Serial coupling of reversed-phase and hydrophilic interaction liquid chromatography to broaden the elution window for the analysis of pharmaceutical compounds

High-efficiency hydrophilic interaction chromatography by coupling 25cm×4.6mm ID×5μm silica columns and operation at 80°C

“…The first observation is that at 1 mL/min (1.4 mm/s), the velocity is above u opt for all solutes at 30°C while close to u opt at 80°C. The fact that higher temperatures generate higher optimal mobile phase velocities is well documented in the literature [22,26,31,35] and the result of the interplay between an increased B-term and a decreased C-term. …”

“…The relationship is described by the Van 't Hoff equation [22,41]. Because retention is an exothermic process (∆H < 0) and generally entropy is lost upon transfer from the mobile to the stationary phase (∆S < 0), an increase in temperature leads to a shift of the equilibrium towards the mobile phase and therefore to a drop in retention [27,31,40,41]. Overall, however, the retention of the test compounds is relatively low at 80°C (k API <2.3).…”

“…HILIC has become a valuable addition to RP-LC in drug analysis, due to its orthogonality to the latter separation mode [9][10][11][12][13]. HILIC is particularly useful for the analysis of highly polar and ionisable compounds [14][15][16][17][18].…”

“…For the pharmaceutical industry, which will certainly still have a plethora of conventional HPLC instrumentation in its good manufacturing practice (GMP) laboratories, kinetic plots can certainly serve a purpose in directing how conventional instrumentation and columns can be used more appropriately in combination with ETLC and long columns. This can help to optimize the use of conventional particle diameters (5 µm) to yield either maximum efficiency and to compared to sub-2µm particles, or to increased the speed of analysis.Since these developments, a chromatographic technique that has been enjoying recent attention in the pharmaceutical industry is hydrophilic interaction chromatography (HILIC).HILIC has become a valuable addition to RP-LC in drug analysis, due to its orthogonality to the latter separation mode [9][10][11][12][13]. HILIC is particularly useful for the analysis of highly polar 4 and ionisable compounds [14][15][16][17][18].…”

Theory and Practice of UHPLC and UHPLC–MS

“…A set of serially coupled columns containing different stationary phases behaves like a new column with modified selectivity, which may enable separations of samples with widely differing properties (Alvarez Segura et al, 2016). An octadecylsilica column in the first dimension serially coupled with a HILIC column in the second dimension, to which a gradient of acetonitrile in water up to the end concentration of 80% or more was applied, allowed separation of a broad range of pharmaceuticals in a single run (Louw et al, 2008). A serial combination of a ZIC-HILIC sulfobetaine column and an amide column with a HILIC gradient run from 95% to 35% aqueous buffer in acetonitrile allowed simultaneous separations…”