We aimed to verify whether vaginal fornix pH is related to endocervical pH and whether or not acid pH could contribute to the prophylactic treatment of vaginal infections. Four groups of healthy women with or without uterus were selected: pre-menopausal (>39 years) and post-menopausal (<61 years). Pre-menopausal non-hysterectomized women were further analysed in proliferative (8–12 days) and secretory phase (18–22 days) of the menstrual cycle. Endocervical and vaginal pH was determined by a digitally pH metro during the gynaecological examination. Vaginal pH is more acid than endocervical pH in pre- and post-menopausal women, 5.1 versus 6 and 5.4 versus 6.4 (P<0.001), respectively. Vaginal pH is related to endocervical pH in proliferative phase of menstrual cycle and post-menopausal women (P=0.009). Pre- and post-menopausal hysterectomized women presented more acid vaginal pH than non-hysterectomized women, 4.35 versus 5.1 and 4.9 versus 5.4 (P<0.05), respectively. Endocervical pH has not presented change during menstrual cycle and menopause, 6 and 6.4, respectively. Vaginal pH is related to endocervical pH in proliferative phase of menstrual cycle phase and post-menopausal women.