This study assesses one possible cause of inter-patient variation in fluorescence spectroscopy of the cervix: the menstrual cycle. Ten patients with no history of an abnormal Pap smear were seen daily throughout 30 consecutive days of their cycle. Fluorescence excitation-emission matrices were measured from three cervical sites on each patient. Principal component analysis was used to determine which spectral regions varied with the day of the cycle. Classification was performed to assess the influence of menstrual cycle on precancer diagnosis. Variations in the principal component scores and the redox ratio values show that the fluorescence emission spectra at 340–380 nm excitation appear to correlate with the cell metabolism of the cervical epithelium throughout the menstrual cycle; these changes do not affect diagnostic classification. The menstrual cycle affects intra-patient variation but does not appear to cause a significant level of inter-patient variation. It does not need to be controlled for in optical detection strategies based on fluorescence spectroscopy. © 2002 Society of Photo-Optical Instrumentation Engineers.