University of Kentucky UKnowledge University of Kentucky Doctoral Dissertations Graduate School 2009 EXPERIMENTAL AND MOLECULAR DYNAMICS SIMULATION STUDIES OF PARTITIONING AND TRANSPORT ACROSS LIPID BILAYER MEMBRANES Ravindra Wadhumal Tejwani University of Kentucky, [email protected] Right click to open a feedback form in a new tab to let us know how this document benefits ou.y Recommended Citation Tejwani, Ravindra Wadhumal, "EXPERIMENTAL AND MOLECULAR DYNAMICS SIMULATION STUDIES OF PARTITIONING AND TRANSPORT ACROSS LIPID BILAYER MEMBRANES" (2009). University of Kentucky Doctoral Dissertations. 738. https://uknowledge.uky.edu/gradschool_diss/738 This Dissertation is brought to you for free and open access by the Graduate School at UKnowledge. It has been accepted for inclusion in University of Kentucky Doctoral Dissertations by an authorized administrator of UKnowledge. For more information, please contact [email protected]. ABSTRACT OF DISSERTATION Ravindra Wadhumal Tejwani The Graduate School University of Kentucky 2009 EXPERIMENTAL AND MOLECULAR DYNAMICS SIMULATION STUDIES OF PARTITIONING AND TRANSPORT ACROSS LIPID BILAYER MEMBRANES ABSTRACT OF DISSERTATION A dissertation submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy in the College of Pharmacy at the University of Kentucky By Ravindra Wadhumal Tejwani Lexington, Kentucky Director: Dr. Bradley D. Anderson, Professor of Pharmaceutical Sciences Lexington, Kentucky 2009 Copyright © Ravindra W. Tejwani 2009 ABSTRACT OF DISSERTATION EXPERIMENTAL AND MOLECULAR DYNAMICS SIMULATION STUDIES OF PARTITIONING AND TRANSPORT ACROSS LIPID BILAYER MEMBRANES Most drugs undergo passive transport during absorption and distribution in the body. It is desirable to predict passive permeation of future drug candidates in order to increase the productivity of the drug discovery process. Unlike drug-receptor interactions, there is no receptor map for passive permeability because the process of transport across the lipid bilayer involves multiple mechanisms. This work intends to increase the understanding of permeation of drug-like molecules through lipid bilayers. Drug molecules in solution typically form various species due to ionization, complexation, etc. Therefore, species specific properties must be obtained to bridge the experiment and simulations. Due to the volume contrast between intra- and extravesicular compartments of liposomes, minor perturbations in ionic and binding equilibria become significant contributors to transport rates. Using tyramine as a model amine, quantitative numerical models were developed to determine intrinsic permeability coefficients. The microscopic ionization and binding constants needed for this were independently measured. The partition coefficient in 1,9-decadiene was measured for a series of compounds as a quantitative surrogate for the partitioning into the hydrocarbon region of the bilayer. These studies uncovered an apparent long-range interaction between the two polar substituents that caused deviations in the microscopic pKa values and partition coefficient of tyramine from the expected values. Additionally the partition coefficients in the preferred binding region of the bilayer were also measured by equilibrium uptake into liposomes. All-atom molecular dynamics simulations of lipid bilayers containing tyramine, 4- ethylphenol, or phenylethylamine provided free energies of transfer of these solutes from water to various locations on the transport path. The experimentally measured partition coefficients were consistent with the free energy profiles in showing the barrier in the hydrocarbon region and preferred binding region near the interface. The substituent contributions to these free energies were also quantitatively consistent between the experiments and simulations. Specific interactions between solutes and the bilayer suggest that amphiphiles are likely to show preferred binding in the head group region and that the most of hydrogen bonds involving solutes located inside the bilayer are with water molecules. Solute re-orientation inside the bilayer lowers the partitioning barrier by allowing favorable interactions. KEYWORDS: Permeability Coefficient, Partition Coefficient, Molecular Dynamics Simulation, Lipid Bilayer, Ionization Constant ____________________________________ Student’s Signature ____________________________________ Date EXPERIMENTAL AND MOLECULAR DYNAMICS SIMULATION STUDIES OF PARTITIONING AND TRANSPORT ACROSS LIPID BILAYER MEMBRANES By Ravindra Wadhumal Tejwani ____________________________________ Director of Dissertation ____________________________________ Director of Graduate Studies ____________________________________ Date RULES FOR THE USE OF DISSERTATIONS Unpublished dissertations submitted for the Doctor's degree and deposited in the University of Kentucky Library are as a rule open for inspection, but are to be used only with due regard to the rights of the authors. Bibliographical references may be noted, but quotations or summaries of parts may be published only with the permission of the author, and with the usual scholarly acknowledgments. Extensive copying or publication of the dissertation in whole or in part also requires the consent of the Dean of the Graduate School of the University of Kentucky. A library that borrows this dissertation for use by its patrons is expected to secure the signature of each user. Name Date ________________________________________________________________________ ________________________________________________________________________ ________________________________________________________________________ ________________________________________________________________________ ________________________________________________________________________ ________________________________________________________________________ ________________________________________________________________________ ________________________________________________________________________ ________________________________________________________________________ ________________________________________________________________________ ________________________________________________________________________ DISSERTATION Ravindra Wadhumal Tejwani The Graduate School University of Kentucky 2009 EXPERIMENTAL AND MOLECULAR DYNAMICS SIMULATION STUDIES OF PARTITIONING AND TRANSPORT ACROSS LIPID BILAYER MEMBRANES DISSERTATION A dissertation submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy in the College of Pharmacy at the University of Kentucky By Ravindra Wadhumal Tejwani Lexington, Kentucky Director: Dr. Bradley D. Anderson, Professor of Pharmaceutical Sciences Lexington, Kentucky 2009 Copyright © Ravindra W. Tejwani 2009 For Amrita and Monica, in the loving memory of my parents Acknowledgements While computers and the internet have shrunk the long distances, working with long distance students can be a challenge. I appreciate the open mindedness of the UKY faculty and especially Dr. Anderson in working with this unconventional arrangement. I am grateful to Dr. Anderson for his guidance throughout the last six years, his teaching of the technical knowledge that I am proud of, and his patience in working with my erratic productivity due to my part time commitment. I am thankful to him for letting me be a part of the research group that is exceptionally bright and knowledgeable. I am grateful to Terrry Stouch for teaching the fundamentals of computer simulations and converting a novice such as me to run large scale MD simulation studies. Initiating and completing three large scale simulations lasting almost three years could not have been possible without Terry’s enthusiasm and expertise. An undertaking this big can only be tackled with programming and the bugs would have kept me at the keyboard a great deal longer if it wasn’t for help from Malcolm Davis. I am indebted to him for teaching me Pearl scripts. Thanks are due to the dissertation committee: Drs. Bummer, Spielmann, Xiang, and Li for their sincere critique of the dissertation and being available for periodic data review meetings through these years. Their patience as well as the time is sincerely appreciated. The multidisciplinary nature of the dissertation topic has required me to tap into expertise of several individuals. In this respect, I would like to thank Dr. Xiang for being available to tap into his knowledge of lipid bilayer related experiments, William Pitts for answering questions regarding chemical syntheses, Greg Stiegler and Saverio Vacca for - iii - computer hardware related help, John Carpenter and Steve Plimpton for Lammps software, Sharon Callahan and Sarah Traeger for answering questions regarding NMR instrumentation, and Catina Rossoll for her help with the graduate school paperwork. I am grateful to my mentors at the Bristol-Myers Squibb Company for continuously encouraging me to obtain the doctoral degree. In particular, thanks are due to Drs. Nancy Barbour, Abu Serajuddin, Sailesh Varia, and Joseph Bogardus for writing testimonials for the admission and fellowship applications. I am particularly thankful to Dr. Srinivasan Venkatesh for suggesting UKY for the degree and for crystallizing the necessary initial steps for obtaining the approvals. I am thankful to the Bristol-Myers Squibb Company for the Doctoral Fellowship and to Dr. Scott Biller for his guidance and support in requesting for the same. Finally I am thankful to the supervisors