17β-Estradiol (E2), estrone (E1) and estriol (E3) are steroid hormones responsible for the regulation of the female reproductive system. Estradiol is planned to be used to feminize eels in aquaculture in order to improve their size and marketability. The residual levels of these hormones in fish tissue must be monitored to meet the requirements of food regulatory agencies. Few studies have studied these hormones in complex biological matrices such as fish tissue.