A multiscale approach spanning from the segmental (subnanometer) up to micrometer level was applied for detailed study of the self-assembly of aliphatic block polyurethane (PU) elastomers. To understand the principles of the self-organization of hard and soft segments in the complex multi-component systems, several two-component model PU samples, that is, the products of 1,6-diisocyanatohexane (HDI) with three diols differing in the length and constitution were also prepared, characterized, and investigated: (i) polycarbonate-based macrodiol (MD), (ii) biodegradable oligomeric diol (DL-L; product of butane-1,4-diol and D,L-lactide), and (iii) butane-1,4-diol (BD). The study (particularly 13C-1H PILGRIM NMR spectra) reveals complex internal organization and interesting (application appealing) behavior of multi-component PUs. Hard segments (HDI+BD products) feature self-assembled and significantly folded chain conformations with interdomain spacing 15–22 nm (small-angle X-ray scattering analysis). The small domains are hierarchically assembled in various structural formations of µm size (spherulites) depending on PU composition, as detected by transmission electron microscopy and atomic force microscopy. © 2014 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2015, 132, 41590.