Poly(ester amide)s from dimethyl sebacate or sebacic acid and 2-aminoethanol or 4-amino-1-butanol were characterized by post-source decay matrix-assisted laser desorption/ionization time-of-flight (PSD-MALDI-TOF) and time-of-flight/time-of-flight tandem mass spectrometry (MALDI-TOF/TOF-MS/MS). Sodiated oligomers were selected as precursor ions for dissociation studies. PSD analysis was performed on dimethyl sebacate, dicarboxylic, carboxylic and amino alcohol, and diamino alcohol terminated oligomers. PSD-MALDI-TOF mass spectra yielded information on the fragmentation mechanisms of the poly(ester amide) chains, showing that the main cleavages proceed through a beta-hydrogen transfer rearrangement. MALDI-TOF/TOF-MS/MS provided structural information concerning ester/amide sequences in the polymer chains. As expected, together with the ions appearing in the PSD-MALDI mass spectrum, several new abundant fragment ions in the low-mass range are present in MALDI-TOF/TOF-MS/MS spectra. These new product ions proved to be diagnostic and made it possible to establish the presence of random sequences of ester and amide bonds in the poly(ester amide)s samples.