Display omitted •A simple and sensitive electrochemical sensor based on RGOQDs was developed by electrodeposited.•RGOQDs could enhance the electrochemical response intensity of UA, X and G with a lower detection limits of 0.024μM.•The RGOQDs/GCE was used to evaluate the cytotoxicity of fluorene on MCF-7 cells for the first time. Fluorene was one of the priority controlled polycyclic aromatic hydrocarbons (PAHs) and distributed abundantly in the environment, which has a great harmful on human body. It was very important to detect the cytotoxicity of fluorene with a rapid and accurate method. In this present work, a simple and ultrasensitive electrochemical biosensor was developed by electrodeposition of reduced graphene oxide quantum dots (RGOQDs) on glassy carbon electrode (GCE) for the first time. The electrochemical behaviors of uric acid (UA), xanthine (X) and guanine (G) were studied on RGOQDs/GCE by differential pulse voltammetry (DPV) with a limit of detection of 0.024μM, respectively. Then the electrochemical biosensor was used to evaluate the cytotoxicity of fluorene for MCF-7 cells based on the variation of G/X in MCF-7 cells solution. The result showed that fluorene exposed to MCF-7 cells at the concentration range of 0–0.05mM could promote cell proliferation, the concentration above 0.05mM exhibited inhibitory effect, which was verified by methyl-thiazolyl-tetrazolium (MTT) assay. The electrochemical biosensor based on RGOQDs/GCE could be used not only for evaluation the cytotoxicity of PAHs, but also for detection the physiological process related to intracellular purine nucleotide metabolism.