This study investigated the significance of particle size differences in the pharmacokinetics of perfluorocarbon (PFC)-based artificial oxygen carrier (aOC) PFC-F13 nanoemulsions and explored methods to minimize the toxicity of these emulsions. Two samples with different particle sizes were used: F13-80 with a particle size of 80 nm and F13-220 with a particle size of 220 nm. Both emulsions satisfied the following criteria for animal experiments: 1) a particle size distribution below 0.300, 2) a zeta potential between -50 mV and -20 mV, 3) a pH between 5.5 and 7.5, and 4) an oxygen saturation of 8 ppm or higher. A toxicity test was conducted in 35 ICR mice via intravenous injection. Body weight and biochemical changes were measured, and tissues were collected for analysis. The results showed significant toxicity in the liver, with the spleen size increasing up to 1.3 times in the F13-220 group, indicating that multiple administrations were not feasible. However, no significant changes were observed in the kidneys or lungs compared to PBS (phosphate-buffered saline). F13-80 showed significantly less damage to the liver than F13-220, suggesting that single administration of F13-80 was possible. To reduce in vivo toxicity, the injection water DIW will be changed to 0.9% Saline in future studies while maintaining the particle size. This will further optimize the first candidate formulation of the next-generation super oxygen carrier.