Passion fruit seed oil (PFSO) is rich in bioactive compounds, which can enhance laying hens' health and performance. The current study was conducted to investigate the effect of increasing PFSO supplementation in laying hens' productive performance, egg quality, relative weight and length of organs, plasma lipid oxidation, antioxidant status, and gene expression of SOD, GPx, CAT, and NRF2 in the liver. One hundred ninety-two 25-week-old Lohmann Whites were randomly divided into three treatments (n = 8 replicates/diet, 8 hens/replicate). The groups were fed a corn-soybean basal diet containing 0.00%, 0.45%, and 0.90% PFSO for 16 weeks. The results indicated that increasing supplementation of PFSO decreased plasma lipid oxidation (n = 8; linear, p = 0.012) and increased CAT gene expression (n = 8; linear, p = 0.001). SOD and NRF2 genes tended to increase linearly, and GPx was not affected (n = 4; p > 0.05). The CAT activity tended to decrease linearly and the SOD and GPx were not affected (n = 8; p > 0.05) by diets. Performance and most egg quality, relative weight, and length of organs did not differ among treatments (n = 8; p > 0.05). Therefore, increasing the supplementation of PFSO in the diet may have positive effects on the laying hens' health by decreasing oxidative stress, stimulating the antioxidant defense system, and sustaining egg production and quality.