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Impact of HILIC Amino-Based Column Equilibration Conditions on the Analysis of Chitooligosaccharides

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Abstract

Obtaining well-defined Chitooligosaccharides (COS) structures is very important as their physicochemical and biological properties depends strongly on their degree of polymerization (DP) and degree of N-acetylation (DA). Hydrophilic Interaction Liquid Chromatography (HILIC), that is commonly used for COS analyses, suffers from several drawbacks, related to their complex structure, leading to broadening/splitting of the chromatogram signals. We investigated, herein, on the role of the equilibration step of an amino-based HILIC column on the separation of COS DP ≤ 5. We demonstrated that COS could be separated according to their DP after equilibrating the column by a NaHCO3 buffer solution (100 mM, pH = 10) and mild elution chromatographic parameters (neutral mobile phase, r.t.); or according to their DA after equilibrating the column by an NH4Ac buffer solution (50 mM, pH = 4.5). Also, the nature of the counterion of the column stationary phase was found to affect both retention times and signal profiles of analyzed COS.

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Acknowledgements

Financial support from Bayer CropScience (La Dargoire, France), the University of Lyon, and the Agence Nationale de la Recherche (“BIOCOS” 10-CD2I-008 research project) are greatly acknowledged. We also thank Mrs. Agnès Crépet for her technical help in HILIC analyses.

Funding

Bayer CropScience (La Dargoire, France), the University of Lyon, and the Agence Nationale de la Recherche (“BIOCOS” 10-CD2I-008 research project).

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  1. Maher Abla

    Present address: Laboratoire d’Innovation Thérapeutique (LIT), Department of Chemistry and Biochemistry, Lebanese University, Faculty of Sciences II, Fanar, Lebanon

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  1. Ingénierie Des Matériaux Polymères, UMR CNRS 5223, Université Claude Bernard Lyon 1, Domaine Scientifique de La Doua, Bâtiment Polytech, 15 bd André Latarjet, 69622, Villeurbanne Cedex, France

    Maher Abla, Catherine Ladavière & Stéphane Trombotto

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MA: methodology/investigation/formal analysis/writing—original draft. ST: conceptualization/methodology/validation/writing—review and editing/funding acquisition/project administration/supervision. CL: conceptualization/methodology/validation/writing—review and editing/funding acquisition/project administration/supervision.

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Abla, M., Ladavière, C. & Trombotto, S. Impact of HILIC Amino-Based Column Equilibration Conditions on the Analysis of Chitooligosaccharides. Chromatographia 85, 55–63 (2022). https://doi.org/10.1007/s10337-021-04109-9

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