Prohydrojasmon (PDJ) is a synthetic jasmonate derivative that is primarily used as a growth regulator, but its mechanism of action is unclear. In this study, we elucidated the effects of PDJ on phytochemical production in red leaf lettuce. The PDJ treatments promoted the accumulation of phenolic compounds in aerial plant parts. An LC-MS analysis revealed that these accumulated compounds were identified as cyanidin-3-O-glucoside, cyanidin-3-O-(6″-O-malonyl)-glucoside and cyanidin-3-O-(6″-O-malonyl)-glucoside methyl ester. The abundance of these compounds in lettuce extracts increased significantly in response to the PDJ treatment. Additionally, the LC-MS analysis also identified the accumulated phenolic compounds in the extracts of PDJ-treated lettuce, including caffeoyltartaric acid, chlorogenic acid, caffeoylmalic acid, chicoric acid, and dicaffeoylquinic acid. Gene expression analyses indicated the PDJ treatments upregulated the expression of PAL, F3H, and ANS genes in lettuce. These results suggest that PDJ treatments enhance the expression of genes involved in the synthesis of anthocyanins and phenolic compounds, resulting in an increase in the quantities of these compounds, which reportedly have various functions affecting human physiology.