Automated Regularized Deconvolution for Eliminating Extra-Column Effects in Fast High-Efficiency Separations

Abstract

With the introduction of ultrahigh efficiency columns and fast separations, the need to eliminate peak deformation contributed by the instrument must be effectively solved. Herein, we develop a robust framework to automate deconvolution and minimize its artifacts, such as negative dips, wild noise oscillations, and ringing, by combining regularized deconvolution and Perona−Malik (PM) anisotropic diffusion methods. A asymmetric generalized normal (AGN) function is proposed to model the instrumental response for the first time. With no-column data at various flow rates, the interior point optimization algorithm extracts the parameters describing instrumental distortion. The column-only chromatogram was reconstructed using the Tikhonov regularization technique with minimal instrumental distortion. For illustration, four different chromatography systems are used in fast chiral and achiral separations with 2.1 and 4.6 mm i.d. columns. Ordinary HPLC data can approach highly optimized UHPLC data. Similarly, in fast HPLC-circular dichroism (CD) detection, 8000 plates were gained for a fast chiral separation. Moment analysis of deconvolved peaks confirms correction of the center of mass, variance, skew, and kurtosis. This approach can be easily integrated and used with virtually any separation and detection system to provide enhanced analytical data.

High-Throughput Capillary Liquid Chromatography Using a Droplet Injection and Application to Reaction Screening

“…The fast separations demonstrated here are limited to analysis of relatively simple mixtures, although coupling to MS or use of peak deconvolution ,,, can further expand the scope. Even with this limitation, numerous applications can be envisioned in addition to the rapid organic reaction screen demonstrated here.…”

“…Separation times of 1–15 s have been demonstrated for several sample types including chiral, achiral, and small biomolecules. − These separations have been achieved by using short HPLC columns (0.5–5.5 cm long), high flow rates (1–8 mL/min), and systems modified to have reduced dead volumes. Although peak capacity of these fast isocratic separations has been modest compared to longer analysis times, the use of peak deconvolution methods can increase information content. ,,, Up to 10 components were resolved in 0.9 s using such approaches . One potential application demonstrated by fast separations was the increased throughput of standard pharmacopeial methods. ,, In principle, separation times of 1–15 s have appealing applications in a variety of high-throughput experiments (HTE) such as screening organic reactions, biocatalyst development, chemical “sensing”, process monitoring, and quality testing of chemical libraries, among others.…”

Minimize Precolumn Band Broadening with Immiscible Solvent Sandwich Injection

“…The sandwich injection technique was also compared with the deconvolution method recently proposed by Handlovic et al It turns out that the latter is not suited to correct the severe band broadening encountered in Figure a for example, as can be noticed from the data shown in Figure S10.…”

Optimization of analytical method greenness scores: a case study of amino acid enantioseparations with carbonated aqueous systems

“…column, the most popular column characteristic for classical HPLC. The increased conceptual understanding of column packing and particle synthesis technology, combined with optimization of instrumentation to reduce extra-column band broadening, 43,44 has allowed smaller columns to produce satisfactory resolution with less solvent use. To demonstrate these improvements in green chemistry, van Demeter plots (Fig.…”

“…This lower efficiency is due to a larger A term from wall effects during packing, 48 and larger efficiency loss from extra-column band broadening. 43,44 Fig. 3B demonstrates the effect of internal diameter changes (4.6 mm, 3.0 mm, and 2.1 mm) on separation kinetics.…”