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Vitamin D supplements have been used to prevent fractures. The effect may be mediated through increased bone mass, but also through reduced falling propensity. The aim of this study was to evaluate the association between 25-hydroxy vitamin D levels (25OHD), fall-associated variables (including tests of functional performance), and fracture in ambulatory women. At baseline 25OHD was measured in 986 women. Fall-associated variables were investigated at baseline. Fractures were recorded during a 3-year follow-up. Four percent of the women had 25OHD levels below 20 ng/ml (50 nmol/l), and 26% had 25OHD levels below 30 ng/ml (75 nmol/l). 25OHD correlated with gait speed (r=0.17, P <0.001), the Romberg balance test (r=0.14, P <0.001), self-estimated activity level (r=0.15, P <0.001), and thigh muscle strength (r=0.08, P =0.02). During the 3-year follow-up, 119 out of the 986 women sustained at least one fracture. The Cox proportional hazard ratio (HR) (95% confidence interval) for sustaining a fracture during the follow-up was 2.04 (1.04-4.04) for the group of women with 25OHD below 20 ng/ml, in which 9 out of 43 women sustained a fracture. Thirty-two of the 256 women with 25OHD levels below 30 ng/ml sustained a fracture during the follow-up, with a non-significant HR of 1.07 (1.07-1.61). This cohort of elderly, ambulatory women had a high mean 25OHD. A low 25OHD was associated with inferior physical activity level, gait speed and balance. A 25OHD level below 30 ng/ml was not associated with an increased risk of fractures in this study. However, a subgroup of women with 25OHD levels below 20 ng/ml had a tendency to an increased risk of fractures, which may be associated with an inferior physical activity and postural stability.