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Comment on “Excited-State Intermolecular Proton Transfer of Lumazine” M. Paula Denofrio,† Andre´s H. Thomas,† Andre´ M. Braun,‡ Esther Oliveros,*,§ and Carolina Lorente*,† Instituto de InVestigaciones Fisicoquı´micas Teo´ricas y Aplicadas (INIFTA), Facultad de Ciencias Exactas, UniVersidad Nacional de La Plata, CCT La Plata-CONICET, Casilla de Correo 16, Sucursal 4, (1900) La Plata, Argentina; Engler-Bunte Institut, Karlsruhe Institute of Technology, D-76128 Karlsruhe, Germany, and Laboratoire des IMRCP, UMR CNRS 5623, UniVersite´ Paul Sabatier (Toulouse III), 118 route de Narbonne, F-31062 Toulouse ce´dex 9, France ReceiVed: April 29, 2010; ReVised Manuscript ReceiVed: July 8, 2010 In their article, Huppert et al.1 reported the emission properties of lumazine (Lum) using steady-state and timeresolved emission techniques. They also report the absorption spectra at different pH and they evaluated the pKa value in water to be about 10.5. Lum belongs to a family of heterocyclic compounds present in biological systems and called pteridines. Pteridines behave as weak acids in aqueous solution, and several acid-base equilibria may be present. Acid-base properties of Lum were reported several decades ago,2-4 but these publications are not cited by Huppert et al.1 In these former studies, pKa values were found to be 7.91, 7.76, and 7.95, respectively. These values correspond to the equilibrium between the neutral form (acid form) and the monoanion (basic form) of Lum (Figure 1). The big difference between the newly reported value (10.5) and the previously found values (7.76-7.95) and the absence of any kind of explanation by Huppert et al.1 incited us to reproduce our own titration measurements (Figure 1) and to determine again the pKa.5 We found a value consistent with Pfleiderer’s value4 (8.0 ( 0.1). The absorption spectra of both acid and base forms, although quite different, have intense bands in the UV-A region (320-400 nm) (Figure 1). Therefore, the pKa value of Lum of 10.5 reported by Huppert et al.1 is in strong contradiction with all previous measurements2-4 and our own results. In their article, the authors do not describe (e.g., in the experimental section) how the pH was measured. At this point, we believe that the pH was not measured in an appropriate way. In addition, our own (unpublished) results on the emission of Lum indicate that the fluorescence of Lum is quenched by hydroxide anions (in agreement with Klein and Tatischeff6). Consequently, a decrease of the fluorescence is observed at pH higher than 10.5 (Figure 2).7 We therefore believe that the evolution (increase) of the fluorescence with the pH, as presented by Huppert et al.1 (Figure 3 in their article), is also erroneous. Moreover, Klein and Tatischeff published in 1987 detailed results on the fluorescence of different forms of Lum.8 The article describes the fluorescence spectra in a wide range of pH values and attributes the four emission maxima observed †
Universidad Nacional de La Plata. Karlsruhe Institute of Technology. § Universite´ Paul Sabatier. ‡
Figure 1. (a) Absorption spectra of Lum at different pH values. Inset: acid-base equilibrium. (b) Spectrophotometric titration curve of Lum (112 µM, HCl and NaOH as titration agents).
to the four ionic species of lumazine (dianion, monoanion, neutral, cation). Moreover, the authors observed that at pH 10 Lum was a mixture of N(1)- and N(3)-deprotonated monoanions and that neutral Lum shows two fluorescence maxima, one with a large Stokes shift. The latter result was explained by the formation of a phototautomer resulting from the N(1) to N(8) water mediated proton transfer in the excited state. Therefore, according to Klein and Tatischeff, the interpretation of the photophysical properties of Lum at different pH values should consider not only the various ionic forms of Lum but also tautomeric species in both the ground and excited states (Figure 3). Surprisingly, Huppert et al. did not refer to this previous publication,8 although the results reported are very close to their own work. Moreover, they ignored the tautomerism of Lum, which implies different atoms participating in proton transfer they also postulate. Indeed, Huppert et al.1 discuss the intermolecular proton transfer in the excited state of Lum, but they
10.1021/jp103900j 2010 American Chemical Society Published on Web 08/04/2010
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Comments
Figure 3. Tautomeric forms of Lum.
analyzed the tautomeric forms of Lum neither in the ground state nor in the excited state. References and Notes
Figure 2. Fluorescence emission spectra of Lum (λEXC ) 350 nm) at (a) 5 < pH < 10 and (b) pH g10. Inset in (b): Stern-Volmer plot.
(1) Presiado, I.; Erez, Y.; Gepshtein, R.; Huppert, D. J. Phys. Chem. C 2010, 114, 3634–3640. (2) Albert, A.; Brown, D. J.; Cheeseman, G. J. Chem. Soc. 1951, 474– 485. (3) Lippert, E.; Prigge, H. Z. Elektrochem. 1960, 64, 662–671. (4) Pfleiderer, W. I. Chem. Ber. 1957, 90, 2582–2587. (5) Spectrophotometric titration; pH of the aqueous solutions was adjusted by adding drops of HCl or NaOH from a micropipette. The concentrations of the acid and base used for this purpose ranged from 0.1 to 2 M. The pH measurements were performed with a pH-meter PHM220 (Radiometer Copenhagen) and a combined pH electrode pHC2011-8 (Radiometer Analytical). (6) Tatischeff, I.; Klein, R. Hoppe-Seyler’s Physiol. Chem. 1984, 365 (S), 1255–1262. (7) Steady-state fluorescence measurements were performed on airequilibrated aqueous solutions using an Edinburgh EAI-FS/FL900 SPC equipment. The quartz cells (1 cm path length) used for the measurements were thermoregulated at 24.0 ( 0.2 °C. Corrected fluorescence spectra obtained by excitation at 350 nm (high pressure Xe lamp, 419 W) were recorded between 360 and 650 nm. (8) Klein, R.; Tatischeff, I. Photochem. Photobiol. 1987, 45, 55–65.
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