Background and Purpose: The in vivo oestrogenicity of genistein and its glycoside genistin is still under debate. The present study aimed to develop a physiologically based kinetic (PBK) model that provides insight in dose-dependent plasma concentrations of genistein aglycone and its metabolites and enables prediction of in vivo oestrogenic effective dose levels of genistein and genistin in humans. Experimental Approach: A PBK model for genistein and genistin in humans was developed based on in vitro metabolic parameters. The model obtained was used to translate in vitro oestrogenic concentration–response curves of genistein to in vivo oestrogenic dose–response curves for intake of genistein and genistin. Key Results: The model predicted that genistein-7-O-glucuronide was the major circulating metabolite and that levels of the free aglycone were generally low [0.5–17% of total plasma genistein at oral doses from 0.01 to 50 mg (kg·bw)−1]. The predicted in vivo benchmark dose for 5% response values for oestrogenicity varied between 0.06 and 4.39 mg kg−1 genistein. For genistin, these values were 1.3-fold higher. These values are in line with reported human data and show that oestrogenic responses can be expected at an Asian dietary and a supplementary intake, while intake resulting from a Western diet may not be effective. Conclusions and Implications: The present study shows how plasma concentrations of genistein and its metabolites and oestrogenic dose levels of genistein in humans can be predicted by combining in vitro oestrogenicity with PBK model-based reverse dosimetry, eliminating the need for human intervention studies.