Many countries and companies are increasingly delving into applying Virtual reality (VR) to lunar rovers. Research institutions including NASA, EAS and the China Space Center have published reports related with online control platforms based on VR technology to control an unmanned lunar rover exploring the moon surface. One of the main research directions of VR human-machine interface is Visual interaction interface. The visual interaction interface will cause visual fatigue more easily. The purpose of this paper is to explore the relationship between asthenopia, teleoperation efficiency and the color of VR interface in a simulated lunar surface environment, to make suggestions for improvement of color selection, to improve VR interface and relieve long hours operation fatigue. 30 participants were recruited and asked to use Likert scale to express. We design a teleoperation interface of unmanned lunar rover to simulate the color contrast of interface including background color and element color (Text and patterns). The hue samples of VR interface were extract from foreground color. Our task is to test user’s ability to recognize objects and environments, make real-time decisions, and control rover movements. The results show that blue and green had the weakest visual fatigue under gray background, while red had the highest visual fatigue. This study explores the visual fatigue level of different hues in the VR display of the moon environment based on the principle of visual perception levels, and finds the color and visual fatigue rules of the VR interface of teleoperating the unmanned lunar rover. The results provide a reference basis for the color level design of the VR interface of the unmanned lunar rover.