The stomatogastric nervous system (STNS) of decapod crustaceans serves as a simple model system to study modulation and control by neuropeptides. The full characterization of STNS neuropeptides in invertebrate neuronal tissues is a necessary step towards understanding the role of these messengers and modulators. Matrix assisted laser desorption/ionization (MALDI) and electrospray ionization (ESI) mass spectrometry play an important role in the characterization of nervous system peptides. Recent studies have demonstrated that MALDI-Fourier transform mass spectrometry (FTMS) presents advantages over MALDI-time of flight MS for the direct analysis of tissue sample in the areas of sensitivity, mass accuracy, and peptide sequencing (Kurtz et al., Anal. Chem. 2004, 76, 5630); however, the longer time frame associated with MALDI-FTMS detection results in peptide fragmentation via small neutral losses and cleavages C-terminal to aspartate residues for charge localized peptides (Stemmler et al., Anal. Chem. 2005, 77 3594). In this study, we describe recent work on the development of a microscale extraction/chemical derivatization procedure for the analysis of single neuronal tissues by MALDI-FTMS. Peptides from single tissues are extracted and separated from phospholipids using a microscale liquid-liquid extraction. Esterification reactions are used to eliminate aspartate cleavages and enhance molecular ion formation. Formation of deuteromethyl esters is used to determine the total number of acidic residues in the peptide sequence. Application of the microscale extraction/derivatization procedure to single commissural ganglia and sinus glands from the Maine lobster, Homarus americanus, and to single pericardial organs from the kelp crab, Pugettia producta, will be presented.
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