Investigations of the retention mechanisms in hydrophilic interaction chromatography

Investigations of the Retention

Mechanisms in Hydrophilic Interaction Chromatography

Ngoc Phuoc Dinh

Department of Chemistry Umeå University

Faculty Opponent: Dr. Andrew J. Alpert, PolyLC Inc.,Columbia, MD 21045 USA

Umeå 2013

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Organisation Document type

Date of publication

Author

Title

Abstract

Keywords:

ISBN:

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v

List of paper

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Abbreviations

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Contents

Abstract ... iii

List of paper ... v

Abbreviations ... vii

Contents ... ix

1 Introduction ... 1

2 Background ... 1

2.1 Liquid chromatography ... 3

2.1.1 Brief history ... 3

2.1.2 General concepts ... 5

2.2 Hydrophilic interaction chromatography ... 7

2.2.1 HILIC stationary phase ... 8

2.2.2 Intermolecular interaction might involve HILIC ... 9

2.3 Characterization methods ... 9

2.3.1 Coulometric KF titration ... 10

2.3.2 Indophenol method ... 11

2.3.3 Specific surface area and porosity measurement ... 11

2.3.4 Multivariate data analysis ... 12

2.3.4.1 Principle component analysis (PCA) ... 12

2.3.4.2 Orthogonal projection to latent structure (OPLS) ... 12

2.3.4.3 Interpretation of biplot ... 13

2.3.4.4 R2 an Q2 ... 14

3 Characterization of water enriched layer ... 14

3.1 Water accumulation... 16

3.1.1 Water adsorption isotherm ... 16

3.1.2 Effect of functional groups ... 17

3.1.3 Effect of mobile phase composition ... 18

3.2 Salt accumulation ... 21

3.2.1 Ammonium acetate adsorption isotherm ...Error! Bookmark not defined. 3.2.1.1 Ammonium adsorption isotherm ... 21

3.2.2 Relationship between adsorptions of ammonium and acetate ions ... 25

4 Investigations of retention mechanism in HILIC ... 24

4.1 Determination of column hold-up volumn ... Error! Bookmark not defined. 4.2 Partitioning vs adsorption ... 25

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4.3 Characterization of interactions involves in retention mechanism ... 27

5 Simple method for HILIC columns characterization ... 30

6 Conclusions and remarks ... 32

7 Acknowledgements ... 34

8 Bibliography ... 36

1

1 Introduction

2 Background

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3

2.1 Liquid chromatography 2.1.1 Brief history

4

5

2.1.2 General concepts

𝑘^′= ^𝑡𝑅′

𝑡𝑀 =^{𝑡𝑅−𝑡𝑀}

𝑡𝑀

𝑘^′= ^𝐾

𝛽

6

𝑁 = 𝑎 (^𝑡𝑅

𝑤)²

𝛼 =^{𝑡𝑅′(𝐵)}

𝑡𝑅′_(𝐴) =^𝑘

′(𝐵) 𝑘^′_(𝐴)

7

2.2 Hydrophilic interaction chromatography

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2.2.1 HILIC stationary phases

≡

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2.2.2 Intermolecular interactions involved in HILIC

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

2.3 Characterization methods

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2.3.1 Coulometric KF titration

3𝐼₂+ 2𝐻₂𝑂 + 𝑆𝑂₂ → 4𝐻⁺+ 2𝐼₃⁻+ 𝑆𝑂₄²⁻

2𝐼₃⁻ → 3𝐼₂+ 2𝑒

𝑛 =^{∫ 𝐼𝑑𝑡}

𝑡 0 𝐹𝑧

11

2.3.2 Indophenol method

2.3.3 Specific surface area and porosity measurements

12

2.3.4 Multivariate data analysis

2.3.4.1 Principal component analysis (PCA)

2.3.4.2 Orthogonal projection to latent structures (OPLS)

13 2.3.4.3 Interpretation of biplots

14

2.3.4.4 R² and Q²

3 Characterization of the water enriched layer

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3.1 Water accumulation

3.1.1 Water adsorption isotherms

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3.1.2 Effect of functional groups

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3.1.3 Effect of mobile phase composition

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3.2 Salt accumulation

3.2.1 Acetate adsorption

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3.2.2 Ammonium adsorption

3.2.3 Effect of water content mobile phase

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3.2.4 Relationship between acetate adsorptions and ammonium adsorption

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4 Investigations of retention mechanism in HILIC

4.1 Partitioning versus adsorption

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4.2 Characterization of interactions involved in the retention mechanism – Column classification

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5 Simple method for HILIC column characterization

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6 Conclusions and remarks

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7 Acknowledgements

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8 Bibliography

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