In this paper, a cellulase pretreatment was studied prior to the acid hydrolysis to decrease the total acid usage during the cellulose nano-crystals (CNC) preparation from a bleached softwood kraft pulp. Cellulase pretreatment facilitates the subsequent acid hydrolysis to produce CNC with similar quality to that of the control, but at a lower sulfuric acid concentration. The underline mechanism is that cellulase pretreatment led to the formation of more carbonyl groups which can be oxidized into carboxyl groups in the subsequent acid hydrolysis, furthermore, more hydroxyl groups are exposed, thus esterification into sulfonic groups can be enhanced. The results showed that with a cellulase dosage of 4.8 u/g (based on dry pulp) in the pretreatment stage, the sulfuric acid concentration can be decreased from 64 to 40 wt% without compromising the quality of resulting CNC particles. Other results from charge properties, Fourier transform infrared spectroscopy (FT-IR) and transmission electron microscopy (TEM) analyses also supported the conclusions.