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Within non-communicable diseases, chronic inflammatory conditions represent one of the biggest challenges for modern medicine. Traditional Chinese Medicine (TCM) has been practiced over centuries and has accumulated tremendous empirical knowledge on the treatment of such diseases. Huangqi Jianzhong Tang (HQJZT) is a famous TCM herbal formula composed of Radix Astragali, Ramulus Cinnamomi, Radix et Rhizoma Glycyrrhizae Praeparata cum Melle, Radix Paeoniae Alba, Rhizoma Zingiberis Recens, Fructus Jujubae and Saccharum Granorum (maltose), which has been used for the treatment of various chronic inflammatory gastrointestinal diseases. However, there is insufficient knowledge about its active constituents and the mechanisms responsible for its effects. The present study aimed at identifying constituents contributing to the bioactivity of HQJZT by combining in vitro cytokine production assays and LC-MS metabolomics techniques. From the HQJZT decoction as well as from its single herbal components, extracts of different polarities were prepared. Phytochemical composition of the extracts was analyzed by means of UPLC-QTOF-MS/MS. The inhibitory effects of the extracts on TNF-α, IL-1β and IFN-γ production were studied in U937 cells. Phytochemical and pharmacological bioactivity data were correlated by orthogonal projection to latent structures discriminant analysis (OPLS-DA) in order to identify those HQJZT constituents which may be relevant for the observed pharmacological activities. The investigations resulted in the identification of 16 HQJZT constituents, which are likely to contribute to the activities observed in U937 cells. Seven of them, namely calycosin, formononetin, astragaloside I, liquiritigenin, 18β-glycyrrhetinic acid, paeoniflorin and albiflorin were unambiguously identified. The predicted results were verified by testing these compounds in the same pharmacological assays as for the extracts. In conclusion, the anti-inflammatory activity of HQJZT could be substantiated by in vitro pharmacological screening, and the predicted activities of the OPLS-DA hits could be partially verified. Moreover, the benefits and limitations of MVDA for prediction pharmacologically active compounds contributing to the activity of a TCM mixture could be detected., Competing Interests: The authors declare no conflict of interest.

Metabolomics as a global analysis of a large number of cellular metabolites relies heavily on the new developments in separation science and technology. None of the existing analytical techniques can simultaneously separate and measure all the cellular metabolites due to complexity of cellular metabolome and, therefore, a combination of analytical techniques must be used. Currently NMR, GC-MS and LC-MS are most often used in metabolomics. Novel separation methods such as supercritical fluid chromatography (SFC), which can increase metabolome coverage while decreasing cost and analysis time, can provide alternative to other analytical techniques. As a result of major improvements in instrumentation and development of a new diverse column chemistries SFC-MS is increasingly used in a variety of biomedical applications and is becoming an attractive compliment to other major analytical platforms in metabolomics. Despite its potential and advantages, SFC-MS application in metabolomics is limited. Here we provide a brief overview of the latest developments of SFC-MS for metabolomics applications., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Prostate cancer (PCa) is the most prevalent cancer amongst men and the second most common cause of cancer related-deaths in the USA. Prostate cancer is a heterogeneous disease ranging from indolent asymptomatic cases to very aggressive life threatening forms. The goal of this study was to identify differentially expressed metabolites and lipids in prostate cells with different tumorigenic phenotypes. We have used mass spectrometry metabolomic profiling, lipidomic profiling, bioinformatic and statistical methods to identify, quantify and characterize differentially regulated molecules in five prostate derived cell lines. We have identified potentially interesting species of different lipid subclasses including phosphatidylcholines (PCs), phosphatidylethanolamines (PEs), glycerophosphoinositols (PIs) and other metabolites that are significantly upregulated in prostate cancer cells derived from distant metastatic sites. Transcriptomic and biochemical analysis of key enzymes that are involved in lipid metabolism demonstrate the significant upregulation of choline kinase alpha in the metastatic cells compared to the non-malignant and non-metastatic cells. This suggests that different de novo lipogenesis and other specific signal transduction pathways are activated in aggressive metastatic cells as compared to normal and non-metastatic cells.

Ultra high resolution SFC-MS (on sub-2μm particles) coupled to mass spectrometry has been evaluated for the metabolic profiling of rat and dog bile. The selectivity of the SFC separation differed from that seen in previous reversed-phase UPLC-MS studies on bile, with the order of elution for analytes such as e.g., the bile acids showing many differences. The chromatography system showed excellent stability, reproducibility and robustness with relative standard deviation of less than 1% for retention time obtained over the course of the analysis. SFC showed excellent chromatographic performance with chromatographic peak widths in the order of 3s at the base of the peak. The use of supercritical fluid carbon dioxide as a mobile phase solvent also reduced the overall consumption of organic solvent by a factor of 3 and also reduced the overall analysis time by a factor of 30% compared to reversed-phase gradient LC. SFC-MS appear complementary to RPLC for the metabolic profiling of complex samples such as bile., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Molecular networking has become a key method to visualize and annotate the chemical space in non-targeted mass spectrometry data. We present feature-based molecular networking (FBMN) as an analysis method in the Global Natural Products Social Molecular Networking (GNPS) infrastructure that builds on chromatographic feature detection and alignment tools. FBMN enables quantitative analysis and resolution of isomers, including from ion mobility spectrometry.

Vangueria agrestis is a shrub indigenous to tropical Africa, belonging to family Rubiaceae and is traditionally used as a decoction for treatment of fever, pain, and malaria. This study was undertaken to investigate the chemical constituents based on precursor exact mass and fragment ion information. The chemical profiling and structural characteristics of chemical constituents from methanolic extracts of dried aerial parts and roots of V. agrestis and dietary supplements were analyzed using ultra‐high‐performance liquid chromatography/ion mobility quadrupole time‐of‐flight mass spectrometry coupled with UNIFI platform and multivariate analysis in both negative and positive ion modes. A non‐targeted ultra‐high‐performance liquid chromatography‐mass spectrometry analysis was carried out to profile the chemical constituents of crude extracts of V. agrestis, and 73 compounds, including reference compounds, were identified. The fragments of flavonoids, monoterpene, and triterpene glycosides revealed the characteristic cleavage of glycosidic linkages, and the fragmentation pattern provided the identity of the sugars. This analytical method provides a quick method for quality assessment of dietary supplements. Finally, a chemometrics approach with multivariate statistical tools was used to visualize the differences between root and aerial parts of plant samples and to find the potential chemical markers that differentiate among these parts of V. agrestis samples and dietary supplements. [ABSTRACT FROM AUTHOR]

Background Controversy exists concerning the beneficial or harmful effects of the presence of ectopic calcification in the coronary arteries. Additionally, further elucidation of the exact pathophysiological mechanism is needed. In this study, we sought to identify metabolic markers of vascular calcification that could assist in understanding the disease, monitoring its progress and generating hypotheses describing its pathophysiology. Methods Untargeted lipid profiling and complementary modeling strategies were employed to compare serum samples from patients with different levels of calcific coronary artery disease (CCAD) based on their calcium score (CS). Subsequently, patients were divided into three groups: no calcification (NC; CS = 0; n = 26), mild calcification (MC; CS:1–250; n = 27) and severe (SC; CS > 250; n = 17). Results Phosphatidylcholine levels were found to be significantly altered in the disease states (p = 0.001–0.04). Specifically, 18-carbon fatty acyl chain (FAC) phosphatidylcholines were detected in lower levels in the SC group, while 20:4 FAC lipid species were detected in higher concentrations. A statistical trend was observed with phosphatidylcholine lipids in the MC group, showing the same tendency as with the SC group. We also observed several sphingomyelin signals present at lower intensities in SC when compared with NC or MC groups (p = 0.000001–0.01). Conclusions This is the first lipid profiling study reported in CCAD. Our data demonstrate dysregulations of phosphatidylcholine lipid species, which suggest perturbations in fatty acid elongation/desaturation. The altered levels of the 18-carbon and 20:4 FAC lipids may be indicative of disturbed inflammation homeostasis. The marked sphingomyelin dysregulation in SC is consistent with profound apoptosis as a potential mechanism of CCAD. [ABSTRACT FROM AUTHOR]

Metabolic profiling studies aim to achieve broad metabolome coverage in specific biological samples. However, wide metabolome coverage has proven difficult to achieve, mostly because of the diverse physicochemical properties of small molecules, obligating analysts to seek multiplatform and multimethod approaches. Challenges are even greater when it comes to applications to tissue samples, where tissue lysis and metabolite extraction can induce significant systematic variation in composition. We have developed a pipeline for obtaining the aqueous and organic compounds from diseased arterial tissue using two consecutive extractions, followed by a different untargeted UPLC-MS analysis method for each extract. Methods were rationally chosen and optimized to address the different physicochemical properties of each extract: hydrophilic interaction liquid chromatography (HILIC) for the aqueous extract and reversed-phase chromatography for the organic. This pipeline can be generic for tissue analysis as demonstrated by applications to different tissue types. The experimental setup and fast turnaround time of the two methods contributed toward obtaining highly reproducible features with exceptional chromatographic performance (CV % < 0.5%), making this pipeline suitable for metabolic profiling applications. We structurally assigned 226 metabolites from a range of chemical classes (e.g., carnitines, α-amino acids, purines, pyrimidines, phospholipids, sphingolipids, free fatty acids, and glycerolipids) which were mapped to their corresponding pathways, biological functions and known disease mechanisms. The combination of the two untargeted UPLC-MS methods showed high metabolite complementarity. We demonstrate the application of this pipeline to cardiovascular disease, where we show that the analyzed diseased groups (n = 120) of arterial tissue could be distinguished based on their metabolic profiles. [ABSTRACT FROM AUTHOR]

• A generic mobile phase has been developed for metabolomic and lipidomic separations • Isopropyl alcohol was replaced by acetonitrile as the organic modifier for lipidomics • Lipidomics on a phenyl-hexyl column increased peak capacity (58 %) and detection (61 %) • RPLC on a C8 stationary phase was used for moderately polar-nonpolar analytes • Polar metabolites were separated by HILIC on an amide stationary phase The untargeted global profiling of endogenous metabolites and lipids has the potential to increase knowledge and understanding in many areas of biology. LC-MS/MS is a key technology for such analyses however, several different LC methodologies, using different mobile phase compositions, are required to cover the diversity in polarity and analyte structure encountered in biological samples. Most notably many lipid screening methods make use of isopropanol (IPA) as a major component of mobile phases employed for comprehensive lipidomic profiling. In order to increase laboratory efficiency, and minimize opportunities for errors, a suite of methods, based on a single acetonitrile (ACN)-aqueous buffer mobile phase combination, has been developed. This mobile phase can be used for hydrophobic interaction liquid chromatography on an amide stationary phase (for polar analytes), reversed-phase (RP) LC analysis on a C8 stationary phase (for moderately polar-non-polar compounds) and RPLC using a CSH phenyl-hexyl bonded column (for lipids). All of these sub 10 minute separations had good throughput and reproducibility with CV's of analyte response <25 % whilst eliminating the need for complex mobile phase preparation and the use of IPA as an organic modifier for lipidomics. Advantages of removing IPA and replacing it with the ACN-based method were a 58 % increase in peak capacity for lipids, with improved resolution for the di- and triglycerides and cholesterol esters compared to current methods. Compared to the IPA-containing solvent system the ACN-based mobile phase also resulted in a 61 % increase in lipid feature detection. The utility of this "universal" mobile phase approach was demonstrated by its application to a rat toxicology study investigating the consequences of methapyrilene administration through on the endogenous metabolite profiles of plasma and urine. Methapyrilene and its metabolites were also profiled in these samples. [ABSTRACT FROM AUTHOR]

Properly implemented, metabolic and lipidomic profiling can provide a deeper understanding of mammalian, plant and bacterial biology. These omics-tools have developed and matured over the last 40-years and are now being deployed to provide valuable information in epidemiological studies, drug toxicology and pharmacology, disease biology and progression and patient stratification. LC/MS has become the technology of choice for both metabolic and lipid profiling, due to its speed, sensitivity and structural elucidation capabilities. In the preceding two decades there have been many technological and methodological advances in LC/MS that have facilitated the evolution of the technology into a rugged, reliable, and easily deployed tool. These advances include, but are not limited to, improvements in chromatography (phases, columns, and delivery system), instruments for mass spectrometry, optimization of sample preparation, the introduction of ion mobility, data analysis tools, metabolite databases, harmonized protocols, and the more widespread use of quality control methods and reference standards/matrices. Here, recent developments and advances in high throughput liquid chromatography/high resolution mass spectrometry for metabolic phenotyping are described. These advances which may provide improved feature detection, increased laboratory efficiency and data quality, as well as "biomarker" identification, are discussed in relation to their potential application to the analysis of large clinical studies, or biobank collections. • High throughput (HT) methods for metabolic phenotyping are needed for large cohorts. • Sample analysis using direct injection or LC/MS can provide HT sample analysis. • UHPLC/MS may provide partial solutions to the HT analysis of large study collections. • The inclusion of ion mobility in HT/UHPLC/MS analysis brings substantial benefits. • Advances in HT untargeted direct injection and LC/MS-based metabolomics are discussed. [ABSTRACT FROM AUTHOR]