Use of Stable Isotopes to Evaluate Bioefficacy of Provitamin A Carotenoids, Vitamin A Status, and Bioavailability of Iron and Zinc

Abstract

The ability of nutrition scientists to measure the status, bioavailability, and bioefficacy of micronutrients is affected by lack of access to the parts of the body through which a nutrient may travel before appearing in accessible body compartments (typically blood or urine). Stable isotope-labeled tracers function as safe, nonradioactive tools to follow micronutrients in a quantitative manner because the absorption, distribution, metabolism, and excretion of the tracer are assumed to be similar to the unlabeled vitamin or mineral. The International Atomic Energy Agency (IAEA) supports research on the safe use of stable isotopes in global health and nutrition. This review focuses on IAEA's contributions to vitamin A, iron, and zinc research. These micronutrients are specifically targeted by the WHO because of their importance in health and worldwide prevalence of deficiency. These 3 micronutrients are included in food fortification and biofortification efforts in low- and middle-income regions of the world. Vitamin A isotopic techniques can be used to evaluate the efficacy and effectiveness of interventions. For example, total body retinol stores were estimated by using 13C2-retinol isotope dilution before and after feeding Zambian children maize biofortified with β-carotene to determine if vitamin A reserves were improved by the intervention. Stable isotopes of iron and zinc have been used to determine mineral bioavailability. In Thailand, ferrous sulfate was better absorbed from fish sauce than was ferrous lactate or ferric ammonium citrate, determined with the use of different iron isotopes in each compound. Comparisons of one zinc isotope injected intravenously with another isotope taken orally from a micronutrient powder proved that the powder increased total absorbed zinc from a meal in Pakistani infants. Capacity building by the IAEA with appropriate collaborations in low- and middle-income countries to use stable isotopes has resulted in many advancements in human nutrition.

FIGURE 1

The 3 isotopic forms of carbon. ¹²C is the most common of the isotopic forms (98.9%) and ¹³C is the stable form (1.1%); ¹⁴C is the radioactive form and is only found in trace amounts in nature.

FIGURE 2

The 4 most common stable isotope–labeled vitamin A structures. These molecules are usually administered as the retinyl acetate compounds in retinol isotope dilution and compartmental modeling studies. Top to bottom: 10, 19, 19, 19-D₄-retinol; 10, 19, 19, 19, 14, 20, 20, 20-D₈-retinol; 14,15–¹³C₂-retinol; and 8, 9, 10, 11, 12, 13, 14, 15, 19, 20-¹³C₁₀-retinol. D, deuterium.

FIGURE 3

For vitamin A status assessment, a dose of vitamin A ester labeled with stable isotope is administered after a baseline blood sample. A period of the dose mixing with the vitamin A body pool is necessary before the follow-up blood sample is taken for analysis by MS (adapted from reference 28). Prediction equations use the data from the mass spectrometer, along with key assumptions about absorption, storage, and catabolism of dose, to estimate total body retinol stores (29).

FIGURE 4

Study design for the paired-isotope dilution method in a highly controlled, 90-d feeding intervention with a high provitamin A–containing food, such as vegetables, a fortified food, or a biofortified staple crop. Negative (placebo) and positive (supplemental preformed vitamin A) control groups are recommended. If both control groups are included, a bioefficacy factor can be estimated for provitamin A–containing foods. RID, retinol isotope dilution.

FIGURE 5

Design used for iron bioavailability studies to compare the quantities of iron absorbed and used between 2 test meals (based on references ^†, ^†, and 99). Two blood samples are required to determine the incorporation of the isotopes into the erythrocytes. After sample preparation, the iron is analyzed with an appropriate mass spectrometer and ratios of the stable isotopes are compared before and after dosing to determine the amount of iron incorporated into the RBCs. ^† following a citation number indicates a work that the International Atomic Energy Agency has supported on stable isotopes.

FIGURE 6

Design for a zinc-bioavailability study to determine the difference in absorption of zinc from typical wheat and a zinc-enhanced variety [based on a study evaluating tortillas (^†)]. Zinc isotopes are administered orally with the test food on days 1 and 2 and intravenously on the first day. Urine samples are collected on days 4 through 8 (2/d) and analyzed with an appropriate mass spectrometer. ^† following a citation number indicates a work that the International Atomic Energy Agency has supported on stable isotopes.