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Cellulose nanocrystals (CNCs), also called cellulose nanowhiskers, cellulose nanorods, or nanocrystalline cellulose, were prepared from Phyllostachys heterocycla using a commercial cellulase for hydrolysis. The enzymatic hydrolysis process and application performance as well as the biocompatibility of the CNCs were investigated. Here, the cellulase hydrolysis conditions were optimized at a cellulase dosage of 0.01 mL/g dried fibers, a hydrolysis temperature of 60 °C, a hydrolysis time of 3 h, and a bamboo fiber concentration of 2 wt%. Under these conditions, the resultant CNCs retained more similarities to the original bamboo fibers than those fabricated by sulfuric acid hydrolysis. The product also demonstrated potential biocompatibility, which expands its applicability in the biopharmaceutical and biomedical fields.