For the development of reference measurement procedures, isotope dilution analysis offers the optimal possibility to quantify analytes in complex matrices as, from the moment in which the so-called spike is added, the spike and the analyte suffer the same fate. This allows possible losses or changes of the analyte during sample preparation to be compensated for.
To make optimal use of this property in species-specific IDMS, spike and analyte have, ideally, the same composition, as in speciation analysis not the whole sample is digested before determination. Due to the complex matrix and the necessary preservation of the structure of the species, sophisticated sample preparation with precipitation steps, derivation and chromatographic separation is often required. This includes the great risk that the analyte will be lost or will react in an undesired way. If the spike is added before any sample preparation and if it has the same properties as the analyte, it also reacts in the same way. As the isotopic ratio in the sample now no longer changes during sample preparation, and as, in the end, only the isotopic ratio is determined in the MS (and not the absolute mass concentrations), losses and changes can be compensated for.
Nevertheless, it is often difficult, especially in clinical chemistry, to synthesise the analyte with changed isotopic composition. If the elements which are used for the detection and quantification are bound in the protein in a covalent way, isotope labelling is possible only by offering the isotopically enriched starting material to the bacterium (for example by Escherichia Coli) in the culture medium. Such a procedure is cost-intensive and requires the DNA sequence to be available as a template and that it can be implemented in the bacterium DNA. For analytes for which the synthesis in an isotopically enriched form is not possible, another form of the IDMS is available: postcolumn IDMS. Here, the spike is added only after separation (i.e., post-column). The disadvantage here is that the yields for each sample preparation step have to be determined exactly, as now losses and conversions cannot be compensated for by the spike. This method is, however, well-suited to characterise spike materials and to determine the column recovery.
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