Chitosan microparticles were prepared for extended release of acetogenins extracted from the leaves of Annona muricata L., known as soursop. The acetogenin extract was encapsulated in chitosan microparticles prepared using sodium tripolyphosphate as the crosslinking agent. The possible interactions between the microparticle components were analyzed based on their FTIR absorption peaks. The effect of the experimental variables (acetogenin to chitosan mass ratio, amount of tripolyphosphate, presence of enzymes) on the 24-h in vitro cumulative release of acetogenins encapsulated in the chitosan microparticles and immersed in simulated gastrointestinal fluids, is presented. The sequential release in simulated (gastric, intestinal, colonic) fluids for 24 h period, facilitated higher cumulative release of acetogenins, indicating the potential of acetogenin-loaded microparticles for anticancer therapy purposes.