Characterization of cladribine and its related compounds by high-performance liquid chromatography/mass spectrometry

High-performance liquid chromatography/mass spectrometer (HPLC/MS) was used to identify and structurally characterize the modified nucleoside cladribine (2-chloro-2'-deoxy-beta-adenosine) and 13 synthesis-related byproducts in bulk drug. Confirmation of compound identity was accomplished by spectral analysis (1H and 13C NMR spectroscopy, mass spectrometry, and UV absorption spectroscopy) of the related compounds as isolated from crude mixtures of the drug substance and by spiking experiments with authentic standards. The use of on-line mass spectrometric analysis (i.e., LC/MS) to augment UV absorption spectra permitted rapid identification of many of the compounds of interest.