The urine collection sessions were divided into two groups, each collecting void by void aliquot samples in pre-acidified tubes. In addition, a 24 hour session pool was collected, which sampled 5ml aliquots and collected before, during, and after bed rest.
Medication and Dietary sessions were collected in two groups by measuring the over the counter medications including pain killers and antacids, and study medications given to the subject as required. Calcium fortified food intake was collected and analyzed, as well as, homogenate samples of each meal in the menu cycle.
How soon after the start of bed rest does a bone loss signal appear in urine and blood? All-subject average urinary Ca isotopes begin to show bone loss on the third day of bed rest. This signal persists throughout the bed rest and recovery periods and becomes statistically significant by day nine of bed rest. Ca isotopes in urine reflect the onset of negative bone mineral balance (BMB) at least ten times faster than radiological techniques like DEXA, which can detect changes in bone mass that require several months of bed rest to produce.
What factors other than BMB can affect soft tissue Ca isotope composition? Several factors other than BMB possibly could affect soft tissue Ca isotope composition. Extraneous factors like changes in dietary Ca isotope composition and differences in geographic and dietary history between subjects can be adequately controlled for under the strict conditions of bed rest studies by normalizing each subjects Ca isotope composition to his or her individual average pre-bed rest value. The former factor—changes in dietary d44/42Ca—is harder to control for, but based on the data does not seem to obscure the bone loss signal under the carefully controlled experimental conditions of bed rest, in which all subjects are fed the same menu on the same ten day rotation. The mass of Ca in soft tissues is large enough to dampen meal-to-meal variations in d44/42Ca of absorbed Ca. This damping effect is enhanced in 24-hour pooled urine samples, which for that reason are ideal for Ca isotope analysis.
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