Joint action and biochemical alteration in the 4^th instar larvae of a laboratory strain of cotton leafworm, Spodoptera littoralis (Boisd.), were evaluated under the laboratory condition against four insecticides from different groups namely: chlorpyrifos, chlorfluazuron, emamectin benzoate, pyrethrins. The toxicity effect for each single compound and binary mixture at levels 1:1, 1:2: and 2:1 from LC₂₅ were evaluated. The study indicated that emamectin benzoate followed by chlorfluazuron proved to be the most effective compounds among all tested insecticides. The highest initial effect (76.66%) was obtained by mixing chlorpyrifos: chlorfluazuron at level 1:2, respectively. while the lowest initial effect (3.33%) was obtained as a result of mixing emamectin benzoate with pyrethrins at level 1:2, respectively, the same effect occurred with chlorpyrifos: pyrethrins at level 1:1. Changing in enzymatic activities as responses of treatment with mixtures at LC₂₅ of all tested compounds, except for pyrethrins was studied at the recommended concentration, after 1and 3 days post treatment. Our study noticed that mixing of pesticides caused antagonism effect except chlorpyrifos plus pyrethrins at the ratio 2:1 caused potentiation effect. Under the effect of the tested insecticides the data showed increasing in total soluble protein and total lipids contents, reduction in carbohydrate hydrolyzing enzymes (amylase, invertase and trehalase), increasing in  transaminase enzymes (AST and ALT) and reduction in phosphatase enzymes (ACP and ALP)  activities .Our data recommend not to mix these insecticides due to the antagonistic effect.