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Structure and Dynamics of Fluids in Confinement : a Case Study of Water, Protein and Ionic Liquid in Reverse Micelles and Microe

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Structure and Dynamics of Fluids in Confinement : a Case Study of Water, Protein and Ionic Liquid in Reverse Micelles and Microe Structure and dynamics of fluids in confinement: A case study of water, protein and ionic liquid in reverse micelles and microemulsions A dissertation submitted for the degree of Dr. rer. nat. (Doctor rerum Naturalium) in the Faculty of Chemistry and Biochemistry at the Ruhr-University Bochum Germany Sangeetha Balakrishnan 2007 ii 1st Examiner: Prof. Dr. Hermann Weingärtner 2nd Examiner: Prof. Dr. Roland Winter Thesis Committee Head: Prof. Dr. Dominik Marx Date of Defense: 10th December, 2007. iii iv Declaration I hereby declare that the dissertation entitled “Structure and dynamics of fluids in confinement: A case study of water, protein and ionic liquid in reverse micelles and microemulsions” is my original work and has been written with no other sources and aids than quoted, and has not been submitted elsewhere for an examination, as thesis or for evaluation in a similar context. Sangeetha Balakrishnan 7 November, 2007. v Acknowledgements I am indebted to many people who have been instrumental in the successful completion of this thesis. First among them is my advisor, Prof. Hermann Weingärtner, whom I would like to thank for the opportunity to pursue this research in the first place; and also for the freedom that he offered me to experiment my ideas. I owe a great deal of gratitude to Prof. Roland Winter, not just for graciously consenting to be my second referee; but more so for the many occasions when he so liberally gave of his time and guidance not just related to this thesis, but regarding research in general as well. I truly value those words of wisdom. For help with many administrative matters, I am grateful for the assistance of Mrs. Gundula Talbot, Mrs. Christel Tönnisen, and Mrs. Ursula Knieper. My colleagues deserve a special round of thanks for the congenial working atmosphere at the lab - Yathrib Ajaj, Mianmian Huang, Sasisanker Padmanabhan, and Peter Romahn - thanks to them all for helping create some wonderful memories. My thanks also to Dr. Holger Nadolny, my former colleague; for having been a good friend over the years. I am grateful to Nadeem Javid (Dortmund) for the successful SAXS collaboration with one of my projects. I would like to thank him and Dr. Sivakumar Sekharan for their help in obtaining some of the literature cited in the thesis. For help in proof-reading a part of the work, my thanks to Dr. Kaushik Chakrabarty. Special thanks to Nilesh Madhu for the many science and non-science discussions and for his support when things were at low ebb. My thanks to the DFG Graduiertenkolleg ‘Structure and Dynamics in Heterogeneous Systems’ for the financial support to carry out this work. Finally, and most importantly, my thanks to my family - my father, Wg. Cdr. A. M. Balakrishnan; my mother, Mrs. Vijayalakshmi Balakrishnan; and my sister, Ms. Revathi Balakrishnan - for all their love, support and the freedom they accorded me to pursue my dream. vi vii Table of contents Declaration iv Acknowledgements v Table of contents vii Preface xi Abbreviations and Symbols xiii Chapter 1 Introduction 1 1.1 The hydrogen bond and its historical background 1 1.2 Classification of hydrogen bonds 2 1.3 Hydrogen bonding in water 4 1.3.1 Structure of liquid water 4 1.3.2 Water clusters 5 1.4 Water in confinement 6 1.5 Proteins in confinement 8 1.6 Ionic liquids in confinement 9 1.6.1 The C − HLF hydrogen bond 10 1.7 Self-assembly 10 1.7.1 The Hydrophobic effect and surfactants in solution 11 Chapter 2 State of the art 17 2.1 AOT – the surfactant 18 2.2 Infrared spectroscopy of AOT reverse micelles 20 2.2.1 The OH stretch region 20 2.2.2 The ‘n-water state’ conundrum 21 2.2.3 NIR spectroscopy in AOT reverse micelles 24 2.2.4 Other vibrational modes in AOT reverse micelles 25 2.3 Dielectric spectroscopy 27 2.4 Small angle x-ray scattering 29 2.5 Other techniques 32 2.5.1 The water libration band and terahertz spectroscopy 32 2.5.2 Non-linear infrared spectroscopy 32 2.5.3 NMR spectroscopy 33 2.5.4 Simulations 35 2.5.5 Molecular probes 36 2.5.6. Calorimetry 37 2.5.7 Other experimental techniques 38 2.6 Non-aqueous reverse micelle interior 38 2.7 Proteins in confinement 39 2.7.1 A first look at macromolecular crowding 40 2.7.2 The biological membrane 41 2.7.2.1 Reverse micelles as membrane mimics 42 2.7.2.2 Encapsulation of proteins in reverse micelles 42 2.7.3 Osmolytes 45 2.7.4 Hydration water 48 2.8 Ionic liquids in confinement 49 2.8.1 Research on confined ILs so far 50 2.8.2 [bmim][BF 4] and Triton X-100 53 2.8.3 Solvatochromism 54 2.8.4 Association constant 55 Chapter 3 Experimental techniques 57 3.1 Near-infrared spectroscopy 57 3.1.1 Principles of NIR spectroscopy 58 3.1.2 Anharmonic vibrations 60 3.1.3 Consequences of anharmonicity 61 3.1.4 Instrumentation 61 3.2 Dielectric relaxation spectroscopy 62 3.2.1 The dielectric constant and polarisation 63 3.2.2 Relaxation Times 64 ix 3.2.3 The Debye equation 65 3.2.4 Instrumentation 66 3.3 Small angle x-ray scattering 68 3.3.1 Theory 69 3.3.2 Instrumentation 71 3.4 Materials 72 3.4.1 Water in confinement 72 3.4.2 α-chymotrypsin in confinement 73 3.4.3 [bmim][BF 4] in confinement 74 Chapter 4 Results and discussion 75 4.1 NIR spectroscopy of bulk water 75 4.2 AOT reverse micelle — the anhydrous system 77 4.3 AOT reverse micelle — the hydrous system: Influence of hydration 78 4.3.1 NIR data 78 4.3.2 SAXS data 84 4.3.3 Dielectric relaxation data 86 4.4 Effect of hydrocarbon medium on AOT reverse micelles 89 4.4.1 NIR data 89 4.4.2 SAXS data 92 4.5 Temperature dependence of hydrogen bonding of micellar water 95 4.5.1 Hydration picture as a function of temperature 99 4.6 NIR analysis of α-CT hydration properties in confinement 99 4.6.1 The basis of the method 101 4.6.1.1 The spectra — origin and resolution 102 4.6.1.2 Resolution of Curve B from the difference spectrum 103 4.6.1.3 Resolution of Curve C from the difference spectrum 103 4.6.2 The kosmotropes 104 4.6.3 The chaotropes 104 4.7 IL in confinement 107 N 4.7.1 Polarity of the confined IL determined by E T(30) and ET solvent x polarity scales 107 4.7.2 Structural changes in the confined ionic liquid 109 4.7.3 Determination of association constant 111 Chapter 5 Conclusions 114 Appendices A List of Figures 117 B List of Tables 120 C Spectral positions 121 D Derivation of the Benesi-Hildebrand equation 122 E List of References 125 F Curriculum Vitae 143 xi Preface The study of fluids in confined geometries and close to interfaces has gained unprecedented momentum over the last decade owing to its applications in diverse fields ranging from geochemistry to astrobiology. A testament to this is the series of Confit workshops held triennially since the year 2000, serving as a confluence for the discussion of confinement effects on the microscopic dynamics of condensed matter. A variety of confined systems like simple organic and inorganic liquids, quantum liquids, water, polymers, biological systems, molecules and atoms; in conjunction with an equally impressive array of confining media like zeolites, carbon nanotubes, reverse micelles, clays, fullerenes, etc. make this a highly interesting field of study. Rapid advances in technology over the years have also contributed to significant developments that have lead to the expansion of the frontiers of confined-fluids research. The present thesis is an exposition of the investigation of the structure and dynamics of water, a protein -- α-chymotrypsin, and an ionic liquid -- 1-butyl-3-methyl-imidazolium tetrafluoroborate ([bmim][BF 4]) confined within the nano pores of reverse micelles and microemulsions formed by two surfactants; Aerosol OT (AOT) and Triton X-100. Right from Lavoisier’s discovery of the composition of water to its detection on Mars and Europa, water has been the most widely investigated substance in laboratories around the world. In nature however, water often appears in interstices or is adsorbed on solid substrates like minerals; and the choice of the first experimental system in this thesis — water in confinement — is an attempt to study the properties of water in an environment that mimics this natural state. To this effect, the hydrogen bonding of water confined in AOT reverse micelles was chosen as the spectral tool to monitor the structure and dynamics of water in accordance with variation in extent of hydration, temperature, and the molecular make-up of the external solvent forming the reverse micelle. Most protein folding and other associated dynamic processes are studied in infinite dilution. In vivo , however, protein dynamics occur in a crowded cellular milieu and in confined spaces such as the chaperonin cavity, the proteosome etc. It thus makes sense to assume that proteins may experience different energy landscapes when folding in vivo than in xii bulk, and this has been the driving force behind the studies involving the second experimental system — α-chymotrypsin in confinement . Several osmolytes were added to this confined protein system and an investigation was undertaken to elucidate the effect of macromolecular crowding on the hydration properties of the confined protein.
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