RESUMEN
UV-A radiation (320-400 nm) induces damage to the DNA molecule and its components through photosensitized reactions. Biopterin (Bip) and its photoproducts 6-formylpterin (Fop) and 6-carboxypterin (Cap) accumulate in the skin of human beings suffering from vitiligo, a depigmentation disorder where the protection against UV radiation fails because of the lack of melanin. This study was aimed to evaluate the photosensitizing properties of oxidized pterins present in the skin and to elucidate the mechanisms involved in the photosensitized oxidation of purine nucleotides by pterins in vitro. For this purpose, steady-state and time-resolved experiments in acidic (pH 5.0-5.8) aqueous solution were performed using Bip, Fop and Cap as photosensitizers and the nucleotide 2'-deoxyguanosine 5'-monophosphate (dGMP) as an oxidizable target. The three pterin derivatives are able to photosensitize dGMP, being Fop the most efficient sensitizer. The reactions proceed through two competing pathways: (1) electron transfer from dGMP to triplet excited-state of pterins (type I mechanism) and (2) reaction of dGMP with (1)O(2) produced by pterins (type II mechanism). Kinetic analysis revealed that the electron transfer pathway is the main mechanism and the interaction of dGMP with the triplet excited-state of pterins and the formation of the corresponding dGMP radicals were demonstrated by laser flash photolysis experiments. The biological implications of the results obtained are also discussed.
Asunto(s)
Biopterinas/química , Nucleótidos de Desoxiguanina/química , Fármacos Fotosensibilizantes/química , Transporte de Electrón , Concentración de Iones de Hidrógeno , Cinética , Oxidación-Reducción , Fotólisis , Oxígeno Singlete/química , Rayos UltravioletaRESUMEN
7,8-Dihydrobiopterin (H(2)Bip) and 7,8-dihydroneopterin (H(2)Nep) belong to a class of heterocyclic compounds present in a wide range of living systems. H(2)Bip accumulates in the skin of patients suffering from vitiligo, whereas H(2)Nep is secreted by human macrophages when the cellular immune system is activated. We have investigated the photochemical reactivity of both compounds upon UV-A irradiation (320-400 nm), the chemical structures of the products and their thermal stability. The study was performed in neutral aqueous solutions. The reactions were followed by UV/Visible spectrophotometry and HPLC and the products were analyzed by means of electrospray ionization mass spectrometry and (1)H-NMR. Excitation of H(2)Bip and H(2)Nep leads to the formation, in each case, of two main isomeric dimers. The latter compounds undergo a thermal process that may consist in a retro [2 + 2]-cycloaddition and hydrolysis to yield the reactant (H(2)Bip or H(2)Nep) and a product that has incorporated a molecule of H(2)O.
Asunto(s)
Biopterinas/análogos & derivados , Neopterin/análogos & derivados , Biopterinas/química , Cromatografía Líquida de Alta Presión , Dimerización , Humanos , Isomerismo , Macrófagos/metabolismo , Espectroscopía de Resonancia Magnética , Neopterin/química , Neopterin/metabolismo , Fotólisis , Espectrometría de Masa por Ionización de Electrospray , Temperatura , Rayos UltravioletaRESUMEN
Pterins belong to a class of heterocyclic compounds present in a wide range of living systems and accumulate in the skin of patients affected by vitiligo, a depigmentation disorder. The study of the emission of 7,8-dihydropterins is difficult because these compounds are more or less unstable in the presence of O(2) and their solutions are contaminated with oxidized pterins which have much higher fluorescence quantum yields (Φ(F)). In this work, the emission properties of six compounds of the dihydropterin family (6-formyl-7,8-dihydropterin (H(2)Fop), sepiapterin (Sep), 7,8-dihydrobiopterin (H(2)Bip), 7,8-dihydroneopterin (H(2)Nep), 6-hydroxymethyl-7,8-dihydropterin (H(2)Hmp), and 6-methyl-7,8-dihydropterin (H(2)Mep)) have been studied in aqueous solution. The fluorescence characteristics (spectra, Φ(F), lifetimes (τ(F))) of the neutral form of these compounds have been investigated using the single-photon-counting technique. Φ(F) and τ(F) values obtained lie in the ranges 3-9 × 10(-3) and 0.18-0.34 ns, respectively. The results are compared to those previously reported for oxidized pterins.
Asunto(s)
Oxígeno/química , Pterinas/química , Agua/química , Biopterinas/análogos & derivados , Biopterinas/química , Neopterin/análogos & derivados , Neopterin/química , Oxidación-Reducción , Teoría Cuántica , Soluciones/química , Espectrometría de FluorescenciaRESUMEN
Breast milk reduces the incidence of necrotizing enterocolitis (NEC). BH4 is a cofactor for endothelial NOS (eNOS). Reduced BH4 levels, or its oxidation to dihydrobiopterin (BH2), uncouple eNOS resulting in formation of reactive oxygen species (ROS) that have been implicated in the pathogenesis of NEC. We evaluated colostrum and mature breast milk, as well as infant formula, BH4 and BH2 content. In addition, we tested the BH4 effect on the newborn rat mesenteric arterial vascular tone. BH4 and BH2 content increased 3-fold in mature breast milk, when compared with colostrum (p < 0.01), without a change in their ratio. Infant formula had a negligible BH4 content and lower biopterins ratio, when compared with breast milk. eNOS is the predominant synthase isoform in newborn rat mesenteric arteries. In the presence of BH4, mesenteric arteries contracted less to thromboxane A2 analog U46619 (p < 0.01) and this effect was abolished following eNOS inhibition. BH4 (10â»6 M) vasorelaxed the newborn rat mesenteric arteries. We conclude that when compared with infant formula, breast milk has a high BH4 content that increases as breastfeeding continues. Given its mesenteric arterial vasorelaxing effect, BH4 may play an important role in the reduced NEC incidence among breast fed infants.
Asunto(s)
Animales Recién Nacidos/anatomía & histología , Biopterinas/análogos & derivados , Arterias Mesentéricas/efectos de los fármacos , Leche Humana/química , Vasodilatación/efectos de los fármacos , Animales , Biopterinas/química , Biopterinas/farmacología , Bovinos , Cromatografía Liquida , Enterocolitis Necrotizante/patología , Enterocolitis Necrotizante/prevención & control , Femenino , Humanos , Fórmulas Infantiles/química , Isoenzimas/metabolismo , Óxido Nítrico Sintasa/metabolismo , Periodo Posparto , Ratas , Ratas Sprague-Dawley , Espectrometría de Masas en TándemRESUMEN
Dihydrobiopterin (H(2)Bip) and its oxidized analogue, biopterin (Bip), accumulate in the skin of patients suffering from vitiligo, a chronic depigmentation disorder in which the protection against UV radiation fails. The photochemistry of H(2)Bip was studied in neutral aqueous solutions upon UV-A irradiation (320-400 nm) at room temperature. The photochemical reactions were followed by UV/vis spectrophotometry, HPLC and enzymatic methods for hydrogen peroxide (H(2)O(2)) determination. Photoproducts were analyzed by means of electrospray ionization mass spectrometry. Under anaerobic conditions, excitation of H(2)Bip leads to the formation of at least two isomeric dimers with molecular masses equal to exactly twice the molecular mass of the reactant. This reaction takes place from the singlet excited state of the reactant. To the best of our knowledge, this is the first time that the photodimerization of a dihydropterin is reported. In the presence of air, the dimers are again the main photoproducts at the beginning of the reaction, but a small proportion of the reactant is converted into Bip. As the reaction proceeds and enough Bip accumulates in the solution, a photosensitized process starts, where Bip photoinduces the oxidation of H(2)Bip to Bip, and H(2)O(2) is formed. As a consequence, the rates of H(2)Bip consumption and Bip formation increase as a function of irradiation time, resulting in an autocatalytic photochemical process. In this process, Bip in its triplet excited state reacts with the ground state of H(2)Bip. The mechanisms involved are analyzed and the biological implications of the results are discussed.
Asunto(s)
Biopterinas/análogos & derivados , Peróxido de Hidrógeno/química , Oxígeno/química , Agua/química , Biopterinas/química , Cromatografía Líquida de Alta Presión , Humanos , Oxidación-Reducción , Soluciones Farmacéuticas/química , Fotoquímica/métodos , Fotoquímica/tendencias , Soluciones , Espectrofotometría UltravioletaRESUMEN
New photochemical studies of the reactivity of biopterin (BPT) and neopterin (NPT) in acidic (pH = 5.5) and alkaline (pH = 10.5) aqueous solutions at 350 nm and room temperature were performed. The photochemical properties of BPT are of particular interest because the photolysis of this compound takes place in the white skin patches of patients affected by vitiligo. The photochemical reactions were followed by UV/VIS spectrophotometry, HPLC, electrochemical measurement of dissolved O(2) and enzymatic methods for hydrogen peroxide (H(2)O(2)) and superoxide anion (O(2)(-)) determinations. When BPT or NPT are exposed to UVA radiation, a red intermediate, very likely 6-formyl-5,8-dihydropterin, is generated in an O(2)-independent process. That product is rapidly oxidized on admission of O(2) to yield 6-formylpterin and H(2)O(2). When the photolysis takes place in aerobic conditions, no additional pathways exist. On the other hand, in the absence of O(2), the intermediate generated is not stable and leads to the formation of many products. O(2)(-) is also generated during photo-oxidation of BPT and NPT. The quantum yields of reactant consumption depends on the O(2) concentration: the higher the O(2) concentration, the lower the quantum yields. This behavior is discussed in connection with the excited state of the pterins.