2-Aminopyridine (2AP) reacts instantaneously with 2,4-dinitrophenol (DNP) in methanol to form yellow proton transfer (PT) complex at absorption maxima from 362–402 nm. The molecular composition of the PT-complex has been identified by Job's and photometric titration methods to be 1 : 1. The PT-complex formation constant (K PT) and molecular extinction coefficient (ε) were estimated by using Benesi–Hildebrand equation. They recorded high values confirming high stability of the formed complex. The stability of the formed complex was interpreted based on the formation of bifurcated and cooperative hydrogen bonds. On the basis of the rapidity of the PT-reactions, a simple and accurate spectrophotometric method for determination 2AP was proposed. Beer's law was obeyed in the concentration range 2–56 µg mL−1 with excellent correlation coefficients (0.99). Furthermore, the recovery percentage recorded 100.93%.