Exocrine pancreatic insufficiency (EPI) causes chronic digestive dysfunction in cats, but its pathogenesis and pathophysiology are poorly understood. Untargeted metabolomics is a promising analytic methodology that can reveal novel metabolic features and biomarkers of clinical disease syndromes. The purpose of this preliminary study was to use untargeted analysis of the serum metabolome to discover novel aspects of the pathobiology of EPI in cats. Serum samples were collected from 5 cats with EPI and 8 healthy controls. The diagnosis of EPI was confirmed by measurement of subnormal serum feline trypsin-like immunoreactivity (fTLI). Untargeted quantification of serum metabolite utilized ultra-high-performance liquid chromatography-tandem mass spectroscopy. Cats with EPI had significantly increased serum quantities of long-chain fatty acids, polyunsaturated fatty acids, mevalonate pathway intermediates, and endocannabinoids compared with healthy controls. Diacylglycerols, phosphatidylethanolamines, amino acid derivatives, and microbial metabolites were significantly decreased in cats with EPI compared to healthy controls. Diacyclglycerols and amino acid metabolites were positively correlated, and sphingolipids and long-chain fatty acids were negatively correlated with serum fTLI, respectively. These results suggest that EPI in cats is associated with increased lipolysis of peripheral adipose stores, dysfunction of the mevalonate pathway, and altered amino acid metabolism. Differences in microbial metabolites indicate that feline EPI is also associated with enteric microbial dysbiosis. Targeted studies of the metabolome of cats with EPI are warranted to further elucidate the mechanisms of these metabolic derangements and their influence on the pathogenesis and pathophysiology of EPI in cats.