This study was performed to investigate whether the circadian variation in urinary pyridinium crosslinks is related to physical activity, age, the menopause, and asymptomatic osteopenia. We measured urinary pyridinoline/creatinine (Pyr/Cr) and deoxypyridinoline/creatinine (D-Pyr/Cr) in 9 healthy premenopausal women in two 27 h studies, before and at the end of 5 days of total bed rest. Both Pyr/Cr and D-Pyr/Cr showed highly significant circadian variations, with the peak at night and the nadir during the day (p < 0.001). The 5 days of complete bed rest produced no changes in the circadian pattern, but a general increase of 28% was observed in pyridinium crosslinks. A group of 12 healthy, early postmenopausal women (aged 55 +/- 2 years), 12 healthy, elderly postmenopausal women (aged 73 +/- 1 years), and 12 elderly osteopenic but otherwise healthy women (aged 73 +/- 1 years) were also studied for 27 h. All three groups showed highly significant (p < or = 0.001) circadian variations in the urinary excretion of pyridinium crosslinks. As expected, both Pyr/Cr (p < 0.05) and D-Pyr/Cr (p < 0.001) increased at the time of menopause, but the circadian variations in Pyr/Cr and D-Pyr/Cr were similar in all groups studied. We conclude that the circadian variation in the urinary excretion of pyridinium crosslinks is independent of physical factors. Furthermore, the circadian variation in pyridinium crosslinks was not related to age, menopausal status, or asymptomatic osteopenia.