Characterization and demonstration of drug compound ring-chain tautomer formation and its impacts on quality control.

Unknown chromatographic peaks, potential impurities, were observed in a series of related compounds. This led to the identification and characterization of tautomeric equilibria. Structural elucidation was required to understand the potential impurity profile, thus impacting method development for quality control. In this work, characterization of the chemical structures, AZ13581258 and AZD5718, and equilibria of the tautomeric forms was performed using a range of advanced analytical techniques such as preparative chromatography, nuclear magnetic resonance (NMR), chromatographic detection by mass spectrometry (MS), MSMS, and ultraviolet spectroscopy (UV). Predictions using density functional theory (DFT) further explains and confirms the tautomer equilibria through predictions of reaction barrier energies, UV-spectra and NMR data. These investigations led to fully understand the impurity profile and to the development of a quality control method for AZD5718 drug substance and drug product. In conclusion, ring-chain tautomeric structures are predominately formed under acidic conditions, and the additional peaks observed in LC during organic impurity determination were found to originate from ring-chain closed tautomers in equilibria with the parent open form compound. Hence, the closed and open tautomer forms should all be considered as the same compound.

HPLC-UV-MS Analysis: A Source for Severe Oxidation Artifacts.

HPLC coupled to both UV and MS is an established setup for purity assessments in many areas. With evolving technology, instrument sensitivity increases, calling for lower sample concentration, while light flux in a commercial UV detector cell is considerably higher than earlier. This evolution has now reached the point where radicals formed by UV light are abundant enough, compared to the analyte levels, to generate unwanted artifact signals in the MS spectrum. In this work we show several examples from pharmaceutical development where UV degradation in the UV detector leads to severely misleading mass spectra in typical day to day samples.