Bill T. G. Ting

Zinc and copper nutritional studies in very low birth weight infants: comparison of stable isotopic extrinsic tag and chemical balance methods

ABSTRACT: Measurements of dietary zinc and copper absorption obtained after administration of a single dose of the extrinsic stable isotopic tags 70Zn and 65Cu were compared to measurements made with standard chemical balance methods in 41 appropriate for gestational age premature infants [body wt 1267 ± 258 g, gestational age 29.8 ± 1.9 wk (mean ± SD), 4 to 83 postnatal d of age]. Fifty studies were performed; 33 with premature formula, five with term formula, seven with preterm human milk (PTHM), and five with fortified-PTHM. The percentages of net zinc and 70Zn absorption were found to be significantly greater from PTHM (66.4 ± 15.2, 68.6 ± 9.8) than from premature formula (14.0 ± 29.9, 31.6 ± 22.4), and term formula (23.6 ± 18.5, 17.6 ± 5.6). The percentages of net copper and 65Cu absorption were also found to be significantly greater from PTHM (61.5 ± 14.0, 69.8 ± 14.0) than from premature formula (16.6 ± 20.6, 39.6 ± 21.6) and term formula (20.6 ± 24.1, 26.5 ± 6.9). The percentages of net zinc and 70Zn absorption (35.9 ± 29.1, 48.4 ± 9.6) and net copper and 65Cu absorption (38.7 ± 10.2 and 57.4 ± 13.1) from fortified PTHM were similar to values from PTHM. Absorption of zinc and copper determined with extrinsic stable isotopic tag and standard nutrient balance methods were significantly correlated. Estimates of endogenous fecal losses of zinc and copper were substantial with each diet, but lower with PTHM. Stepwise, multiple linear regression analysis accounted for, at most, 58% of the variability in the measures of zinc and copper availability. We conclude that extrinsic 70Zn and 65Cu tags can be used to study absorption of dietary zinc and copper by very low birth wt infants.

Erythrocyte Incorporation of Ingested 58-Iron by Infants

ABSTRACT: The least abundant stable isotope of iron, 58Fe (natural abundance 0.322 weight %), was administered orally to infants to explore the feasibility of using a stable rather than a radioisotope in studies of iron absorption. The dose of 58Fe was given between feedings at age 126 days. The mass isotope ratio, 58Fe/57Fe, was determined in blood by inductively coupled plasma mass spectroscopy and at ages 140, 168, and 196 days. The percentage of the 58Fe dose entering the circulation (3.2 to 16.0%) was inversely correlated with serum ferritin concentration (r = −0.867, p < 0.01). For individual infants the SD of the percentage of administered dose of iron appearing in the circulation ranged from 0.22 to 1.28. We conclude that the method is likely to be suitable for within-subject comparisons of iron availability from foods. Because of the large between-subject variation, we are pessimistic for this age group about the usefulness of study designs based on group comparisons.

Iron absorption and incorporation into red blood cells by very low birth weight infants : studies with the stable isotope 58Fe

Summary Measurements of iron absorption and incorporation into RBCs were obtained with the stable isotope 58Fe, administered as a reference dose, in 11 premature infants with birth weights between 780 and 1,520 g and gestational ages between 24 and 33 weeks. Each study included a timed stool and urine collection, nasogastric tube administration of a single dose of about 228 μmlg of 58Fe/kg of body weight (as FeSO4, with 10 mg/kg of vitamin C) between feedings, and blood samples before 58Fe (day 1) and then 2 weeks (day 15) later. Gastrointestinal absorption of the 58Fe dose as measured by fecal isotope balance was 41.6 ± 17.6% (mean ± SD). However, only 12.0 ± 9.6% of the 58Fe dose (28.7 ± 22.3% of the absorbed 58Fe dose) was incorporated into RBCs on day 15. 58Fe absorption and 58Fe incorporation into RBCs on day 15 were significantly correlated with the hemoglobin concentration and reticulocyte count on day 1. Transfusion history did not affect 58Fe absorption or 58Fe incorporation into RBCs. We conclude that concurrent measurement of 58Fe absorption with fecal monitoring and of 58Fe incorporation into RBCs permits a better understanding of the fate of iron ingested by premature infants than either measurement alone.

Isotopic determination of selenium in biological materials with inductively coupled plasma mass spectrometry

A method for the isotopic determination of selenium in biological matrices is described. The method is based on hydride generation inductively coupled plasma mass spectrometry (ICP-MS). The development is specifically related to the requirements of stable isotope tracer studies in human subjects. The method is based on isotope dilution using 82Se as the in vitro spike and can quantify the 74Se and 77Se contents of samples. It involves wet oxidation (HNO3 - H2O2 or HNO3 - HClO4) of the 82Se-spiked matrix, reduction to selenite by boiling with HCl followed by measurement of the isotope ratios (82Se/77Se and 74Se/77Se) in the gas stream (H2Se) generated from on-line reduction of the sample selenite with NaBH4. Compared with the isotopic signal resulting from a selenite solution containing 5 ng ml-1 of Se, the total sample blank contributions at m/z = 74, 77 and 82 were less than 5% of the respective isotope signal. Worst-case absolute detection limits were 0.2-0.9 ng of Se, depending on the isotope used. Ion beam intensity ratios were measured with an over-all precision [relative standard deviation (RSD)] of 1% for both isotope pairs. Measured ratios (MRa/b) were stable during a given days operation within the expected precision of the measurements but varied for different days. The magnitude of MRa/b was generally independent of the nature of the matrix. Highly linear relationships were found between ion beam intensity ratios (MRa/b) and the corresponding true isotope ratios for calibration solutions whose isotope ratios had been altered by as much as one order of magnitude.(ABSTRACT TRUNCATED AT 250 WORDS)

Optimisation of instrumental parameters for the precise measurement of isotope ratios with inductively coupled plasma mass spectrometry

The effect of several instrumental operating parameters (IOPs) on the precise and accurate measurement of isotope ratios was investigated. The IOPs of interest were: dwell time (DT) and cycle time (CT) of the software control parameters, the r.f. power level, the analyte solution flow-rate and the argon gas plasma, auxiliary and nebuliser flow-rates. Two measurement criteria were evaluated for each isotope pair (a,b) in relation to these independent variables: the precision (RSD) of the isotope ratio measurements and the deviation (Δ) of the measured isotope ratio (MRa/b) from a reference value (MR0a/b). The isotope pairs of interest were: 6Li : 7Li, 57Fe : 54Fe, 58Fe : 54Fe, 65Cu: 63Cu, 67Zn : 68Zn and 70Zn : 68Zn.It was shown that the best precision (lowest RSD) and most stable measurements (lowest Δ) were generally obtained over a relatively wide range of r.f. powers, while the range of r.f. power corresponding to the maximum ion-beam intensity was much narrower. The optimum r.f. power for the best results was Li, 1170; Fe, 1240; Cu, 1250; and Zn, 1290 W. Argon gas flow-rates corresponding to the optimum values for %RSD and %Δ were: plasma flow-rate 15 for Li and 12 l min–1 for the other elements, auxiliary flow-rate 1.7 for Li and 1.8–1.9 l min–1 for the other elements, and nebuliser pressure 42 lb in–2 for all elements. The analyte solution flow-rate giving the best results was investigated for Cu only (best value 0.91 ml min–1).Employing the optimum settings of the IOPs, the long-term stability (5 h of continuous operation) of the measurements was investigated. It was shown that the ion-beam intensities were stable to better than 8%. In contrast, values of isotope ratios (MRa/b) always stayed within the expected measurement precision [RSD<1% for all except Li (<1.5%)].It was concluded that inductively coupled plasma mass spectrometry (ICP-MS) permitted the precise and accurate measurement of stable isotope ratios for routine application in the studies of trace-element metabolism in man employing the concept of stable isotope tracers.

Accurate measurement of stable isotopes of magnesium in biological materials with inductively coupled plasma mass spectrometry

A method is described for the accurate isotopic determination of magnesium (24Mg, 25Mg, 26Mg) in biological materials, which is based on inductively coupled plasma mass spectrometry (ICP-MS). The analytical performance of the method was examined with respect to the requirements of stable isotope tracer studies. When applied to the measurement of base-line isotope ratios (MRa/b) in rat tissue the following results were obtained (data are given in terms of MR25/24 and MR26/24± 1RSD, n= 4 or 5): standard solution of Mg (0.05 µg ml–1), 0.1394 ± 0.6, 0.1697 ± 0.8; bone, 0.1401 ± 0.5, 0.1706 ± 0.8; brain, 0.1404 ± 0.2, 0.1720 ± 0.3; kidney, 0.1392 ± 0.5, 0.1702 ± 0.6; liver, 0.1388 ± 0.4, 0.1696 ± 0.5; muscle, 0.1396 ± 0.2, 0.1716 ± 0.4; plasma, 0.1385 ± 0.3, 0.1691 ± 0.3; red cells, 0.1383 ± 0.1, 0.1694 ± 0.2; and urine, 0.1402 ± 0.1, 0.1721 ± 0.3. The measurement precision for replicate analyses of each matrix was in the range 0.1–1.0% and the mean value of the isotope ratio for different matrices agreed with the corresponding ratio for the standard solution to within 1.5%. When measured over a 10-h period, the isotope ratios appeared to be independent of the observed drifts in the ion beam intensities. The instrument blank contribution to the ion beam intensities was about 0.1% of the values obtained for a solution containing 0.05 µg ml–1 of natural Mg. The absolute detection limit for Mg (based on the experimental standard deviation of blanks run over 10 h) was <2 ng for all three stable isotopes. The ICP-MS phase of the analysis was capable of making 50–150 separate measurements of both ratios in a 10-h period, depending on the desired level of measurement precision within the range 0.1–1.0%. A complete analytical scheme for the accurate determination of the three stable isotopes of Mg, which is based on precipitation with ammonium phosphate, is described. The accuracy of the method was tested using the standard reference materials Bovine Liver (NBS 1577a, 600 ± 15 µg g–1 of Mg) and Animal Bone (IAEA H-5, 3550 ± 90 µg g–1). The proposed method provided the following data (µg g–1): SRM 1577a, 617 ± 4 and IAEA H-5, 3585 ± 16. For other biological matrices of interest, the accuracy of the method was compared with atomic absorption spectrometry. The complete analytical procedure, up to the point of mass spectrometric measurement, can be performed on about 20 samples per working day.

Accurate analysis of stable isotopes 68Zn, 70Zn, and 58Fe in human feces with neutron activation analysis

Accurate isotopic analysis of human fecal samples is discussed in relation to mineral bioavailability studies of human subjects. It is shown that for bioavailability investigations of Zn and Fe, the method of neutron activation analysis is suitable for routine application to clinical experiments; and the human fecal samples resulting from diet suitably enriched in isotopes 70Zn and 58Fe can be routinely analyzed with relative precisions of about 1% for the isotopes 68Zn, 70Zn and 58Fe.

Selenium absorption and retention by very-low-birth-weight infants : studies with the extrinsic stable isotope Tag 74Se

Measurements of dietary selenium absorption and retention were obtained after administration of a single dose of the extrinsic stable isotope tag 74Se in 20 appropriate for gestational age premature infants with birth weights between 720 and 1,630 g and gestational ages between 26 and 33 weeks. Infants were assigned randomly to receive a standard premature formula (1.34 μg of Se/dl) or a selenium-supplemented version of that formula (2.03 μg of Se/dl). Each study consisted of one feeding that had been extrinsically labeled with 74Se (1.03 μg/kg) and a timed stool and urine collection. The percent 74Se absorption was 91.2 ± 5.4% (mean ± SD) from the standard formula and 86.2 ± 3.0% from the selenium-supplemented formula (p < 0.05), but the percent of the absorbed 74Se retained was not different, i.e., 96.6 ± 2.1% and 95.0 ± 2.8%, respectively. The percent net absorption and net retention were also not different between the standard and selenium-supplemented formulas; net absorption was 72.7 ± 18.1% vs. 67.8 ± 18.8% and net retention was 57.2 ± 17.6% vs. 53.3 ± 20.2%, respectively. The percent 74Se absorption and true selenium absorption were significantly correlated with the percent net selenium absorption and net selenium absorption, respectively. We conclude that an extrinsically administered dose of 74Se can be used to study selenium nutrition in growing premature infants.

Measurement of stable isotopes of bromine in biological fluids with inductively coupled plasma mass spectrometry

A method is reported for the accurate measurement of the two stable isotopes of bromine in biological fluids of interest in human metabolic studies. The method is based on inductively coupled plasma mass spectrometry (ICP-MS).It is shown that the background ion beam intensities at m/z= 79 and 81 are typically in the range 70–335 and 600–7200 ions s–1, respectively, when de-ionised water is aspirated into the plasma. The corresponding range for 1.0 µg ml–1 of natural Br is 9700–18 500 ions s–1 at m/z= 79. The detection limit (3[graphic omitted]) for Br is in the range 2–5 ng ml–1. A method is given for automatic correction of the argon contribution at m/z= 81.Data are presented which show that the isotope ratio 81Br/79Br can be measured routinely with a precision (relative standard deviation) of 1% or better. The measured ratio is independent of the Br concentration in the range 3–20 µg ml–1. Linear regression equations are obtained for stable isotope calibration graphs over the range 0.997–5.322 (MIR81/79). However, the slopes of these plots deviate considerably from the expected value of one.Two chemical separation schemes are described, Scheme I, based on cation exchange and Scheme II, based on distillation from acidified solutions. The former is applicable to plasma (and possibly saliva) samples whereas the latter is successful for urine. The presence of large amounts of sulphate produces significant enhancement of the ion intensity at m/z= 81 (due to 32S16O31H+). Distillation permits the required separation of Br from sulphate, whereas precipitation with Ba(NO3)2 does not appear to be satisfactory.Application of the method of standard additions and stable isotope dilution analysis to samples of urine from several subjects indicates that this method permits quantitative analysis of bromine to be carried out with a precision (and accuracy) of about 2%.

Determination of thorium and uranium in urine with inductively coupled argon plasma mass spectrometry

An accurate and simple method has been developed for the determination of thorium and uranium in urine using inductively coupled argon plasma mass spectrometry (ICP-MS). Determination of thorium and uranium was by external calibration using matrix matched standards and high-purity spiking materials. Aliquots of each urine specimen were diluted (1 + 9) with 0.2 mol l–1 nitric acid containing iridium as an internal standard. The counts at m/z 232 (thorium), 238 (uranium) and 193 (iridium) were measured, and ratios of the counts at m/z 232 or 238 to those at m/z 193 were calculated. These ratios were compared with those from urine-based calibration standards to calculate the thorium and uranium concentrations in unknown specimens. The concentrations of thorium and uranium were calculated as µg l–1 in the sample and also corrected for dilution via creatinine measurement, expressed as µg g–1 of creatinine. The method has been evaluated by determination of reference materials from the Los Alamos National Laboratory, as well as of those from the Oak Ridge National Laboratory. The proposed method provides the basis of an accurate method for determining thorium and uranium in unexposed subjects as well as in those considered to be exposed to thorium or uranium through environmental or other pathways. About 40 specimens, excluding blanks, calibration standards and quality-control materials, can be processed in an 8 h day.