Abstract: Examining transfersomes as a potential transdermal delivery mechanism for the poorly soluble medication was the objective of the current investigation. Ketoprofen, in order to get around the problems with taking it orally. The rotational vacuum evaporation and sonication process, along with a non-ionic surfactant (span 80) and phosphatidyl choline, were used to create several transfersomal formulations. The finished products were examined using a variety of techniques, including light microscopy, particle size analysis, turbidity, drug content, rheology, surface charge and density, penetration ability, in vitro release, ex vivo permeation, skin irritation/sensitivity study, and stability studies. We used the paw edema test paradigm to see whether the improved formulation had any anti-inflammatory effects on rats. The drug was found to be compatible with the excipients in FTIR and DSC investigations. Findings showed that all formulations effectively captured Ketoprofen with consistent drug content. Because it demonstrated the highest levels of drug entrapment (95.69±0.17%) and cumulative percent drug release (74.34±0.34), the optimal formulation (F4-S1-R4) was determined to be a transfersomal formulation consisting of 85% phosphatidyl choline of the medication and 15% span 80. We evaluated the ex vivo permeation profiles of the various transfersomal formulations with F4-S1- R4. The F4-S1-R1 transfersomal formulation outperformed the F4-S1-R2, F4-S1-R3, F4-S1-R4, and F4-S1-R5 in terms of cumulative drug permeation and flux, as well as lag time (p<0.05). From the perspective of its skin application, the transfersomal formulation F4-S1-R4, which had a lower flux, may be preferred over other formulations owing to its longer impact and greater viscosity. Better application and the absence of skin irritation were further contributing factors. As a result, the research proved that the transfersomal formulation is a viable substitute for the traditional Ketoprofen formulations that had improved transdermal penetration.Keywords: In vivo study, Ketoprofen, Permeation studies, Transdermal, Transfersomes