Net deposition of calcium into bone requires that calcium intake is greater than obligatory calcium losses, which can occur in urine, feces, and the whole-body dermis. Women are at risk for calcium insufficiency, mainly because of inadequate intake (11). Although serum calcium concentration is tightly regulated through a vitamin D and parathyroid hormone-mediated homeostatic regulatory system, this mechanism is unable to fully compensate for low calcium intakes or high calcium losses. Obligatory losses of calcium in urine and endogenous excretion are substantial (> 200 mg·d−1). Sweat calcium loss in adults has been estimated from the difference between whole-body 47Ca retention and 47Ca excretion as an average of 63 mg·d−1 (6).
A few studies in men have reported much higher calcium losses during periods of vigorous physical activity. Sweat samples collected during practice from cotton T-shirts of basketball players during 3 d of two practices per day lasting two or more hours averaged a calcium loss of 247 mg, which was sufficient to translate to a skeletal loss of calcium (12). In 42 males undergoing fire-fighting training, sweat calcium loss averaged 107 mg per session (18). These studies in men undergoing strenuous activity do not predict calcium losses that might occur in women participating in strenuous exercise programs. These studies also did not measure total calcium balance and looked only at dermal losses from a small proportion of the total skin surface.
Several studies have included measurements of calcium losses during exercise under short-term, controlled conditions or have measured 24-h calcium losses without studying the effects of exercise on calcium losses. Arm bags were used to measure sweat calcium losses of men in environmental chambers of various temperatures for 7.5 h that included 100 min·d−1 of exercise (9). Sweat calcium losses averaged 8.1 mg·h−1 at 21°C. In a second experiment, three men participating in moderate exercise for 30 min at 23.9°C lost an average of 3 mg·h−1 calcium in sweat (9). Whole-body dermal calcium losses during 40 min of intense exercise in 16 young men ranged from 18 to 31 mg (8). We have previously reported whole-body 24-h dermal calcium losses of 103 ± 22 mg·d−1 (~4.3 mg·h−1) in six young women not participating in an exercise intervention (19). The lower calcium losses in this study compared with the studies involving exercise could be an exercise effect on dermal calcium loss, overestimates from partial body measurements compared with whole-body measurements, or a gender difference in dermal calcium loss.
The aims of the present study were to assess the effects of moderate-intensity exercise on dermal calcium losses during abstinence from exercise versus strenuous exercise in the context of overall daily calcium retention, and to determine the role of calcium supplementation on calcium homeostasis in premenopausal physically active sportswomen. We hypothesized that calcium supplementation could correct exercise-induced calcium losses.
METHODS
Subjects.
Healthy, premenopausal women (N = 26, 20-40 yr) who had regular menstrual cycles (21-35 d) were recruited through flyers posted at local exercising facilities. Aerobically trained women were included as assessed during screening who participated in more than three exercise sessions per week and who had maximal O2 uptake capacities (V˙O2peak) of > 36.7 mL·kg−1·min−1. Each woman's V˙O2peak was estimated using the YMCA multistage cycle ergometry protocol and a Monark Ergomedic bike, Model 818E. Women were included with a BMI between 18.5 and 29.9 kg·m−2. General health was assessed by a clinical chemistry profile and a screening instrument. Exclusion criteria included pregnancy, lactation, amenorrhea, unwillingness to stop dietary supplements, use of medications affecting calcium metabolism, use of prescription products for osteoporosis, known intolerance to study materials, history of renal or hepatic disease or eating disorders, or clinical laboratory values greater than or less than 10% of normal ranges.
Weight in light clothing was measured with a calibrated electronic scale and height, without shoes, was measured with a wall-mounted stadiometer. Prestudy dietary intake was determined by a 3-d diet record following instructions given by study personnel. The study protocol was approved by the Purdue University institutional review board, and all subjects provided written, informed consent.
Sample size was determined by power calculations. Power calculations were made with a univariate, one-way repeated-measures ANOVA with constant correlation design with a significance of 0.95, three treatment periods, and an assumption of correlation of 0.75 between periods for each end point. For dermal calcium loss, assuming a within-treatment standard deviation of 22.3 mg·d−1 (19) and a between-treatment standard deviation of 37.5 mg·d−1, a sample size of 21 was determined to have > 90% power to detect a difference of 15 mg·d−1 between two treatments. For calcium retention, assuming a within-treatment standard deviation of 104 mg·d−1 (22) and a between-treatment standard deviation of 506 mg, a sample size of 21 had 85% power to detect a 50-mg·d−1 difference between two treatments.
Protocol.
For at least 2 wk before the beginning of the intervention, subjects consumed one Geritol Complete daily containing 148 mg of Ca and 400 IU of vitamin D; they continued this throughout the complete study. Subjects participated in three 8-d intervention phases in a randomized-order, crossover design. The three intervention phases were 1) placebo/no exercise (control), 2) placebo/exercise, and 3) calcium supplementation/exercise. Each phase was separated by a 7- to 30-d washout period. During each intervention period, subjects consumed a controlled diet provided by the research staff. The diet consisted of a 3-d cycle menu, which provided 2400-2500 kcal, 60-65 g of protein, 25-30% kcal as fat, 400 g of carbohydrate, and 450-500 mg of calcium (including Geritol calcium). Our targeted daily calcium intake was 600 mg·d−1, but the software program's analysis of diet estimated the calcium content to be about 150 mg higher than was found when the food was analyzed. Uneaten food was returned for determination of actual calcium intake.
Calcium supplementation was provided as calcium carbonate (TUMS Ultra); each tablet contained 400 mg of elemental calcium and two tablets were taken daily with meals for a total daily supplementation of 800 mg of elemental calcium. Calcium supplements and matching placebos were provided by Glaxo Smith Kline Consumer Health Care (Parsippany, NJ). Unused tablets were returned to assess compliance. During the supplemented phase of the study, total calcium consumption was equal to approximately 1250-1350 mg·d−1.
The exercise intervention consisted of daily supervised 1-h sessions of cycling in a temperature (67-69°F) and humidity (~30%)-controlled facility on a cycle ergometer at 65-70% of heart rate reserve. This level of exercise intensity was chosen as it is within the 55-90% range of V˙O2max recommended by the American College of Sports Medicine to promote cardiorespiratory fitness in healthy adults, and 1 h was chosen because it is the upper limit recommended for cardiorespiratory health promotion (2). Intensity was monitored by heart rate using a watch dial heart rate monitor (Heart Meter Impulse, Sports Beat, Inc.) at 5-min intervals for 15 min then at 10-min intervals for 45 min. During the no-exercise phase, subjects participated in 1 h of supervised rest in the same facility. The remainder of the 24-h·d−1 period was not supervised. Subjects recorded hours and intensity of all daily exercise outside of the monitored session. During the no-exercise phase, participants were instructed to eliminate all exercise. If they could not comply with that guideline, they were to reduce both time and intensity of exercise as much as possible.
Twenty-four-hour urine and fecal collections were taken throughout the intervention phase, which ended with rising collections on day 9. The first 4 d were considered the adjustment period, and the next 4 d were used to determine calcium balance.
On day 7 of each 8-d phase, 24-h whole-body dermal calcium losses were determined as previously described (19). After the supervised exercise or rest period on day 7, subjects had a whole-body scrubdown to remove sweat, dirt, and exfoliated skin. They dressed in pretreated cotton pajamas and an external paper suiting, which covered the whole body except the head and hands. After the supervised exercise or rest period on day 8, which corresponded to 24 h in the cotton pajamas, the pajamas and rinses from a second whole-body scrubdown were collected for extraction and determination of calcium losses. This procedure includes sweat plus any dermal losses from exfoliated skin during the 24-h period from the whole body minus hands and head.
Fasting blood was drawn on days 1 and 7 for determination of serum calcium, 25-hydroxyvitamin D, 1, 25-hydroxyvitamin D, and parathyroid hormone.
Safety of the treatments was ensured by using both verbal and written query reports of adverse events. Nonpregnant status was verified at the beginning of each phase (Quick Vue ± One Step hCG Combo test, Pacific Biotech, Inc). Resting blood pressure and heart rate were measured at the beginning of each exercise session. Subjects were not allowed to exercise if blood pressure was outside of normally accepted values (90-140/50-90 mm Hg) after three consecutive measures.
Analysis.
Duplicate composites of each cycle menu, urine, and stools were processed and analyzed for calcium by inductively coupled plasma spectrophotometry (Optical Emission Spectrometer, Optima 4300 DV, Perkin Elmer, Shelton, CT) as previously described (4). Uneaten food that was returned was weighed, and the calcium content was calculated using Food Processor v. 7.4 (ESHA Research, Salem, OR). Resulting milligrams of calcium not consumed were subtracted from known dietary calcium content. Calcium retention was determined through mass balance as intake minus (dermal + fecal + creatinine adjusted urinary losses). Balance for each subject was calculated from 4 d of urine and fecal collections, but only 1 d of sweat collection during each period. All serum biochemical assays were conducted by a certified lab (Quest Diagnostics, Clinical Trials, Van Nuys, CA).
Statistical analysis.
The 95% confidence intervals were computed for the differences in dermal calcium loss through sweat and calcium retention between each pair of regimens. Least squares means and their variances were used to compute the confidence intervals. The means were obtained from a general linear model that regressed critical end-point variables on treatment, sequence, subject nested in sequence, study period, and carryover effect (SAS Institute, Inc., v. 9, Cary, NC).
RESULTS
Subject characteristics are shown in Table 1. The average V˙O2peak represents the 75th percentile for women between the ages of 20 and 29 yr (2). Habitual calcium intakes averaged almost 1000 mg·d−1.
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TABLE 1: Subject characteristics (N = 26).
Eighty-three women were screened for participation in this study. Thirty women met the screening criteria and began the first phase of the study. Subsequently, four subjects withdrew from the study because of lack of time to complete the protocol. Compliance with calcium supplementation was 100% according to pill count. Compliance with the supervised 1-h·d−1 exercise intervention was 100%. Although subjects were requested to avoid physical activity during the no-exercise period, activity logs indicated that there were no significant differences in reported activity (in METS) among the three intervention periods outside the 1-h·d−1 supervised exercise or no-exercise intervention. During days of sweat collection, subjects remained mostly sedentary while wearing the sweat collection suits, except during the 1 h of supervised exercise intervention. Diet intake compliance was adjusted by analysis of collected uneaten food. Incomplete or missed fecal or urine collections were reported, but were < 5% or < 2.5%, respectively, of total collections. No significant adverse events from the treatment were observed.
The effect of treatment on calcium losses and retention is given in Table 2. Exercise had a modest effect on whole-body 24-h dermal calcium loss. Urinary and fecal calcium excretion increased with calcium supplementation, but there was no effect of exercise on these losses. Net calcium retention was negative on low-calcium diets, but it was quite positive during the period of calcium supplementation. The increase in calcium retention with supplementation was 304 ± 225 mg·d−1.
TABLE 2: Effect of exercise and dietary calcium on calcium loss and retention in physically active, premenopausal women (N = 26).
There were no significant differences in serum PTH, 25-hydroxyvitamin D, 1,25-dihydroxyvitamin D, or calcium concentrations among the three intervention periods or change from day 1 to day 7 within each phase.
DISCUSSION
This study demonstrated that healthy, exercising premenopausal sportswomen who exercise strenuously for 1 h·d−1 have a small but significant increase in dermal calcium loss. The 13-mg additional dermal calcium loss with exercise would require consumption of an additional 40 mg·d−1 of calcium to make up for these dermal losses. The effect of this loss of calcium on bone is dependent on the calcium status of the woman. If her daily calcium intake is below the total calcium loss, then she will be in negative calcium balance and calcium will be released from bone to maintain normal serum calcium levels. In this study, participants consumed prepared diets that contained about 450 mg·d−1 of calcium, and we found that even without partaking in the exercise program, the women were in negative calcium balance. In contrast, supplementation with 800 mg·d−1 of calcium ensured sufficient calcium so that the additional loss in sweat resulting from the 1 h of exercise in addition to low calcium intakes did not put these women in negative calcium balance.
The modest dermal loss of calcium seen in our population is very similar to the less than 20 mg·d−1 reported by Lentner et al. (14), even though their population included postmenopausal women with osteoporosis who consumed about 900 mg·d−1 of calcium. Our data are similar to the findings of Rianon et al. (20), who found that dermal calcium losses were very similar for individuals when they were either at rest or exercising, with dermal losses of approximately 35 mg·d−1. The effect of exercise on dermal calcium loss in our study was less than expected on the basis of the much greater losses (> 100 mg per session) reported in men participating in rigorous exercise-that is, professional basketball practice and firefighting training (12,18). The losses were more in line with those reported in men participating in moderate exercise (8,9) at room temperature. The difference may be partly attributable to environment, because calcium losses in sweat increase with heat (9,21). Calcium losses through sweat increased from 111 mg·d−1 at 21°C to 201 mg·d−1 at 37.8°C, as estimated by arm-bag collections during 7.5-h periods that included 100 min of exercise (9). Differences in estimates of exercise-induced dermal loss may also relate to the error associated with projections using surface-area extrapolations from regional sweat collections through arm bags or patches to whole-body losses. For example, projections from arm and leg patches overestimated whole-body calcium dermal losses by three- and fourfold from patches on the upper back (19). When eight patches were used to provide a more representative sample of whole-body surface area, the error in projecting to whole-body dermal calcium losses reduced to 1.6-fold.
The effect of 1-h strenuous exercise on dermal calcium losses was too small to be reflected in the daily net calcium retention, which is largely influenced by fecal excretion. The variability in calcium retention is three to four times that of dermal loss, which greatly reduces power to observe differences. Urinary calcium excretion was not affected by exercise in our study, indicating lack of compensation for dermal losses. Urinary calcium excretion is little affected by exercise or heat (5,9).
Calcium supplementation improved calcium retention by threefold. Although urinary and fecal calcium losses increased with calcium supplementation, net calcium retention was dramatically improved. The effect of calcium supplementation on dermal calcium loss cannot be determined independently from exercise from our study design. Calcium intake from the basal diet in the groups receiving placebo was low, just under half of the adequate intake of 1000 mg·d−1 recommended for this age group by the Institute of Medicine (11). This is below the mean of U.S. intakes for women of similar age from the 1994 USDA CFS II of 647 mg·d−1 (17) and by the 1999-2000 NHANES III of 797 mg·d−1 (25). However, it is comparable with mean intake of women in many populations (15) and for 25% of U.S. women (17). The low level of calcium intake put the women in our study in negative calcium balance. When total intake was approximately 1300 mg·d−1, the women were in positive calcium balance, even with 1 h of moderate exercise. The average usual calcium intake in this group of women was near the adequate intake, but the range was broad; one fourth of the subjects consumed < 600 mg·d−1.
We have previously reported positive calcium balance in young women aged 18-31 yr of 73 ± 107 mg·d−1 on calcium intakes similar to those achieved with calcium supplementation in the present study (22). Fecal calcium losses were similar at 1061 ± 142 mg·d−1, but urinary calcium excretion was more than twice as high in that study (204 ± 73 mg·d−1). Dermal calcium losses were not measured. In that study, the subjects were not sportswomen. Sportswomen may be better able to use calcium in the diet when it is adequate. Clinical and laboratory animal studies have shown that exercise is associated with higher bone mass (16) and higher calcium absorption (13) and retention (13,26) in rats. Calcium and vitamin D supplementation decreased stress fractures by 27% relative to a placebo control group in female military recruits during basic training (Lappe, personal communication, 2006).
In this study, if we assume that the sportswomen had average total-body calcium levels of 1000 g (11), and if this relative decrement in calcium retention was sustained, it would translate into a skeletal calcium difference between the period of supplementation versus the loss seen when they were on low calcium diets of 11.7% per year. It is possible that this difference will not likely be sustained for one year as it is expected that the low calcium intakes will result in increases in calcium absorption efficiency and decreased urinary losses (23). However, full adaptation is unlikely, because calcium homeostasis does not efficiently adapt to low calcium intakes even during puberty when bone is rapidly growing (1). Calcium from supplements and calcium enriched milk have been shown to benefit bone in postmenopausal women who have usual calcium intakes of 450-750 mg·d−1 (7,10), but not in premenopausal women aged 23.1 ± 2.7 yr (3). The authors argue that failure to see a benefit of calcium supplementation on bone in their study of premenopausal women was likely attributable to the increase in calcium intake in the placebo group to 824 ± 213 mg over the course of the study, use of a calcium supplement with half the expected bioavailability, and lack of power due to attrition.
Strengths of our study include the use of a crossover design to reduce subject to subject variation (and thus, the measurement of dermal calcium and other parameters three times/subject, once in each arm of the study), a carefully controlled intervention that included the provision of all meals during the intervention and analysis of calcium content, use of a matched placebo, and rigorous whole-body 24-h dermal calcium collections. A limitation of this study is the lack of no exercise + calcium supplementation group to clarify if calcium supplementation partially offset exercise-induced dermal calcium losses. Other limitations include that dermal calcium losses were only measured once during each period, and that subjects were free living, so exercise, consumption of nonstudy food, and urine and fecal sample collections were not monitored when subjects were away from the research facility. However, participants did keep diaries, and recordings by participants of certain snacks provide some assurance of the accuracy of our analyses.
Although 1 h of strenuous exercise had modest effects on dermal calcium loss, achieving recommended dietary calcium intakes may be particularly important in physically active women. The large benefit of calcium supplementation on net calcium retention in premenopausal sportswomen observed in this study suggests that this practical lifestyle habit can correct compromised calcium balance attributable to low calcium intakes and dermal losses from exercise.
This study was funded by a grant from Glaxo Smith Kline Consumer Health Care, Parsippany, NJ. The authors acknowledge the clinical expertise and support of Dr. Adrianne Bendich from GSK. The results of the present study do not constitute endorsement of TUMS Ultra by the authors or ACSM.
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