The formation of cyclic dipeptides (DKPs, 2,5-diketopiperazines) from dipeptides having proline (Pro) as the fixed N-terminal amino acid was investigated by analytical pyrolysis with a heated platinum filament coil. Glutamic acid (Glu), aspartic acid (Asp), glutamine (Gin), lysine (Lys) or arginine (Arg) was the terminal amino acid. Products evolved from off-line pyrolysis at 500 degrees C were analysed by GC-MS as such or after trimethylsilylation. The structure of a novel DKP from the pyrolysis of Pro-Glu, namely hexahydrodipyrrolo[1,2-a:1',2'-d]pyrazine-3,5,10(10aH)-trione, was established by ESI-MS/MS and extensive NMR analysis. This DKP could be identified in the pyrolysates of dipeptide Pro-Gln, tripeptide Pro-Glu-Leu, collagen and bovine serum albumin (BSA). Pyrolysis of Pro-Lys and Lys-Pro-Leu afforded the cyclo(Pro-Lys) tentatively identified by interpretation of its trimethylsilyl (TMS) derivative mass spectra and the DKPs resulting from the deamination of the lateral chain. The dipeptide Pro-Asp yielded isomeric cyclo(Pro-Asp) revealed as TMS derivatives and cyclo( Pro-Ala) from the decarboxylation of the side chain. Analytical pyrolysis of the above peptides as well as of Pro-Tyr, collagen and BSA enabled the compilation of GC (retention times relative to an internal standard) and MS data (characteristic ions) of over eighty DKPs including their TMS derivatives, the latter ones useful in the identification of thermal degradation products of proteinaceous materials in several matrices. (C) 2012 Elsevier B.V. All rights reserved.