Agave tequilana and barley by-products from the Mexican food industry were characterized to determine their chemical composition and morphology by scanning electron microscopy (SEM). Cellulose was then extracted by carrying out alkali and bleaching treatments and cellulose nanocrystals (CNC) were isolated by an acid hydrolysis. Structural analyses and thermal stability of the isolated CNC were studied by X-ray diffraction (XRD) and thermogravimetric analysis (TGA), respectively. atomic force microscopy (AFM) and diffusion light scattering (DLS) were used to investigate the morphology of the CNC. The results indicate that hemicellulose and lignin were removed from the extracted cellulose. The isolated CNC from bagasse were found to be longer than those from microcrystalline cellulose (MCC), with a length of 322 ± 112 and 329 ± 123 nm for A. tequilana and barley CNC, respectively, compared to 218 ± 56 nm for CNC from MCC. The degradation temperature of A. tequilana and barley CNC was at least 30 °C higher than CNC from MCC. These values are promising for the value of by-products for the isolation of CNC and their use in reinforced-polymer manufacturing.