Determination of Fe isotopes in blood by Multicollector ICP-MS: a feasibility study

In this study, we used a Thermo Neptune MC-ICP-MS to evaluate the optimal analytical conditions for Fe isotope measurements in blood. Isobaric interferences and definition of routines were evaluated to obtain the mass resolution in this instrument necessary to obtain results with adequate precision to investigate variations in the abundances of Fe isotopes with a view to characterizing clinical conditions of anemia. Almost all the iron in the body is located in hemoglobin. Hemoglobin is the component of red blood cells that allows them to transport oxygen and distribute it to the body's tissues. To achieve this objective, we used the commercially available reference material IRMM-014 (JM Fe) commercially available with high purity, easily soluble and available in large quantities supplied by the Institute for Reference Materials and Measurements (Geel, Belgium). It is enriched in the 57Fe isotope and it was used in this investigation to optimize instrumental conditions and evaluate instrumental reproducibility.

The Thermo Finnigan Neptune MC-ICP-MS is a dual-focus sector mass spectrometer equipped with nine Faraday detectors. Varying the entrance slit width allows for two different degrees of increased mass resolution (“medium” and “high” mass resolution), thus enabling measurements of the precise Fe isotopes abundances. Mass resolution is a measure of the relative width of an ion beam and, as such, the resolving power of an instrument of particular geometry to physically separate two ions of similar masses along their flight paths. In this sense, accurate corrections for even small amounts of isobaric elementary interferences (e.g., 54Cr+ in 54Fe+, 58Ni+ in 58Fe+, 64Ni+ in 64Zn+, etc.) were performed using the medium resolution mode. Using these criteria, we determined the external reproducibility for 56Fe, 57Fe and 58Fe with accurately adequate values, based on variable sample volumes and concentrations, transport gas flow and plasma intensity (power between 1000 and 1300 W). In addition, other instrumental parameters that were investigated during the test runs were focus lens settings, cycle integration times, total number of cycles during acquisition, and wash times. Optimal running conditions, including stability and signal intensity, ensure reproducibility of the 56Fe, 57Fe and 58Fe values of the samples and the JM Fe standard.

Blood samples were collected and prepared as follows: the first collection was performed on patients before ingestion of the solution with Fe isotopes enriched in mass 57Fe (JM Fe). The periodicity of blood collection after standard ingestion was 7 days, 13 days, 25 days and 35 days. The samples underwent hemoglobin concentration and dilution in 5% nitric acid. The Fe isotope data measured over a period of 35 days was plotted in a 57Fe/54Fe versus time diagram where the “natural” composition of the patients' blood could be observed before ingesting the solution with enriched 57Fe isotope, while the results of blood samples collected after ingestion of the enriched solution can be observed in increasing abundances until the 30th day, when it begins to decline. This pattern is interpreted as that of a healthy patient, demonstrated by the body's ability to absorb the Fe element from the food supply.

                These preliminary results point to the possibility of tests with different analysis purposes, with the collection of samples taken from patients in a non-invasive way, such as samples of hair, nails, urine and blood (among others), serving as a control group, or in research of population-based healthcare. In this sense MC-ICP-MS can be an alternative choice to analyze liquid samples with a rich matrix that predominantly needs to be mineralized and diluted before analysis, which could reduce sensitivity, prolong procedure time and increase the risk of contamination of the sample. Such samples can be successfully analyzed directly by MC-ICP-MS for different isotopes and in different matrices.