Brain NMR Metabolomics Using Extracted Liquid Samples vs Focused Beam Microwave Fixed Solid Tissues

Abstract

Magic Angle Spinning (MAS) Nuclear Magnetic Resonance (NMR) Spectroscopy is known for its ability to produce sharper, more resolved peaks compared to traditional liquid chemical extraction NMR. The broad, poorly resolved peaks typically observed in liquid NMR average out in solid-state MAS, resulting in higher quality data, especially when applied to complex samples such as brain metabolomics. A key step in achieving this is microwave irradiation, which fixes the brain metabolome in place, preventing the degradation of sensitive organic structures caused by postmortem conditions. In this study, both liquid chemical extraction and solid-state MAS NMR were used to analyze brain samples and compare the results. Initially, the brains were subjected to liquid extraction and separated into lipid and water bilayers for analysis. The lipid layer produced high-quality Lorentzian peaks that were suitable for analysis, while the water layer generated poor-quality data, contaminated by noise. To evaluate the effect of irradiation and solid-state MAS, microwave fixation was applied using a repaired Muromachi microwave fixation machine, halting metabolic decay. Two eight-month-old mouse brains were tested: one as a control (without fixation) and one subjected to microwave fixation. The results demonstrated that microwave fixation did not destroy metabolites and, importantly, that MAS NMR provided higher quality data. The spectra from the fixed brain sample showed sharper peaks and narrower signals associated with brain metabolites, in contrast to liquid chemical extraction NMR. This comparison underscores the advantage of MAS NMR in providing more precise and reliable data for brain metabolomics analysis.

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