Most animals, including humans, produce natural sex hormones such as estrogens: 17β-estradiol (E2) and estrone (E1). These compounds are able to disrupt the reproductive systems of living organisms at trace concentrations (ng·L − 1). This experiment tests the hypothesis that 1% slow pyrolysis biochar-amended sandy soil could retain significant amount of estrogens (E1, E2) from poultry manure in its second year of application. The experiment was conducted over 46 days and consisted of a series of lysimeters containing sandy soil with biochar-amended topsoil. The application rate of poultry manure was kept at 2.47 kg·m − 2. The biochar held a significant concentration of hormone during the first year of its application. However, in the following year (current study), there was no significant retention of hormones in the biochar-amended soil. During the first year after application, the biochar was fresh, so its pores were available for hydrophobic interactions and held significant concentration of hormones. As time passed there were several biotic and abiotic changes on the surface of the biochar so that after some physical fragmentation, pores on the surface were no longer available for hydrophobic interactions. The biochar started releasing dissolved organic carbon, which facilitated greater mobility of hormones from poultry manure down the soil profile.