The conformation and aggregation behavior of synthetic Alzheimer's amyloid peptides (Aβ) has been investigated using hydrogen-deuterium exchange measured by electrospray ionization mass spectrometry and matrix-assisted laser desorption/ionization mass spectrometry. Mass spectrometric fragmentation of deuterated Aβ peptides was carried out by collision-induced dissociation, inlet fragmentation, and post-source decay. In contrast to the C-terminally truncated peptides Aβ(1-40) and Aβ(1-36) showing full hydrogen-deuterium exchange, Aβ(1-42) and the pyroglutamyl peptide Pyr3-Aβ(3-42) produced more complex signal patterns resulting from the formation of β-sheet-structured oligomers having 18–20 strongly protected protons. Using mass spectrometric fragmentation the results show that the reduced isotope exchange of Aβ(1-42) can be attributed to the central part of the chain comprising residues 8-23. This confirms involvement of the hydrophobic binding domain LVFFA in the course of Aβ aggregation and demonstrates that hydrogen-deuterium exchange in combination with mass spectrometry is well suited for structural analysis of monomeric and reversibly associated amyloid peptides using picomole quantities of material. Copyright © 2002 John Wiley & Sons, Ltd.