Abstract Biochemical properties of the aci-GTH prepared by gel filtration and DEAE-C chromatography were studied. This material had a sedimentation coefficient of 2.4–2.8. In analytical electrophoresis acrylamide gel at pH 8.9 it gave two main bands, a and b ( R f = 0.27 and 0.31) both of them active in the frog spermiation test. Preparative acrylamide electrophoresis led to the separation of two active fractions, of which one contained only band a. Amino acid composition of this fraction was determined. Dissociation agents strikingly modified the properties of aci-GTH. Sedimentation rate was decreased to a value of 1.5. Biological activity was less than one-tenth of that of the native hormone. Analytical electrophoresis showed the formation, from band a, of two more acidic bands ( R f = 0.53 and 0.59). These results strongly indicate that aci-GTH was, in these conditions, dissociated into two subunits. Aci-GTH is different from carp GTH in amino acid composition, electrophoretic behavior, and level of the equilibrium between the hormone and its subunits in several experimental conditions.