The object of this study was the acaricidal activities of acetophenone (AP) and its derivatives for their potentials as natural acaricides using fumigant and contact toxicity bioassays against Dermatophagoides farinae, D. pteronyssinus, and Tyrophagus putrescentiae. Based on the LD50 values of AP derivatives against D. farinae, 3’-methoxyAP (0.41 ㎍/㎠) was 89.9 times more toxic than DEET (36.87 ㎍/㎠), followed by 4’-methoxyAP (0.52 ㎍/㎠), 2’-methoxyAP (0.75 ㎍/㎠), 2’-hydroxy-5’-methoxyAP (1.03 ㎍/㎠), 2’-hydroxy -4’-methoxyAP (1.29 ㎍/㎠), AP (1.48 ㎍/㎠), 2’-hydroxyAP (1.74 ㎍/㎠), 2’,5’-dimethoxyAP (1.87 ㎍/㎠), 2’,4’-dimethoxyAP (2.10 ㎍/㎠), and benzyl benzoate (9.92 ㎍/㎠). With regard to structure-activity relationships between acaricidal activity and functional groups (hydroxyl and methoxy groups) on the AP skeleton, a mono-methoxy group (2’-, 3’-, and 4’-methoxyAP) on the AP skeleton was more toxic than the other groups (2’,4’- and 2’,5’-dimethoxyAP, 2’- and 4’-hydroxyAP, 2’-hydroxy-4’-methoxyAP, 2’-hydroxy-5’-methoxyAP, and 4’-hydroxy-3’-methoxyAP). These results indicated that acaricidal activity against three mite species was changed with the introduction of functional radicals (hydroxyl and methoxy groups) onto the AP skeleton.