Characterization of stationary phases by a linear solvation energy relationship utilizing supercritical fluid chromatography.

Supercritical fluid chromatography was utilized in combination with the Abraham model of linear solvation energy relationship to characterize 11 different HPLC stationary phases. System constants were determined at one supercritical fluid chromatography condition for each stationary phase. The results indicate that several types of silica columns, including type B silica, type C silica, and fused core silica, are very similar in their retention behavior. Several aromatic stationary phases were characterized and it was found that, in contrast to the other phases studied, all of the aromatic stationary phases had positive contributions from the dispersion/cavity (v) term of the linear solvation energy relationship. Several aliphatic phases were characterized and there were several linear solvation energy relationship constants that differentiated the phases from each other, mainly the polar terms (dipolarity and hydrogen bonding). One stationary phase, a fused core pentafluorophenyl (PFP) phase, had very poor regression quality. The column volume of this phase was lower than the others in the study, which may have had some impact on the results of the regression.

Comparison of the sensitivity of evaporative universal detectors and LC/MS in the HILIC and the reversed-phase HPLC modes.

It was hypothesized that the hydrophilic interaction liquid interface chromatography (HILIC) mode should produce more response than the reversed-phase HPLC mode on detectors with an evaporative component to the detection process. HILIC mobile phases are mostly composed of polar organic solvent and are more volatile than reversed-phase mobile phases. Therefore the more easily evaporated HILIC mobile phases should produce greater sensitivity for those detectors that remove mobile phase by evaporation. The responses of 12 compounds were measured in the reversed-phase mode and the HILIC mode with three detectors: evaporative light scattering detector (ELSD), corona charged aerosol detector (cCAD), and electrospray mass spectrometry (ESI-MS). The compounds studied were very polar compounds that were retained in the HILIC mode. Generally, the HILIC mode was able to achieve greater sensitivity than the reversed-phase mode for these compounds. The increases in sensitivity observed can be attributed to the more volatile HILIC mobile phase. For the ELSD, the HILIC mode produced slightly greater sensitivity than the reversed-phase mode. The cCAD was approximately 10 times more sensitive in the HILIC mode and the ESI-MS was approximately 5-10 times more sensitive in the HILIC mode. There was one instance in the study where a compound produced more response in the reversed-phase mode. Thymine yielded more sensitivity in the reversed-phase mode with the ESI-MS detector. In a given mode of operation, there was significant variation in the measured response factors for all compounds on each detector. While this is not unexpected for the ESI-MS detector, variation in the response factors between compounds indicates that the cCAD and ELSD are not truly universal detectors in the sense that all compounds have identical responses.

Comparison of the factors that contribute to retention on immobilized polysaccharide-based chiral stationary phases and macrocyclic glycopeptide chiral stationary phases with the Abraham model.

The Abraham model of linear solvation energy relationship (LSER) was utilized to characterize three recently commercialized chiral stationary phases (CSPs), the Chiralpak IA, IB and IC. Normal phase system constants were determined by HPLC for these three CSPs and compared to literature values for the Chirobiotic T and V CSPs. The results indicate that the Chirobiotic T and V CSPs participate in more polar interactions, such as hydrogen bonding and dipolar interactions, than the three immobilized derivatized polysaccharide CSPs. Additionally, differences were noted for the e and b terms of the Abraham model (polarizable interactions and hydrogen bond acidity) for the IA and IB CSPs, which are nominally similar CSPs in their chemical makeup.

Normal phase and reverse phase HPLC-UV-MS analysis of process impurities for rapamycin analog ABT-578: application to active pharmaceutical ingredient process development.

ABT-578, an active pharmaceutical ingredient (API), is a semi-synthetic tetrazole derivative of the fermented polyene macrolide rapamycin. Reverse phase (RP)-HPLC-UV-MS and normal phase (NP)-HPLC-UV-MS methods employing an LC/MSD trap with electrospray ionization (ESI) have been developed to track and map all significant impurities from the synthetic process. Trace-level tracking of key impurities occurring at various process points was achieved using complimentary methodologies, including a stability indicating reverse phase HPLC method capable of separating at least 25 starting materials and process-related impurities from the API (YMC-Pack Phenyl column, UV-MS, 210 nm) and a targeted reverse phase HPLC method capable of separating very polar compounds from crude reaction mixtures (Phenomenex Synergi Polar RP column, UV, 265 nm). In addition, a normal phase HPLC method condition with post-column modifier infusion is described for the separation of epimeric impurities, and analysis of aqueous-sensitive reactive species (YMC-Pack SIL column, UV-MS, 278 nm). Process control strategies were established with these combinations of analytical technologies for impurities analyses to enable a rich understanding of the ABT-578 process.