In order to investigate the bactericidal conditions of cold atmospheric plasma (CAP) on Pseudomonas fluorescen in modified atmosphere packaging (MAP) and the bactericidal mechanism of CAP in MAP， three factor and three levels orthogonal test was used to optimize the conditions of CAP treatment. The inhibitory mechanism of CAP against Pseudomonas fluorescens was studied by cell activity， cell wall permeability， cell membrane integrity， antioxidant enzyme activity and microstructure. The results showed that the best sterilization conditions were as follows: Treatment voltage as 50 kV， treatment time as 180 s， modified atmosphere volume ratio as V(N2)∶V(CO₂) = 3∶2， under these conditions， the sterilization rate was 99.49%. The inhibitory mechanism of CAP on Pseudomonas fluorescens showed that cold atmospheric plasma treatment reduced cell activity. The integrity of cell wall and cell membrane was damaged， and the leakage of nucleic acid and protein increased， Na+/K+-ATPase (ATP) content decreased by 13.10 U/mg prot， Propidium iodide (PI) increased， and alkaline phosphatase(AKP) activity decreased to 12.09 King unit/g prot. The activities of antioxidant enzymes superoxide dismutase(SOD) and catalase (CAT) in cells decreased. It changes the shape of the cells， making them stick and sag. In conclusion， cold atmospheric plasma combined with modified atmosphere packaging could effectively inhibit Pseudomonas fluorescens， resulting in cell inactivation or death， and the longer the treatment time， the better the effect.