Possibilities of new generation columns packed with 1.3 μm core–shell particles in gradient elution mode

The aim of this work was to evaluate the practical possibilities in gradient elution mode of a column packed with 1.3 μm core–shell particles recently released on the market. For this purpose, two types of analytes possessing different diffusion coefficients were selected (small molecule and peptide). It appears that the new 1.3 μm material was particularly well suited for fast separations, compared to other existing core–shell particle dimensions in gradient mode. The new material systematically outperforms the other existing ones for peak capacity up to 300 for small molecules and 700 (corresponding to t0 = 15 min) for peptides. Based on these cut-off values, the advantage of column packed with 1.3 μm was much more obvious for peptides vs. small molecules analysis. Further improvements in terms of column mechanical stability and system upper pressure capability could expand the limits of separation speed and efficiency to a different level. Again, because of the current pressure limitation and low permeability, a column length of more than 5–8 cm is never desired for small molecules analysis in gradient elution. On the contrary, longer columns were useful for peptide analysis. As example, a column of 28 cm packed with 1.3 μm particles provides a peak capacity of 1000 in the case of peptides analysis. All the predicted values were experimentally confirmed using a standardized extract of Ginkgo biloba and a tryptic digest of a monoclonal antibody (Panitumumab). For the plant extract, the better performance was always achieved with a 5 cm long column (P = 267 and 268 for the 5 and 15 cm, respectively, using a gradient time of 10 and 40 min, respectively). Finally, in the case of peptide mapping, a 15 cm long column packed with 1.3 μm particles was the best choice (P = 176 and 311 for the 5 and 15 cm, respectively, using a gradient time of 10 and 40 min, respectively).