Due to its renewability, availability and high cellulose content (≈45%), Alfa fibers (Stipa tenacissima) have been identified as a sustainable source for cellulose microfibers (CMF) and cellulose nanocrystals (CNC) production. Subjecting raw Alfa fibers to alkali, bleaching and sulfuric acid hydrolysis treatments allowed producing CMF and CNC with high yields. The fluorescence microscopy confirmed that CMF, with average diameter of 10 μm, were successfully obtained after bleaching treatments. TEM and AFM showed that the CNC exhibit needle-like shape with an average diameter and length of 5 ± 3 nm and 330 ± 30 nm, respectively, giving rise to an aspect ratio of about 66. XPS measurement confirmed the presence of sulfate groups on the surface of CNC with 2.04 sulfate groups per 100 anhydroglucose units, confirming the negatively charged surface of CNC, with zeta potential value of − 47.39 mV. XRD studies showed that CMF and CNC exhibit cellulose I structure with crystallinity index of 71% and 90%, respectively. FTIR and TGA analyses were used to identify the chemical composition and thermal stability changes during different chemical treatments, suggesting that all non-cellulosic compounds were removed after alkali and bleaching treatments. The obtained CNC were dispersed into three different biopolymer matrices, e.g. chitosan, alginate, and k-carrageenan, at various CNC loadings (1, 3, 5 and 8 wt%), to evaluate their ability to enhance the tensile properties of biopolymers and, at the same time, to produce new biopolymer-based nanocomposite films. It was found that the tensile properties of the as-produced nanocomposite films were largely improved with addition of CNC, resulting in mechanically strong and flexible ecofriendly nanocomposite films.