The rising environmental concerns have triggered film technology to find a bio-based polymer for environmental sustainability. Thus, this research provides a sustainable route for the preparation of new thermoplastic elastomer film based on non-polar natural rubber (NR) and poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (P(3HB-co-3HHx) which could be employed in film separation application. To modify the new film properties, graft copolymerization technique was employed, and benzoyl peroxide (BPO) was selected as a radical initiator. Besides that, the influence of various polymer compositions was investigated, and the physico-chemical characteristics of the NR graft P(3HB-co-3HHx) film were characterized. NR graft P(3HB-co-3HHx) exhibited homogeneous and porous nature with improved thermal stability as the grafting interaction between the polymer was achieved. Besides that, swelling test has shown the excellent resistance and affinity towards polar solvents such as ethanol, and it showcases lower affinity towards non-polar solvents as n-hexane was observed. Therefore, the NR graft P(3HB-co-3HHx) films can potentially be used for biofilm can be used for environmental remediation applications as a replacement to the synthetic polymer films. © 2023 Elsevier Ltd