We aimed to investigate how assimilation of nutrients affects the postprandial responses of GLP-1 and GIP, and to evaluate the effect of pancreatic enzyme substitution (PES) on insulin secretion in patients with chronic pancreatitis (CP) and pancreatic exocrine insufficiency (PEI). Eight male patients with CP and PEI were studied. Blood was sampled frequently on two separate days after ingestion of a liquid meal with and without PES, respectively. Eight healthy male subjects served as a control group. Beta-cell responsiveness was estimated as changes in insulin secretion rates in response to changes in postprandial plasma glucose (PG). There was no difference in the PG incremental area under curve (AUC) for patients with and without PES (406±100 vs. 425±80 mM · 4h (mean±SEM); P = 0.8). The response of total GLP-1 was higher after PES (AUC: 7.8±1.2 vs. 5.3±0.6 nM · 4h; P = 0.01), as was the response of total GIP (AUC: 32.7±7.5 vs. 21.1±8.3 nM · 4h; P = 0.01). Concurrently, both plasma insulin, plasma C-peptide, and total insulin secretion increased after PES (AUC: 17.7±4.2 vs. 13.6±2.9 nM · 4h; P = 0.02, 237±31.4 vs. 200±27.4 nM · 4h; P = 0.005, and 595±82 vs. 497±80 pmol · kg^-1 · 4h; P = 0.01, respectively). Beta-cell responsiveness to glucose was not significantly different on the two study days for patients with CP. These results suggest that the secretion of GLP-1 and GIP is under influence of the assimilation of nutrients in the small intestine, and that PES increases insulin secretion.