Due to the increasing number of outbreaks of Pseudomonas infections in hospitals, it becomes necessary to set up a sanitation program for use of appropriate chemical agents in the disinfection of the hospital environment. Aim: To evaluate the activities of commonly used hospital disinfectants on multidrug-resistant biofilm-forming Pseudomonas species. Methods: Isolation of Pseudomonas spp. from the hospital environment by applying different culture media. Screening of antimicrobial sensitivity for isolated Pseudomonas spp. was done by Kirby-Bauer disc-diffusion method according to CLSI, 2017, before and after exposure to different chemical disinfectants. Detection of the ability of isolates for biofilm production before and after exposure to a different chemical agent was evaluated by Microtiter plate and Congo red method. Determination of minimum inhibitory concentration (MIC) of chemical disinfectant. Result: A total of 125 Pseudomonas spp. were isolated from different sites in the hospital environment. Antibiotic sensitivity showed resistance to Minocycline and sensitivity to Tobramycin. MICs for the disinfectants used showed that sodium hypochlorite used at a concentration between 0.312% and 0.15625% and Povidone-iodine 10 % used at a concentration of 0.625%. Antibiotic sensitivity pattern and biofilm production were changed after exposure to disinfectants as following that resistance to some antibiotics changed in to be sensitive and strong biofilm isolates converted to be non-biofilm. Conclusion: the most effective disinfectants were sodium hypochlorite 5%, Povidone-iodine 10 % at contact time after 30 minutes. It is crucial to use disinfectant at appropriate concentrations at specific contact time to overcome biofilm multidrug-resistant Pseudomonas.