Mining user’s learning preference is one of the key issues in the personalized online learning system, which is of great significance technology for modern educational. In this paper, using the hierarchical characteristics of the knowledge points in the course domain, we defined the equivalence relation and equivalence of knowledge points, and defined the structure of the knowledge points quotient space. Then, the functions of support, pheromone concentration and preference were defined on various levels, and an improved ant colony optimization was proposed to handle the multi granularity data structure of quotient space. An algorithm of multi-granularity Learning Preference Mining based on Ant Colony Optimization (ACO-LPM) was proposed to address the problems about too many learning knowledge points and too few user’s test data in the online personalized learning system. The pheromone has the characteristic of dynamic evaporation, so, the preference patterns mined by ACO-LPM can be changed with the change of user interest in real time. The experimental results show that the algorithm can mining the user’s learning preferences in online learning system effectively and efficiently.