Evaluation of UV-HPLC and mass spectrometry methods for specific activity determination.

Abstract

The specific activity (SA) values determined using two different methods were compared for a set of tritium-labeled and carbon-14-labeled compounds. The methods employed were as follows: (a) liquid chromatography/mass spectrometry (LC/MS) isotopic peak intensity distribution, and (b) determination of the tracer mass concentration using ultraviolet-high-performance liquid chromatography analysis coupled with the radioactive solution concentration measured by liquid scintillation counting. In general, at lower SA, the accuracy and or precision of the LC/MS-determined SA value decreased significantly. Because of this decrease in accuracy, a rough guideline of ~10% of the theoretical maximum SA is recommended as the lower cutoff for MS-based SA measurements. If the tracer contains heteroatoms that possess significant percentages of heavy isotopes at natural abundance (e.g. Cl and Br), then the MS-based SA cutoff recommendation is approximately 25-30% of the fully labeled compound in the tracer mixture. Additionally, IsoPat(2) was found to be the preferred calculation method for LC/MS-based SA determination because SA values via this program were more consistent with those obtained by ultraviolet concentration calibration with solution count.