A Brief Introduction to Using Stable Isotopes
For the analysis of real-world samples, one can envision that there are a variety of competing effects in the ionization source due to the compounds present in the matrix (i.e. blood, urine) that compete with the target analytes for ionization. These may either enhance or suppress the signal due to the target analyte and would result in artificially reduced or elevated quantitative measurements, respectively. Some of these matrix effects are due to the presence of ionic species, buffer ions, and high concentrations of non-target analytes. Use of a stable-isotope internal standard is the best method to correct for such competing ionization processes. A known concentration of stable-isotope internal standard (for example a stable-isotope 2H or 13C labeled derivative of the target analyte) is spiked into the sample matrix (or solution to be analyzed) before analysis. The only thing that differentiates the stable-isotope internal standard from the target analyte is a small mass difference due to the degree of labeling. Generally, 13C labeling is preferred as there is always the possibility that hydrogen exchange with deuterium can occur in deuterated internal standards. Ideally, there should be no overlap between the natural abundances of the isotopic m/z ions in the target analyte and the labeled internal standard. For instance, testosterone produces a precursor ion corresponding to [M+H]+ at 289 m/z while the internal standard testosterone-[13C3] produces a unique precursor ion corresponding to [M+H]+ at 292 m/z.
Precursor ion spectra for testosterone
Precursor ion spectra for testosterone-[13C3]
Anthony Lagalante, Ph. D.
Consultant for IsoSciences, LLC
Associate Professor of Chemistry, Villanova University
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