HRMS as a Tool for Targeted Stabilization and Improved LLOD

Description: Presented are two case studies encountered during novel method development and validation in recent months. In both cases, high-resolution mass spectrometry is used as a critical tool in evaluating and successfully validating methods that contain complex interferences and stability challenges by allowing for biotransformation identification, lower limits of detection, improved signal to noise, and allow for targeted stabilization efforts by identifying the mechanisms of instability. In case study 1, we observe poor prodrug stability but are able to target and identify the degradation product from accurate mass data in addition to improving the S/N ratio of the analyte from triple quadrupole data to allow for proper quantitation of the targeted lower limit of quantitation. In case study 2, we utilize HRMS to simultaneously monitor a very complex set of ex-vivo reactions involving auto-redox of a quinone and hydroquinone pair and instability of in-vivo glucuronides to successfully chart a path to validation and maintaining the equilibrium of samples from the moment they are collected from the live-phase site. Both methods, while outwardly appearing as a conventional small molecule validation, required intense development and refinement that would not have been possible without the use of HRMS technology and highlight new and complex challenges being encountered in the industry.