Esterase-mediated metabolic resistance more or less happens in all insects and all classes of insecticides in ester group. In the current study, the inhibitory effect of oxydemeton-methyl and pirimicarb on acetylcholinesterase (AChE) activity and their insecticides and imidaclopride extracted from two populations of Aphis gossypii (cotton aphid and melon aphid) on carboxylesterase activity were compared. Results showed that cotton aphid esterase activity against two substrates including α- and β-naphthyl acetate was 124.6 and 71.6 U/mg and for melon aphid 33.7 and 53 U/mg, respectively. Native electrophoresis showed that cotton aphid has more esterase bands than melon aphid (5 versus 3 bands). Cholinesterase inhibitory test showed that 50% of the enzyme activity of cotton aphid inhibited by 776.2 and 958.7 ppm of oxydemeton-methyl and pirimicarb, respectively, while 50% of the enzyme activity of melon aphid inhibited by 225.8 and 741.3 ppm of oxydemeton-methyl and pirimicarb, respectively. Thus, cotton aphid is more resistant to selected insecticides than melon aphid.