Mental rotation is generally analyzed based on event-related potential (ERP) in a time domain with several characteristic electrodes, but neglects the whole spatial-temporal brain pattern in the cognitive process which may reflect the underlying cognitive mechanism. In this paper, we mainly proposed an approach based on microstates to examine the encoding of mental rotation from the spatial-temporal changes of EEG signals. In particular, we collected EEG data from 11 healthy subjects in a mental rotation cognitive task using 12 different stimulus pictures representing left and right hands at various rotational angles. We applied the microstate method to investigate the microstates conveyed by the event-related potential extracted from EEG data during mental rotation, and obtained four microstate modes (referred to as modes A, B, C, D, respectively). Subsequently, we defined several measures, including microstate sequences, topographical map, hemispheric lateralization, and duration of microstate, to characterize the dynamics of microstates during mental rotation. We observed that (1) the microstates sequence had a specified progressing mode, i.e., A→B→A; (2) the activation of the right parietal occipital region was stronger than that of the left parietal occipital region according to the hemispheric lateralization of the microstates mode A; and (3) the duration of the second microstates mode A showed the shorter duration in the vertical stimuli, named “angle effect”.