These are scientific publications for Targeted Metabolomics using an Agilent GC/MS system. Please note that the links below will take you to outside of Agilent’s website to PubMed.
1. 1H NMR and GC/MS metabolomics of earthworm responses to sub-lethal DDT and endosulfan exposure
McKelvie JR , Yuk J, Xu Y, Simpson AJ and Simpson MJ
Metabolomics 2009 5:84–94
The metabolic response of the earthworm Eisenia fetida to two pesticides, dichlorodiphenyltrichloroethane (DDT) and endosulfan, was characterized in contact tests using proton nuclear magnetic resonance (1H NMR) and principal component analysis (PCA). PCA loading plots suggested that maltose, leucine and alanine were important metabolites contributing to the differences in dosed and control earthworms for both compounds at doses of 0.5, 1.0 and 2.0 μg/cm2. Gas chromatography/mass spectrometry (GC/MS) was used to quantify the metabolites identified in E. fetida and determine if the changes in maltose, leucine and alanine following exposure to DDT and endosulfan (at 0.5 and 1.0 μg/cm2) were reproducible and greater than the natural variability. Quantification by GC/MS suggested that maltose was not a reliable biomarker since it both increased and decreased in earthworms exposed to DDT and increased by just 3% with exposure to endosulfan. Leucine was not stable with the GC/MS derivitization method used in this study and could not be confirmed as a reliable biomarker. However, alanine consistently increased for both DDT and endosulfan exposed E. fetida. Alanine showed considerable variability in control earthworms (±41.6%), yet the variability in alanine to glycine ratios was just ±10.5%. Increases in the alanine to glycine ratio were statistically significant at the P = 0.05 level for the 1.0 μg/cm2 DDT dose and both the 0.5 and 1.0 μg/cm2 endosulfan doses, suggesting that deviations from the normal homeostatic ratio of 1.5 for alanine to glycine is a potential biomarker of DDT and endosulfan exposure warranting further study.
2. Microbial metabolomics with gas chromatography/mass spectrometry.
Koek MM, Muilwijk B, van der Werf MJ, Hankemeier T.
Anal Chem. 2006 Feb 15;78(4):1272-81.
An analytical method was set up suitable for the analysis of microbial metabolomes, consisting of an oximation and silylation derivatization reaction and subsequent analysis by gas chromatography coupled to mass spectrometry. Microbial matrixes contain many compounds that potentially interfere with either the derivatization procedure or analysis, such as high concentrations of salts, complex media or buffer components, or extremely high substrate and product concentrations. The developed method was extensively validated using different microorganisms, i.e., Bacillus subtilis, Propionibacterium freudenreichii, and Escherichia coli. Many metabolite classes could be analyzed with the method: alcohols, aldehydes, amino acids, amines, fatty acids, (phospho-) organic acids, sugars, sugar acids, (acyl-) sugar amines, sugar phosphate, purines, pyrimidines, and aromatic compounds. The derivatization reaction proved to be efficient (>50% transferred to derivatized form) and repeatable (relative standard deviations <10%). Linearity for most metabolites was satisfactory with regression coefficients better than 0.996. Quantification limits were 40-500 pg on-column or 0.1-0.7 mmol/g of microbial cells (dry weight). Generally, intrabatch precision (repeatability) and interbatch precision (reproducibility) for the analysis of metabolites in cell extracts was better than 10 and 15%, respectively. Notwithstanding the nontargeted character of the method and complex microbial matrix, analytical performance for most metabolites fit the requirements for target analysis in bioanalysis. The suitability of the method was demonstrated by analysis of E. coli samples harvested at different growth phases.
3. A metabolomic study of substantial equivalence of field-grown genetically modified wheat.
Baker JM, Hawkins ND, Ward JL, Lovegrove A, Napier JA, Shewry PR, Beale MH.
Plant Biotechnol J. 2006 Jul;4(4):381-92.
The 'substantial equivalence' of three transgenic wheats expressing additional high-molecular-weight subunit genes and the corresponding parental lines (two lines plus a null transformant) was examined using metabolite profiling of samples grown in replicate field trials on two UK sites (Rothamsted, Hertfordshire and Long Ashton, near Bristol) for 3 years. Multivariate comparison of the proton nuclear magnetic resonance spectra of polar metabolites extracted with deuterated methanol-water showed a stronger influence of site and year than of genotype. Nevertheless, some separation between the transgenic and parental lines was observed, notably between the transgenic line B73-6-1 (which had the highest level of transgene expression) and its parental line L88-6. Comparison of the spectra showed that this separation resulted from increased levels of maltose and/or sucrose in this transgenic line, and that differences in free amino acids were also apparent. More detailed studies of the amino acid composition of material grown in 2000 were carried out using gas chromatography-mass spectrometry. The most noticeable difference was that the samples grown at Rothamsted consistently contained larger amounts of acidic amino acids (glutamic, aspartic) and their amides (glutamine, asparagine). In addition, the related lines, L88-6 and B73-6-1, both contained larger amounts of proline and gamma-aminobutyric acid when grown at Long Ashton than at Rothamsted. The results clearly demonstrate that the environment affects the metabolome and that any differences between the control and transgenic lines are generally within the same range as the differences observed between the control lines grown on different sites and in different years.