understanding the pathophysiology of depres- sion and assisting in clinical diagnosis of depression and is also a valuable tool for studying the essence of Chinese medicine's syndrome theory and therapeutic effect mechanism of TCM. © 2011 Elsevier Ireland Ltd. All rights reserved.

A metabolomic investigation of chronic unpredictable mild stress (CUMS) rats was carried out. Plasma obtained from Sprague-Dawley (SD) rats treated by CUMS was analyzed using gas chromatography/mass spectrometry. Thirty-seven metabolites were identified among the detected compounds. Subsequent data analysis using the t test and principal component analysis (PCA) revealed significant metabolic changes in the rats' plasma after CUMS treatment. Clear separation between the model and control group was achieved, and the level of twelve metabolites, including amino acids, sugar, organic acids and fatty acids, were significantly different between plasma samples from the controls and CUMS group. These observations suggested that the depressed state may be associated with perturbation of amino acid metabolism, energy metabolism and glycometabolism. The study suggested that the metabolomics approach could be used as a potential powerful tool to investigate the biochemical change in certain physiopathological conditions, such as depression, as an early diagnostic means.

A high-performance liquid chromatographic (HPLC) method was established to analyze 36 Chaihu (Radix Bupleuri) samples collected from three species (Bupleurum chinense DC., B. scorzonerifolium Willd. and B. smithii Wolff.). Addition of trifluoroacetic acid into the mobile phase resulted in fingerprint chromatograms with stable baselines. There were thirty-two characteristic peaks in the standard fingerprint of B. chinense DC. Different recognition pattern methods, including similarity analysis (SA), hierarchical cluster analysis (HCA), principal component analysis (PCA) and partial least squares-discrimination analysis (PLS-DA) were utilized to analyze the 36 samples based on the contents of chemical constituents. Consistent results from SA, HCA and PCA analysis illustrated the rationalisation for why B. smithii Wolff. was not quoted in the Chinese Pharmacopoeia and classified samples were in agreement with their species. PLS-DA loading plots showed the chemical markers which had the most influences on the separation among different species. However, SA, HCA and PCA could not differentiate between wild and cultivated B. chinense DC. as well as between samples from different provinces. HPLC fingerprint in combination with chemometric techniques provided a very flexible and reliable method for homogeneity evaluation and quality assessment of traditional Chinese medicine.

Ethnopharmacology Xiaoyaosan, a famous Chinese prescription, composed of Poria (Poria cocos (Schw.) Wolf), Radix Paeoniae Alba (Paeonia lactiflora Pall.), Radix Glycyrrhizae (Glycyrrhiza uralensis Fisch.), Radix Bupleuri (Bupleurum chinense DC.), Radix Angelicae Sinensis (Angelica sinensis (Oliv.) Diels), Rhizoma Atractylodis Macrocephalae (Atractylodes macrocephala Koidz.), Herba Menthae (Mentha haplocalyx Briq.), and Rhizoma Zingiberis Recens (Zingiber officinale Rosc.), has been widely used in the clinic for treating mental disorders. Behavior and biochemical analyses indicate xiaoyaosan has obvious anti-depression activity. However, there is no report on the effects of xiaoyaosan using a metabolomics approach. Aim of the study A urinary metabolomics method was applied to evaluate the efficacy of xiaoyaosan on rat model of chronic unpredictable mild stress. Material and methods Rats were divided into 6 groups and drugs were administered during the 21-day model building period. Urine was measured using GC–MS, processed with XCMS and Microsoft Excel and analyzed by SIMCA-P and SPASS software. Variable importance in projection statistics and loading plot were used to find biomarker ions. Results Clear separation between model and each drug group was achieved. High dose group of xiaoyaosan was much closer to control group than middle dose group and amitriptyline group. The time-dependent recovery tendency in high dose group was obtained. Conclusions In term of anti-depression effect, high dose xiaoyaosan was the most effective and amitriptyline equaled middle dose xiaoyaosan as shown by metabolomics strategy and behavior tests. Some common and characteristic metabolites on the anti-depression of xiaoyaosan and amitriptyline were obtained. The work showed metabolomics is a valuable tool in studying the efficacy and potential biomarkers of therapeutic effect of complex prescriptions.

To determine the contents of saikosaponin a, d and saikosaponins in Radix Bupleuri from different and assess their quality.Saikosaponin a and d were used as the chemical reference substance to establish RP-HPLC and visible-spectrophotometry methods for determination of contents of saikosaponin a, d and saikosaponins.The quality of B. chinense was better than that of B. scozonerifolium and B smishii; The contents of a, saikosaponin d and total saikosaponin in B. chinense were 0.100%-0.330%, 0.200%-0.400% and 1.00%-2.00% , respectively.It is to suggest that the minimal contents requirements of saikosaponin a, d and saikosaponins in B. chinense should be set at 0. 100% , 0.200% and 1.00% , respectively; the best suitable planting areas are Taigu in the middle of Shanxi province, Ruicheng, Pinglu and Wanrong in the south of Shanxi province, and Longxi, Lixian in Gansu province.

The root, stem and leaf of Salvia miltiorrhiza were digested with HNO3-HClO4 (4:1), and the contents of five metallic elements such as K, Cu, Zn, Fe and Mg were detected by flame atomic absorption spectrophotometry. The recovery ratio by standard addition is 99.7%-104.4% and RSD is less than 2%. It was found that the content sequence of metallic elements is as follows: KMgFeZnCu. K, Cu, Zn, Fe and Mg in the leaf are clearly richer than those in the root and stem. The content ratio of Zn to Cu is relatively lower than that in the terrestrial angiosperms. The results provide a useful data for discussing the relationship between the contents of these elements in Chinese medicinal herb Salvia miltiorrhiza and the medical effects. Furthermore they also provide a data for the distribution of these elements in different parts of Salvia miltiorrhiza.

To investigate the expression of T cell early activation marker (CD(69)) on peripheral CD(4)(+) and CD(8)(+) lymphocytes and serum levels of soluble tumor necrosis factor receptor 1 and 2 (sTNF-R1 and sTNF-R2) in serum and bone marrow in patients with aplastic anemia (AA) and their pathophysiological significance.In vitro activation of T lymphocytes was carried out by whole blood cell culture containing PHA (20 micro g/ml). The CD(69) expressions on CD(4)(+) and CD(8)(+) lymphocytes at 0 h and 4 h after PHA exposure were analyzed by two-color flow cytometry. The levels of sTNF-R1 and sTNF-R2 in serum and bone marrow were measured by ELISA.The CD(69) expression rates of CD(4)(+) and of CD(8)(+) cells in SAA patients were (8.96 +/- 7.23)% and (10.67 +/- 7.58)%, respectively, and that of CD(8)(+) cells in CAA patients was (7.36 +/- 5.49)% before PHA stimulation. The CD(69) expression rates of CD(4)(+) and of CD(8)(+) cells in SAA patients were (71.73 +/- 11.91)% and (61.74 +/- 13.44)% and in CAA (59.35 +/- 10.15)% and (48.78 +/- 8.25)% respectively, and were significantly elevated after PHA stimulation. CD(69) expression on CD(4)(+) cells was much higher than that on CD(8)(+) cells after stimulation. The levels of the two sTNF-R (sTNF-R1 and sTNF-R2) in peripheral blood and bone marrow of SAA patients were elevated and in the bone marrow of CAA patients were also increased. The serum levels of sTNF-R2 were positively related to the CD(69) expression rates of CD(8)(+) cells before PHA stimulation.Increased early activation and activated potentials of T lymphocytes, along with abnormally elevated immunologically active molecules might play a major role in the pathogenesis of AA.

BackgroundThe dried root of Polygala tenuifolia, named Radix Polygalae, is a well-known traditional Chinese medicine. Triterpenoid saponins are some of the most important components of Radix Polygalae extracts and are widely studied because of their valuable pharmacological properties. However, the relationship between gene expression and triterpenoid saponin biosynthesis in P. tenuifolia is unclear.Methodology/findingsIn this study, ultra-performance liquid chromatography (UPLC) coupled with quadrupole time-of-flight mass spectrometry (Q-TOF MS)-based metabolomic analysis was performed to identify and quantify the different chemical constituents of the roots, stems, leaves, and seeds of P. tenuifolia. A total of 22 marker compounds (VIP>1) were explored, and significant differences in all 7 triterpenoid saponins among the different tissues were found. We also observed an efficient reference gene GAPDH for different tissues in this plant and determined the expression level of some genes in the triterpenoid saponin biosynthetic pathway. Results showed that MVA pathway has more important functions in the triterpenoid saponin biosynthesis of P. tenuifolia. The expression levels of squalene synthase (SQS), squalene monooxygenase (SQE), and beta-amyrin synthase (β-AS) were highly correlated with the peak area intensity of triterpenoid saponins compared with data from UPLC/Q-TOF MS-based metabolomic analysis.Conclusions/significanceThis finding suggested that a combination of UPLC/Q-TOF MS-based metabolomics and gene expression analysis can effectively elucidate the mechanism of triterpenoid saponin biosynthesis and can provide useful information on gene discovery. These findings can serve as a reference for using the overexpression of genes encoding for SQS, SQE, and/or β-AS to increase the triterpenoid saponin production of P. tenuifolia.

The roots of Rehmannia glutinosa Libosch, Rehmanniae Radix, also known as Dihuang in China, are an important herbal drug in Traditional Chinese Medicine (TCM). According to the theory of TCM, the raw and processed Rehmanniae Radix are used quite differently. In this study, the chemical change of Rehmanniae Radix induced by processing is investigated by 1H NMR spectroscopy based fingerprinting coupled with multivariate data analysis. The results indicated that big chemical change had occurred after the processing of Rehmanniae Radix. Due to the high concentration of sugars, liquid–liquid partition approach was used to eliminate the interfering effect from sugars, in which more metabolites could be detected, and phenethylalcohol glycosides were concentrated in the EtOAc fractions, while the iridoid glycoside were enriched in the n-BuOH part. In addition, various statistical methods were used to find the differential metabolites between the groups. The results obtained in this study suggest that more than one extraction and statistical method should be used for comprehensive metabolic profiling of medical plants, which contained various metabolites in different concentrations.

The purpose of this work was to investigate the anti-depressant effect of genipin and its mechanisms using (1)H-NMR spectroscopy and multivariate data analysis on a chronic unpredictable mild stress (CUMS) rat model. Rat serum and urine were analyzed by nuclear magnetic resonance (NMR)-based metabonomics after oral administration of either genipin or saline for 2 weeks. Significant differences in the metabolic profile of the CUMS-treated group and the control group were observed, which were consistent with the results of behavioral tests. Metabolic effects of CUMS included decreases in serum trimetlylamine oxide (TMAO) and β-hydroxybutyric acid (β-HB), and increases in lipid, lactate, alanine and N-acetyl-glycoproteins. In urine, decreases in creatinine and betaine were observed, while citrate, trimethylamine (TMA) and dimethylamine were increased. These changes suggest that depression may be associated with gut microbes, energy metabolism and glycometabolism. Genipin showed the best anti-depressive effects at a dose of 100 mg/kg in rats. These results indicate that metabonomic approaches could be powerful tools for the investigation of the biochemical changes in pathological conditions or drug treatment.