Dialkyl phthalates have been suggested to function as xenoestrogen. To explore the structural essentials, a series of ring and alkyl-chain isomers of dialkyl phthalates C 6 H 4 (COOC n H m ) 2 were examined for their ability to displace [ 3 H]17β-estradiol in the recombinant human estrogen receptor expressed on Sf9 vaculovirus. Compounds with an alkyl chain of more than C 3 (n= 3) exhibited a distinct full receptor binding in a dose-dependent manner. When the ring isomers of C 3 -diallyl (-CH 2 -CH=CH 2 ) derivatives, namely diallyl phthalate, diallyl isophthalate, and diallyl terephthalate, were examined, theorthoisomer of diallyl phthalate was most potent to bind to the estrogen receptor. The interaction with the estrogen receptor was optimized with dibutyl phthalates of C 4 . The conformational studies by 1 H-NMR measurements andab initiomolecular orbital calculations have suggested that the structure mimics the interface of steroid A and B/C rings of 17β-estradiol. Dicyclohexyl phthalate bound to the estrogen receptor with a biphasic binding curve, suggesting the compound discriminates two different receptor conformations.