LC-MS metabonomics

Warrack, B., Hnatyshyn, S., Zhang, H., and Sanders, M. (2005). Strategies for the use of mass spectrometry for LC/MS metabonomics profiling How much resolution is needed and in what dimension In Proceedings of the 53rd ASMS Conference on Mass Spectrometry and Allied Topics, San Antonio, TX. 

In another research article, Wang et al. [20] successfully applied LC-MS metabonomic techniques to the metabolite profiling of plasma phospholipids in type 2 diabetes mellitus (DM2) patients. Diabetes mellitus is associated with a metabolic disorder of lipid or fatty acid in phospholipids. The authors were not only able to differentiate between the samples from the DM2 patients and the healthy subjects but also identified a number of phospholipid molecular species that could be used as potential biomarkers for a differentiation of DM2 patients from the healthy individuals and perhaps even an early detection of DM2. 

In a GC-MS or LC-MS metabonomics experiment each file represents a single biological sample. The basic goal of data processing is to transform raw data files to a standard and uniform format so that statistical analyses can be carried out. A variety of instrument vendors utilize different proprietary data formats. Thus, the first step in data processing requires a conversion of such raw proprietary data into common raw data format such as ASCII text or binary netCDF. More recently, a universal mzXML format has become more popular. Many vendor instrument software packages contain scripts to allow for such a file conversion [76],... 

Shipkova, P.A. et al., Sample preparation approaches and data analysis for non-targeted LC/MS metabonomic profiling of plasma and urine. Proceedings of the 55th ASMS Conference, Indianapolis, IN, 2007. 

Hnatyshyn S, Sanders M, Shipkova P, Luk E, Warrack B, Reily M, Poster 375 Automated Mass Spectra Interpretation Approach to Data Reduction for LC-MS Metabonomics Analysis, Poster presented at the American Society for Mass Spectrometry Conference, June 5, Denver, CO, 2008... 

Description of LC-MS, including free articles. This site has subdivisions such as proteomics, metabonomics, 2D-electrophoresis, and sample preparation. 

LC-MS has only very recently started to be used extensively for metabonomics analyses and the amount of recently published reports in this area is extensive [110— 116], In many ways LC-MS is an ideal technique for metabolic profiling as many biological fluids such as urine and plasma upon PP can be directly injected onto an... 

Luc et al. [86] evaluated a number of different columns and gradients for LC-MS-based nontargeted metabonomics analyses of rat plasma and urine samples. The authors compared equivalent UPLC and HPLC columns of the same size and with the same packing material. The only difference was in the particle size, which was 1.7 and 3.5 pm for UPLC and HPLC columns, respectively. Luc et al. concluded that although the UPLC peaks had improved peak widths (FWHM) and overall intensity,... 

Luc, C.E. et al., Evaluation of columns and gradients for LC/MS-based non-targeted metabonomics. Proceedings of the 56th ASMS Conference, Denver, CO, 2008. 

Waybright, T.J. et al., LC-MS in metabonomics Optimization of experimental conditions for the analysis of metabolites in human urine, J. Liquid Chromatogr. Related Technol., 29(17), 2475, 2006. 

Theodoridis, G., Gika, H.G., and Wilson, I.D., LC-MS based methodology for global metabolite profiling in metabonomics/metabolomics, Trends Anal. Chem., 27(3), 251, 2008. 

Wagner, S. et al., Metabonomics and biomarker discovery LC-MS metabolic profiling and constant neutral loss scanning combined with multivariate data analysis for mercap-turic acid analysis, Anal. Chem., 78(4), 1296, 2006. 

Sun J, Von Tungeln LS, Hines W, Beger RD. Identification of metabolite profiles of the catechol-O-methyl transferase inhibitor tolcapone in rat urine using LC/MS-based metabonomics analysis. J Chromatogr B Anal Technol Biomed Life Sci 2009 877 2557-2565. 

Wagner S, Scholz K, Sieber M, Kellert M, Voelkel W. Tools in metabonomics An integrated validation approach for LC-MS metabolic profiling of mercapturic acids in human urine. Anal Chem 2007 79 2918-2926. 

This approach differs from those discussed above in that measured analytes have been selected a priori, usually to address certain specific biological questions within a study. Furthermore, since the exact analytes are known, it is possible and desirable to measure their absolute concentrations with appropriate use of internal standards and calibration curves. The lines between targeted metabolomics and traditional LC—MS-based assay development are arguably ill defined. Certainly, no one would equate an MS assay developed to measure alanine with targeted metabonomics. However, a multiplex assay to measure 13 or 14 organic acids in a urine sample probably qualifies. Often, targeted approaches can be developed for specific compoimd classes such as... 

Designing a successful metabonomics experiment is truly a multidisciplinary exercise, and one must consider details of the in-life portion, sample collection, sample preparation, analytical data generation, data processing, analysis, and interpretation. The in-life portion involves study subjects selection, acclimatization, dosing or treatment, and sample collection and has been discussed in detail elsewhere (Robertson et al., 2002). The following sections will focus on the remaining steps in this process as it pertains to LC—MS. 

An LC—MS chromatogram is essentially a three-dimensional data set with separate chromatographic, mass-to-charge (m/z) and ion count (intensity) dimensions. The combination of chromatography and mass spectral approaches can take on many incarnations and are dependent on the nature of the metabonomics investigation. Because of the complexity of most metabonomics samples, the final solution is often a compromise between good science and expediency since the ideal analytical conditions for 1000+ metabolome components is an um-ealistic goal. In this section, we will discuss each of these elements separately with an emphasis on biomedical metabonomic applications. 

Lu X, et al. LC-MS-based metabonomics analysis. J Chromatogr B Anal Technol Biomed Life Sci 2008b 866 64—76. 

NMR plays a key role in metabonomics as the science is typically studied by measuring the metabolite pools in various biofluids or tissues using NMR spectroscopy, although other technologies such as MS or LC-MS can be employed. 

Plumb, R. and Wilson, 1. 2004. High throughput and high sensitivity LC/MS-OA-TOF and UPLC/TOF-MS for the identification of biomarkers of toxicity and disease using a metabonomics approach. Afof/ Pap. Am. Chem. Soc. 228 189. 

In order to enable holistic metabolic assessment of living organisms, one requires methods that can acquire metabolic profiles in a rapid, reproducible and comprehensive manner, without bias towards compound classes. NMR meets these requirements effectively, and can assess metabolites down to the tens of pM level . This may seem high when compared to more targeted methods such as LC(MS) and GC(MS), but NMR has the imique advantage that hundreds of metabolites can be assessed in a reproducible and quantitative manner in a single shot. Metabolite identification is a notorious bottleneck in metabonomics. In anticipation of this challenge, we built a pH dependent AMIX H NMR spectral database for gut polyphenols fermentation products for which literature provided clues on their abundance in faeces and urine. 

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