This study aimed to optimize the processing conditions to minimize the initial particle size of perfluorocarbon-based artificial oxygen carrier emulsion formulations. Previous research has shown that excessive particle size in nanoemulsions can induce cytotoxicity. Therefore, to ensure safe systemic oxygen delivery, it is crucial to design a process that achieves a particle size below 100 nm. Initially, the settings of the homogenizer and microfluidizer were adjusted, and the rotation speed (rpm) and operation time of the homogenizer, identified as key factors for particle size reduction, were fixed. Subsequently, based on relevant literature, the unit of the microfluidizer was changed from ‘cycles’ to ‘minutes’, and nanoemulsions were prepared by varying the pressure of the microfluidizer. Dynamic light scattering was used to measure the particle size, polydispersity index (PDI), and zeta potential at six time points: 5, 10, 20, 30, 40, and 60 minutes. Additionally, oxygen saturation was measured using NeoFox. The results showed that a particle size of 98.7 nm was achieved at the 60-minute point, where the microfluidizer was operated for the longest duration, confirming that the rate of particle size reduction is proportional to the processing time of the microfluidizer. Based on these findings, we plan to further analyze the stability of the emulsifier over time to evaluate the effects of long-term storage.