Development of Mass Spectrometric Strategies for Lipid Characterization and Mapping
Abstract
Lipids are a group of biomolecules which play crucial roles in many biological functions such as cell membrane construction, energy storage, and signal transduction. The aberrant metabolism of lipids is involved in the pathogenesis of many human diseases. The study of lipid structures and their spatial distributions in tissue samples is of great importance to describe lipid metabolism, discover disease biomarkers and understand disease mechanism.
Lipids display remarkable structural diversity, which is caused by many factors such as variable acyl chain lengths, unsaturation levels, substitutional positions and geometries. These factors allow the existence of various lipid isomers, which results in a significant challenge of accurate and rapid lipid structural identification. High resolution mass spectrometry (MS) coupled with tandem MS can readily differentiate chain length isomers and head group isomers. However, the structural differences between isomers can be very subtle, although they have dramatically different biochemical consequences, and cannot be directly identified through conventional MS.
To this end, the first project in the thesis describes the development of a mass spectrometric strategy that uses an alternating current to initiate i) electrospray (ES) for mass spectrometric analysis, and ii) electro-epoxidation of double bonds in unsaturated lipids when positive voltage is applied to the electrode. The formed epoxides can generate diagnostic fragments in tandem MS to locate double bonds and can be detected in both positive and negative ion modes. The feasibility of this method has been illustrated using a lipid mixture with both positively-charged and negatively-charged lipids. The results show the successful identification of over 30 different lipid isomers in the lipid extracts of a mouse brain.
A sufficient characterization of lipids in a tissue sample requires not only structural identification but also their spatial distribution. The classic bulk “bind and grind” sample preparation for MS analysis homogenizes the lipids in tissue samples, and loses the spatial information of lipids in tissues. In the second project, desorption electrospray ionization-mass spectrometry imaging (DESI-MSI) was used to spatially map lipids in the mouse brain from an alcoholism model. Assisted by partial least squares-discriminant analysis (PLS-DA), a potential lipid biomarker at m/z 841.5 (tentatively identified as triacylglycerol 51:3) has been identified in dorsal striatum that shows lower intensity in the alcoholism mouse group compared with the normal one.
Lipids with low abundance or low ionization efficiency may play an important role in biological process, while the lack of sensitive and specific methods limits the studies of them. The third project in the thesis develops a derivatization method for the detection of 2-arachidonoylglycerol (2-AG) using phenylboronic acid in nano-electrospray ionization-MS analysis. 2-AG is an important lipid in endocannabinoid system with low abundance and low ionization efficiency. The charge tag formed by reacting with phenylboronic acid can be added to cis-diol of 2-AG, which facilitates its mass analysis. Additionally, the specificity has been largely improved compared to protonation method, which is shown by the differentiation of 2-AG in tissue samples from its isomers in the background.
Citation
Fan, Licheng (2020). Development of Mass Spectrometric Strategies for Lipid Characterization and Mapping. Master's thesis, Texas A&M University. Available electronically from https : / /hdl .handle .net /1969 .1 /192267.