37
Original Article
Zinc Supplementation and Serum Zinc During Diarrhea Abdullah H. Baqui, Robert E. Black, Christa L. Fischer Walker, Shams Arifeen, Khalequz Zaman, Mohammad Yunus, Mohammad A. Wahed and Laura E. Caulfield Johns Hopkins Bloomberg School of Public Health, Department of International Health, Baltimore, Maryland (AHB, REB, CFW, LEC) and International Centre for Diarrheal Diseases Research, Bangladesh (ICDDR,B), Dhaka, Bangladesh (AHB, SA, KZ, MY, MAW)
ABSTRACT Objective. Zinc deficiency is very common in developing countries and is more pronounced during an episode of diarrhea. Supplementation with zinc improves diarrhea and might correct zinc deficiency in both the short and longer term. Method. We conducted a nested study within a cluster randomized treatment trial. Fifty children with diarrhea living in the zinc treated clusters, 50 children with diarrhea living in control clusters, and 50 healthy children living in the control clusters were enrolled. We assessed serum zinc at the start of the diarrhea episode, which was 1-3 days after supplementation began in zinc treated children, and again one week after the diarrhea ended and supplementation ceased. Baseline characteristics and serum zinc concentration were assessed. Results. Serum zinc was low in 44% of healthy children at the first blood draw. Compared to healthy controls, serum zinc was 3.1 mmol/L higher among children with diarrhea who were supplemented with zinc at first blood draw and 1.3mmol/L higher 3 weeks later. Conclusion. Zinc supplementation enhances serum zinc concentration when given as a treatment for diarrhea and helps children maintain a more adequate zinc status during the convalescent period. [Indian J Pediatr 2006; 73 (6) : 493-497] E-mail :
[email protected]
Key words : Zinc; Diarrhea; Supplementation
Zinc deficiency is highly prevalent in children in developing countries and is one of the major risks factors for poor child health accounting for over 800 000 child deaths per year, including 10% of all diarrhea deaths.1, 2 Dietary zinc insufficiency is exacerbated during a diarrhea episode by net zinc loss in the stool. 3 Zinc supplementation during diarrhea reduces the duration and severity of the episode and the risk of subsequent diarrhea and ALRI morbidity.4-6 It is now recommended that all children under 5 receive 20 mg supplemental zinc for 10-14 days as part of clinical treatment of diarrhea.7 How zinc exerts these therapeutic and preventive effects is not fully understood. Zinc is an essential mineral for human cell growth, differentiation, and DNA synthesis 8 , and it is known to improve water and electrolyte absorption,9, accelerate the regeneration of the gut epithelium, 10 increase the activity of brush border
enzymes11 and enhance overall immune function. 12 A better understanding of the mechanism of protection may be possible if more is known about the effect of short-term supplementation on zinc status, as described by serum zinc. Studies to date have assessed serum zinc prior to supplementation during diarrhea and then again on day 14 in both zinc-supplemented and control children generally showing higher zinc concentration at the end of supplementation.13-15 In a study nested within a cluster randomised trial of zinc therapy for diarrhea we examined the serum zinc concentrations of children with diarrhea in the 1-3 days after the initial supplement was given as well as the lasting effect one week after a 14-day course of zinc supplementation to determine if the higher serum zinc concentrations continued after supplementation had been stopped. MATERIAL AND METHODS
Correspondence and Reprint requests : Dr. Abdullah H. Baqui Johns Hopkins Bloomberg School of Public Health 615 North Wolfe St. Rm E8138 Baltimore, MD 21205 Phone : 410-955-3850 Fax: 410-614-1419 (attn: A.Baqui)
Indian Journal of Pediatrics, Volume 73—June, 2006
We have previously reported the details of the study design and impact of two weeks of zinc therapy for diarrhea on acute lower respiratory infections (ALRI) and 493
38
Abdullah H. Baqui et al diarrhea morbidity and mortality in a community-based cluster randomised trial 4 in the Matlab field area of ICDDR,B: Centre for Health and Population Research. Thirty service areas (clusters) around Matlab ICDDR,B Hospital, each with about 200 children 3-59 months old, were randomly allocated to intervention or comparison areas. Each cluster was served by one community health worker (CHW). From November 1998 to October 2000, all children 3-59 months old were included. CHWs and Bari Mothers (BMs - community volunteers) distributed oral rehydration solution (ORS) packets and adviced on feeding to parents who sought treatment for a child with diarrhea in both intervention and control areas. In the intervention clusters, CHWs and BMs also distributed zinc syrup and instructed the mothers on how to give the syrup to her child. The syrup was a 20 mg dose of zinc acetate prepared in 5 ml syrup to be given once a day for 14 days regardless of the duration of diarrhea. Both ORS and zinc were provided free. In the comparison communities, CHWs and BMs distributed ORS and provided advice on feeding only. To assess the differences in serum zinc concentrations among children with diarrhea supplemented with zinc, children with diarrhea who were not supplemented with zinc, and children without infection, blood specimens were collected in 3 groups of children: (i) children with acute diarrhea in the zinc treatment group after 1-3 days of zinc supplementation, and on day 21 (1 week after
completing 14 days of treatment); (ii) children with acute diarrhea in the control area on day 1-3 of diarrhea and 21 days after the first blood draw; (iii) healthy children in the control areas, defined as those not having diarrhea for at least 2 weeks prior to the blood draw. The sample size was calculated to see an 11% difference (80% power and 95% confidence) in mean serum zinc after 14 days of supplementation between zinc-supplemented children and non-zinc-supplemented children with diarrhea. 16 Fifty children with acute diarrhea from the villages receiving zinc supplementation were selected over the study year. As each child from one of these villages was enrolled, an age (within 1 month) and gender-matched child with acute diarrhea from a control village and an age and gender-matched healthy child also from a control village were selected. The children from the control areas were chosen within one week of the selection of the child in the treatment group to also account for seasonal influences. The larger cluster-randomised trial and the selection of subjects for this nested study are described in Fig. 1. All selected children were brought to the Matlab hospital for blood drawing. Approximately 3 ml of blood was collected via venipuncture using plastic tubes carefully washed to make them zinc free. The blood samples were centrifuged immediately, frozen at –20 °C, and transported on ice to the ICDDRB laboratory in Dhaka where serum zinc concentrations were measured
30 Clusters
15 Intervention Clusters with 3974 partially or fully complete follow-up resulting in 5866 observable child years
50 Children with diarrhoea
50 Children with diarrhoea
13 loss to follow-up/ bad blood work
15 Control Clusters with 4096 partially or fully complete follow-up resulting in 6015 observable child years
50 Healthy Controls with 1 successful blood draw
8 loss to follow-up/ bad blood work 37 with 2 successful blood draws
42 with 2 successful blood draws Fig. 1. Trial Profile
494
Indian Journal of Pediatrics, Volume 73—June, 2006
39
Zinc Supplementation and Serum Zinc During Diarrhea using atomic absorption spectrophotometry (AAS, Perkin Elmer model 3100). A certified trace element control serum was used daily to ensure accuracy and precision (UTAK Lab Inc., California, USA). All lab analyses were blinded to diarrhea and intervention status of the children sampled. Of the 50 children per group, 45 children in the zinc treatment group and 43 children with acute diarrhea in the control group had a second blood draw (Fig 1). Of the 238 blood draws, 11 of them were either contaminated, insufficient or grossly haemolyzed. Samples from the index diarrhea children and diarrhea controls which did not have matching first and second blood draws were excluded. Thus, 37 index diarrhea children, 42 diarrhea controls, and 50 healthy controls were included in the analysis. The study was approved by the ethical review committees of ICDDR,B and the Johns Hopkins Bloomberg School of Public Health. We obtained verbal consent from village leaders and parents for this nested study. Statistical analyses were done with STATA 8.0 statistical software. Mean differences in baseline characteristics between zinc-supplemented and control children were analyzed by analysis of variance. Differences in proportions were assessed by chi-squared analysis. The mean serum zinc concentration for each blood draw in each group of children was calculated, as well as the proportion of children in each group with concentrations less than 9.9 µmol/L, a recommended cut point for zinc deficiency in young children.1 Comparisons of mean serum zinc at each time point were assessed by multiple linear regression controlling for the sex of the child. Although there were no differences in mean serum zinc by sex, we adjusted for sex to reduce the variance in the serum zinc measures. A p value of <0.05 was considered statistically significant. For comparison of mean serum zinc concentrations after the diarrhea episode, the duration of the diarrhea was added to the regression. RESULTS There were no differences in characteristics of the study children by group at baseline (Table 1). All children with diarrhea from the zinc intervention and control villages had their first blood draw within 1-4 days of starting diarrhea (1-3 days after starting zinc in the intervention villages). Healthy controls had a mean serum zinc concentration of 10.0 µmol/L (+ 1.7), the suggested lower threshold for zinc deficiency; 44% had a serum zinc concentration lower than 9.9 µmol/L. Children with diarrhea who were not supplemented with zinc had a mean serum zinc concentration similar to healthy controls (Table 2). Children with diarrhea who were supplemented with Indian Journal of Pediatrics, Volume 73—June, 2006
TABLE 1. Baseline Characteristics of Children Selected Characteristics
Children With Diarrhoea Zinc (n=37)
Male (%) Age in months (at 1st
blood collection) [Mean (SD)]
Birth Order [median (range)]
Mother’s education (%)
None 1-5 years > 6 years Household size [median (range)]
No Zinc (n=42)
Healthy Controls (n=50)
51.4
61.9
58.0
28.2 (13.3)
27.6 (13.5)
27.2 (13.7)
2 (1, 8)
3 (1, 7)
3 (1, 7)
54.0 37.5 29.7 40.0 16.2 22.5 5 (2, 11) 5 (2, 16)
23.7
61.7
14.8
5 (2, 15)
** There were no statistically significant differences between groups TABLE 2. Mean Serum Concentration (µ µ mol/L) and Number of
Children with Low Serum Zinc Concentration*
Children with Diarrhoea Zinc (n=37)
Healthy Controls No Zinc (n=42) (n=50)
First Blood Draw 13.1 ± 4.3a,b Mean + SD 8 (21.6) Number deficient (%) Second Blood Draw Mean + SD 11.3 ± 2.0a Number deficient (%) 10 (27.0)
9.4 ± 1.9 29 (69.1)
10.0 ± 1.6 22 (44.0)
10.5 ± 1.8 15 (35.7)
(a) Compared to healthy controls by linear regression analysis adjusted for sex (p <0.05) (b) Compared to children with diarrhea not receiving zinc at first blood draw by linear regression analysis adjusted for sex (p = 0.000) *Low serum zinc defines as <9.9 µmol/L [1].
zinc had a significantly higher mean serum zinc concentrations at the first blood draw (13.1 µmol/L) and 1 week after supplementation ended (11.3 µmol/L) than healthy controls after controlling for sex (Table 2). Duration of diarrhea did not influence serum zinc concentration of the second blood draw when added into the regression analysis. Zinc-supplemented children had an overall lower duration of diarrhea (4.5 + 3.6 days) compared to children with diarrhea who did not receive zinc (7.0 + 5.7 days). DISCUSSION This study investigated the immediate and sustained biochemical response of zinc supplementation on serum zinc concentration within the context of a randomised controlled trial of zinc therapy for diarrhea. Given the community-based design, it was not possible to collect a blood sample from children with diarrhea prior to zinc treatment; however, given random assignment of villages, 495
40
Abdullah H. Baqui et al it is reasonable to make post-treatment comparisons regarding changes in zinc status with a 14-day regimen of zinc supplementation. The results indicate that zinc deficiency is common in otherwise healthy children in Bangladesh; 44%, based on the 2.5 th percentile cut off in population studies; 1 this indicates poor individual zinc status of these children and reflects the high prevalence of zinc deficiency among children under 5 in Bangladesh. Children with diarrhea who were not treated with zinc had lower serum zinc concentrations (69% were below that threshold), significantly more than the healthy children (p<0.05) which could either be due to more zinc deficient children being at greater risk of diarrhea. 17 or shifts in zinc from blood to the liver during infections.18 These results are congruent with previous studies in Bangladesh of children presenting with diarrhea.16, 19 After 1-3 days of supplementation, children treated with zinc had significantly higher serum zinc concentrations than either unsupplemented children with diarrhea or healthy controls. Although compliance was not specifically measured in this subset, children living in the zinc clusters on average consumed 7 tablets per treatment.4 In absolute terms, the difference was 3.1-3.7 µmol/L, and when considered in relation to sample variability, was about 1 SD. One week after finishing the 14-day zinc regimen, the serum zinc concentrations were lower than during supplementation, but were still significantly higher than those of healthy controls (1.3 µmol/L or 0.77 SD). They were higher than those of control children with diarrhea (0.8 µmol/L or 0.38 SD), although the difference was not statistically significant. This may have been due to the small sample size. The effects on zinc status persisted beyond the delivery of the supplement. These results are largely consistent with the study by Raquib et al19 in which Bangladeshi children with shigella dysentery were randomized to receive 20 mg of zinc in a multivitamin mix or a multivitamin mix alone for 14 days in addition to antibiotic treatment, and were evaluated at baseline, 7 days, and 30 days (16 days after supplementation). In that study, serum zinc concentrations were 0.76 µmol/L higher at 7 days, and 1.22 µmol/L higher than the control group after 30 days. As was observed here, zinc rose over time in the zinc and control groups, but the serum zinc concentrations continued to rise even 16 days after supplementation. One week after supplementation, the serum zinc concentration of our study children showed evidence of decline from those during supplementation, but in the study by Raquib et al 19 serum zinc concentrations continued to rise and remained higher than baseline even 16 days after supplementation. Thus the persistence of higher zinc concentrations following supplementation in young children may not be universal; mediating factors for this are yet to be identified. Zinc supplementation has been shown to continue to benefit the child after supplementation has ended. Short 496
course treatment trials have shown a decrease in both incidence and prevalence of diarrhea and acute lower respiratory infections in the months following the treatment of one episode of diarrhea. 6 It may be that a persisting biochemical response to zinc therapy as shown here is partly responsible for these effects. Concerns remain about the utility of plasma or serum zinc concentration as an indicator of individual zinc status. Strand et al20 found that serum zinc was influenced by the presence of other clinical symptoms or illnesses in children with diarrhea. The children bled in this study did not have additional illnesses. In a recent study plasma zinc correlated well with changes in dietary zinc intake and kinetic indicators of zinc status in adult men with no underlying infection.21 On a population level, serum zinc is a useful indicator of overall zinc status and shifts in zinc nutriture. This study showed that zinc treatment for childhood diarrhea replaced zinc stores immediately after supplementation and kept zinc status elevated in the weeks following illness. Further studies are needed to define the relation between zinc concentration and whole body zinc status during illness in order to understand the mechanisms by which short regimens of zinc are effective as therapeutic agents for childhood diarrhea and other common infections, and continue to have benefits after the supplementation period. Acknowledgements The study has been supported by the Johns Hopkins Family Health and Child Survival Co-operative Agreement and ICDDR,B Co operative Agreement (#HRN-A-OO-06-90005-00), with funding from the US Agency for International Development.
REFERENCES 1. Brown, KH et al. Assessment of the risk of zinc deficiency in populations. Food and Nutrition Bulletin 2004; 25(1): S130-S162. 2. WHO, World Health Report 2002: Reducing risks, promoting healthy life. 2002, World Health Organization: Geneva. 3. Castillo-Duran C, Vial P. and R. Uauy. Trace mineral balance during acute diarrhea in infants. J Pediatr 1988; 113(3): 452-457. 4. Baqui, AH et al. Effect of zinc supplementation started during diarrhoea on morbidity and mortality in Bangladeshi children: community randomised trial. BMJ 2002; 325(7372): 1059. 5. Bhutta ZA et al. Therapeutic effects of oral zinc in acute and persistent diarrhea in children in developing countries: pooled analysis of randomized controlled trials. Am J Clin Nutr 2000; 72(6): 1516-1522. 6. Bhutta ZA et al. Prevention of diarrhea and pneumonia by zinc supplementation in children in developing countries: pooled analysis of randomized controlled trials. Zinc Investigators’ Collaborative Group. J Pediatr 1999; 135(6): 689-697. 7. WHO and UNICEF. WHO-UNICEF Joint statement on the clinical management of acute diarrhea. in World Health Assembly. 2004. Geneva. 8. Sandstead HH, Zinc deficiency. A public health problem? Am J Dis Child 1991; 145(8): 853-859. 9. Ghishan FK. Transport of electrolytes, water, and glucose in zinc deficiency. J Pediatr Gastroenterol Nutr 1984; 3(4): 608-612. 10. Roy SK, et al. Impact of zinc supplementation on intestinal
Indian Journal of Pediatrics, Volume 73—June, 2006
41
Zinc Supplementation and Serum Zinc During Diarrhea
11.
12. 13.
14.
15.
permeability in Bangladeshi children with acute diarrhoea and persistent diarrhoea syndrome. J Pediatr Gastroenterol Nutr 1992; 15(3): 289-296. Jones PE and TJ Peters, Oral zinc supplements in nonresponsive coeliac syndrome: effect on jejunal morphology, enterocyte production, and brush border disaccharidase activities. Gut 1981; 22(3): 194-198. Ibs, K.H. and L. Rink, Zinc-altered immune function. J Nutr 2003; 133(5 Suppl 1): 1452S-6S. Bhatnagar S et al. Zinc with oral rehydration therapy reduces stool output and duration of diarrhea in hospitalized children: a randomized controlled trial. J Pediatr Gastroenterol Nutr 2004; 38(1): 34-40. Bhutta ZA, S.Q. Nizami, and Z. Isani, Zinc supplementation in malnourished children with persistent diarrhea in Pakistan. Pediatrics 1999; 103(4): e42. Roy SK et al. Impact of zinc supplementation on persistent diarrhoea in malnourished Bangladeshi children. Acta Paediatr 1998; 87(12): p. 1235-1239.
Indian Journal of Pediatrics, Volume 73—June, 2006
16. Roy SK et al. Randomised controlled trial of zinc supplementation in malnourished Bangladeshi children with acute diarrhoea. Arch Dis Child 1997; 77(3): 196-200. 17. Bhandari N et al. Substantial reduction in severe diarrheal morbidity by daily zinc supplementation in young north Indian children. Pediatrics 2002; 109(6): e86. 18. Brown, K.H., Effect of infections on plasma zinc concentration and implications for zinc status assessment in low-income countries. Am J Clin Nutr 1998; 68(2 Suppl): 425S-429S. 19. Raqib R et al. Effect of zinc supplementation on immune and inflammatory responses in pediatric patients with shigellosis. Am J Clin Nutr 2004; 79(3): 444-450. 20. Strand TA et al. Predictors of plasma zinc concentrations in children with acute diarrhea. Am J Clin Nutr 2004; 79(3): 451 456. 21. Lowe, NM et al. Kinetic parameters and plasma zinc concen tration correlate well with net loss and gain of zinc from men. J Nutr 2004; 134(9): 2178-2181.
497
