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This article represents data regarding a study published in Toxicology in vitro entitled “ in vitro CYP-mediated drug metabolism in the zebrafish (embryo) using human reference compounds” (Saad et al., 2017) [1]. Data were acquired with ultra-performance liquid chromatography – accurate mass mass spectrometry (UPLC-amMS). A full spectrum scan was conducted for the testosterone (TST) metabolites from the microsomal stability assay in zebrafish and humans. The microsomal proteins were extracted from adult zebrafish male (MLM) and female (FLM) livers, whole body homogenates of 96 h post fertilization larvae (EM) and a pool of human liver microsomes from 50 donors (HLM). Data are expressed as the abundance from the extracted ion chromatogram of the metabolites.