The main macroscopic characteristics of Hydrotreated Vegetable Oil (HVO) spray in both injection and post-injection periods are investigated via computational fluid dynamics (CFD) in this research. A 2D CFD work employing the Wave breakup model and the KHRT breakup model are validated by the experimental data from a Constant Volume Vessel (CVV). Spray tip penetration and cone angle are obtained by the CFD model under various conditions, where the rail pressure, fuel temperature, ambient pressure and ambient temperature are independently varying. Results demonstrate that the Wave model has overall higher precision in predicting the spray tip penetration and the average cone angle than the KHRT model. By the End of Injection (EOI), spray tip penetration is significantly increased by increasing rail pressure and decreasing ambient pressure. While the average cone angle is larger at high ambient pressure but not sensitive to rail pressure at the cold ambient condition. The average cone angle during injection can be enlarged by high ambient temperature, especially when the rail pressure is also high. Nevertheless, spray tip penetration can only be slightly promoted by high ambient temperature. Fuel temperature has no comparable impact on spray tip penetration and cone angle during injection. In the post-injection period (after the EOI), ambient temperature becomes dominant and spray tip penetration can be reduced by either ambient temperature or fuel temperature. An empirical model is also correlated via Design of Experiments (DoE) and has high precision in predicting spray tip penetration after the breakup time.