Detailed efficiency analysis of columns with a different packing quality and confirmation via total pore blocking

Highlights

• Columns with 4 distinct types of band broadening quality were analyzed.

• Retention and peak parking measurements showed identical B-, C_m-, C_s-term dispersion.

• Differences could be fully attributed to eddy-dispersion.

• Total pore blocking confirmed packing heterogeneity as main difference between columns.

Abstract

We report on a systematic study involving columns with a clearly different efficiency (4 distinct quality groups) obtained by packing the columns that were C₁₈ bonded and endcapped with a different carbon loading. Using B-term analysis (via peak parking) and theoretical models to estimate the magnitude of the C_m- and C_s-term contributions, it could be concluded that the difference in efficiency among the groups was entirely due to a difference in eddy dispersion. As such, the columns provided an ideal testing ground to verify how well the total pore blocking (TPB)-method can be used to probe differences in packing heterogeneity. In agreement with earlier literature observations, it turns out the TPB-method is much more sensitive to packing heterogeneities than the eddy dispersion (H_eddy)-contribution measured under open-pore conditions via B- and C- term subtraction. Typically, differences in H_eddy on the order of 0.1–0.5μm translate into a difference on the order of 0.5–2μm in the TPB mode. This confirms the TPB as a powerful technique to make very sensitive measurements of the homogeneity of packed beds.

Introduction

In the last 15 years, the column technology in liquid chromatography has witnessed two important evolutions. The first was the introduction of sub-2 μm particles at the beginning of this century, readily followed by the introduction of UHPLC instrumentation capable to deliver the high pressures needed to operate these columns at reasonable flow rates [[1], [2], [3], [4], [5]]. The second evolution was the (re-)introduction of core-shell particles, producing reduced plate heights which are 20–30% lower than the traditional h_min = 2-value which was up till then considered as the performance limit for fully porous particles [[5], [6], [7]]. Originally promoted for their reduced diffusion distance in the porous shell and thus lower mass transfer resistance (C-term), it was later found that the decrease in H was mainly due to a lower longitudinal diffusion (B-term) and a reduced eddy dispersion (A-term). Whereas the underlying reasons for the lower B-term of core-shell particles are well understood, the reason for the strong reduction in A-term is still under debate. One hypothesis is that the smaller A-term contribution is due to the narrower particle size distribution (PSD) of core-shell particles (in turn being a consequence of their production method) [[8], [9], [10], [11], [12]], although this was disputed in other publications [[13], [14], [15]]. In other studies, it was claimed that higher surface roughness results in smaller variations in packing density [11,16] or even lower film mass transfer resistance [13,16]. Supporting the PSD hypothesis however, is that, stimulated by the success of core-shell particles, it was also attempted to produce fully porous particles with a narrower PSD. Doing so, it was indeed found that reduced plate heights between 1.6 and 1.9 (depending on column dimensions) can be obtained for columns packed with fully porous particles [[17], [18], [19]]. Obviously, the PSD cannot be driven to zero as this would lead to regions in the column that are packed into a crystalline configuration, coexisting with randomly packed regions, and this coexistence of poorly and normally permeated regions can lead to a dramatic increase in band broadening.

In the present study, we compared the efficiency of columns packed with C₁₈-derivatized and endcapped silica particles prepared with coating mixtures with increasing theoretical carbon load, i.e., with a composition theoretically leading to a coverage of 2,3,4 or 5 μmol/m². For each of the four coverage types, 2 columns were tested, except for the 5 μmol/m², for which 3 columns were tested. The different coverage types are referred to in the present study as TC2, TC3, TC4, TC5, using the digit in the abbreviation to represent the theoretical surface coverage in μmol/m². Although the packing method for all columns was optimized in the same manner, a systematic and marked difference in overall plate height and eddy-dispersion was observed. To investigate this in detail, a full plate height analysis was made, including testing the columns under total pore blocking (TPB) conditions. This method has been introduced in 2007 [20] and provides indisputably the best measure of packing heterogeneity because, with the particles being completely inaccessible by the analytes, the only remaining dispersion source originates from the heterogeneous network of interstitial flow-through pores. Since its introduction, the TPB method has been applied in a large number of packing heterogeneity studies [16,[21], [22], [23]]. The possible impact of high surface coverage on efficiency has been recently investigated by other authors in chiral chromatography [24,25]. Previous work on the impact of C₁₈ surface coverage on retention and adsorption behavior, as well as on the mass transfer kinetics, can be found in [[26], [27], [28], [29], [30]].