Cytochrome P450 monooxygenases (P450s) are a large class of enzymes that play essential roles in metabolic processes such as hormone synthesis and the catabolism of toxins and other chemicals in insects. In the present study, we identified 82 P450 genes using comprehensive RNA sequencing in the flea beetle Agasicles hygrophila, and all of the sequences were validated by cloning and sequencing. Phylogenetic analysis showed that the P450 genes in A. hygrophila fell into the mitochondrial clan, CYP2 clan, CYP3 clan and CYP4 clan and were classified into 20 families and 48 subfamilies. Most A. hygrophila P450 genes had high sequence homology with those from other coleopteran insects. To understand the effects of high temperatures on the metabolic processes of female and male adults, we studied the effects of two temperature regimes (constant temperature of 28 °C for 20 h with a 4-h period of high temperatures of 30 °C and 39 °C) on the expression levels of P450 genes in A. hygrophila using RT-PCR and qRT-PCR. The results showed that there were no differences in expression in 30 P450 genes between the control and high-temperature-treated A. hygrophila adults, while 22 P450 genes showed up-regulated expression and 19 P450 genes were down-regulated in A. hygrophila female adults after high-temperature treatment. For A. hygrophila male adults exposed to high temperatures, we found that 8 P450 genes had higher expression levels and 12 P450 genes had lower expression levels under the same conditions. The P450 genes are candidates that showed significantly different expression levels after high-temperature treatments in A. hygrophila adults, and further studies are needed to determine their possible roles in metabolic processes during the response to elevated temperatures.