To explore how the stiffness index of the calcaneus is related to lifestyle factors, we examined the associations of lifestyle factors, age, and years since menopause, with stiffness index of the calcaneus among 573 community-dwelling Japanese women aged 40-89 years. Stiffness index of the calcaneus was obtained from quantitative ultrasound measurement. Body height and weight were measured, from which body mass index (BMI) was calculated. Physical activity index was calculated using a validated questionnaire. Dietary calcium intake and amount of ingested alcohol were estimated by a semiquantitative food frequency questionnaire. Current smoking status was obtained by questionnaire. In univariate analysis, stiffness index was positively related to physical activity index and BMI, and inversely to age and years since menopause, but not to dietary calcium intake. Stiffness index of drinkers and current smokers was not significantly different from that of non-drinkers and non-smokers, respectively. Among the natural menopausal women, multiple regression analysis showed that advancing age and years since menopause were associated with lower stiffness index, while higher physical activity index and greater BMI were associated with higher stiffness index. In conclusion, higher physical activity, in combination with adequate nutrition for weight maintenance, should be emphasized as a part of a healthy lifestyle in order to maintain bone health among middle-aged and elderly Japanese women.