Photochemistry and Photobiology, 2005, zyxwvut 81 : 793-801 zyxwvu Symposium-in-Print: Vlll ELAFOT, La Plata, Argentina, 2004 Photochemical Behavior of 6-Methylpterin in Aqueous Solutions: Generation of Reactive Oxygen Speciesq Franco M. Cabrerizo', Carolina Lorente2, Mariana Vignoni', Romina Cabrerizo', Andres H. Thomas*' and Albert0 L. Capparelli*' ' lnstituto de lnvestigaciones FisicoquimicasTeoricas y Aplicadas (INIFTA), Facultad de Ciencias Exactas, Universidad Nacional de La Plata, CONICET, La Plata, Argentina *CHIDECAR-CONICET, Departamento de Quimica Organica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Buenos Aires, Argentina Received 29 November 2004; accepted 11 February 2005 ABSTRACT zyxwvutsrq Pterins are a family of heterocyclic compounds present in a wide range of living systems that participate in relevant bio- logical functions and are involved in different photobiological processes. 6-Methylpterin (MPT) was investigated for its effi- ciency of singlet-oxygen zyxwvutsrq (lo2) production and quenching in aqueous solution. The quantum yields of '02 production (@a) was determined by measurements of the '02 luminescence in the near-infrared upon continuous excitation of the sensitizer. Values of @A were found to be 0.10 k 0.02 and 0.14 k 0.02 in acidic and alkaline media, respectively. Studies of the photo- oxidation of MPT in acidic (pH = 5.0-6.0) and alkaline (pH = 10.2-10.8) aqueous solutions at 350 nm and room temperature have been performed. The photochemical reactions were followed by UV-visible spectrophotometry, high-performance liquid chromatography and an enzymatic method for HzOz determination. MPT is not light sensitive in the absence of oxygen, but it undergoes a photooxidation reaction in the presence of oxygen, yielding several nonpteridinic products. The quantum yields of MPT disappearance were determined and values of 2.4 (&0.5) X and 8.1 (k0.8) X were obtained in acidic and alkaline media, respectively. Hz02 was detected and quantified in irradiated solutions of MPT. The rate constant of the chemical reaction between '02 and MPT zyxwvut (k,) was determined to be 4.9 X lo6 zyxwvut A4-l s-l in alkaline TPosted on the website on 18 Feb 2005 *To whom correspondence should be addressed: INIFTA, Casilla de Correo 16, Sucursal4, (1900) La Plata, Argentina. Fax: 0054-221-424- 2333; AHT e-mail: athomas@inifta.unlp.edu.ar ALC e-mail: alcappa@dalton.quimica.unlp.edu.ar This work was presented at the VIII ELAFOT, La Plata, Argentina. Abbreviations: BF'T, 6-biopterin; CPT, 6-carboxypterin; ED, experimental difference; FA, folic acid FPT, 6-formylpterin; HPLC, high-performance liquid chromatography; HF'T, 6-hydroxymethylpterin; IR, infrared; MPT, 6-methylpterin; NED, normalized experimental difference: NIR, near- infared; '02, singlet-oxygen; NPT, neopterin; F'T, pterin; RB, rose bengal; ROS, reactive oxygen species; Sens, sensitizer. zyxwvut 0 2005 American Society for Photobiology 0031-8655/05 medium and the role of '02 in the photooxidation of MPT is discussed. INTRODUCTION Pterins are heterocyclic compounds widespread in biological sys- tems, participating in relevant biological functions. Folic acid, a conjugated pterin, acts as a coenzyme in reactions related to the synthesis of puric and pirimidinic bases (1). Tetrahydrobiopterin acts as a coenzyme in hydroxylation reactions of the metabolism of some amino acids (2) and is also relevant in the metabolism of nitric oxide (3,4). Some pterin derivatives zyx (e.g. xantopterin, leucopterin) are present in butterflies as natural pigments zyx (5). 6-Substituted pterins, the most common pterin derivatives found in nature, behave as weak acids in aqueous solutions. As reported by Albert (6) for several compounds, at pH higher than 5, pterins present an acid-base equilibrium that involves an amide group (acid form) and a phenolate group (basic form) (Fig. 1). The pK, of this equilibrium is around 8 for different studied pterin derivatives (6-9). Other functional groups of the pterin moiety (e.g. 2-amino group or ring nitrogen atoms) have pK, values lower than 2 (6). Light sensitivity of pterins has been known for several decades (10,ll). Interest in photochemistry and photophysics of this group of compounds has been shown since participation of pterins in different photobiological processes has been proven or suggested. Pterins may play some role in photoreception processes that take place in plants (12,13). 5,1O-Methenyltetrahydrofolate, a folk acid de- rivative, is present as the light-harvesting antenna in DNA photo- lyases (14,15), enzymes involved in DNA repair processes after UV light irradiation. Pterins are also interesting as possible photo- sensitizers in photochemical processes that take place in organisms exposed to UV-A radiation; e.g. pterins participate as sensitizers in photochemical reactions that induce DNA damage (16,17). Singlet molecular oxygen (02 ['A,], denoted throughout as lo2), the lowest electronic excited state of molecular oxygen, is an im- portant reactive oxygen species (ROS) and one of the main activated species responsible for the damaging effects of light on biological systems (photodynamic effects) (1 8). This activated metastable state is much more reactive than the ground triplet state zy (02 r3A;], denoted throughout as 302) and has been attracting interest in both 793