Peptides with substance P-like immunoreactivity, neurokinin A-like immunoreactivity and neurotensin-like immunoreactivity were isolated in pure form from an extract of the intestine of the Burmese python (Python molurus). The primary structure of python substance P (Arg-Pro-Arg-Pro-Gln-Gln-Phe-Tyr-Gly-Leu- Met-NH2) shows one amino acid substitution (Phe8-->Tyr) compared with chicken/alligator substance P and an additional substitution (Lys3-->Arg) as compared with mammalian substance P. The neurokinin A-like immunoreactivity was separated into two components. Python neuropeptide gamma (Asp-Ala-Gly-Tyr- Ser-Pro-Leu-Ser-His-Lys-Arg-His-Lys-Thr-Asp-Ser-Phe-Val-Gly-Leu-Met-NH2 shows three substitutions (Gly5-->Ser, Gln6-->Pro and Ile7-->Leu) compared with alligator neuropeptide gamma and an additional substitution (His4-->Tyr) compared with mammalian neuropeptide gamma. Python neurokinin A (His-Lys-Thr-Asp-Ser-Phe-Val-Gly- Leu-Met.NH2) is identical to human/chicken/alligator neurokinin A. Python neurotensin (pGlu-Leu-Val-His-Asn-Lys-Ala-Arg-Pro-Tyr-Ile-Leu) is identical to chicken/alligator neurotensin. The data are indicative of differential evolutionary pressure to conserve the amino acid sequences of reptilian gastrointestinal peptides.