Background As an integral part of computer-assisted surgery, virtual surgical planning (VSP) leads to significantly better surgery results, such as for oral and maxillofacial reconstruction with microvascular grafts of the fibula or iliac crest. It is performed on a 2D computer desktop screen (DS) based on preoperative medical imaging. However, in this environment, VSP is associated with shortcomings, such as a time-consuming planning process and the requirement of a learning process. Therefore, a virtual reality (VR)–based VSP application has great potential to reduce or even overcome these shortcomings due to the benefits of visuospatial vision, bimanual interaction, and full immersion. However, the efficacy of such a VR environment has not yet been investigated. Objective This study aimed to demonstrate the possible advantages of a VR environment through a substep of VSP, specifically the segmentation of the fibula (calf bone) and os coxae (hip bone), by conducting a training course in both DS and VR environments and comparing the results. Methods During the training course, 6 novices were taught how to use a software application in a DS environment (3D Slicer) and in a VR environment (Elucis) for the segmentation of the fibula and os coxae, and they were asked to carry out the maneuvers as accurately and quickly as possible. Overall, 13 fibula and 13 os coxae were segmented for each participant in both methods (VR and DS), resulting in 156 different models (78 fibula and 78 os coxae) per method (VR and DS) and 312 models in total. The individual learning processes in both environments were compared using objective criteria (time and segmentation performance) and self-reported questionnaires. The models resulting from the segmentation were compared mathematically (Hausdorff distance and Dice coefficient) and evaluated by 2 experienced radiologists in a blinded manner. Results A much faster learning curve was observed for the VR environment than the DS environment (β=.86 vs β=.25). This nearly doubled the segmentation speed (cm3/min) by the end of training, leading to a shorter time (P<.001) to reach a qualitative result. However, there was no qualitative difference between the models for VR and DS (P=.99). The VR environment was perceived by participants as more intuitive and less exhausting, and was favored over the DS environment. Conclusions The more rapid learning process and the ability to work faster in the VR environment could save time and reduce the VSP workload, providing certain advantages over the DS environment.