The polymeric based membrane technology has been an attractive alternative option among other gas separation technologies due to its ease of operation, good efficiency and low operational cost. However, a few concerns have been expressed about the polymeric based membrane technology for GS applications, such as the permeability-selectivity trade-off, mechanical stability and its environmental impacts. Therefore, the main objective of this research is to investigate the effects of polymer concentration on a biopolymer-PVA asymmetric membrane morphology, its mechanical properties and gas transport behaviour in order to optimize it for O2/N2 separation using the non-solvent induced phase separation (NIPS) technique. For this purpose, initially, a theoretical solubility model and cloud point phase diagram analysis were conducted to study the solubility and demixing behaviour of the PVA/Water/THF ternary system. Afterwards, these membranes were prepared at different polymeric concentrations from 10 to 14 weight percent (wt.%) and then characterized using scanning electron microscopy (SEM), porosity measurement test, universal testing machine (UTM) and gas permeation test to analyse their physical structure, mechanical properties and GS performance. The analysis results showed that a lower polymer concentration of 10 wt.% facilitated larger pore sizes of 2.486 ± 1.2 μm with a higher porosity of 77.73 ± 15.26% having higher O2 permeance but lower O2/N2 selectivity and mechanical properties. Whereas, a higher polymer concentration of 12 wt.% promoted smaller pore sizes of 2.096 ± 0.5 μm with a porosity of 56.31 ± 3.6%, having better O2/N2 separation performance and higher mechanical properties. However, increasing the polymeric concentration to 14 wt.% resulted in a densified membrane structure having voids and small pore sizes of 1.447 ± 0.9 μm, with a porosity of 31.35 ± 11.98%, and lower elasticity causing membrane rigidified, making it unsuitable for GS applications. Therefore, the intermediate concentration of 12 wt.% PVA asymmetric membrane represents the most optimum morphology and mechanical properties for better O2/N2 separation.