Biofilm of Pseudomonas aeruginosa, an opportunistic pathogen, can cause serious health problems, such as chronic infections, especially in immunocompromised patients. Many studies have suggested administration of new generation of antibiotics, as P. aeruginosa biofilms have developed high resistance to antimicrobial drugs. This study reports the inhibitory effect of three medicinal plant extracts and an essential oil on biofilm formation by a clinical isolate of P. aeruginosa. In this study biofilm formation of P. aeruginosa strain 214 was determined in presence of three plant extracts, Cyclamen coum, Dianthus orieltalis and Origanum majorana, and Zataria multiflora Bio essential oil. Minimum Biofilm Inhibitory Concentrations (MBICs) were determined by microdilution techniques and XTT assay. The C. coam extract and Z. multiflora Bio essential oil inhibited biofilm formation completely at concentrations<0.062 mg/ml and 4 µl/ml, respectively. The D. orientalis and O. majorana extracts did not inhibit biofilm formation at the used concentrations (0.003 – 8 mg/ml). The results of this study indicate that some plant extracts at low concentrations may provide a complementary medication for biofilm-associated infections. Further evaluations are required to validate the antibiofilm effect of these medicinal plants.