| Direct analysis of tissue from both the central and peripheral nervous systems of control rats and rats administered the potential neurotoxin dichloroacetate (DCA) was investigated using an intermediate-pressure matrix-assisted laser desorption/ionization (IP-MALDI) source coupled to a linear ion trap (LIT) mass spectrometer. The MALDI matrix, 2,5-dihydroxybenzoic acid, was applied to the tissue using a novel automated inkjet printer system. The MS/MS capabilities of the LIT allowed identification of lipids desorbed directly from tissue. In some cases, a marked decrease is observed in the intensity of lipid ions in spinal cord and sciatic nerve from rats exposed to DCA. The results also demonstrate the rapid, sensitive and semi-quantitative capabilities of this method.;Further quantitative capabilities of MALDI were examined through the use of an internal standard. A non-endogenous phospholipid, PC(10:0,10:0), was chosen because it does not interfere with ions typically found in the lipid mass range (m/z 700--900) as well as having similar ionization and extraction efficiency to the endogenous lipids of interest. Three different application methods of the internal standard were evaluated; results show that, for quantitative purposes, application on top of dry tissue is the most desirable method. Utilizing an internal standard allows the calculation of the relative concentration of endogenous lipids with a precision of better than 5% standard error.;A hybrid linear ion trap/orbitrap mass spectrometer was used to perform MS/MS experiments and high resolution mass analysis of lipids desorbed from nerve tissue. A dramatic improvement in mass spectral resolution and a decrease in background are observed in the spectra collected from the orbitrap, which allows generation of more accurate mass spectrometric images of the distribution of lipids within nerve tissue. Employment of both mass analyzers provides a rapid and reliable means of compound identification based on MS/MS fragmentation and HRMS accurate mass.;Finally, three different ionization methods were approached for the analysis of lipids in nerve tissue on the hybrid linear ion trap/orbitrap mass spectrometer. Extracted lipids were analyzed by both direct infusion nanospray and MALDI, and direct interrogation of intact tissue was performed using MALDI. Nanospray experiments produced protonated ions that were identified as PCs, PEs, Cers, and SPMs. MALDI of intact tissue with additional Na resulted in the formation of [M+H], [M+Na], and [M+K] ions. Molecular species were identified as Cers, SPMS, PCs, PEs, and PSs. Lipid extracts analyzed by MALDI resembled a combination of the data obtained in the other two experiments. A comparison of the three ionization techniques used here also demonstrated that lipid ions tend to more fragile when produce by MALDI compared to nanospray. The direct analysis of tissue proved to be more sensitive than the tissue extract experiments, and also affords the opportunity to provide localization of lipids within tissue. |
