Separation of gases from air mixture is one of the most challenging and laborious separations due to the remarkably uniform molecular size of gas molecules. Therefore, the present study aimed to synthesize polyacrylonitrile-based nanofibers mat(NM) impregnated with p-diethanolaminomethylcalix[4]arene (PAN/p-DEAC4 NM) for the separation of two crucial gases O2 and N2. The affinity of the prepared PAN/p-DEAC4 NM for O2 was examined by optimizing the loading concentration of p-DEAC4 in the range from 5% to 20% (w/v). The results showed remarkable performance of the PAN/p-DEAC4 NM for O2/N2 separation with a superior O2/N2 selectivity of 12.75 and excellent permeance of 10.2 GPU for O2 and 0.8 GPU for N2 at 2 bar. The PAN/p-DEAC4 NM followed a facilitated transport mechanism for the separation of gases and it was revealed that the p-DEAC4 platform in the PAN NM is facilitating the transport of O2 due to its greater affinity towards O2. BET analysis revealed that the prepared NM possesses non-porous morphology with a surface area of 12.69 m2 g-1. SEM micrographs also confirmed the formation of defect-free NM. Thus, this study presents a unique perspective and direction for fabricating highly permeable nanofiber mats for O2/N2 separation.