This study examined whether menstrual cycle phase was associated with performance on the Primary Mental Abilities Test of Spatial Relations, a test of mental rotation, in undergraduate students (N = 82). As cortisol levels also vary across the menstrual cycle under conditions of stress and influence cognitive performance, saliva samples were obtained before and after the test session to examine whether cortisol levels were related to between- and within-group differences in spatial performance. Men scored higher on the spatial test than all the groups of women, although the difference between men and women in the menstrual phase was not significant. Women in the luteal phase scored lower than the menstrual, follicular, and oral contraceptive user groups of women. There were no sex or menstrual cycle differences in cortisol levels, and no association between cortisol levels and spatial performance. The poorer performance of women in the luteal phase was not related to differences in ratings of perceived stress, perceived success on the test, or mood. Although menstrual cycle phase accounted for a significant proportion of the variance (15%) in performance on the spatial test, this does not explain why men outperformed women regardless of the phase of the cycle. Thus, there are clearly several other variables, sociocultural and physiological, involved in mediating individual differences in spatial performance.