Two series of segmented thermoplastic polyurethanes were synthesized having 33 wt % hard segment based on 4,4'-dicyclohexyl methane diisocyanate with different trans-trans isomer contents and 1,3-propanediol chain extender. The soft segments were based on poly(hexamethylene-pentamethylene carbonate)diol and poly(butylene sebacate)diol, amorphous and semicrystalline polyol, respectively. 4,4'-Dicyclohexyl methane diisocyanate with different trans-trans isomer contents were obtained by fractional crystallization of commercial diisocyanate and were characterized by differential scanning calorimetry and nuclear magnetic resonance spectroscopy. 4,4'-Dicyclohexyl methane diisocyanate trans-trans isomer lead to some interesting properties in the synthesized polyurethanes, due to the more ordered hard domains formed by packing of trans-trans 4,4'-dicyclohexyl methane diisocyanate. Thereby, as 4,4'-dicyclohexyl methane diisocyanate trans-trans isomer content increased, a better phase separated structure was observed.