Effective control of biofouling is necessary for sustainable application of reverse osmosis and nanofiltration in seawater desalination as well as water and wastewater treatment. In this study, the effectiveness and mechanisms of d-tyrosine in controlling biofouling of a polyamide nanofiltration (NF) membrane by a model biofilm bacterium, Pseudomonas aeruginosa, were investigated. The impact of d-tyrosine on cell attachment and biofilm formation of P. aeruginosa was studied in both batch and continuous flow filtration systems. d-tyrosine was found to strongly inhibit P. aeruginosa attachment and biofilm formation on the NF membrane without impacting bacterial growth. When continuously supplemented to the membrane feed water in a bench scale NF system, d-tyrosine prevented irreversible biofouling by prohibiting adhesion and biofilm formation of P. aeruginosa on the NF membrane. Such inhibitive effect, however, may not be reflected in the membrane flux behavior during one single filtration cycle when the bacterial concentration in the system is high due to the dominant effect of cake enhanced osmotic pressure. The effective concentration of d-tyrosine was as low as 3 μM, although the effectiveness of biofouling control by d-tyrosine strongly depended on the ratio of d-amino acid concentration to bacterial cell number.