Development of a compartmental model of human zinc metabolism: identifiability and multiple studies analyses.

A compartmental model of zinc metabolism has been developed from stable isotope tracer studies of five healthy adults. Multiple isotope tracers were administered orally and intravenously, and the resulting enrichment was measured in plasma, erythrocytes, urine, and feces for as long as 3 wk. Data from total zinc measurements and model-independent calculations of various steady-state parameters were also modeled with the kinetic data. A structure comprised of 14 compartments and as many as 25 unknown kinetic parameters was developed to adequately model the data from each of the individual studies. The structural identifiability of the model was established using the GLOBI2 identifiability analysis software. Numerical identifiability of parameter estimates was evaluated using statistical data provided by SAAM. A majority of the model parameters was estimated with sufficient statistical certainty to be considered well determined. After the fitting of the model and data from the individual studies using SAAM/CONSAM, results were submitted to SAAM extended multiple studies analysis for aggregation into a single set of population parameters and statistics. The model was judged to be valid based on criteria described elsewhere.

Abnormalities in zinc homeostasis in young infants with cystic fibrosis.

Low plasma zinc concentrations have been reported in approximately 30% of young infants with cystic fibrosis identified by newborn screening. The objective of this study was to examine zinc homeostasis in this population by application of stable isotope methodology. Fifteen infants with cystic fibrosis (9 male, 6 female; 7 breast-fed, 8 formula-fed) were studied at a mean (+/-SD) age of 1.8 +/- 0.7 mo. On d 1, 70Zn was administered intravenously, and 67Zn was quantitatively administered with all human milk/formula feeds during the day. Three days later, a 3-d metabolic period was initiated, during which time intake was measured and complete urine and fecal collections were obtained. Fractional zinc absorption, total absorbed zinc, endogenous fecal zinc, and net absorbed zinc were measured; fecal fat excretion was also determined. Fractional absorption was significantly higher for the breast-fed infants (0.40 +/- 0.21) compared with the formula-fed group (0.13 +/- 0.06) (p = 0.01), but with the significantly higher dietary zinc intake of the formula-fed group, total absorbed zinc was higher for those receiving formula (p = 0.01). In 1 infants with complete zinc metabolic data, excretion of endogenous zinc was twofold greater for the formula-fed infants (p < 0.05); net absorption (mg zinc/d) was negative for both feeding groups: -0.04 +/- 0.52 for breast-fed; -0.28 +/- 0.57 for formula-fed. Endogenous fecal zinc losses correlated with fecal fat excretion (r = 0.89, n = 9, p = 0.001), suggesting interference with normal conservation of endogenously secreted zinc. These findings indicate impaired zinc homeostasis in this population and suggest an explanation for the observations of suboptimal zinc status in many young infants with cystic fibrosis prior to diagnosis and treatment.

Estimation of doubly labeled water energy expenditure with confidence intervals.

Bivariate regression is used to estimate energy expenditure from doubly labeled water data. Two straight lines are fitted to the logarithms of the enrichments of oxygen-18 and deuterium simultaneously as a bivariate regression, so that the correlations between the oxygen and deuterium regression coefficients can be estimated. Maximum likelihood methods are used to extend bivariate regression to unbalanced situations caused by missing observations and to include replicate laboratory determination from the same urine samples, even if one of the replicates is missing. Use of maximum likelihood allows the determination of a confidence interval for the energy expenditure based on the log likelihood surface rather than use of the propagation of variance methods for nonlinear transformations. The model is extended to include the subject's deviations from the two lines as a bivariate continuous-time first-order autoregression to allow for serial correlation in the observations. The analysis of data from two subjects, one without apparent serial correlation and one with serial correlation, is presented.

Zinc homeostasis in healthy infants fed a casein hydrolysate formula.

BACKGROUND: The results of earlier, nonquantitative studies suggested that absorption of zinc from a semielemental (casein hydrolysate) formula was inferior to absorption from a cow's milk-based formula. The objective of this study was to compare fractional, total and net zinc absorption, and fecal excretion of endogenous zinc in the same healthy young infants when fed a casein hydrolysate versus cow's milk-based formula. METHODS: Fractional absorption of zinc and fecal excretion of endogenous zinc were determined from measurement of cumulative fecal excretion of unabsorbed tracer and by an isotope dilution technique, respectively, after oral administration of a 70Zn tracer with all formula feedings for 1 day. Six infants were assigned randomly to receive the test or control formula, and the other formula was administered 2 to 5 weeks later. RESULTS: Mean (+/-SD) fractional absorption of zinc from the casein hydrolysate formula (0.47 +/- 0.17) was double that from the cow's milk-based formula (0.22 +/- 0.04; P = 0.01) with a correspondingly greater total zinc absorption (3.23 +/- 1.67 mg Zn/day vs. 1.55 +/- 0.55 mg Zn/day; P = 0.05). Because the excretion of endogenous zinc in the feces did not differ between formulas (0.90 +/- 0.44 mg Zn/day vs. 0.91 +/- 0.29 mg Zn/day), net absorption of zinc was also higher with the casein hydrolysate formula (2.33 +/- 1.65 mg Zn/day vs. 0.81 +/- 0.67 mg Zn/day; P = 0.02). CONCLUSIONS: Retention of zinc appeared to be adequate to meet the needs for growth during feeding with cow's milk-based formula and was more than adequate during short-term feeding with the casein hydrolysate formula.

Human zinc metabolism: advances in the modeling of stable isotope data.

Compartmental modeling is a useful tool for investigating metabolic systems and processes. We and others have applied it to the study of zinc metabolism in humans. Because existing models could not be accurately fitted to our data, we have developed a new model of human zinc metabolism based on stable isotope tracer data from studies of five healthy adults. Multiple isotope tracers were administered orally and intravenously and the resulting enrichment measurement in plasma, erythrocytes, urine, and feces. These tracer kinetic data, along with other measured and calculated tracee and steady-state data, were used to develop the model. A single model structure composed of fourteen compartments was found to be suitable for all subjects. Model development and fitting of data and model for each subject were accomplished using the SAAM/CONSAM computer programs. The model development and fiting processes are described and exemplified using data from one of the subjects. While identifiability could not be demonstrated a priori due to the model's complexity, parameter statistics for the fitted models did show most parameters to be adequately identified a posteriori.

Elevated intakes of zinc in infant formulas don not interfere with iron absorption in premature infants.

BACKGROUND: Zinc and iron may share common pathways for absorption and compete for uptake into mucosal cells. We determined whether elevated ratios of zinc to iron would interfere with erythrocyte incorporation of iron in premature infants both during and between feeds. METHODS: In the first experiment, five premature infants (<2500 g birth weight) were enrolled, once receiving full oral feeds by nasogastric tube. They received either high (1200 microg/kg, ratio 4:1) or low (300 microg/kg, ratio 1:1) doses of oral zinc sulfate, together with 300 microg/kg oral 58Fe as chloride in saline with 10 mg/kg vitamin C, between designated feeding periods. Each infant served as its own control and randomly received either high or low doses of zinc or iron and then the alternate dose after 2 weeks. In the second experiment, nine additional premature infants were assigned to the same zinc:iron intake protocol except zinc and iron were given with usual oral feeds (premature formula or human milk) equilibrated before feeding. Iron absorption was measured by the erythrocyte incorporation of 58Fe. RESULTS: High doses of zinc given between feeds significantly inhibited erythrocyte incorporation of iron. 58Fe incorporation (%) with the 1:1 ratio of zinc:iron intake was 7.5 (5.7, 10; geometric mean, -I SD, +1 SD). The percentage of 58Fe incorporation on the 4:1 ratio of zinc:iron intake was 3.6 (2.6, 5.1). Given with feeds, the percentage of 58Fe incorporation on low zinc:iron intake was 7.0 (2.6, 19). Finally, the percentage of 58Fe incorporation on high zinc:iron intake was 6.7 (2.5, 19). CONCLUSION: Elevated intakes of zinc do not interfere with erythrocyte incorporation of iron in premature formulas.

Microwave method for preparing erythrocytes for measurement of zinc concentration and zinc stable isotope enrichment.

A microwave digestion method to prepare human erythrocytes for measurement of Zn concentration by atomic absorption spectrophotometry and stable isotope enrichment by mass spectrometry is described. Also described is a process for purifying digested erythrocyte samples enriched with Zn stable isotope for analysis by fast atom bombardment mass spectrometry. Microwave digestion was investigated as a way to increase sample throughput by replacing a more time-consuming conventional oven ashing/hot plate wet digestion method. Pooled red blood cells and NIST bovine liver standard reference material were digested by the two different methods and zinc recoveries compared by atomic absorption spectrophotometry. Microwave and conventional methods yielded 11.7 +/- 0.1 and 11.7 +/- 0.2 micrograms/g (wet wt), respectively, for the pooled erythrocytes, and Zn recovery from NIST bovine liver standard (certified 123 +/- 8 micrograms/g) was 128.2 +/- 1.2 and 127.4 +/- 1.3 micrograms/g, p > or = 0.282, respectively. Microwave digestion improved the processing of erythrocytes for atomic absorption spectrophotometry and mass spectrometry by reducing digestion time from 1 week to 2 h. In addition, a procedure for purifying digested erythrocyte samples by either extraction and ion-exchange chromatography in preparation for mass spectrometry analysis of Zn stable isotope enrichment is outlined.

Zinc homeostasis in breast-fed infants.

The needs for dietary zinc are adequately met by most fully breast-fed infants despite relatively low zinc intakes in relation to estimated requirements. The objective of this study was to use stable isotope techniques to evaluate how zinc retention is achieved in normal fully breast-fed infants. Nine male infants, aged 2-5 mo, were fed expressed human milk labeled with 70Zn over a 24-h period. Complete fecal collections were obtained for 8 d. On d 4-7, a metabolic period was initiated which included test weighing and milk sampling, to measure zinc intake, and daily urine collections. Isotopic enrichment of fecal and urine samples was determined by fast atom bombardment mass spectrometry. Results included a mean (+/- SD) dietary zinc intake of 17.8 +/- 6.6 mumol/d; fractional absorption of 0.54 +/- 0.075; and total absorbed zinc of 9.5 +/- 3.5 mumol/d. Mean endogenous fecal zinc, determined on seven infants by isotope dilution, was 4.7 +/- 2.3 mumol/d, which resulted in a mean net absorption of 4.8 +/- 3.4 mumol/d. The results of the study indicated that, for fully breast-fed infants, it is the combination of a relatively high fractional absorption and efficient conservation of intestinal endogenous zinc that results in zinc retention adequate to meet the demands of growth in the face of modest intake.

Zinc absorption in premature infants: comparison of two isotopic methods.

The fractional absorption of an oral dose of zinc can be measured in adults when given simultaneously with an intravenous dose and subsequently measuring the ratio of the double isotopic enrichment of urine. To test this method in very-low-birth-weight (VLBW) premature infants [n=5 females and 7 males, 1160 +/- 290 g (chi +/- SD) birth weight, 29 +/- 4 wk gestational age], an oral dose of either 300 or 1200 micrograms 68Zn.kg(-1).d(-1) was equilibrated with formula or human milk and administered simultaneously with either 50 or 100 micrograms 70Zn.kg(-1).d(-1) given intravenously 35 +/- 3 wk postconception. Urine and fecal samples were collected for 3-6 d and analyzed by inductively coupled plasma mass spectrometry. Endogenous fecal zinc (EFZ) was determined from isotopic enrichment, whereas net absorption and retention were calculated by traditional methods. The mean fractional absorption calculated from urine was 0.22 +/- 0.09 and from feces it was 0.25 +/- 0.07. Zinc intake averaged 1821 +/- 330, fecal excretion 1637 +/- 419, and urinary excretion 67 +/- 30 micrograms.kg(-1).d(-1). EFZ averaged 390 +/- 270 micrograms.kg(-1).d(-1) and ranged from 48 to 889 micrograms.kg(-1).d(-1). Net absorption was 220 +/- 316 micrograms.kg(-1).d(-1) and net retention was 131 +/- 334 micrograms.kg(-1).d(-1). True absorption was 373 +/- 161 micrograms.kg(-1).d(-1). Fecal collection is difficult, tedious, and often incomplete, and may be replaced by urine collection for the fractional absorption of zinc in groups of premature infants.

Zinc absorption and intestinal losses of endogenous zinc in young Chinese women with marginal zinc intakes.

The objective of this study was to determine fractional absorption of exogenous zinc and intestinal excretion of endogenous zinc in women of childbearing age whose habitual dietary zinc intake was marginal. The target population (L group) comprised residents of a remote farming village in northeast China and the control subjects (M group) were residents of Beijing. Mean (+/-SE) calculated dietary zinc intakes were 5.2 +/- 0.2 and 8.1 +/- 0.2 mg/d, respectively. The phytate-zinc molar ratio in the diet of both groups was approximately 10:1. 70Zn was administered intravenously before breakfast and 67Zn orally with three main meals in 1 d. Subsequently, all feces were collected quantitatively until the second visible marker had been excreted and 12-h urine samples were collected on days 3-9. Fractional absorption was determined by measuring cumulative fecal excretion of nonabsorbed 67Zn and endogenous fecal zinc by isotope-dilution technique (70Zn). Fractional absorption values for L and M groups, respectively, were 0.31 +/- 0.03 and 0.34 +/- 0.03 (P=0.45). Corresponding figures for endogenous fecal zinc were 1.30 +/- 0.07 and 2.34 +/- 0.20 mg Zn/d (P<0.001). Both the estimated total size of the pools of zinc that exchange with zinc in plasma within 2 d (r=0.762, P<0.001) and the excretion of endogenous zinc in the feces (r=0.706, P<0.0001) were positively correlated with calculated total daily zinc absorption. We conclude that fractional absorption of zinc does not differ between women consuming marginal and adequate quantities of zinc in their diets, but endogenous zinc is conserved effectively by the intestine in women whose habitual dietary zinc is marginal.

Occupation and visual/vestibular interaction in vestibular rehabilitation.

Otolaryngologists often prescribe head movement exercise programs for patients with vestibular disorders, although the effectiveness of these programs and the critical features of the exercises are poorly understood. Because many patients who dislike exercising do not follow through with their exercises, alternatives to the traditional repetitive exercises would be useful. Subjects diagnosed with vestibular disorders were treated for 6 weeks with either an outpatient exercise program that incorporated interesting, purposeful activities or a simple home program of head movements, comparable with the exercises otolaryngologists often give their patients when they do not refer to rehabilitation. Both treatments incorporated repetitive head movements in all planes in space, graduated in size and speed. Subjects were all tested before and after treatment with standard measures of vestibulo-ocular reflex and balance, level of vertigo, gross motor skills, and self-care independence. Subjects in both groups improved significantly on the functional measures, with slightly greater improvements in the occupational therapy group. The results were maintained 3 months after the cessation of intervention. These data suggest that graded purposeful activities are a useful alternative for treating this patient population and that the essential factor in any exercise program is the use of repetitive head movements.

Vestibular rehabilitation with graded occupations.

These three case reports are examples of the use of graded, purposeful activities in remediating the symptoms of vestibular disorders. Therapists can design individualized treatment plans for each patient by incorporating activities of interest to the patient. The treatment activities must include the particular head movements and positions that elicit vertigo during assessment. Additionally, activities must be interesting to the individual patient. Use of interesting activities may sustain the patient's interest and motivation for the treatment program and enable the patient to relate the learning process to real life experience. The principle that graded, repetitive head movement exercise is efficacious in reducing vertigo and disequilibrium in patients with vestibular disorders is now well accepted among physicians and therapists who treat these patients. The principle of adding meaning or purpose to otherwise rote exercise is well accepted among occupational therapists and is supported by evidence from the empirical literature (Heck, 1988; Kircher, 1984; Yoder, Nelson, & Smith, 1989). This concept of adding purpose to repetitive exercise may now be extended to the treatment of patients with peripheral vestibular disorders.

Size of the zinc pools that exchange rapidly with plasma zinc in humans: alternative techniques for measuring and relation to dietary zinc intake.

A relatively simple technique was used to estimate the size of the combined pools of zinc with which plasma zinc exchanges within 2 d (EZP). EZP size was determined from the amount of isotope introduced into the plasma and the coefficient of the simple exponential decay function fitting enrichment data between d 3 and 9 after isotope administration. Using data from 11 healthy adults, comparisons were made of EZP size estimations using oral and intravenous isotopes (r = 0.93 using urine) and urine and plasma enrichment (r = 0.85 for intravenous). EZP size estimations from urine and plasma enrichment following intravenous isotope administration were 2.35 +/- 0.84 and 2.83 +/- 0.86 mmol, respectively (mean +/- SD, P < 0.01). EZP size correlated with habitual dietary zinc intake (partial r = 0.74, P < 0.02). Cumulative declines in EZP size in two healthy adults after 3 wk of consuming a moderately zinc-restricted diet followed by 1 wk of severe zinc restriction were 26 and 32%. These results indicated that EZP size is dependent on dietary intake. We conclude that this technique is adequate to demonstrate EZP differences that are of nutritional and physiological interest. EZP size estimates can be obtained using orally or intravenously administered isotope and using plasma or urine enrichment data.

Development of techniques for the isolation of iron from biological material for measurement of isotope ratios by fast atom bombardment mass spectrometry.

Stable isotopes have gained prominence in nutrition and trace element research. Fast atom bombardment mass spectrometry (FAB-MS) measurement of iron isotope ratios has an accuracy of > or equal to 99.7% with the stable isotopes (54)Fe, (56)Fe, and (58)Fe with relative standard deviations (RSDs) of < or equal to 0.9%. The isolation of iron from biological matrices can be accomplished in 4 h with almost total removal of isobaric interferences caused by (39)KOH, (39)K.H2O, (40)CaOH, and/or (40)Ca.H2O. FAB-MS isotope enrichment measurements from this method compare favorably to predicted absorption/enrichment levels.

Influence of a meal and incremental doses of zinc on changes in zinc absorption.

The objective of this study was to determine the effect of increasing quantities of zinc, administered with and without a meal, on zinc absorption. Fractional absorption of incremental quantities of zinc in four normal adults was determined by measuring fecal excretion of unabsorbed isotope on 3 consecutive days by using three different stable isotopes of zinc (67Zn, 68Zn, and 70Zn). Isotopes were administered in the post-absorptive state and, on a subsequent occasion, with a standard zinc-free breakfast. In the postabsorptive state, fractional absorption was not affected by the quantity of zinc ingested until this exceeded 5 mg. When the zinc was administered with a meal, however, fractional absorption of 3 and 5 mg was less than for 1 mg. These results are compatible with the hypothesis that exogenous dietary zinc has to compete for absorption with endogenous zinc that has been secreted into the lumen of the gastrointestinal tract in response to a meal.

The analysis of stable isotopes in urine to determine the fractional absorption of zinc.

We measured isotopic enrichment in urine after oral and intravenous administration of stable isotopes of zinc to determine fractional absorption (FA). 68Zn and 70Zn were administered orally and intravenously to four normal adults. Subsequently, urine and fecal samples were collected for 7 and 14 d, respectively, ashed, and passed through ion-exchange columns to separate zinc from other elements. Samples were analyzed by fast-atom-bombardment mass spectrometry. From 32 h onwards the enrichment of 68Zn and 70Zn in urine declined proportionately so that FA could be determined as follows: FA = enrichment (oral/iv) x dose (iv/oral). FA determinations from urine and feces (cumulative excretion) were, respectively, for subject ZK1, urine 0.79 +/- 0.03 and feces 0.70 +/- 0.01; ZK2, 0.79 +/- 0.05 and 0.69 +/- 0.02; ZK3, 0.26 +/- 0.01 and 0.25 +/- 0.01; and ZK4, 0.41 +/- 0.02 and 0.37 +/- 0.02. ZK1 and ZK2 received the oral isotope while fasting whereas ZK3 and ZK4 received the oral isotope with meals. FA of zinc can be determined by measurement of isotope enrichment in urine.