Role of Hydrophobic Effect on the Noncovalent Interactions Between Salicylic Acid and a Series of -Cyclodextrins Elena Junquera, David Ruiz, and Emilio Aicart 1 Departamento de Quı ´mica Fı ´sica I, Facultad de Ciencias Quı ´micas, Universidad Complutense de Madrid, 28040-Madrid, Spain Received January 25, 1999; accepted April 14, 1999 The molecular complexation of salicylic acid (o-hydroxybenzoic acid) by -cyclodextrin (-CD) and/or two of its most used deriv- atives, 2,6-di-O-methyl--cyclodextrin (DIMEB) and hydroxypro- pyl--cyclodextrin (HPBCD), was studied from pH potentiomet- ric measurements. The role of the hydrophobic effect was evaluated by studying the influence of the presence of different constant amounts of a series of alcohols (methanol, ethanol, pro- panol, and butanol) on the CD:guest interaction at 25°C. The study was carried out by measuring the pH of the hydroalcoholic solutions of the guest, whose concentration is kept constant, as a function of cyclodextrin concentration. The dissociation constant of salicylic acid and the binding constants of the inclusion com- plexes formed by the CD and both the nonionized (HSA) and ionized (SA  ) forms of the guest were simultaneously determined at all alcohol concentrations by using a model previously derived by us. The carboxylic forms were found to bind the CD with higher affinities than the carboxylate partners, irrespective of the polarity of the medium and the cyclodextrin used. The ratio K CD:HSA /K CD:SA  is a constant value characteristic of the cyclodextrin, which points to the hydrophobic effect as one of the main forces involved in the association. A clear influence of the solvent polarity on the affinity of binding was found, in the sense that, as long as the medium becomes more apolar, the interaction between the drug and the cyclodextrin is weakened. A phenomenological limit association curve is proposed to define the limiting conditions for association in the presence of an alcohol as a cosolvent. © 1999 Academic Press Key Words: binding constant; -cyclodextrins; salicylic acid; inclusion complex; hydrophobic effect; solvent effect. INTRODUCTION Molecular complexation in artificial model systems is an interesting tool widely used in the interpretation of a number of biological mechanisms based on molecular recognition pro- cesses (1, 2). Many groups have devoted a great deal of effort to the study of the driving forces governing molecular com- plexation ((3), and references therein). However, in spite of the increased interest and the promising results, many questions still remain. The origin for this apparent confusion is that a receptor normally associates with a substrate as an overall result of the contribution of a series of noncovalent intermo- lecular forces: hydrophobic interactions, solvation/desolvation processes, van der Waals contacts, conformational energy, hydrogen bonds, dipole– dipole and ion– dipole interaction, and solvent effects. It is not an easy task to separate and quantify these contributions or to elucidate which is responsible for the overall free energy of binding. It is nevertheless well known that the hydrophobic effect and solvophobic forces play a relevant role in the association processes of apolar molecules in aqueous media (3, 4). It is also well accepted that among the many artificial receptors used in the study of molecular recog- nition events with model systems, cyclodextrins (CDs) are considered the most suitable host molecules for the recognition in aqueous media of guest molecules with hydrophobic parts (i.e., drugs, surfactants, dyes, pesticides, etc.) via noncovalent inclusion in their apolar cavities (5). Actually, these cyclic oligosaccharides constituted by (1 3 4) linked glucopyranose units, have been widely used for the improvement of the bioavailability of apolar drugs with serious problems of side effects, limited aqueous solubilities, or instabilities ((5) and references therein). Our group has been involved during past years in the study of the different factors which govern and affect the inclusion of a series of drug molecules within the cavity of various CDs (6). Recently, we analyzed the effect of the temperature and the type and/or position of substituents in both the CD and the guest molecule on the association of a series of hydroxyben- zoic acids and -cyclodextrin (-CD) and its derivatives (7). It was found that the monohydroxybenzoic acids are encapsu- lated by the CDs studied in 1:1 stoichiometries; the carboxylic forms of the guests bind cyclodextrin with higher affinities than the carboxylate partners, independently of the temperatures and of the substituents of both host and guest molecules. The association processes were found to be enthalpy-driven, with affinities of binding decreasing with increasing temperature, indicating the contribution of van der Waals contacts, the hydrophobic effect, and solvent reorganization, as the main driving forces governing the interactions. The present work is focused on the study of the role of the solvent and the hydro- phobic effect on these association processes. We have chosen 1 To whom correspondence should be addressed. Fax: 34-91-3944135. E-mail: aicart@eucmax.sim.ucm.es. Journal of Colloid and Interface Science 216, 154 –160 (1999) Article ID jcis.1999.6290, available online at http://www.idealibrary.com on 154 0021-9797/99 $30.00 Copyright © 1999 by Academic Press All rights of reproduction in any form reserved.