One of the urgent tasks in understanding endocrine disruptors (EDs) is to compile a list of suspected substances among the huge number of chemicals by using the screening test method. We developed a simple and rapid screening method using the yeast two-hybrid system based on the ligand-dependent interaction of nuclear hormone receptors with coactivators. To date, we have tested the estrogenic activity of more than 500 chemicals including natural substances, medicines, pesticides, and industrial chemicals. 64 compounds were evaluated as positive, and most of these demonstrated a common structure; phenol with a hydrophobic moiety at the para-position without bulky groups at the ortho-position. These results are expected to facilitate further risk assessment of chemicals.