Polyurethane (PU) films are regarded as an important polymeric class in coating sectors. PU exhibits various advantageous features like coatings to shield metals and wooden objects, adhesives, sealants, and elastomeric properties in a broader perspective to boost the appearance and packaging. In the medical field, PU shows bactericidal qualities for real-world uses, but efforts are required to strengthen the antibacterial characteristics in polyurethane films. The present research is intended to develop five (05) different novel polyurethane films with excellent anti-bacterial properties. Polyurethane films were prepared using isophorone diisocyanate (IPDI) and hexamethylene diisocyanate (HDI) as aliphatic diisocyanate, 1, 4-Butanediol (BDO) as chain extender, and Polytetra-methylene glycol-1000 (PTMG-1000) and Polytetra-methylene glycol-2000 (PTMG-2000) as glycol. The reaction was conducted in a polymerization chamber using 2,2-Dimorpholinodiethylether (DMDEE) and 4–4, bioxdimorphline as a catalyst. Surface morphology and characteristics, thermal stability, tensile and viscoelastic properties were characterized using SEM, XRD, FTIR, contact angle, TMA, tensile and rheometry testing. Furthermore, an anti-bacterial test was performed to confirm PU film usage as an occlusive dressing. FTIR results of PU films based on PTMG2000 & IPDI, PTMG1000 & IPDI, and PTMG2000 & HDI showed an increase in hydrogen bonding with the intensity of the C = O band when the NCO: OH concentration was increased. But due to incompatibility of the (OH) soft segment of glycol with the hard (NCO) segment of isocyanate caused a drop in the peak strength of PU films based on PTMG2000 & HDI and PTMG1000 & IPDI. The tensile strength is trending upward for these PU samples PTMG2000 & IPDI, PTMG1000 & IPDI, and PTMG2000 & HDI. While the PTMG2000 & HDI polyurethane sample has the highest tensile strength (105 MPa) and the lowest elongation (12.2 %) at break point. (XRD and (SEM) Results showed that the crystallinity of the synthesized polyurethane films was affected by varying the NCO: OH content and this content favored the formation of ordered structure as higher peak intensities in these PU samples PTMG2000 & IPDI, PTMG1000 & IPDI, and PTMG2000 & HDI.