The European red mite, Panonychus ulmi Koch (Chelicerata: Acari: Tetranychidae) is one of the most important pests of apple orchards that can cause severe damages on apple trees every year. Chemical acaricides are often used extensively to control P. ulmi in Iran. The biopesticide Requiem®EC is a commercial formulation based on the essential oil of Chenopodium ambrosioides. This botanical product is a mixture of monoterpenes (mainly α-terpinene, p-cymene, and limonene) that formulated for control of insect and mite pests. In this study, the acaricidal activity of Requiem®EC against adult females of two P. ulmi populations was assessed in four replications by serially diluted concentrations (10, 5, 2.5, 1.25, 0.62, and 0.31 ml/l). The biopesticide showed toxic action when a concentration series from 0.31 to 10 ml/l was directly sprayed by Potter spray tower (1.5 ml, 1bar, and 1.46±0.05 mg deposit cm2) on the adult females. The LC50 values of Requiem® were estimated to be 1.23 (0.78-1.87) and 2.18 (1.24-3.02) ml/l for Mianeh2 and Marand populations, respectively. The activity of glutathione-S-transferase (GST), cytochrome P450 (P450), and carboxylesterase (CarE) in Mianeh2 population were 932.47±51 (nmol glutathione conjugated min-1 mg-1 protein), 8.30±0.38 (equivalent units of cytochrome P450 mg-1 protein) and 4642.16±238 (nmol 1-naphthol min-1 mg-1 protein), respectively. Also, the activity of GST, P450, and CarE in the Marand population were 391.10±30 (nmol glutathione conjugated min-1 mg-1 protein), 6.80±0.18 (equivalent units of cytochrome P450 mg-1 protein) and 5274.84±315 (nmol 1-naphthol min-1 mg-1 protein), respectively. Apparently, the difference in detoxification enzyme activities especially GSTs is one of the reasons of different susceptibility to Requiem® in Mianeh2 and Marand populations. The LC95 values of Requiem® were 8.21 (6.98-10.61) and 9.86 (6.91-20.24) ml/l on Mianeh2 and Marand populations, respectively. The recommended concentration of Requiem® is 4-10 ml/l based on the label. Considering the estimated LC95 values and the recommended field dose of Requiem®, it seems that application of this biopesticide at 10 ml/l concentration in the apple orchards could be effective against P. ulmi.