Previously, we isolated six heterocyclic compounds (1-6) from the fruits of mulberry trees (Morus alba L.) and determined that loliolide affords rat pancreatic islet β-cell (INS-1) protection against streptozotocin‑induced cytotoxicity. In the present study, we further investigated the effect of the six heterocyclic compounds (1-6) on glucose-stimulated insulin secretion (GSIS) in INS-1 cells. Among them, (R)‑5‑hydroxypyrrolidin‑2‑one(1) and indole (6) increased GSIS without inducing cytotoxicity. Additionally, compounds 1 and 6 enhanced the phosphorylation of total insulin receptor substrate-2, phosphatidylinositol 3-kinase, and Akt, and activated pancreatic and duodenal homeobox-1, which play a crucial role in β-cell functions related to insulin secretion. Collectively, these findings indicate that (R)‑5‑hydroxypyrrolidin‑2‑one(1) and indole (6), isolated from M. alba fruits, may be beneficial in managing type 2 diabetes.