Here, we report a facile and effective one-pot approach to prepare uniform amylose-based polymeric microparticles (PMPs) through enzymatic synthesis of short-chain amylose (SCA) followed by spontaneous self-assembly of the SCA in the presence of lecithin. The effect of lecithin on nucleation and growth kinetics of amylose microparticles was investigated by monitoring the turbidity of reaction solution and the size of particles over the course of the self-assembly process. The results suggest that lecithin played a critical role in controlling the self-assembly kinetics to form uniform amylose microparticles through steric stabilization of the growing particles and diffusion-limited growth effect. The crystallinity of amylose microparticles was not affected by lecithin, implying that lecithin did not disrupt the crystal structure within the particle and would mainly be present on the surface of the microparticles. Considering its biodegradable and biocompatible nature, the amylose-based microparticles would find a range of useful applications in the area of food, cosmetics, medicine, chromatography and other related materials sciences.