tRNA-derived small RNAs (tsRNAs) are non-coding small RNAs that are produced through the precise cleavage of tRNA molecules under specific conditions. tsRNA has multiple functions, including inhibiting translation, acting in association with classical small RNA effector mechanisms, or acting in conjunction with Argonaute proteins that affect cell proliferation, migration, cycle, and apoptosis. Recent studies have revealed the clinical potential of tsRNAs in numerous diseases. This article aims to provide a comprehensive and up-to-date review of the classification and biological function of tsRNAs in gastrointestinal diseases. Furthermore, this review explores the underlying mechanisms by which tsRNAs are believed to exert their effects in both tumor and non-tumor digestive tract diseases. Therefore, specific tsRNAs prove promising for disease diagnosis, prognosis prediction, and therapeutic interventions as novel biomarkers for digestive tract diseases.