It is important to understand the effects of temperatures on Egyptian cotton leafworm Spodoptera littoralis (Boisd.) (Lepidoptera: Noctuidae) which damages a wide variety of crops in the Middle East and is one of the major economic pests of cotton in Egypt. Therefore, the investigating was designed to study the effect of fluctuation temperatures on the field strain in Abnob city, Assuit Governorate during the 2017 and 2018 cotton growing seasons. On the other hand, the results compared with the laboratory strain under constant temperature (25°C). The biochemical parameters were measured on the 4^th larval instars for both strains. Biochemical analyses were performed to determine the effects of fluctuation and constant temperatures on field and laboratory strain on total proteins, lipids, carbohydrates and free amino acids as well as metabolic digestive, neural and detoxification enzymes. It's clear from the results that the GST, GOT, AchE, Alpha esterases, Beta esterases as well as, the activity levels were increased significantly by field strain. While the free amino acids, total proteins, proteases, lipases, amylase were decreased significantly by field strain and no significant changes were detected between field and laboratory strains in total carbohydrates, total lipids, trehalase and GPT.