A monitoring method of biofouling in reverse osmosis (RO) system was proposed based on the fluorescent signal of resorufin,which is reduced by nicotinamide adenine dinucleotide released from viable cells during aerobic respiration.The fluorescent signal of resorufin reduced by planktonic cells and microorganisms of biofilm showed linearity,indicating its feasibility to monitor biofouling in a RO system.For the application of the method to the lab-scale RO system,the injection concentration of resazurin and the injection flow rate were optimized.Biofilm on RO membranes continuously operated in a lab-scale RO system was estimated by resorufin fluorescence under optimized detection condition.As a result,resorufin fluorescence on RO membrane showed a significant increase in which the permeability of RO system decreased by 30.48％.Moreover,it represented the development of biofilm as much as conventional biofilm parameters such as adenosine triphosphate,extracellular polymeric substances,and biofilm thickness.The proposed method could be used as a sensitive and low-cost technology to monitor biofouling without autopsy of membranes.