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Multi-modal applicability of a reversed-phase/weak-anion exchange material in reversed-phase, anion-exchange, ion-exclusion, hydrophilic interaction and hydrophobic interaction chromatography modes

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Abstract

We recently introduced a mixed-mode reversed-phase/weak anion-exchange type separation material based on silica particles which consisted of a hydrophobic alkyl strand with polar embedded groups (thioether and amide functionalities) and a terminal weak anion-exchange-type quinuclidine moiety. This stationary phase was designed to separate molecules by lipophilicity and charge differences and was mainly devised for peptide separations with hydroorganic reversed-phase type elution conditions. Herein, we demonstrate the extraordinary flexibility of this RP/WAX phase, in particular for peptide separations, by illustrating its applicability in various chromatographic modes. The column packed with this material can, depending on the solute character and employed elution conditions, exploit attractive or repulsive electrostatic interactions, and/or hydrophobic or hydrophilic interactions as retention and selectivity increments. As a consequence, the column can be operated in a reversed-phase mode (neutral compounds), anion-exchange mode (acidic compounds), ion-exclusion chromatography mode (cationic solutes), hydrophilic interaction chromatography mode (polar compounds), and hydrophobic interaction chromatography mode (e.g., hydrophobic peptides). Mixed-modes of these chromatographic retention principles may be materialized as well. This allows an exceptionally flexible adjustment of retention and selectivity by tuning experimental conditions. The distinct separation mechanisms will be outlined by selected examples of peptide separations in the different modes.

A silica-based material functionalized with alkyl-strands having polar embedded groups and terminal anion-exchange site provides exceptional operational flexibility and can be used in reversed-phase, anion-exchange, ionexclusion, hydrophilic interaction and hydrophobic interaction chromatography as well as mix-modes thereof

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Abbreviations

AEX:

Anion-exchange

HIC:

Hydrophobic interaction chromatography

HILIC:

Hydrophilic interaction chromatography

IEC:

Ion-exclusion chromatography

IEX:

Ion-exchange

LSS:

Linear solvent strength (theory)

RPLC:

Reversed-phase liquid chromatography

RP/WAX:

Mixed-mode reversed-phase/weak anion-exchange material

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Acknowledgments

The financial support of the Christian-Doppler Research Society and the industry partners piCHEM (Graz, Austria), Merck KGaA (Darmstadt, Germany), Sandoz (Kundl, Austria), Fresenius Kabi Austria (Graz, Austria), and AstraZeneca (Mölndal, Sweden) is gratefully acknowledged.

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  1. Department of Analytical Chemistry, Faculty of Chemistry, University of Vienna, Waehringer Strasse 38, 1090, Vienna, Austria

    Michael Lämmerhofer, Raquel Nogueira & Wolfgang Lindner

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  1. Michael Lämmerhofer
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  2. Raquel Nogueira
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  3. Wolfgang Lindner
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Correspondence to Michael Lämmerhofer.

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Published in the special issue Analytical Sciences in Austria with Guest Editors G. Allmaier W. Buchberger, and K. Francesconi.

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Lämmerhofer, M., Nogueira, R. & Lindner, W. Multi-modal applicability of a reversed-phase/weak-anion exchange material in reversed-phase, anion-exchange, ion-exclusion, hydrophilic interaction and hydrophobic interaction chromatography modes. Anal Bioanal Chem 400, 2517–2530 (2011). https://doi.org/10.1007/s00216-011-4755-3

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