In this study, we identified major aryl hydrocarbon receptor (AhR) agonists in the sediments from Yeongil Bay (n = 6) using effect-directed analysis. Using the H4IIE-luc bioassays, great AhR-mediated potencies were found in aromatic fractions (F2) of sediment organic extracts from silica gel column chromatography and sub-fractions (F2.6–F2.8) from reverse phase-HPLC. Full-scan mass spectrometric analysis using GC-QTOFMS was conducted to identify novel AhR agonists in highly potent fractions, such as F2.6–F2.8 of S1 (Gumu Creek). Selection criteria for AhR-active compounds consisted of three steps, including matching factor of NIST library (≥70), aromatic structures, and the number of aromatic rings (≥4). Fifty-nine compounds were selected as tentative AhR agonist candidates, with the AhR-mediated activity being assessed for six compounds for which standard materials were available commercially. Of these compounds, 20-methylcholanthrene, 7-methylbenzaanthracene, 10-methylbenzapyrene, and 7,12-dimethylbenzaanthracene exhibited significant AhR-mediated potency. Relative potency values of these compounds were determined relative to benzoapyrene to be 3.2, 1.4, 1.2, and 0.2, respectively. EPA positive matrix factorization modeling indicated that the sedimentary AhR-active aromatic compounds primarily originated from coal combustion and vehicle emissions. Potency balance analysis indicated that four novel AhR agonists explained 0.007% to 1.7% of bioassay-derived AhR-mediated potencies in samples. Display omitted •Novel AhR agonists were identified in industrial sediments using EDA combined with FSA.•7,12DbA, 10MbA, 7MbA, and 20MC had significant AhR-mediated potencies.•Relative potency values of novel AhR agonists compared to benzoapyrene were obtained.•Novel AhR agonists explained from 0.007 to 1.7% of total induced AhR potencies.