Fluorescent carbon quantum dots (CQDs) have been emerged in countless applications due to their interesting optical, surface properties, and the simplicity of their synthetic routes. On the other hand, pyridine is toxic that could inter into the human living environment causing many health problems, e.g. asthmatic breathing and laryngitis. Here, water-soluble 2.75 nm size N-doped CQDs have been synthesized from folic acid and used as chemo-sensor for the determination of pyridine. It was found that pyridine can quench the fluorescence of N-CQDs effectively. The quenching is explained in terms of Stern–Volmer relation and confirmed by fluorescence lifetime measurements to have a dynamic character with a predetermined electron transfer. The detection limit for pyridine was found to be 18 nM. To the best of our knowledge, such a small detection limit is more than three orders of magnitude smaller than that obtained so far for pyridine detection using any other facile methods.