Article

Monitoring Dynamic Changes in Lymph Metabolome of Fasting and Fed Rats by Electrospray Ionization-Ion Mobility Mass Spectrometry (ESI-IMMS)

Washington State University, Pullman, Washington 99164, USA.
Analytical Chemistry (Impact Factor: 5.83). 10/2009; 81(19):7944-53. DOI: 10.1021/ac901030k
Source: PubMed

ABSTRACT Ambient pressure ion mobility time-of-flight mass spectrometry (IMMS) has recently emerged as a rapid and efficient analytical technique for applications to metabolomics. An important application of metabolomics is to monitor metabolome shifts caused by stress due to toxin exposure, nutritional changes, or disease. The research presented in this paper uses IMMS to monitor metabolic changes in rat lymph fluid caused by dietary stresses over time. Extracts of metabolites found in the lymph fluid collected from dietary stressed rats were subjected to analysis by electrospray (ESI) IMMS operated both in positive and negative ion detection mode. Metabolites detected were tentatively identified based on their mass to charge ratio (m/z). In one sample, 1180 reproducible tentative metabolite ions were detected in negative mode and 1900 reproducible tentative metabolite ions detected in positive mode. Only biologically reproducible ions, defined as metabolite ions that were measured in different rats under the same treatment, were analyzed to reduce the complexity of the data. A metabolite peak list including m/z, mobility, and intensity generated for each metabolome was used to perform principle component analysis (PCA). Dynamic changes in metabolomes were investigated using principle components PC1 and PC2 that described 62% of the variation of the system in positive mode and 81% of the variation of the system in negative mode. Analysis of variance (ANOVA) was performed for PC1 and PC2 and means were statistically evaluated. Profiles of intensities were compared for tentative metabolite ions detected at different times before and after the rats were fed to identify the metabolites that were changing the most. Mobility-mass correlation curves (MMCC) were investigated for the different classes of compounds.

1 Bookmark
 · 
107 Views
  • [Show abstract] [Hide abstract]
    ABSTRACT: Breast cancer is well-known to broadly impact cellular metabolism and aberrant metabolism in breast cancer tumors has been widely studied by both targeted and untargeted analyses to characterize the affected metabolic pathways. In this work, we utilize ultra-performance liquid chromatography (UPLC) in tandem with ion mobility-mass spectrometry (IM-MS), which provides chromatographic, structural, and mass information, to characterize the aberrant metabolism associated with breast diseases such as cancer. In a double-blind analysis of matched control (n=3) and disease tissues (n=3), tissues were homogenized, polar metabolites were extracted, and the extracts were characterized by UPLC-IM-MS/MS. Principle component analysis revealed a strong separation between disease tissues, with one diseased tissue clustering with the control tissues along PC1 and two others separated along PC2. Using post-ion mobility MS/MS spectra acquired by data-independent acquisition, the features giving rise to the observed grouping were determined to be biomolecules associated with aggressive breast cancer tumors, including glutathione, oxidized glutathione, thymosins β4 and β10, and choline-containing species. Pathology reports revealed the outlier of the disease tissues to be a benign fibroadenoma, whereas the other disease tissues represented highly metabolic benign and aggressive tumors. This IM-MS-based workflow bridges the transition from untargeted metabolomic profiling to tentative identifications of key descriptive molecular features using data acquired in one analysis, with additional experiments performed only for validation. The ability to resolve cancerous and non-cancerous tissues at the biomolecular level demonstrates UPLC-IM-MS/MS as a robust and sensitive platform for metabolomic profiling of tissues.
    Molecular BioSystems 09/2014; 10(11). DOI:10.1039/C4MB00250D · 3.35 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Metabolomics is a rapidly-evolving analytical approach in life and health sciences. The structural elucidation of the metabolites of interest remains a major analytical challenge in the metabolomics workflow. Here, we investigate the use of ion mobility as a tool to aid metabolite identification. Ion mobility allows for the measurament of the rotationally-averaged collision cross-section (CCS), which gives information about the ionic shape of a molecule in the gas phase. We measured the CCSs of 125 common metabolites using travelling-wave ion mobility-mass spectrometry (TW-IM-MS). CCS measuraments were highly reproducible on instruments located in three independent laboratories (RSD <5% for 99%). We also determined the reproducibility of CCS measurements in various biological matrices including urine, plasma, platelets, and red blood cells, using UPLC coupled with TW-IM-MS. The mean RSD was <2% for 97% of the CCS values, compared to 80% of retention times. Finally, as proof of concept, we used UPLC/TW-IM-MS to compare the cellular metabolome of epithelial and mesenchymal cells, an in vitro model used to study cancer development. Experimentally-determined and computationally-derived CCS values were used as orthogonal analytical parameters in combination with retention time and accurate mass information, to confirm the identity of key metabolites potentially involved in cancer. Thus, our results indicate that adding CCS data to searchable databases and to routine metabolomics workflows will increase the identification confidence compared to traditional analytical approaches.
    Analytical Chemistry 03/2014; 86(8). DOI:10.1021/ac500405x · 5.83 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Despite recent advances in analytical and computational chemistry, lipid identification remains a significant challenge in lipidomics. Ion-mobility spectrometry provides an accurate measure of the molecules' rotationally averaged collision cross-section (CCS) in the gas phase and is thus related to ionic shape. Here we investigate the use of CCS as a highly specific molecular descriptor for identifying lipids in biological samples. Using travelling-wave, ion-mobility mass spectrometry (MS), we measured the CCS values of over 200 lipids within multiple chemical classes. CCS values derived from ion mobility were not affected by instrument settings or chromatographic conditions, and they were highly reproducible on instruments located in independent laboratories (inter-laboratory RSD<3% for 98% of molecules). CCS values were used as additional molecular descriptors to identify brain lipids using a variety of traditional lipidomic approaches. The addition of CCS improved the reproducibility of analysis in a liquid chromatography-MS workflow, and maximized the separation of isobaric species and the signal-to-noise ratio in direct-MS analyses (e.g.,"shotgun" lipidomics and MS imaging). These results indicate that adding CCS to databases and lipidomics workflows increases the specificity and selectivity of analysis, thus improving the confidence in lipid identification compared to traditional analytical approaches. The CCS/accurate-mass database described here is made publicly available.
    Analytical Chemistry 12/2014; DOI:10.1021/ac503715v · 5.83 Impact Factor

Full-text (2 Sources)

Download
55 Downloads
Available from
May 27, 2014