AbstractAlthough trimetylammonium-2-hydroxypropyl-hydroxyethylcellulose (QHEC) is a well-known polysaccharide material, some of its properties have not been previously studied in detail. Therefore, we applied a combination of multidimensional nuclear magnetic resonance (NMR) in D2O on hydroxyethylcellulose (HEC) and HEC derivatives, and size-exclusion chromatography with multi-angle laser light scattering (SEC-MALS) in carbonate buffer at pH 10; while Fourier-transform infrared spectroscopy (FTIR), the X-ray diffraction (XRD), the thermogravimetry/differential/differential thermogravimetry (TG/DTG/DTA) and atomic force microscopy (AFM) in film form. SEC-MALS revealed various water-solubility of prepared derivatives: HEC (98%) > CHEC (72%) > QHEC (26%) > QCHEC (14%). Due to its substituents, the HEC macromolecule forms coil structures with varying gyration radii: QHEC (38–260 nm) > QCHEC (10–230 nm) > CHEC (21–100 nm) > HEC (23–50 nm). FTIR analysis of all prepared films confirmed their identical structure compared to that observed in liquid form in D2O. Onset temperatures (OT) of films degradation decreased in order: HEC (222 °C) > QCHEC (162 °C) > CHEC (142 °C) > QHEC (141 °C). The X-ray diffraction confirmed residual crystallinity of cellulose II (CII) in all four types of prepared films and was linked to ~ 2% water-insolubility of HEC derivatives revealed by SEC-MALS. Atomic force microscopy (AFM) showed significant differences in surface morphology among the four prepared films with surface roughness of: HEC (25 nm) > QCHEC (5.8 nm) > QHEC (4.2 nm) > CHEC (2.8 nm). Various spherical particles were found in case of HEC, circular depressions/holes approximately 4 μm in diameter were observed in case of QHEC. Except for the HEC, all other films showed granular surface probably due to insoluble components. Based on the mechanisms of quaternization and crosslinking and the results of analysis on water-soluble part and films we could assume that there are similarities in structures between the soluble and insoluble products of the reactions.