Federal Aviation Administration (FAA)’s 20 years of research and development with 200 unleaded blends and full-scale engine tests on 45 high-octane unleaded blends has not found a “drop-in” unleaded replacement for aviation gasoline (AVGAS) 100 low lead (100LL) fuel. In this study, analysis of compatibility via optimization of Lycoming O-320 engine fuelled with RON 97, RON 98, RON 100, and AVGAS was conducted using the Response Surface Methodology (RSM). Test fuels were compositionally characterized based on Gas Chromatography (GC) analysis and were categorized based on types of Hydrocarbon (HC). Basic fuel properties of fuels in this research were analyzed and recorded. For optimization analysis, engine speed and fuel were considered as the input parameters. The output responses were Brake Horsepower (BHP), Brake Thermal Efficiency (BTHE), Brake-Specific Fuel Consumption (BSFC), Exhaust Gas Temperature (EGT), Carbon Dioxide (CO2), Carbon Monoxide (CO), HC, and Nitrogen Oxides (NOx). The engine speed (RPM) was varied at 2000-2700, and the fuels were varied at four (04) levels, RON 97, RON 98, RON 100, and AVGAS. The design matrix was selected based on one factor of RSM with 28 experimental runs. Analysis of Variance (ANOVA) was performed on the models. Values of “Prob > F” less than 0.05, differences between “predicted R2” and “adjusted R2” of less than 0.2, and “Adequate Precision” ratios greater than 4 were used to validate the significance of the model tested. Desirability approach was applied to measure the desirability function. Input parameters, engine speed and type of fuel set to be in range, BHP and BTHE were maximized while BSFC, EGT, and all emission responses were minimized. To confirm that the model can predict actual outcomes at the optimal settings determined from the analysis, confirmation test was carried out. Results indicated that when the engine was run with a speed of 2279.064 RPM, RON 97 fuel gave optimum solution of all tested fuels, and the corresponding values of BHP, BTHE, BSFC, EGT, CO2, CO, HC, and NOx were found to be 146.669 HP, 27.7%, 0.270 Kg/kW-hr, 382.008°C, 7.162%, 7.201%, 199.460 ppm, and 51.296 ppm, respectively, with a desirability index of 0.755. Results of this study indicate that lower octane fuels are favorable in this type of engine with lower compression ratio (CR). Matching engine design and fuel octane rating plays a significant and dominant role in the performance and exhaust emission.