Liquid chromatography-mass metabolite profiling

The system relies upon preliminary fractionation of the microbial crude extract by dualmode countercurrent chromatography coupled with photodiode array detection (PDA). The ultraviolet-visible (UV-Vis) spectra and liquid chromatography-mass spectrometry (LC-MS) of biologically active peaks are used for identification. Confirmation of compound identity is accomplished by nuclear magnetic resonance (NMR). Use of an integrated system countercurrent chromatography (CCC) separation, PDA detection, and LC-MS rapidly provided profiles and structural information extremely useful for metabolite identification (dereplication, Figure 14.1). 

Rochat B et al (2008) Imatinib metabolite profiling in parallel to imatinib quantification in plasma of treated patients using liquid chromatography-mass spectrometry. J Mass Spectrom... 

Kerns, E.H. Rourick, R.A. Volk, K.J. Lee, M.S. Buspirone metabolite structure profile using a standard liquid chromatography-mass spectrometric protocol. J. Chromatogr. B 1997, 698, 133-145. 

Overall, it is well understood that one method cannot cover the whole metabolome, so the current state-of-the-art demonstrates the need for the application of a number of analytical techniques such as reversed-phase liquid chromatography-mass spectrometry ((RP)LC-MS) untargeted as a general profiling tool, another RPLC-MS method to profile the lipids (on the C30 or C18 column), GC-MS to profile the content of volatiles, and one or two LC-MS or CE-MS methods (targeted and untargeted) to profile the polar metabolites (see Section 5.1). 

Gu, M. Lim, H.K. An Intelligent Data Acquisition System for Simultaneous Screening of Microsomal Stability and Metabolite Profiling by Liquid Chromatography/Mass Spectrometry, 7. Mass Spectrom. 36(9), 1053-1061 (2001). 

Gu M, Lim HK. An intelligent data acquisition system for simultaneous screening of microsomal stability and metabolite profiling by liquid chromatography/mass spectrometry. J Mass Spectrom 2001 36 1053-1061. 

Hong, Y-J, Mitchell E. 2005. Metabolic profiling of flavonol metabolites in human urine by liquid chromatography and tandem mass spectrometry. J Agric Food Chem 52 6794-6801. 

Superior sensitivity, efficiency, and specificity have made high-performance liquid chromatography coupled with tandem mass spectrometry (HPLC-MS/MS), the predominant analytical technique for characterization and quantitative analysis of metabolites (Kostiainen et al., 2003 Ma et al., 2006 Prakash et al., 2007). Ion trap, triple-quadrupole, and quadmpole time-of-flight (Q-TOF) mass spectrometers are routinely used to profile and characterize metabolites in plasma and excreta (Ma et al., 2006). The combination of scan types and features available on mass spectrometers of different design (product ion, MS", neutral loss, precursor ion scans, accurate mass measurements) allows identification and characterization of putative and unexpected metabolites with or without little prior knowledge of biotransformation pathways of a given dmg molecule. 

Link M, Hakala KS, Wsol V, et al. Metabolite profile of sibutramine in human urine a liquid chromatography-electrospray ionization mass spectrometric study. J Mass Spectrom 2006 41 1171-1178. 

Fig. 5 Discovery metabolite profiling of brain tissue, where mass ion intensity ratios (FAAH / / FAAH+/+) of metabolites are presented on three-dimensional surface plots. Global view of the relative levels of metabolites in FAAH / and FAAH+/+ brains, plotted over a mass range of 200-1,200 m/z and liquid chromatography retention times of 0-105 min (plot shown for negative ionization mode). FAAH / brains possessed highly elevated levels of A-acyl ethanolamines (NAEs) (lipid group 4) and an unknown class of lipids (group 5), identified as A-acyl taurines (NATs). Other lipids, e.g., free fatty acids (group 1), phospholipids (group 2), and ceramides (group 3) were unaltered in these samples...

Cordell, R.L. et al. Quantitative profiling of nucleotides and related phosphate-containing metabolites in cultured mammalian cells by liquid chromatography tandem electrospray mass spectrometry. J. Chromatogr. B. 2008, 871,115-124. 

Feistner, G.J. Profiling of Bacterial Metabolites by Liquid Chromatography-Electrospray Mass Spectrometry A Perspective, Am. Lab. 32L-32Q (Sept. 1994). 

Gao, H. et al., Method for rapid metabolite profiling of drug candidates in fresh hepa-tocytes using liquid chromatography coupled with a hybrid quadrupole linear ion trap, Rapid Commun. Mass Spectrom., 21, 3683, 2007. 

Metabolomics is the newest of the functional genomic technologies and also the least well defined in terms standard experimental approaches. A number of different spectroscopic methods can be used to obtain metabolite profiles from tissue, cellular, or extracellular fluid samples. These include NMR, mass spectroscopy, liquid chromatography, and optical spectroscopic techniques [26]. Of these, NMR approaches, such as magic angle spinning NMR, which can be used with intact cells and tissues, and mass spectrometric approaches appear to be the most promising. 

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