The fall armyworm Spodoptera frugiperda (J. E. Smith) (Lepidoptera: Noctuidae) is a major agricultural pest. Low temperature is an important factor that limits S. frugiperda survival and reproduction. The physiological cold tolerance indexes of S. frugiperda larvae from October 2022 to February 2023 were investigated. The results showed that the cold-resistant substances in S. frugiperda differed significantly among different months. The water content was the lowest in January. Lipids were the highest in December and January, trehalose and protein contents were the highest in December, and glycerol content was the highest in January. The contents of lipid, protein, trehalose, and glycerol were the highest in the 6th instar larvae, whereas the water content did not differ among the larva stages. The functions of two genes, including endothelial lipase (Eol1) and gastric lipase (Epl1) that were extracted from the transcriptome analysis of S. frugiperda in the cold resistance were assessed. After 24 h of cold stress, the mRNA levels of Eol1 and Epl1 RNA interference were significantly down-regulated by 60.76 % and 82.53 %, and the survival rate of S. frugiperda larvae decreased. Additionally, Eol1 and Epl1 silencing significantly decreased the lipid content and pupal weight of S. frugiperda. The results indicate that S. frugiperda larvae can change the contents in winter to response the cold conditions and the lipids plays an important role in the cold resistance of S. frugiperda.