Ionization threshold energies of 26 substituted polyacetylenes were determined by ultraviolet photoelectron spectroscopy. Examination of the data obtained, along with data for other polymers, and the energy of the lowest energy electronic excitation revealed that the threshold energies correspond to the photoemission from the highest occupied orbitals in the π-conjugated main chains. For the majority of the polymers, the main origin of the significant variation of threshold energy and electronic excitation energy is ascribed to the change of the degree of π-conjugation due to steric hindrance of substituents. Calculations with Hückel theory gave semiquantitative agreement with the observed results. Polymers with Cl substituents at the main chain also showed a large inductive effect by the electron-withdrawing Cl atom. The HOMO of these polymers was found to be derived from the π-conjugated main chain.