Abstract

Achieving resolution in liquid chromatography (LC) separations has been achieved by improving efficiency and applying mobile phase optimisation. However, given the increasing complexity of pharmaceutical molecules, the exploitation of stationary phase selectivity has come more into play in the form of generic column screening approaches, stationary phase optimised selectivity and two-dimensional liquid chromatography (2D-LC). Despite this, the major limitation of these is the limited number of HPLC stationary phases that are sufficiently orthogonal from one another. To this end, the general aim of this study was to investigate the potential for new charge transfer phases based on electron-deficient aromatics as an additional “orthogonal stationary phase”.

In this context, a charge transfer dinitrobenzamido- (DNBA) column had not only already been shown to be useful in normal-phase (NP) for a limited number of applications but also, much more recently, in reversed phase (RP), it had been shown to have different selectivity to C18 phases in screening drugs of abuse. This suggested that this fairly simple phase might have much more general applicability. When assessed here in RP mode, it was shown that the DNBA phase was not only orthogonal in selectivity to the C18 (r2 = 0.1212) but was also shown to be orthogonal to other so-called charge transfer phases (r2 = 0.1196 and 0.7853). Therefore, the DNBA- phase was evaluated for use not only for applications where charge transfer interactions would clearly be helpful but also for a panel of “challenging separations” that were problematic when using other phases. In particular, it was used successfully to separate the related substances of baclofen, dothiepin, paroxetine and pizotifen. However, for the phase to have the potential to be commercially viable, it would need to be stable under RP, reproducible and durable under extreme conditions. This was shown to be the case both over this initial test at extremes and throughout the entire RP applications study. Overall, it was clear that there was sufficient promise to progress with further development of the dinitrobenzamido- phase based on modern particle technology.

The subsequent objective was to prepare a DNBA phase based on 2.6-µm superficially porous particles (spp). This material showed the expected improved efficiency, better mass transfer at high flow rates and similar selectivity to the original material. The phase was effective in NP and RP applications and maintained retention when switching from NP to RP and vice versa. The band broadening found in some instances associated with strong π−π interactions was, in most cases, more than offset by the improvements in selectivity. Also, the retention shifts were strong enough to suggest that it might be compatible with use as a solid phase extraction (SPE) sorbent. In supercritical fluid chromatography (SFC) operations, it was proved to be useful in separating a mixture of thymine, theophylline and theobromine, as well as a PAH mixture.