Metabolomics and metabolite profiles for phenotyping of individuals.Application of Lipidomics
Rob J. Vreeken and Thomas Hankemeier1
1Netherlands Metabolomics Centre, Leiden University, Leiden, the Netherlands
Abstract
Metabolomics involves the comprehensive quantitative and qualitative analysis of all small molecules in cells, tissues, and body fluids. Metabolic processes are at the core of physiology. Consequently, metabolomics is ideally positioned to distinguish between health and disease. Comparing metabolic profiles of patients and healthy controls, including repeated measurements within patients, is a powerful novel tool to unravel the biochemical pathways involved in multi-factorial disorders and to determine the efficacy of pharmacological and/or nutritional interventions.
Differences in genetic background and/or environmental exposure among individuals are expected to give rise to differences in measurable characteristics, or phenotypes. The metabolome reflects many of the system properties and is therefore an important part of the phenotype. Lipids are a subclass of the metabolome and play an important role in physiology, as they have a wide range of functions. The systems-level analysis of lipids is also called lipidomics. Due to the complexicity of the composition of lipids, analysis of a wide range of lipids classes is complicated. We developed a method for lipid profiling and applied it to the phenotyping of healthy twins. Different Mass Spectrometric approaches, viz. full-scan as well as Selected Reaction Monitoring analysis using Quadrupole and Ion-Trap systems were used in a quantitative manner ands were evaluated.
The lipid profiling method comprised a reversed-phase liquid chromatography (RPLC)-MS method using electrospray ionization. An Ascentis Express C8 column (fused core particles) and a gradient comprising water, acetonitrile and isopropanol were used. Extraction of the blood sample was obtained using water, methanol and dichloromethane. The method was validated by adding non-endogenous lipids to the blood sample at different concentration levels. For the sample preparation it was investigated whether a normal-phase based lipid class separation prior to the RPLC-MS was improving the overall performance. Methods for the acquisition of MSn spectra using an LTQ Orbitrap were developed to investigate to which extent differences in the structures of lipids can be revealed by MSn spectra.
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