Bisphenol A (BPA) is a widely used plastic material, and halogenated BPA derivatives are formed either by synthesis or environmental processes. However, the effect of halogenated bisphenols on steroidogenesis remains unclear. The aim of this study was to compare inhibition of 6 BPA derivatives on gonadal 3β-hydroxysteroid dehydrogenases (3β-HSDs) in three species (human, rat, and mouse). The inhibition on human 3β-HSD2 was tetrabromo BPA (TBBPA, IC50, 1.01 μM)>trichloro BPA (TrCBPA, 3.95 μM)>tetrachloro BPA (TCBPA, 4.14 μM)>monochloro BPA (MCBPA, 4.74 μM)>others with TrCBPA of competitive, TBBPA of noncompetitive and MCBPA/TCBPA of mixed inhibition. The inhibition on rat 3β-HSD1 was TCBPA (1.68 μM)>TrCBPA (1.72 μM)>MCBPA (2.80 μM)>BPA>others with mixed inhibition. The inhibition on mouse 3β-HSD6 was TrCBPA (1.59 μM) >MCBPA (3.36 μM)>TCBPA (3.72 μM)>others with mixed inhibition. Molecular docking analysis showed that TBBPA, TrCBPA, and TCBPA bind to steroid active sites, contacting with catalytic residue Tyr154 of human 3β-HSD2. MCBPA, TrCBPA, and TCBPA bind to steroid active site of rat 3β-HSD1. MCBPA and TrCBPA bind to active site of mouse 3β-HSD6. Regression of lowest binding energy values with Ki values revealed a significant negative linear regression (P < 0.05). In conclusion, halogenated BPA derivatives are more potent inhibitors of three 3β-HSDs than BPA and there is structure-dependent inhibition. Chlorinated bisphenol derivatives after water chlorination process and other halogenated bisphenols effectively inhibit human and rat 3β-HSD activity, thereby leading to steroid hormone deficiency. Display omitted •Chlorinated BPA derivatives potent inhibit human, rat, and mouse gonadal 3β-HSD.•Tetrabromobisphenol A is the most potent inhibitor of human 3β-HSD2.•Tetrabromobisphenol A is ineffective to inhibit rat and mouse gonadal 3β-HSD.•Chlorination significantly increases the inhibitory potency of BPA derivatives.