Polylactide acid (PLA) is a biocompatible sustainable material with notable characteristics due to its good mechanical properties and low environmental impact. The present study investigated the effects of PLA-based green membranes on gas separation and identified the best factor condition for the membrane. Prepared membranes were tested to determine oxygen (O2) and carbon dioxide (CO2) gas permeability properties. Oxygen gas permeability of the PLA/PEG/HA membrane obtained by drying for two days was increased from 100 kPa to 400 kPa; the permeability value of this membrane increased by 15%. On the other hand, the oxygen permeability value of the membrane prepared by dry for three days under the same pressure conditions and ambient temperature increased by 5%. This result indicates that the permeabilities of prepared membranes for O2 gas increase with increasing feed pressures. On the other hand, it was observed that the CO2 permeability decreased by 38.83% with the increase in pressure and drying time due to the plasticizing and swelling effect of carbon dioxide on the membrane. A factorial design was also constructed from experimental data and applied to determine the interactions of experimental parameters. All of the parameter interactions were of statistical significance for permeability. It is further argued that molecular weight has a significant positive effect on permeability, while dry time and pressure have just a slight negative effect. This study could contribute to further studies by reducing the number of tests necessary to understand the characteristics and gas separation performance of green materials.