Received: 4 June 2009, Revised: 12 January 2010, Accepted: 15 January 2010, Published online in Wiley Online Library: 26 March 2010 Polymethine cyanine dyes in b-cyclodextrin solution: multiple equilibria and chemical oxidation y Teresa C. Barros a , Sergio H. Toma b , Henrique E. Toma b , Erick L. Bastos c * and Mauricio S. Baptista** Absorption and fluorescence spectroscopy, electrochemical techniques, and semiempirical calculations were employed to characterize the multiple complexation equilibria between two polymethine cyanine dyes (IR-786 and Indocyanine green-ICG, S) and b-cyclodextrin (b-CD, L), as well as the chemical reactivity of the complexed and uncomplexed species against the oxidizing agents hypochlorite (HC) and hydrogen peroxide (HP). IR-786 dimerization is favored with the increase in b-CD concentration in the form of (SL) 2 complexes. In the case of ICG, free dimers (D) and SL complexes are favored. Both IR-786 and ICG react and discolor in the presence of HC and HP. For IR-786, the reaction with HP and HC proceeds with observed rate constants of 10 S3 and 0.28 s S1 and second-order rate constants (k 2 ) of 10 S3 and 10 4 M S1 s S1 , respectively. The intermediate species observed in the bleaching reactions of IR-786 and ICG were shown, by cyclic voltammetry and VIS absorption, to result from one electron oxidation. IR-786 complexed with b-CD is protected against bleaching in the presence of HP and HC by factors of 20 and 4, respectively. This protection was not observed in ICG complexes. Superdelocalizability profile of both dyes and frontier orbital analysis indicates that b-CD does not protect ICG from oxidation by HP or HC, whereas the 2:2 IR-786/b-Cd complex is able to avoid the oxidation of IR-786. We concluded that the decrease in the chemical reactivity of the dyes against oxidant agents in the presence of b-CD is due to the formation of (SL) 2 complexes. Copyright ß 2010 John Wiley & Sons, Ltd. Supporting information may be found in the online version of this paper. Keywords: chemical oxidation; cyclodextrin; inclusion complex; inclusion complex; near-infrared dyes INTRODUCTION Indocyanine green (ICG) and IR-786 (Scheme 1) are polymethine cyanine (PC) dyes with large extinction coefficients in the near-infrared (NIR) region. [1,2] Such dyes find increasing interest in biotechnology and medical applications because light in the NIR region is poorly absorbed by biomolecules, and thus can penetrate deeply into tissues. [3,4] ICG was early recognized as a suitable NIR fluorogenic label, being used in several analytical and medical applications, including photodynamic therapy (PDT). [1,2,5–10] IR-786 is widely used in enzymatic and immu- noassays, DNA sequencing applications and it offers potential to target mitochondria due to its positive charge. [1,11–17] ICG and IR-786, as well as several NIR cyanine dyes, contain a conjugate imidinium system with an odd number of methine groups. [18] The charge alternation along the chromophoric chain and charge inversion upon photoexcitation determine high efficiency of light absorption in the visible and IR spectra regions and strong tendency to aggregation. [5,18–21] In fact, Van der Waals forces and dispersion interactions of p-conjugated systems play a major role in the formation of dimers. [22] Therefore, ICG and IR-786 are known to form H-dimers in concentrated aqueous solutions, resulting in blue shift in the absorption maximum as well as fluorescence suppression. [23–27] PC dyes also show high chemical reactivity with weak oxidizing agents. [28] Actually, both dyes (as well as most of the PC family) completely lose their colors few seconds after getting contact with tap water. Oxidation leads to the breakdown of the p-electron system resulting in structural and spectroscopic changes (dye bleaching) with loss of activity and formation of smaller molecules. As a result, dye bleaching in biological fluids is perhaps the most important factor preventing the widespread use of these molecules. [29] (wileyonlinelibrary.com) DOI 10.1002/poc.1692 Research Article * Correspondence to: E. L. Bastos, Centro de Cie ˆncias Naturais e Humanas, Av. dos Estados, 5001, Bloco B, sala 1023, 09210-170 Santo Andre ´, SP, Brazil. E-mail: erick.bastos@ufabc.edu.br ** Correspondence to: M. S. Baptista, Department of Biochemistry, Av. Prof. Lineu Prestes, 748, Bloco 12S, sala 1262, 05513-970 Sa ˜o Paulo, SP, Brazil. E-mail: baptista@iq.usp.br a T. C. Barros Instituto de Cie ˆncias da Sau ´de, Universidade Paulista, Bauru, SP, Brazil b S. H. Toma, H. E. Toma, M. S. Baptista Instituto de Quı ´mica, Universidade de Sa ˜o Paulo, Sa ˜o Paulo, SP, Brazil c E. L. Bastos Centro de Cie ˆncias Naturais e Humanas, Universidade Federal do ABC, Santo Andre ´, SP, Brazil y This article is published in Journal of Physical Organic Chemistry as a special issue on Tenth Latin American Conference on Physical Organic Chemistry, edited by Faruk Nome, Dept de Quimica, Universidade Federal de Santa Catarina, Campus Universitario – Trindade 88040-900, Florianopolis-SC, Brazil. J. Phys. Org. Chem. 2010, 23 893–903 Copyright ß 2010 John Wiley & Sons, Ltd. 893