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INTRODUCTION |
Zinc metabolism is subject to strong homeostatic regulation (Taylor et al. 1991
). Body zinc nutritional status (King 1990), dietary zinc intakes (Bear 1984, Wada 1985) and some dietary components such as Fe, Ca, protein, dietary fiber and phytate (Sandstead 1981) affect zinc balance. Zinc losses via the whole body surface and hair are often neglected in the estimation of zinc requirement (Milne 1983). These factors should be considered in the estimation of zinc requirements whether determined by balance studies or by factorial calculation of endogenous losses (Food and Nutrition Board 1989).
We studied the zinc metabolism and requirement of Chinese preschool children consuming different diets by using a balance method and determination of zinc losses through the whole body surface and hair.
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MATERIALS AND METHODS |
Subjects.
Eleven subjects (six males and five females) ranging in age from 5.5 to 6.5 y with a mean age of 6 y were chosen from a full-time kindergarten. Each of the children's parents consented in writing to participation in the study after receiving a written description of all methods to be used and of diets to be consumed throughout the study. They were given oral explanations as necessary. The following comprehensive criteria were used to evaluate zinc nutritional status: hair zinc concentration above 1.68 µmol/g, zinc concentration of whole blood above 76.9 µmol/L (Yu et al. 1989), the main indicators of growth, such as height and weight, normal (Chen et al. 1989), clinical examinations showing no signs of zinc malnutrition and related diseases (Chen et al. 1989). Judged by the above criteria, zinc status of each subject was normal.
Experimental design.
The 27-d study was divided into three phases. All subjects consumed all the meals and lived in the kindergarten throughout the study. In the first 3-d phase (pretest metabolic period), all subjects consumed the kindergarten's original diets, with no limit on water intake, snacks and daily activities. A nutritional survey was taken; food consumption of each subject including water intake and snacks was recorded in detail. Zinc concentration of foods, hair and whole blood were determined. Zinc losses via the whole body surface, urine, feces and hair were measured. In the second phase, all subjects consumed a balanced diet designed for this study for 3 wk. Nutrients compositions are shown in Table 1. Different kinds of foods were bought from markets to form diet menus that kept nutrient compositions constant but the food compositions different. Seven days was defined as a rotation period. A sample menu is shown in Table 2. In the third 3-d phase (posttest metabolic period), all subjects continued to consume the balanced diet with no limit on water intake and daily activities, but snacks were forbidden. All determinations were taken again.
Indicators and methods.
Precautions against environmental zinc contamination were taken for all diet, blood, hair and excreta collections and analyses. Only double distilled deionized water and ultra high-purity reagents were used in sample and standards preparations. All glassware was soaked in 1.0 mol/L HCl for at least 20 h and rinsed four times with deionized water. All containers were washed in a detergent containing EDTA (Shanghai First Chemical Reagent Factory, Shanghai, China) and rinsed four times with deionized water. All zinc determinations were made using a Shimadzu AA-680 (Shimadzu Corporation, Tokyo, Japan) flame atomic absorption spectrophotometer (AAS).
Height, weight and clinical examination were performed using current methods for nutritional surveys (Chen et al. 1989). Blood hemoglobin was measured by the cyanhematin extinction coefficient method (Yu et al. 1989).
Blood zinc concentration.
Thirty minutes before breakfast blood samples were drawn from the antecubital vein of each subject into trace element-free tubes containing heparin. Blood (1 ml) was then digested by mixed acid (concentrated HNO3:HClO4 = 4:1, all mixed acid used in this study is the same), and zinc was determined by AAS (Yu et al. 1989).
Hair zinc concentration.
On the first day of the study, hair 2 cm in length was collected from the scalp in the occipital area and stored in tared envelopes. On each occasion the hair was thoroughly washed with hair shampoo (Shanghai Lever Co. Ltd., Shanghai, China), rinsed with deionized water, and dried. A 0.1-g hair sample was digested using mixed acid, and zinc concentration was determined by using AAS (Yu et al. 1989). At the end of the third phase, all the hair growing in the same area was collected, and zinc concentration was determined by following the same procedure.
Hair zinc losses.
All hair that grew in the 27-d period was collected by hiring the same barber to cut the hair and by having the children keep the same hair style. The cut hair was weighed. The zinc concentration of hair sampled at the end of the third phase times total hair weight was considered to be the hair zinc losses in the 27-d period (Yu et al. 1989).
Fecal zinc concentration.
Fifteen minutes before breakfast on the first day of each metabolic period and next day to each metabolic period each subject took two tablets of active carbon that was used as a marker. Fecal samples of each subject were separately collected when the marker appeared for the first time, and collection was stopped when the marker appeared the second time. All fecal samples were weighed and frozen at 
20°C until analyzed. Before sampling all fecal samples were homogenized with a constant volume of deionized water. About 50 g of the homogenized feces were dried, smashed and passed through a 0.833-mm mesh sieve. The dried fecal sample (0.1 g) was digested by mixed acid, and zinc concentration was determined by AAS (Yu et al. 1989).
Urine zinc concentration.
In the first and third phases, 24-h urine samples were collected daily in a zinc metal-free plastic container and treated with a drop of concentrated HNO3. Ten percent of each daily urine collection was reserved and mixed for further analyses after its volume was measured. Ten milliliters of the urine was digested by mixed acid, and zinc concentration was determined by AAS (Yu et al. 1989).
Food composition.
The weighing method and recording method were combined in this diet survey. In the first and third phases, the pretest and the posttest diets were recorded in detail. Four to five servings of each food were weighed, and the average weight was recorded. Two to three servings (every kind of food consumed in terms of servings in the two periods was collected as a separate sample) were collected in zinc-free plastic containers as a sample and frozen at 20°C until analyzed. The proper volume of double distilled deionized water was added, and then each sample was homogenized, dried, smashed and passed through a 0.833-mm mesh sieve. These samples were used for further analyses. Nutrients such as protein, fat, carbohydrates, calcium, phosphorus, phytate, iron and zinc were measured in our laboratory (Yu et al. 1989). The other nutrients and nutrients of snacks were calculated by using a program in basic language of our department. The nutrient compositions of every kind of food were calculated in terms of the average value of one serving.
Nutrient intakes.
In each metabolic period, each subject's food consumption and water intake were recorded in detail (in terms of servings). The number of consumed servings of a kind of food times the nutrient compositions of per serving is the nutrient intake from this kind of food. Nutrient intakes were calculated in terms of the average value per child per day.
Whole body surface zinc loss.
Whole body sweat samples were obtained and determined at every metabolic period by using a modified procedure (Chen et al. 1995, Milne et al. 1983). Before the experiment, pure cotton underwear, bath towels and plastic shower basins were washed in a detergent containing EDTA, rinsed with deionized water and dried. At the beginning of the metabolic periods, the subjects showered in the usual way with zinc-free soap (Shanghai Lever Co. Ltd., Shanghai, China). After rinsing with tap water, the children rinsed thoroughly with deionized water, dried with zinc-free bath towels and then put on two pairs of underwear. The inner pair was used to collect sweat samples, while the outer was used to avoid environmental contamination. Seventy-two hours later, each disrobed subject showered with 4 L deionized water in a zinc-free plastic basin. After rinsing with deionized water, they were dried with a zinc-free bath towel. Proper volume of bath water was reserved for analyses after its volume was measured and stirred completely. The inner pair of underwear and bath towels of each subject were collected and extracted in 4 L deionized water with a detergent containing EDTA for 20 h. A volume of the extraction water was reserved for further analyses. Sample blanks were obtained by following the same extraction procedure. The zinc content of extraction liquid of pure cotton underwear and bath towel plus the zinc content of bath water minus blank was considered the zinc loss through the whole body surface measured by this method. Since the pure cotton underwear just cover 78% of whole body surface, zinc excretion through whole body surface should be the results measured by our method times 1.28.
In a separate pilot study to validate the method, bath water with a known zinc concentration was sprayed on zinc-free pure cotton underwear. The pure cotton underwear was dried. The zinc concentrations were determined by our method. The recovery rate of this method averaged 94.1% (92.4 ~ 96.7%); variance was 2.6% (2.2 ~ 3.1%).
Foxbase+ (Multi-user Foxbase+ Chinese version 2.10, Foxsoftware, Beijing, 1988) was used to create the database, and SAS (SAS version 6.04, SAS Institute, Cary, 1989) was used to do paired t tests and correlation analyses. Significance was established at P < 0.05.
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RESULTS |
Indicators of zinc nutritional status were all in the normal range both in pre- and posttest diet period (Table 3). No significant differences were observed between the periods.
Compared with nutrient intakes in the pretest period, intakes of fat, calcium, zinc and retinol were greater during the posttest period (P < 0.05), and those of carbohydrates, dietary fiber and phytate were lower (P < 0.05) (Table 4). Dietary zinc intakes increased from 5.38 to 7.12 mg/d (P < 0.05, Table 5). Zinc excretions via feces and urine also increased (P < 0.05), but whole body surface zinc loss, zinc retention and zinc absorption were not affected. Hair zinc loss averaged 5.26 ± 2.49 µg/d. Zinc absorption averaged 25.8%. If all the zinc losses via urine, feces, the whole body surface and hair were considered, the absorbed zinc during the posttest diet period was 1.84 ± 0.47 mg/d.
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Table 6.
Comparison of dietary zinc requirements in children as estimated using the balance method and the factorial calculation method
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During the pretest diet period, zinc excretions via urine (r = 0.69, P < 0.05) and the whole body surface (r = 0.64, P < 0.05) were positively related to dietary zinc intake (Fig. 1). After children consumed the balanced diet for 3 wk, zinc losses via feces (r = 0.88, P < 0.05), urine (r = 0.72, P < 0.05) and the whole body surface (r = 0.68, P < 0.05) were positively related to zinc intake.
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| Fig 1.
Relationships between zinc intakes and zinc losses via the whole body surface, urine and feces, and zinc retention in the pre- and posttest diet phases in children.
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DISCUSSION |
Zinc requirement and RDA are often determined by the metabolic balance method. It has been suggested that substantial quantities of zinc are lost via the whole body surface (including sweat, lipid and desquamated epithelial cells) (Milne et al. 1983), hair and other routes. In this study, a modified procedure was used to determine the whole body surface zinc losses. The average value of whole body zinc loss of Chinese preschool children who were in normal zinc nutritional status and fed a balanced diet was 0.27 ± 0.09 mg/d, accounting for 5% of total zinc losses, second in magnitude to fecal zinc losses. This value was much lower than that reported by Ritchey et al. (1979). These losses are not negligible and should be considered in the estimation of zinc requirements and the RDA.
Zinc balance can be maintained at different levels through a strong homeostatic mechanism, and zinc status is determined by the relationship between the zinc requirement and dietary intake (King et al. 1990). In preschool children, the zinc absorbed from diets is comparatively large. It should not only meet the functional needs of organs, but also fulfill the needs for body development. Their zinc metabolism should always be in a positive state. The absorbed zinc of those who are in normal zinc nutritional status and positive zinc balance can be regarded as their zinc requirements (Liu et al. 1987).
During the pretest diet period, the height and weight of all subjects reached or surpassed the average level of their age. Their zinc concentrations of hair and whole blood were all within normal range, and clinical examination showed no signs of zinc deficiency. Judged by our comprehensive criteria, the zinc nutriture of each subject was good. Zinc intakes were comparatively low, averaging 5.38 mg/d. Zinc excretions via urine and whole body surface were positively correlated to dietary zinc intakes (P < 0.05), while zinc retention and fecal zinc excretion were not. This suggested that the amount of zinc excreted, predominantly through the intestine, is roughly proportional to dietary intake and to zinc status. The children were in good zinc status; they absorbed the limited dietary zinc efficiently and excreted less zinc through feces so as to maintain their zinc metabolic balance. In the second phase, diets were designed to supply the children with sufficient dietary zinc to ensure that these children's zinc nutritional status was optimal. After they consumed balanced diets for 3 wk, their average dietary zinc intake increased to 7.12 mg/d (P < 0.05). Intakes of nutrients such as Ca, dietary fiber and phytate also changed significantly. These dietary factors affect zinc availability (Sandstead 1982). In theory, zinc absorption of the subjects in this study may be changed significantly. In fact, zinc excretions via both feces and urine significantly increased compared to those of the pretest period (P < 0.05). Post-treatment, zinc losses via feces, urine and the whole body surface were positively correlated with dietary zinc intake (P < 0.05), while zinc retention was not. Absorbed zinc did not change. It suggested that with the significant increase in dietary zinc intake, some parts of increased dietary zinc were excreted through feces and urine, body zinc metabolic balance was maintained by strong homeostasis. All of these metabolic changes confirmed that after all subjects consumed the balanced diet for 3 wk their zinc metabolism did actually reach a stable and positive state. Their absorbed zinc can be regarded as their zinc requirement. When the children consumed a balanced diet, their zinc requirement was 1.84 ± 0.47 mg/d, slightly higher than Tech reports of WHO in 1973 (Table 6).
In this study, zinc absorption of these children was 25.8% when they consumed a balanced diet. It was the same as reported by others (Yu et al. 1991). Assuming that the zinc availability is 20%, the zinc requirement in daily diet of Chinese preschool children should be 9.23 ± 2.35 mg/d (6.88-11.58 mg/d).
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Abstract
Introduction
Abstract
