Peter Kastenmayer

Interlaboratory trial on the determination of the In Vitro iron dialysability from food

An in vitro method for the estimation of iron bioavailability was subjected to an interlaboratory trial. The method involved a simulated gastrointestinal digestion using pepsin for the gastric stage followed by pancreatin and bile salts for the intestinal stage. The proportion of iron diffused through a semipermeable membrane (molecular mass cut-off 10 kDa) was used to measure the iron dialysability. An interlaboratory trial between nine laboratories was conducted to evaluate the repeatability and reproducibility of the agreed method. The reproducibility of the method among the participating laboratories was 20-30% and depended on the content of dialysable iron. Several factors contributing to the variation in the in vitro dialysability among laboratories are discussed. The pH adjustment in the intestinal digestion was identified as one of the critical parameters. The present in vitro method was used to evaluate the iron dialysability from three meals. The dialysability data were in reasonable agreement with human absorption data. The usefulness of the in vitro dialysability method is discussed.

A double stable isotope technique for measuring iron absorption in infants

A stable isotope technique has been developed which uses 57Fe and 58Fe as labels and which enables the simultaneous measurement of Fe absorption from two test meals in infants. The method was evaluated by measuring Fe absorption from a commercial whey-adjusted infant formula in nine healthy infants aged 13-25 weeks. Each infant was fed 210 ml formula, labelled with either 57Fe or 58Fe, on four consecutive mornings, in random order. The total Fe content in each feed was 2.5 mg Fe; either as 2.5 mg 57Fe, or 0.6 mg 58Fe plus 1.9 mg Fe with normal isotopic composition. Isotopic enrichment of Fe in erythrocytes was measured by thermal ionization mass spectrometry 14 d after the last administration, and Fe absorption was calculated based on isotope ratio shifts, total circulating Fe and intake of each isotope. Geometric mean absorption for the 57Fe and 58Fe labels was 6.72 and 6.58% respectively, and the absorption of the two isotopes was not significantly different (Students paired t test). By this technique, paired comparisons of Fe absorption can be obtained and systematic studies of the influence of dietary factors on Fe absorption during infancy can be conducted.

Iron Bioavailability Studied in Infants: The Influence of Phytic Acid and Ascorbic Acid in Infant Formulas Based on Soy Isolate

ABSTRACT: The influence of phytic acid and ascorbic acid content of soy formula on iron (Fe) bioavailability was investigated in infants by analysis of the incorporation of stable isotopes of Fe into red blood cells 14 d after administration using a double stable isotope technique. Paired comparisons were made with each infant acting as his or her own control. The geometric mean fractional Fe incorporation into red blood cells increased from 5.5 to 6.8% (p<0.05) when soy formula with the native content of phytic acid was compared with a 83% dephytinized formula. A more pronounced effect was shown with soy formula containing no phytic acid; the mean fractional Fe incorporation increased from 3.9 (native phytic acid) to 8.7% (zero phytic acid; p < 0.001). A significant (p < 0.01) effect was also demonstrated when the Fe:ascorbic acid molar ratio in the native phytate-containing formula was increased from 1:2.1 to 1:4.2; mean fractional Fe incorporation increased from 5.9 to 9.6%. These results demonstrate that the Fe bioavailability from soy-based infant formulas can be similarly increased by either removing phytic acid or increasing the ascorbic acid content.

Influence of lactoferrin on iron absorption from human milk in infants.

ABSTRACT: Lactoferrin (Lf) is a major iron (Fe)-binding protein in human milk and has been proposed to facilitate Fe absorption. The potential effect of Lf on Fe absorption was investigated by measuring Fe absorption in infants fed breast milk (with its native content of Lf) and the same milk from which Lf had been removed (>97%) by treatment with heparin-Sepharose. Eight breast-fed infants (2–10 mo; mean age 5 mo) were fed 700 to 1000 g of each milk in a randomized, cross-over design with each child acting as his/her own control. The milk was labeled with 8.6 Mmol (0.5 mg) of 58Fe and Fe absorption was measured by quantifying the incorporation of the isotope into red blood cells 14 d after intake using thermal ionization mass spectrometry. Fractional Fe absorption was significantly lower (p < 0.05) from breast milk than from Lf-free breast milk. The geometric mean (range) was 11.8% (3.4–37.4%) for breast milk and 19.8% (8.4–72.8%) for Lf-free breast milk. These results do not support a direct role for Lf in the enhancement of Fe absorption from human milk at this age. In addition, Fe absorption (11.8%) from human milk fed over several feeds was lower than that previously reported for single feed studies.

A comparison of iron absorption in adults and infants consuming identical infant formulas

Fe absorption was estimated in adults and infants from the erythrocyte incorporation of Fe isotopes added to infant formula. Fe absorption was measured in adults using radioisotopes, and in infants with a stable-isotope technique. In adults, the geometric mean Fe absorption from a ready-to-feed soya formula with its native phytic acid content was 2.4%. This increased to 6.0% (P < 0.05) after almost complete dephytinization. In infants, mean Fe absorption values were 3.9 and 8.7% respectively from the same products (P < 0.05). In adults, mean Fe absorption from a spray-dried soya formula containing 110 mg ascorbic acid/l was 4.1%, increasing to 5.3% (P < 0.05) when ascorbic acid was doubled to 220 mg/l. In infants, mean Fe absorption values were 5.7 and 9.5% (P < 0.05) from the same products. Mean Fe absorption from a milk-based formula was 6.5% in adults compared with 6.7% in infants. All meals in the adult and infant studies were fed using an identical meal size of 217 g. Increasing the meal size threefold in adults did not change fractional Fe absorption. Mean Fe absorption values for each meal were lower in adults than in infants but the relative inhibitory effect of phytic acid and the enhancing effect of ascorbic acid were similar. We conclude that Fe absorption studies in adults can be used to assess the influence of enhancers and inhibitors of Fe absorption in infant formulas fed to infants. Further studies, however, are required to extend these findings to weaning foods and complete meals.

Mass spectrometric methods for studying nutrient mineral and trace element absorption and metabolism in humans using stable isotopes : a review

Mass spectrometric methods for determining stable isotopes of nutrient minerals and trace elements in human metabolic studies are described and discussed. The advantages and disadvantages of the techniques of electron ionization, fast atom bombardment, thermal ionization, and inductively coupled plasma and gas chromatography mass spectrometry are evaluated with reference to their accuracy, precision, sensitivity, and convenience, and the demands of human nutrition research. Examples of specific applications are described and the significance of current developments in mass spectrometry are discussed with reference to present and probable future research needs.

Zinc and calcium apparent absorption from an infant cereal: a stable isotope study in healthy infants.

Fractional apparent absorption of Zn and Ca from a wheat-milk-based infant cereal was studied in six healthy infants (18-30 weeks old). Mineral absorption was measured by a stable-isotope technique based on faecal excretion of the isotopes. Each test meal (40 g cereal) was extrinsically labelled with 70Zn and 42Ca before intake. All faecal material passed during the 21 d following intake of the labelled test meal was collected on trace-element-free nappies. Individual stool samples were analysed for their content of 70Zn and 42Ca by thermal ionization mass spectrometry. Apparent absorption was calculated as intake minus total faecal excretion of the isotopes over 68-92 h after administration. The fractional apparent absorption values for Zn and Ca were 33.9 (SD 16.4) % (range 19.2-63.9%) and 53.5 (SD 12.6) % (range 36.7-71.7%) respectively. Re-excretion of absorbed 70Zn (> 68-92 h to 21 d after intake of the labelled meal) was 0.44 (SD 0.38) % of administered dose while only one infant re-excreted detectable amounts of 42Ca (1.74% of administered dose). The analysis of individual stool samples confirmed that 72 h is a sufficient time period for complete collections of non-absorbed isotopes in faecal material from infants during the weaning period and that re-excretion of initially absorbed 70Zn and 42Ca (> 68-92 h to 21 d after intake of the labelled meal) is negligible.

Erythrocyte incorporation of iron by infants: iron bioavailability from a low-iron infant formula and an evaluation of the usefulness of correcting erythrocyte incorporation values, using a reference dose or plasma ferritin concentrations.

Bioavailability of iron (Fe) from a low-Fe infant formula was determined by erythrocyte incorporation of 58Fe 14 d after administration in ten healthy, non-Fe-deficient infants. Two feeding protocols were compared, with each infant acting as his/her own control. At 140 and 154 d of age, infants were fed 1000 g of 58Fe-labelled formula (1.44 mg total Fe/1000 g) as six feeds over 24 h (Protocol A) or as two feeds/day on three consecutive days (Protocol B). A water solution with 57Fe and ascorbic acid was given separately as a reference dose in both study protocols. Erythrocyte incorporation of 58Fe and 57Fe was determined by thermal ionisation mass spectrometry. Geometric mean 58Fe incorporation was 7.6% (range 3.3-13.5%) with Protocol A as compared to 10.6% (range 6.7-18.6%) with Protocol B (P = 0.05); paired t test. Inter-individual variability of 58Fe was not reduced by correcting for the incorporation of 57Fe from the reference dose, or by correcting for plasma ferritin concentration. Fractional erythrocyte incorporation of Fe from low-Fe infant formula was in the same range as our earlier published data on erythrocyte incorporation of Fe from human milk extrinsically labelled with 58Fe (Davidsson et al. 1994a). The methodological evaluations included in this study clearly indicate the importance of using standardised study protocols when evaluating Fe bioavailability in infants. Corrections of erythrocyte incorporation data based on plasma ferritin or erythrocyte incorporation of Fe from a reference dose were not found to be useful.

Influence of the consumption pattern of magnesium from magnesium-rich mineral water on magnesium bioavailability

It is generally considered that the absorption of Mg is inversely related to the ingested dose. The objective of the present study was to determine if the mode of administration (bolus v. consumption throughout the day) could influence Mg bioavailability from Mg-rich natural mineral water comparing the same nutritional Mg amount (126 mg). Using a 2 d cross-over design, twelve healthy men were asked to drink 1·5 litres Mg-rich mineral water either as 2 × 750 ml or 7 × 212 ml throughout the day. Two stable isotopes ((25)Mg and (26)Mg) were used to label the water in order to distinguish both regimens. Fractional apparent Mg absorption was determined by faecal monitoring and Mg retention was determined by measuring urinary excretion of Mg isotopes. Higher Mg absorption (50·7 (SD 12·7) v. 32·4 (SD 8·1) %; P = 0·0007) and retention (47·5 (SD 12·9) v. 29·0 (SD 7·5) %; P = 0·0008) from Mg-rich mineral water were observed when it was consumed in seven servings compared with larger servings. Thus, regular water consumption throughout the day is an effective way to increase Mg bioavailability from Mg-rich mineral water.

Magnesium enrichment and distribution in plants

Food products enriched with stable isotopes are used in nutrition to study the metabolic fate of nutrients in humans. This study reports on the labeling of green beans, white beans, soybeans and wheat with a stable isotope of magnesium (25Mg) obtained in greenhouse conditions for further studies on magnesium bioavailability. Soybean and green bean are the most efficient plant species to obtain large amounts of edible parts rapidly with a minimum loss of labeled Mg in other parts of the plants. The results obtained showed that a relatively high percentage of the magnesium found in seeds (grains/beans) can come from the redistribution of magnesium previously accumulated in other organs.