We examined the fragmentation of the electrospray-produced [M–H]− and [M–2H]2− ions of a number of peptides containing two acidic amino acid residues, one being aspartic acid (Asp) or glutamic acid (Glu), and the other being cysteine sulfinic acid [C(SO2H)] or cysteine sulfonic acid [C(SO3H)], on an ion-trap mass spectrometer. We observed facile neutral losses of H2S and H2SO2 from the side chains of cysteine and C(SO2H), respectively, whereas the corresponding elimination of H2SO3 from the side chain of C(SO3H) was undetectable for most peptides that we investigated. In addition, the collisional activation of the [M–H]− ions of the C(SO2H)-containing peptides resulted in the cleavage of the amide bond on the C-terminal side of the C(SO2H) residue. Moreover, collisional activation of the [M–2H]2− ions of the above Asp-containing peptides led to the cleavage of the backbone N-Cα bond of the Asp residue to give cn and/or its complementary [zn–H2O] ions. Similar cleavage also occurred for the singly deprotonated ions of the otherwise identical peptides with a C-terminal amide functionality, but not for the [M–H]− ions of same peptides with a free C-terminal carboxylic acid. Furthermore, ab initio calculation results for model cleavage reactions are consistent with the selective cleavage of the backbone N–Cα bond in the Asp residue. Copyright © 2006 John Wiley & Sons, Ltd.