ABSTRACT Porcine epidemic diarrhea virus (PEDV) is a pig coronavirus that causes severe diarrhea and high mortality in piglets, threatening the global pig industry. Clinically effective medicines against this virus are still not available. To find an optional natural feed compound, viral titers were detected, and the intestinal contents of specific pathogen-free pigs more effectively blocked PEDV invasion than those of the other two breeds investigated. Based on proteomic and metabolic analyses of the intestinal content, 10 metabolites were selected for further investigation, and linoleic acid (LA) was the most promising according to the selectivity index. By detecting viral gene expression, LA inhibited viral replication and release from Vero-E6 cells, mainly by influencing the PI3K pathway and, in particular, inhibiting AKT phosphorylation. In addition, LA bound to viral NSP5 and attenuated NSP5-activated AKT phosphorylation. In the in vivo pig experiment, oral administration of the higher dose of LA protected two-thirds of pigs from death, both of which completely recovered from the infection, while the lower dose of LA protected one-third of pigs from death but with the induction of severe diarrhea. In conclusion, LA can be used as a candidate medicine for the clinical prevention and treatment of PEDV, and its mechanism of inhibiting the PI3K signaling pathway provides some data support for the subsequent exploration of antiviral drugs for coronavirus infections. IMPORTANCE Porcine epidemic diarrhea virus (PEDV) is a pig coronavirus that causes severe diarrhea and high mortality in piglets, but as no effective drugs are available, this virus threatens the pig industry. Here, we found that the intestinal contents of specific pathogen-free pigs effectively blocked PEDV invasion. Through proteomic and metabolic analyses of the intestinal contents, we screened 10 metabolites to investigate their function and found that linoleic acid (LA) significantly inhibited PEDV replication. Further investigations revealed that LA inhibited viral replication and release mainly by binding with PEDV NSP5 to regulate the PI3K pathway and, in particular, inhibiting AKT phosphorylation. In vivo experiments illustrated that orally administered LA protected pigs from PEDV challenge and severe diarrhea. These findings provide strong support for exploring antiviral drugs for coronavirus treatment.