This research article deals with the determination of optimal conditions of extraction parameters (e.g. temperature (60-100 degrees C), pressure (200-350 bar), particle size (0.5-1.0 mm), flow rate-CO2 (5-15 g/min), and the % of co-solvent (0.0-10% of flow rate-CO2) resulting to the optimal cumulative extraction yield during the supercritical fluid extraction of Argemone mexicana (L.) seed oil with and without a modifier (ethanol) using a supercritical carbon dioxide as solvent. A "five-factors-three-levels" Box-Behnken design under the response surface methodology was used to show independent and interactive effects of extraction parameters. A mathematical regression model was expressed properly by a quadratic second-order polynomial equation. The maximum oil yield (42.86%) from A. mexicana seeds was obtained with the optimal conditions (85 degrees C, 305 bar, 0.75 mm, 11 g/min, and 9% of flow rate-CO2) of extraction parameters. The fatty acids analysis of the seed oil was done using gas chromatography and found its suitability as bio-fuel.