J Nutr Health Aging
THE JOURNAL OF NUTRITION, HEALTH & AGING©
DO CALCIUM SUPPLEMENTS INCREASE SERUM AND URINE CALCIUM LEVELS IN POST-MENOPAUSAL WOMEN? M.N. SAMOZAI, A.K. KULKARNI Department of Physiology, MediCiti Institute of Medical Sciences, Ghanpur, Medchal Mandal, R. R. Dist., Hyderabad – 501 401, A.P., India. Corresponding author: A.K. Kulkarni, Department of Physiology, MediCiti Institute of Medical Sciences, Ghanpur, Medchal Mandal, R. R. Dist., Hyderabad – 501401, Andhra Pradesh, India; Telephone: +91-9849845967 Email: [email protected]; [email protected]
Abstract: Objectives: The frequent prescription of calcium supplements with vitamin D by health practitioners is a topic of concern globally. The present study was designed to find out whether the calcium supplements with vitamin D really affect serum and urinary calcium levels in post-menopausal women. Design, Settings & Participants: The age-matched comparative study was performed among postmenopausal women who were already on and those who were not on calcium supplements with vitamin D for a period of time in relation to the estimation of the serum calcium and the urine calcium levels. Sixty healthy postmenopausal women were enrolled, with thirty among them forming the study group (SG) - who were on calcium supplements with vitamin D for a period of a month, three months, and up to twelve months - were studied and compared the results obtained with the age-matched control group (CG) of thirty post–menopausal women who were not on calcium supplements. Measurements: The serum and the urinary calcium levels were estimated by using appropriate biochemical methods and the data were analysed using relevant statistical methods. Results: The serum calcium levels did not vary significantly in SG in spite of consuming calcium supplements over a period of time, whereas the urinary calcium levels increased progressively (p value < 0.005) in those who have taken calcium supplements for a year compared to those who have taken for a month. Conclusion: Thus, the calcium supplements were of little significance on the serum calcium levels, but have a significant effect on the urinary calcium levels in post-menopausal women. Key words: Calcium supplements, serum calcium, urine calcium, post-menopausal women.
Introduction
(BMD) (5-7), and therefore the sufficient quantity of calcium need to be obtained in postmenopausal women to preserve the bone health (8). Recently there has been an increase in the prescription of calcium, and consequently the increased abuse of calcium as therapeutic agents has been implicated to many complications such as increase of gall stones, renal stone, increased incidence of myocardial infarction and various fibrotic conditions(9). Ordinary calcium supplements taken by the old age people to strengthen bones may boost the risk of heart attacks, suggesting for reconsideration of the role of calcium in the treatment of osteoporosis. Calcium tablets have been commonly prescribed to boost skeletal health, but recent studies (10-11) suggested that they might increase the rates of myocardial infarction and cardiovascular events in healthy older women. Dr John Cleland of the University of Hull and colleagues in Britain (12) pointed out that regardless of possible impacts on heart attack rates; calcium supplements were probably not very efficient in reducing fractures in any case. In community-dwelling postmenopausal women with no symptoms of bone disease such as osteoporosis, daily supplementation with 1,000 mg or less of calcium and 400 IU or less of vitamin D did not help to prevent bone fractures (13). Calcium balance has also been reported to be significantly influenced by the changes in the quantity of calcium excreted in the urine (14) and studies (15, 16) reported the association of urinary calcium with intake of calcium and hence the urinary calcium can be utilized as an index of calcium intake. Though there have been studies on intestinal calcium absorption,
Calcium plays a vital function in many cellular processes, including hormone secretion, muscle contraction, nerve conduction, exocytosis and activation and inactivation of many enzymes (1). Calcium also serves as an intracellular second messenger by carrying information from the cell membrane into the interior of the cell. It is, therefore, not surprising that the body very carefully regulates the plasma concentration of free ionized calcium (the physiologically active form of the ion) and maintains plasma calcium within normal range between (1.0 and 1.3 mM) or (4.0 – 5.2 mg/dL). Calcium is one of the principle components of hydroxyl apatite crystals [Ca10(PO4) (OH)2], which constitute by far the major portion of the mineral phase of bone (2). Not all calcium consumed is actually absorbed in the gut. The humans consume about 30% of the calcium in foods, but this varies depending upon the type of food consumed (3). Other factors also affect calcium absorption, including (i) the amount consumed: the efficiency of absorption decreases as calcium intake increases, and (ii) age and life stage: net calcium absorption is as high as 60% in infants and young children, who need substantial amounts of the mineral to build bone (3, 4). On account of the concern in the community about calcium deficiency in their bone health, there has been an increased usage of calcium supplements by the postmenopausal women. Particularly, the low dietary intake of calcium in elderly women has been reported to be reducing the bone mineral density Received April 17, 2014 Accepted for publication September 1, 2014
1
J Nutr Health Aging
CALCIUM SUPPLEMENTATION IN POST-MENOPAUSAL WOMEN calcium metabolism, and calcium balance, the data lacks a simple, observational comparative study on calcium supplementation in postmenopausal women. Hence, we have performed an age-matched comparative study of the calcium supplementation in relation to the serum calcium and the urinary calcium levels among the supplemented and nonsupplemented postmenopausal women. This study is an observational comparative study, not time consuming, with no follow-up, only with estimation of serum and urine calcium levels, and the factors like vitamin D and PTH hormonal assays, serum estrogen levels being the limitations of the present study. Materials and Methods Subjects The subjects enrolled in the study were postmenopausal women attending the out-patient block, Orthopaedics, Osmania General Hospital, Afzal Gunj Hyderabad, Andhra Pradesh, India. The participants for this study were chosen from the OP register who were already on calcium supplements with vitamin D. They took supplements from the hospital pharmacy (dicalciium phosphate with vitamin D). The subjects for this study were 60 (30 study group and 30 control group) postmenopausal women aged between 48-60 years old. Women with a history of diseases such as diabetes, hypertension, malignancies, and renal diseases were ineligible as being those with any other medical conditions that affect calcium metabolism. Care has been taken to select women who were not on hormone replacement therapy, not with bone fractures, and not had been the regular users of anabolic steroids, glucocorticoids, anticonvulsants or any other drugs known to affect calcium metabolism within the past twelve months. The healthy postmenopausal women were recruited based on history and other routine clinical investigations and assigned to calcium study group (SG) on calcium supplementation and the control group (CG) not on calcium supplementation. The SG consisted of thirty postmenopausal women, those who were already on calcium supplements with vitamin D for a month, up to three months, and up to 12 months (dosage: each uncoated tabletdicalcium phosphate IP- 0.5 gm; vitamin D3 IP – 500 international units; 2 tablets OD). Each subject in SG had received the same supplement dosage from the hospital pharmacy. While the age-matched control group (CG) of thirty subjects were not given any placebo and advised to continue with their routine diet and lifestyle.
Data Collection The subjects were assigned using our own structured validated questionnaire at the start of the study. Written consent has been taken from all the subjects after explaining them about the purpose and use of study. The study protocol was approved by the independent local ethical committee. Sample collection The blood and urine samples were collected from both the SG and the CG for estimation of the serum calcium and the urine calcium. The subjects were studied while consuming their usual diet without specific dietary recommendations, but were asked to keep on fasting from 9 pm the day before sample collection. The blood samples and 24 hour urine samples were collected the following morning at 8 am in clean, sterile dry containers, and stored at -20 °C. Estimation of Serum Calcium Levels Serum levels of calcium were measured by using calcium-kit method (17-19) (calorimetric method using Arsenazo III), which was based on the principle that, calcium with Arsenazo III at neutral pH yields a blue coloured complex. The intensity of the colour formed was directly proportional to calcium concentrations. Estimation of Urinary Calcium Levels The 24 hourly collected urine samples were assigned for estimation of urinary calcium by using calorimeter based calcium-kit method by using Arsenazo III, which was based on the principle that, calcium with Arsenazo III at neutral pH yields a blue coloured complex. The intensity of the colour formed was directly proportional to calcium concentrations. Statistical Methods The statistical analyses were performed by comparing the data in various groups of SG and CG and the measures of serum calcium and urinary calcium were performed by using analysis of variance for multiple measurements. The biostatistical evaluation was carried out using the statistical package, SPSS version 18.0 (PASW statistics) for windows, (SPSS, Inc.; Chicago, II). Results and Discussion The laboratory values, based on the duration of calcium intake, obtained for the serum calcium and the urinary calcium of the subjects are presented in Table 1a for the study group and in Table 1b for the control group. The relative data for serum calcium and urine calcium among the study group and the control groups are presented in Table 2. The mean and the standard deviation values for the serum calcium in both the control group and the study group exhibited little variation. There was also little variation in values of serum calcium within the study group itself in spite of supplementation for the duration of more than 3 months. The results for analysis of
Ethics Approval Ethical approval was received from the local ethical committee of Osmania Medical College, Hyderabad, prior to commencement. Informed written consent was obtained from each participant prior to participating in the study.
2
J Nutr Health Aging
THE JOURNAL OF NUTRITION, HEALTH & AGING© variance (Table 3) showed little significance for serum calcium in both SG and CG. In the case of the study group, even though there was a constant supply of the same quantity of calcium, there was no substantial change of serum calcium. Therefore, it is suggested that in the study group and the control group of the post-menopausal women in the same age and with the same inclusion and exclusion criteria, the effects of hormones on calcium metabolism is presumed to be the same. Such a view is consistent with a reported study (20) that, no obvious symptoms were brought about in the short term with inadequate intakes of dietary calcium from food and supplements, as the circulating blood levels of calcium were tightly regulated and did not fluctuate with little alterations in dietary calcium intakes. The similar presumed hormonal effects on calcium metabolism from our data also corroborate the fact (21) that with respect to calcium, the homeostatic system was sufficiently flexible to maintain blood ionized calcium within the normal range without being interrupted by wide fluctuations in dietary calcium intake and changing rates of bone mineralization.
Table 1b Laboratory Findings of Control Group (without Calcium Supplementation)
Control Group
Table 1a Laboratory Findings of Study Group (with Calcium Supplementation) based on Duration of Calcium Intake Duration of Calcium Intake (months)
Age
Serum Calcium (mg/dl)
Urine Calcium (mg/day)
54 50 49 50 51 60 50 52 60 49 50 55 48 50 55 49 51 49 52 50 51 49 52 49 49 52 50 52 50 50
9.1 8.2 9 8.3 8.6 8.3 8.5 8.4 8.3 8.9 8.4 9 8.4 8.4 8.6 8.9 8.3 8.3 8.9 8.3 8.9 8.4 8.0 9.6 9.4 8.5 8.9 8.3 9.0 9.1 8.64± 0.38
220 172 150 129 84 75 150 150 100 180 130 155 175 150 130 210 185 200 180 170 200 200 252 250 245 225 225 250 220 230 179.73± 48.20
Study Group
Up to 1 month
Up to 3 months
Up to 12 months
Average ± SD
Average ± SD
Age
Serum Calcium (mg/dl)
Urine Calcium (mg/day)
50 48 49 49 50 60 52 49 50 49 52 54 60 49 50 48 52 50 49 50 52 50 60 58 50 48 52 51 49 51
8.5 8.9 9.3 9 9 8.5 8.4 8.8 9.1 9 8.5 8.9 8.4 9 8.5 8.9 8.5 8.4 8.5 9 8.6 8.5 8.3 8.2 8.3 9 8.9 8.6 9 8.6 8.7 ± 0.3
120 119 110 150 130 100 125 120 120 155 120 130 110 150 130 125 130 140 135 150 140 130 120 125 130 120 140 150 150 140 130±14
The urine calcium levels progressively increased in the study group as compared to those in the control group. Even in the study group, the urine calcium levels were high in those subjects who have consumed calcium supplements for twelve months. The analysis of variance results (Table 3) also showed a statistical significance (p < 0.05) for urinary calcium in SG. In the case of CG, there was no statistically significant variation of urinary calcium. However, only two subjects among the thirty of the SG exhibited the low levels of urinary calcium (Table 1a). This finding of urinary calcium only in two subjects may not be of much significance to the overall study. We really didn’t understand the low levels of the urinary calcium in these two patients. This might probably be due to that they might not have taken the calcium supplements, or could be due to the influence of PTH and Vit D. There was a steady increase in urinary calcium levels corresponding to the duration of intake of calcium supplements (i.e. more urinary calcium levels in subjects taking calcium supplements for more than three months compared to those taking for one month). These increased levels in urinary 3
J Nutr Health Aging
CALCIUM SUPPLEMENTATION IN POST-MENOPAUSAL WOMEN Table 2 Comparison of duration of intake of calcium between control and the subject groups Control Group (No Calcium Supplementation)
Age Serum Calcium (mg/dl) 24 hrs Urinary calcium (mg/dl)
Study Group (Calcium Supplementation)
Mean
Standard Deviation
51 8.7 130
4 0.3 14
1 month Mean Standard Deviation 52 8.6 143
4 0.3 38
Duration of intake of calcium 3 months Mean Standard Deviation 50 8.6 192
12 months Mean Standard Deviation
1 0.3 14
51 8.9 237
1 0.5 13
Table 3 Mean (SEM) values of serum calcium and urinary calcium measures, and results of analyses of variance Control Group (No Calcium Supplementation)
Mean
Serum Calcium (mg/dl)
24 hrs urinary calcium (mg/dl)
Study Group (Calcium Supplementation)
Standard Deviation
Between Groups Within Groups Total Between Groups Within Groups Total
1 month Mean Standard Deviation
Duration of intake of calcium 3 months Mean Standard Deviation
12 months Mean Standard Deviation
Sum of Squares
df
Mean Square
F*
Sig.
0.542 6.380 6.922 83710.9 27974.5 111685.4
3 56 59 3 56 59
0.181 0.114
1.585
0.203
27903.6 499.5
55.9
0.000
*Asymptotically F distributed
excretion of calcium may indirectly reflect the rise of the renal threshold for excretion. As the urine sample was collected after 24 hours of intake of last dose of calcium supplements, the amount of absorbed calcium and later spilling in urine is questionable, giving way to various complications such as deposition in the soft tissues, hence enabling the scope for curbing the indiscriminate use of calcium supplements in postmenopausal women.
Acknowledgements: The authors thank to Prof. B. Ram Reddy and Prof. Ch. N. Rajkumari of Osmania Medical College, Hyderabad, Andhra Pradesh, India for their valuable suggestions and advice for the present work. Conflict of Interest: The authors declare they have no conflict of interest. Financial support: This research work received no specific grant from any funding agency, commercial or not-for-profit sectors, and had not involved any sponsors Statement of authorship: MNS initiated and designed the study, participated in the data acquisition, contributed in statistical analysis and in reviewing the manuscript. AKK performed the statistical analysis, conceived the study, in its design and coordination, drafted the manuscript and edited the manuscript. All authors have read and approved the final manuscript.
Conclusion
References
There was no significant change in serum calcium levels in the study group even after consumption of calcium supplements for a period of one year, whereas the urine calcium levels progressively increased only in the study group and not in the control group. This indicates prolonged calcium supplementation has little effect on serum calcium levels but significantly affecting the urinary calcium levels in postmenopausal women.
1. 2.
3.
4.
4
Nordin BEC. Calcium and osteoporosis. Nutrition 1997;13: 664-686. Eugene , Paula B. The Parathyroid glands and vitamin D. In: Boron WF, Boulpaep E (eds). Medical Physiology, 2nd edn. W B Saunders Publishing, Philadelphia, 2003;pp 1094-1103. Committee to Review Dietary Reference Intakes for Vitamin D and Calcium, Food and Nutrition Board, Institute of Medicine. Dietary Reference Intakes for Calcium and Vitamin D. Academic Press, Washington DC, 2010. National Institutes of Health. Optimal calcium intake: NIH Consensus Statement. 1994;12: 1-31.
J Nutr Health Aging
THE JOURNAL OF NUTRITION, HEALTH & AGING© 5.
6.
7.
8.
9.
10. 11.
12.
Dawson-Hudges B, Dallal GE, Krall EA, Sadowski L, Sahyoun N, Tannenbaum S. A controlled trial of the effect of calcium supplementation on bone density in postmenopausal women. N Engl J Med 1990;323: 878-883. Prince RL, Devine A, Dick I et al. The effects of calcium supplementation (milk powder or tablets) and exercise on bone density in postmenopausal women. J Bone Min Res 1995;10: 1068-1075. Reid IR, Ames RW, Evans MC, Gamble GD, Sharpe SJ. Long-term effects of calcium supplementation on bone loss and fractures in postmenopausal women – a randomized controlled trial. Am J Med 1995;98: 331-335. McKane WR, Khosla S, Egan KS, Robins SP, Burritt MF, Riggs BL. Role of calcium in modulating age-related increases in parathyroid function and bone resorption. J Clin Endocrin Metab 1996;81:1699-1703. Bolland MJ, Avenell A, Baron JA, Grey A, MacLennan GS, Gamble GD et al. Effect of calcium supplements on the risk of myocardial infarction and cardiovascular events: meta-analysis. Brit Med J 2010;341: c3691 doi:10.1136/bmj.c3691. Reid IR, Mason B, Horne A, Ames R, Reid HE, Bava U et al. Randomized controlled trial of calcium in healthy older women. Am J Med 2006;119: 777-785. Bolland MJ, Barber PA, Doughty RN, Mason B, Horne A, Ames R et al. Vascular events in healthy older women receiving calcium supplementation: randomized controlled trial. Brit Med J 2008;336: 262-266. doi: 10.1136/bmj. Cleland JGF, Klaus W, Sue S. Calcium supplements in people with osteoporosis (2010) Brit Med J 2010;341: c3856. doi: http://dx.doi.org/10.1136/bmj.c3856.
13.
14. 15.
16.
17. 18. 19. 20.
21.
5
Moyer VA. Vitamin D and calcium supplementation to prevent fractures in adults: U.S. Preventive Services Task Force recommendation statement. Ann Int Med 2013;158: 691-696. doi:10.7326/0003-4819-158-9-201305070-00603. Heaney RP. Calcium. In: Bilezikian JP, Raisz GA, Rodan GA (eds). Principles of Bone Biology. Academic Press, New York, 1996;pp 1007-1018 Yoshida M, Fukuwatari T, Sakai J, Tsuji T, Shibanda K. Correlation between mineral intake and urinary excretion in free-living Japanese young women. Food Nutr Sci 2012;3: 123-128 doi:10.4236/fns.2012.31018]. Heinrich T, Heidt H, Hafner V, Schmidt-Gayk H, Jenetzky E, Walter-Sack I et al. Calcium load during the administration of calcium carbonate or sevelamer in individuals with normal renal function. Neph Dial Transpl 2008;23, 2861-2867. doi: 10.1093/ndt/gfn151. Smith HG Jr, Baucer PJ. Light-induced permeability changes in sonicated bovine disks: arsenazo III and flow system measurements. Biochem 1979;18: 5067-5073. Budesinky B. Analyt Chem. Chelñates 1969;18: 5067-5073. Cadwell PC. Calcium chelation buffers. In: Cuthbert AW (ed). Calcium and Cellular Function, Mackmillan Publishing Co Inc, London, 1970;pp 10-16. Dawson-Hughes B, Harris SS, Palermo NJ, Castaneda-Sceppa C, Rasmussen HM, Dallal GE. Treatment with potassium bicarbonate lowers calcium excretion and bone resorption in older men and women. J Clin Endocrin Metab 2009;94: 96-102. Goldman's Cecil Medicine. Elsevier, Philadelphia, 2007.
THE JOURNAL OF NUTRITION, HEALTH & AGING©
DO CALCIUM SUPPLEMENTS INCREASE SERUM AND URINE CALCIUM LEVELS IN POST-MENOPAUSAL WOMEN? M.N. SAMOZAI, A.K. KULKARNI Department of Physiology, MediCiti Institute of Medical Sciences, Ghanpur, Medchal Mandal, R. R. Dist., Hyderabad – 501 401, A.P., India. Corresponding author: A.K. Kulkarni, Department of Physiology, MediCiti Institute of Medical Sciences, Ghanpur, Medchal Mandal, R. R. Dist., Hyderabad – 501401, Andhra Pradesh, India; Telephone: +91-9849845967 Email: [email protected]; [email protected]
Abstract: Objectives: The frequent prescription of calcium supplements with vitamin D by health practitioners is a topic of concern globally. The present study was designed to find out whether the calcium supplements with vitamin D really affect serum and urinary calcium levels in post-menopausal women. Design, Settings & Participants: The age-matched comparative study was performed among postmenopausal women who were already on and those who were not on calcium supplements with vitamin D for a period of time in relation to the estimation of the serum calcium and the urine calcium levels. Sixty healthy postmenopausal women were enrolled, with thirty among them forming the study group (SG) - who were on calcium supplements with vitamin D for a period of a month, three months, and up to twelve months - were studied and compared the results obtained with the age-matched control group (CG) of thirty post–menopausal women who were not on calcium supplements. Measurements: The serum and the urinary calcium levels were estimated by using appropriate biochemical methods and the data were analysed using relevant statistical methods. Results: The serum calcium levels did not vary significantly in SG in spite of consuming calcium supplements over a period of time, whereas the urinary calcium levels increased progressively (p value < 0.005) in those who have taken calcium supplements for a year compared to those who have taken for a month. Conclusion: Thus, the calcium supplements were of little significance on the serum calcium levels, but have a significant effect on the urinary calcium levels in post-menopausal women. Key words: Calcium supplements, serum calcium, urine calcium, post-menopausal women.
Introduction
(BMD) (5-7), and therefore the sufficient quantity of calcium need to be obtained in postmenopausal women to preserve the bone health (8). Recently there has been an increase in the prescription of calcium, and consequently the increased abuse of calcium as therapeutic agents has been implicated to many complications such as increase of gall stones, renal stone, increased incidence of myocardial infarction and various fibrotic conditions(9). Ordinary calcium supplements taken by the old age people to strengthen bones may boost the risk of heart attacks, suggesting for reconsideration of the role of calcium in the treatment of osteoporosis. Calcium tablets have been commonly prescribed to boost skeletal health, but recent studies (10-11) suggested that they might increase the rates of myocardial infarction and cardiovascular events in healthy older women. Dr John Cleland of the University of Hull and colleagues in Britain (12) pointed out that regardless of possible impacts on heart attack rates; calcium supplements were probably not very efficient in reducing fractures in any case. In community-dwelling postmenopausal women with no symptoms of bone disease such as osteoporosis, daily supplementation with 1,000 mg or less of calcium and 400 IU or less of vitamin D did not help to prevent bone fractures (13). Calcium balance has also been reported to be significantly influenced by the changes in the quantity of calcium excreted in the urine (14) and studies (15, 16) reported the association of urinary calcium with intake of calcium and hence the urinary calcium can be utilized as an index of calcium intake. Though there have been studies on intestinal calcium absorption,
Calcium plays a vital function in many cellular processes, including hormone secretion, muscle contraction, nerve conduction, exocytosis and activation and inactivation of many enzymes (1). Calcium also serves as an intracellular second messenger by carrying information from the cell membrane into the interior of the cell. It is, therefore, not surprising that the body very carefully regulates the plasma concentration of free ionized calcium (the physiologically active form of the ion) and maintains plasma calcium within normal range between (1.0 and 1.3 mM) or (4.0 – 5.2 mg/dL). Calcium is one of the principle components of hydroxyl apatite crystals [Ca10(PO4) (OH)2], which constitute by far the major portion of the mineral phase of bone (2). Not all calcium consumed is actually absorbed in the gut. The humans consume about 30% of the calcium in foods, but this varies depending upon the type of food consumed (3). Other factors also affect calcium absorption, including (i) the amount consumed: the efficiency of absorption decreases as calcium intake increases, and (ii) age and life stage: net calcium absorption is as high as 60% in infants and young children, who need substantial amounts of the mineral to build bone (3, 4). On account of the concern in the community about calcium deficiency in their bone health, there has been an increased usage of calcium supplements by the postmenopausal women. Particularly, the low dietary intake of calcium in elderly women has been reported to be reducing the bone mineral density Received April 17, 2014 Accepted for publication September 1, 2014
1
J Nutr Health Aging
CALCIUM SUPPLEMENTATION IN POST-MENOPAUSAL WOMEN calcium metabolism, and calcium balance, the data lacks a simple, observational comparative study on calcium supplementation in postmenopausal women. Hence, we have performed an age-matched comparative study of the calcium supplementation in relation to the serum calcium and the urinary calcium levels among the supplemented and nonsupplemented postmenopausal women. This study is an observational comparative study, not time consuming, with no follow-up, only with estimation of serum and urine calcium levels, and the factors like vitamin D and PTH hormonal assays, serum estrogen levels being the limitations of the present study. Materials and Methods Subjects The subjects enrolled in the study were postmenopausal women attending the out-patient block, Orthopaedics, Osmania General Hospital, Afzal Gunj Hyderabad, Andhra Pradesh, India. The participants for this study were chosen from the OP register who were already on calcium supplements with vitamin D. They took supplements from the hospital pharmacy (dicalciium phosphate with vitamin D). The subjects for this study were 60 (30 study group and 30 control group) postmenopausal women aged between 48-60 years old. Women with a history of diseases such as diabetes, hypertension, malignancies, and renal diseases were ineligible as being those with any other medical conditions that affect calcium metabolism. Care has been taken to select women who were not on hormone replacement therapy, not with bone fractures, and not had been the regular users of anabolic steroids, glucocorticoids, anticonvulsants or any other drugs known to affect calcium metabolism within the past twelve months. The healthy postmenopausal women were recruited based on history and other routine clinical investigations and assigned to calcium study group (SG) on calcium supplementation and the control group (CG) not on calcium supplementation. The SG consisted of thirty postmenopausal women, those who were already on calcium supplements with vitamin D for a month, up to three months, and up to 12 months (dosage: each uncoated tabletdicalcium phosphate IP- 0.5 gm; vitamin D3 IP – 500 international units; 2 tablets OD). Each subject in SG had received the same supplement dosage from the hospital pharmacy. While the age-matched control group (CG) of thirty subjects were not given any placebo and advised to continue with their routine diet and lifestyle.
Data Collection The subjects were assigned using our own structured validated questionnaire at the start of the study. Written consent has been taken from all the subjects after explaining them about the purpose and use of study. The study protocol was approved by the independent local ethical committee. Sample collection The blood and urine samples were collected from both the SG and the CG for estimation of the serum calcium and the urine calcium. The subjects were studied while consuming their usual diet without specific dietary recommendations, but were asked to keep on fasting from 9 pm the day before sample collection. The blood samples and 24 hour urine samples were collected the following morning at 8 am in clean, sterile dry containers, and stored at -20 °C. Estimation of Serum Calcium Levels Serum levels of calcium were measured by using calcium-kit method (17-19) (calorimetric method using Arsenazo III), which was based on the principle that, calcium with Arsenazo III at neutral pH yields a blue coloured complex. The intensity of the colour formed was directly proportional to calcium concentrations. Estimation of Urinary Calcium Levels The 24 hourly collected urine samples were assigned for estimation of urinary calcium by using calorimeter based calcium-kit method by using Arsenazo III, which was based on the principle that, calcium with Arsenazo III at neutral pH yields a blue coloured complex. The intensity of the colour formed was directly proportional to calcium concentrations. Statistical Methods The statistical analyses were performed by comparing the data in various groups of SG and CG and the measures of serum calcium and urinary calcium were performed by using analysis of variance for multiple measurements. The biostatistical evaluation was carried out using the statistical package, SPSS version 18.0 (PASW statistics) for windows, (SPSS, Inc.; Chicago, II). Results and Discussion The laboratory values, based on the duration of calcium intake, obtained for the serum calcium and the urinary calcium of the subjects are presented in Table 1a for the study group and in Table 1b for the control group. The relative data for serum calcium and urine calcium among the study group and the control groups are presented in Table 2. The mean and the standard deviation values for the serum calcium in both the control group and the study group exhibited little variation. There was also little variation in values of serum calcium within the study group itself in spite of supplementation for the duration of more than 3 months. The results for analysis of
Ethics Approval Ethical approval was received from the local ethical committee of Osmania Medical College, Hyderabad, prior to commencement. Informed written consent was obtained from each participant prior to participating in the study.
2
J Nutr Health Aging
THE JOURNAL OF NUTRITION, HEALTH & AGING© variance (Table 3) showed little significance for serum calcium in both SG and CG. In the case of the study group, even though there was a constant supply of the same quantity of calcium, there was no substantial change of serum calcium. Therefore, it is suggested that in the study group and the control group of the post-menopausal women in the same age and with the same inclusion and exclusion criteria, the effects of hormones on calcium metabolism is presumed to be the same. Such a view is consistent with a reported study (20) that, no obvious symptoms were brought about in the short term with inadequate intakes of dietary calcium from food and supplements, as the circulating blood levels of calcium were tightly regulated and did not fluctuate with little alterations in dietary calcium intakes. The similar presumed hormonal effects on calcium metabolism from our data also corroborate the fact (21) that with respect to calcium, the homeostatic system was sufficiently flexible to maintain blood ionized calcium within the normal range without being interrupted by wide fluctuations in dietary calcium intake and changing rates of bone mineralization.
Table 1b Laboratory Findings of Control Group (without Calcium Supplementation)
Control Group
Table 1a Laboratory Findings of Study Group (with Calcium Supplementation) based on Duration of Calcium Intake Duration of Calcium Intake (months)
Age
Serum Calcium (mg/dl)
Urine Calcium (mg/day)
54 50 49 50 51 60 50 52 60 49 50 55 48 50 55 49 51 49 52 50 51 49 52 49 49 52 50 52 50 50
9.1 8.2 9 8.3 8.6 8.3 8.5 8.4 8.3 8.9 8.4 9 8.4 8.4 8.6 8.9 8.3 8.3 8.9 8.3 8.9 8.4 8.0 9.6 9.4 8.5 8.9 8.3 9.0 9.1 8.64± 0.38
220 172 150 129 84 75 150 150 100 180 130 155 175 150 130 210 185 200 180 170 200 200 252 250 245 225 225 250 220 230 179.73± 48.20
Study Group
Up to 1 month
Up to 3 months
Up to 12 months
Average ± SD
Average ± SD
Age
Serum Calcium (mg/dl)
Urine Calcium (mg/day)
50 48 49 49 50 60 52 49 50 49 52 54 60 49 50 48 52 50 49 50 52 50 60 58 50 48 52 51 49 51
8.5 8.9 9.3 9 9 8.5 8.4 8.8 9.1 9 8.5 8.9 8.4 9 8.5 8.9 8.5 8.4 8.5 9 8.6 8.5 8.3 8.2 8.3 9 8.9 8.6 9 8.6 8.7 ± 0.3
120 119 110 150 130 100 125 120 120 155 120 130 110 150 130 125 130 140 135 150 140 130 120 125 130 120 140 150 150 140 130±14
The urine calcium levels progressively increased in the study group as compared to those in the control group. Even in the study group, the urine calcium levels were high in those subjects who have consumed calcium supplements for twelve months. The analysis of variance results (Table 3) also showed a statistical significance (p < 0.05) for urinary calcium in SG. In the case of CG, there was no statistically significant variation of urinary calcium. However, only two subjects among the thirty of the SG exhibited the low levels of urinary calcium (Table 1a). This finding of urinary calcium only in two subjects may not be of much significance to the overall study. We really didn’t understand the low levels of the urinary calcium in these two patients. This might probably be due to that they might not have taken the calcium supplements, or could be due to the influence of PTH and Vit D. There was a steady increase in urinary calcium levels corresponding to the duration of intake of calcium supplements (i.e. more urinary calcium levels in subjects taking calcium supplements for more than three months compared to those taking for one month). These increased levels in urinary 3
J Nutr Health Aging
CALCIUM SUPPLEMENTATION IN POST-MENOPAUSAL WOMEN Table 2 Comparison of duration of intake of calcium between control and the subject groups Control Group (No Calcium Supplementation)
Age Serum Calcium (mg/dl) 24 hrs Urinary calcium (mg/dl)
Study Group (Calcium Supplementation)
Mean
Standard Deviation
51 8.7 130
4 0.3 14
1 month Mean Standard Deviation 52 8.6 143
4 0.3 38
Duration of intake of calcium 3 months Mean Standard Deviation 50 8.6 192
12 months Mean Standard Deviation
1 0.3 14
51 8.9 237
1 0.5 13
Table 3 Mean (SEM) values of serum calcium and urinary calcium measures, and results of analyses of variance Control Group (No Calcium Supplementation)
Mean
Serum Calcium (mg/dl)
24 hrs urinary calcium (mg/dl)
Study Group (Calcium Supplementation)
Standard Deviation
Between Groups Within Groups Total Between Groups Within Groups Total
1 month Mean Standard Deviation
Duration of intake of calcium 3 months Mean Standard Deviation
12 months Mean Standard Deviation
Sum of Squares
df
Mean Square
F*
Sig.
0.542 6.380 6.922 83710.9 27974.5 111685.4
3 56 59 3 56 59
0.181 0.114
1.585
0.203
27903.6 499.5
55.9
0.000
*Asymptotically F distributed
excretion of calcium may indirectly reflect the rise of the renal threshold for excretion. As the urine sample was collected after 24 hours of intake of last dose of calcium supplements, the amount of absorbed calcium and later spilling in urine is questionable, giving way to various complications such as deposition in the soft tissues, hence enabling the scope for curbing the indiscriminate use of calcium supplements in postmenopausal women.
Acknowledgements: The authors thank to Prof. B. Ram Reddy and Prof. Ch. N. Rajkumari of Osmania Medical College, Hyderabad, Andhra Pradesh, India for their valuable suggestions and advice for the present work. Conflict of Interest: The authors declare they have no conflict of interest. Financial support: This research work received no specific grant from any funding agency, commercial or not-for-profit sectors, and had not involved any sponsors Statement of authorship: MNS initiated and designed the study, participated in the data acquisition, contributed in statistical analysis and in reviewing the manuscript. AKK performed the statistical analysis, conceived the study, in its design and coordination, drafted the manuscript and edited the manuscript. All authors have read and approved the final manuscript.
Conclusion
References
There was no significant change in serum calcium levels in the study group even after consumption of calcium supplements for a period of one year, whereas the urine calcium levels progressively increased only in the study group and not in the control group. This indicates prolonged calcium supplementation has little effect on serum calcium levels but significantly affecting the urinary calcium levels in postmenopausal women.
1. 2.
3.
4.
4
Nordin BEC. Calcium and osteoporosis. Nutrition 1997;13: 664-686. Eugene , Paula B. The Parathyroid glands and vitamin D. In: Boron WF, Boulpaep E (eds). Medical Physiology, 2nd edn. W B Saunders Publishing, Philadelphia, 2003;pp 1094-1103. Committee to Review Dietary Reference Intakes for Vitamin D and Calcium, Food and Nutrition Board, Institute of Medicine. Dietary Reference Intakes for Calcium and Vitamin D. Academic Press, Washington DC, 2010. National Institutes of Health. Optimal calcium intake: NIH Consensus Statement. 1994;12: 1-31.
J Nutr Health Aging
THE JOURNAL OF NUTRITION, HEALTH & AGING© 5.
6.
7.
8.
9.
10. 11.
12.
Dawson-Hudges B, Dallal GE, Krall EA, Sadowski L, Sahyoun N, Tannenbaum S. A controlled trial of the effect of calcium supplementation on bone density in postmenopausal women. N Engl J Med 1990;323: 878-883. Prince RL, Devine A, Dick I et al. The effects of calcium supplementation (milk powder or tablets) and exercise on bone density in postmenopausal women. J Bone Min Res 1995;10: 1068-1075. Reid IR, Ames RW, Evans MC, Gamble GD, Sharpe SJ. Long-term effects of calcium supplementation on bone loss and fractures in postmenopausal women – a randomized controlled trial. Am J Med 1995;98: 331-335. McKane WR, Khosla S, Egan KS, Robins SP, Burritt MF, Riggs BL. Role of calcium in modulating age-related increases in parathyroid function and bone resorption. J Clin Endocrin Metab 1996;81:1699-1703. Bolland MJ, Avenell A, Baron JA, Grey A, MacLennan GS, Gamble GD et al. Effect of calcium supplements on the risk of myocardial infarction and cardiovascular events: meta-analysis. Brit Med J 2010;341: c3691 doi:10.1136/bmj.c3691. Reid IR, Mason B, Horne A, Ames R, Reid HE, Bava U et al. Randomized controlled trial of calcium in healthy older women. Am J Med 2006;119: 777-785. Bolland MJ, Barber PA, Doughty RN, Mason B, Horne A, Ames R et al. Vascular events in healthy older women receiving calcium supplementation: randomized controlled trial. Brit Med J 2008;336: 262-266. doi: 10.1136/bmj. Cleland JGF, Klaus W, Sue S. Calcium supplements in people with osteoporosis (2010) Brit Med J 2010;341: c3856. doi: http://dx.doi.org/10.1136/bmj.c3856.
13.
14. 15.
16.
17. 18. 19. 20.
21.
5
Moyer VA. Vitamin D and calcium supplementation to prevent fractures in adults: U.S. Preventive Services Task Force recommendation statement. Ann Int Med 2013;158: 691-696. doi:10.7326/0003-4819-158-9-201305070-00603. Heaney RP. Calcium. In: Bilezikian JP, Raisz GA, Rodan GA (eds). Principles of Bone Biology. Academic Press, New York, 1996;pp 1007-1018 Yoshida M, Fukuwatari T, Sakai J, Tsuji T, Shibanda K. Correlation between mineral intake and urinary excretion in free-living Japanese young women. Food Nutr Sci 2012;3: 123-128 doi:10.4236/fns.2012.31018]. Heinrich T, Heidt H, Hafner V, Schmidt-Gayk H, Jenetzky E, Walter-Sack I et al. Calcium load during the administration of calcium carbonate or sevelamer in individuals with normal renal function. Neph Dial Transpl 2008;23, 2861-2867. doi: 10.1093/ndt/gfn151. Smith HG Jr, Baucer PJ. Light-induced permeability changes in sonicated bovine disks: arsenazo III and flow system measurements. Biochem 1979;18: 5067-5073. Budesinky B. Analyt Chem. Chelñates 1969;18: 5067-5073. Cadwell PC. Calcium chelation buffers. In: Cuthbert AW (ed). Calcium and Cellular Function, Mackmillan Publishing Co Inc, London, 1970;pp 10-16. Dawson-Hughes B, Harris SS, Palermo NJ, Castaneda-Sceppa C, Rasmussen HM, Dallal GE. Treatment with potassium bicarbonate lowers calcium excretion and bone resorption in older men and women. J Clin Endocrin Metab 2009;94: 96-102. Goldman's Cecil Medicine. Elsevier, Philadelphia, 2007.
