Reactions of Manganese Porphyrins with Peroxynitrite and Carbonate Radical Anion
2003; Elsevier BV; Volume: 278; Issue: 30 Linguagem: Inglês
10.1074/jbc.m213302200
ISSN1083-351X
AutoresGerardo Ferrer‐Sueta, Darío A. Vitturi, Ines Batinić‐Haberle, Irwin Fridovich, Sara Goldstein, Gidon Czapski, Rafael Radí,
Tópico(s)Electrochemical Analysis and Applications
ResumoWe have studied the reaction kinetics of ten manganese porphyrins, differing in their meso substituents, with peroxynitrite (ONOO–) and carbonate radical anion (CO3·¯ ) using stopped-flow and pulse radiolysis, respectively. Rate constants for the reactions of Mn(III) porphyrins with ONOO– ranged from 1 × 105 to 3.4 × 107m–1 s–1 and correlated well with previously reported kinetic and thermodynamic data that reflect the resonance and inductive effects of the substituents on the porphyrin ring. Rate constants for the reactions of Mn(III) porphyrins with CO3·¯ ranged from 2 × 108 to 1.2 × 109m–1s–1 at pH ≤ 8.5 and increased with pH as a consequence of the ionization of the complexes. Mn(II) porphyrins reacted with CO3·¯ with rate constants ranging from 1 × 109 to 5 × 109m–1s–1 at pH 10.4. Hence, fast scavenging of ONOO– and CO3·¯ by manganese porphyrins could occur in vivo because of the catalytic reduction at the expense of a number of cellular reductants. Additionally, we determined the pK a of the axial water molecules of the Mn(III) complexes at pH 7.5–13.2 by spectrophotometric titration. Results were consistent with two acid-base equilibria for most of the complexes studied. The pK a values also correlated with the resonance and inductive effects of the substituents. The correlations of E½ with the rate constants with ONOO– and with the pK a values display a deviation from linearity when N-alkylpyridinium substituents included N-alkyl moieties longer than ethyl, which is interpreted in terms of a decrease in the local dielectric constant. We have studied the reaction kinetics of ten manganese porphyrins, differing in their meso substituents, with peroxynitrite (ONOO–) and carbonate radical anion (CO3·¯ ) using stopped-flow and pulse radiolysis, respectively. Rate constants for the reactions of Mn(III) porphyrins with ONOO– ranged from 1 × 105 to 3.4 × 107m–1 s–1 and correlated well with previously reported kinetic and thermodynamic data that reflect the resonance and inductive effects of the substituents on the porphyrin ring. Rate constants for the reactions of Mn(III) porphyrins with CO3·¯ ranged from 2 × 108 to 1.2 × 109m–1s–1 at pH ≤ 8.5 and increased with pH as a consequence of the ionization of the complexes. Mn(II) porphyrins reacted with CO3·¯ with rate constants ranging from 1 × 109 to 5 × 109m–1s–1 at pH 10.4. Hence, fast scavenging of ONOO– and CO3·¯ by manganese porphyrins could occur in vivo because of the catalytic reduction at the expense of a number of cellular reductants. Additionally, we determined the pK a of the axial water molecules of the Mn(III) complexes at pH 7.5–13.2 by spectrophotometric titration. Results were consistent with two acid-base equilibria for most of the complexes studied. The pK a values also correlated with the resonance and inductive effects of the substituents. The correlations of E½ with the rate constants with ONOO– and with the pK a values display a deviation from linearity when N-alkylpyridinium substituents included N-alkyl moieties longer than ethyl, which is interpreted in terms of a decrease in the local dielectric constant. Metalloporphyrins catalyze numerous redox reactions (1Meunier B. Chem. Rev. 1992; 92: 1411-1456Crossref Scopus (2095) Google Scholar); in particular, manganese porphyrins have been used as redox catalysts in several model systems relevant to biochemistry, for instance, as superoxide dismutase (2Batinic-Haberle I. Methods Enzymol. 2002; 349: 223-233Crossref PubMed Scopus (64) Google Scholar, 3Spasojevic I. Batinic-Haberle I. Reboucas J.S. Idemori Y.M. Fridovich I. J. Biol. Chem. 2003; 278: 6831-6837Abstract Full Text Full Text PDF PubMed Scopus (65) Google Scholar) and catalase (4Day B.J. Fridovich I. Crapo J.D. Arch. Biochem. Biophys. 1997; 347: 256-262Crossref PubMed Scopus (268) Google Scholar) mimics. Some of the N-alkylpyridinium substituted complexes afforded protection of superoxide dismutase-deficient Escherichia coli from O2 toxicity (5Batinic-Haberle I. Spasojevic I. Hambright P. Benov L. Crumbliss A.L. Fridovich I. Inorg. Chem. 1999; 38: 4011-4022Crossref Scopus (241) Google Scholar) and in several rodent models of transient brain ischemia (6Mackensen G.B. Patel M. Sheng H. Calvi C.L. Batinic-Haberle I. Day B.J. Liang L.P. Fridovich I. Crapo J.D. Pearlstein R.D. Warner D.S. J. Neurosci. 2001; 21: 4582-4592Crossref PubMed Google Scholar, 7Sheng H. Enghild J.J. Bowler R. Patel M. Batinic-Haberle I. Calvi C.L. Day B.J. Pearlstein R.D. Crapo J.D. Warner D.S. Free Radic. Biol. Med. 2002; 33: 947-961Crossref PubMed Scopus (98) Google Scholar), diabetes (8Piganelli J.D. Flores S.C. Cruz C. Koepp J. Batinic-Haberle I. Crapo J. Day B. Kachadourian R. Young R. Bradley B. Haskins K. 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Moreover, MnIIITCPP 1The abbreviations used are: MnIIITCPP, manganese(III) meso-tetrakis(4-carboxylatophenyl)porphyrin; MnIIITM-2-PyP, manganese-(III)meso-tetrakis((N-methyl)pyridinium-2-yl)porphyrin; MnIIITM-4-PyP, manganese(III)meso-tetrakis((N-methyl)pyridinium-4-yl)porphyrin; MnIIITSPP, manganese(III)meso-tetrakis(4-sulfonatophenyl)-porphyrin; MnIIITE-2-PyP, manganese(III)meso-tetrakis((N-ethyl)pyridinium-2-yl)porphyrin; MnIIITnPr-2-PyP, manganese(III)meso-tetrakis((N-n-propyl)pyridinium-2-yl)porphyrin; MnIIITnBu-2-PyP, manganese(III)meso-tetrakis((N-n-butyl)pyridinium-2-yl)porphyrin; MnIIITnHex-2-PyP, manganese(III)meso-tetrakis((N-n-hexyl)pyridinium-2-yl)porphyrin; MnIIITnOct-2-PyP, manganese(III)meso-tetrakis((N-n-octyl)pyridinium-2-yl)porphyrin; MnIIITM-3-PyP, manganese(III)meso-tetrakis((N-methyl)pyridinium-3-yl)porphyrin. has been effective in a number of model studies of oxidative stress-mediated injury (for a review see Ref. 11Patel M. Day B.J. Trends. Pharmacol. Sci. 1999; 20: 359-364Abstract Full Text Full Text PDF PubMed Scopus (252) Google Scholar) despite having low superoxide dismutase and catalase activities. Our group (12Ferrer-Sueta G. Ruiz-Ramirez L. Radi R. Chem. Res. Toxicol. 1997; 10: 1338-1344Crossref PubMed Scopus (45) Google Scholar, 13Ferrer-Sueta G. Batinic-Haberle I. Spasojevic I. Fridovich I. Radi R. Chem. Res. Toxicol. 1999; 12: 442-449Crossref PubMed Scopus (157) Google Scholar, 14Ferrer-Sueta G. Quijano C. Alvarez B. Radi R. Methods Enzymol. 2002; 349: 23-37Crossref PubMed Scopus (72) Google Scholar) and others (15Lee J. Hunt J.A. Groves J.T. Bioorg. Med. Chem. Lett. 1997; 7: 2913-2918Crossref Scopus (59) Google Scholar) have studied the capability of manganese porphyrins for the catalytic reduction of peroxynitrite (ONOOH/ONOO–), a powerful oxidant that can be formed in vivo by the reaction of O2·¯ with ·NO (16Beckman J.S. Beckman T.W. Chen J. Marshall P.A. Freeman B.A. Proc. Natl. Acad. Sci. U. S. A. 1990; 87: 1620-1624Crossref PubMed Scopus (6718) Google Scholar, 17Radi R. Peluffo G. Alvarez M.N. Naviliat M. Cayota A. Free Radic. Biol. Med. 2001; 30: 463-488Crossref PubMed Scopus (675) Google Scholar). A significant fraction of the oxidative biochemistry of peroxynitrite is derived from the rapid reaction of ONOO– with CO2 (k = 5.8 × 104m–1s–1 at 37 °C (18Radi R. Cosgrove T.P. Beckman J.S. Freeman B.A. Biochem. J. 1993; 290: 51-57Crossref PubMed Scopus (348) Google Scholar, 19Lymar S.V. Hurst J.K. J. Am. Chem. Soc. 1995; 117: 8867-8868Crossref Scopus (495) Google Scholar, 20Denicola A. Freeman B.A. Trujillo M. Radi R. Arch. Biochem. Biophys. 1996; 333: 49-58Crossref PubMed Scopus (508) Google Scholar)), which produces the carbonate radical anion (CO3·¯ ) and nitrogen dioxide (·NO2) with about 33% yield (see Equation 1, below), with the remaining yielding carbon dioxide and nitrate, where k 1b/k 1a ≈ 2 (21Bonini M.G. Radi R. Ferrer-Sueta G. Ferreira A.M. Augusto O. J. Biol. Chem. 1999; 274: 10802-10806Abstract Full Text Full Text PDF PubMed Scopus (281) Google Scholar, 22Goldstein S. Czapski G. J. Am. Chem. Soc. 1998; 120: 3458-3463Crossref Scopus (167) Google Scholar, 23Lymar S.V. Hurst J.K. Inorg. Chem. 1998; 37: 294-301Crossref Scopus (166) Google Scholar, 24Hodges G.R. Ingold K.U.J. J. Am. Chem. Soc. 1999; 121: 10695Crossref Scopus (123) Google Scholar). x0026;x0026;ONOO-+CO2x0026;↗x0026;CO3·¯+·NO2x0026;x0026;x0026;↘x0026;CO2+NO3-(Eq. 1) Given the ubiquity of CO2, its high concentration (1 to 2 mm in human tissues), and the reactivity of CO3·¯ (25Augusto O. Bonini M.G. Amanso A.M. Linares E. Santos C.C. De Menezes S.L. Free Radic. Biol. Med. 2002; 32: 841-859Crossref PubMed Scopus (476) Google Scholar), a useful peroxynitrite scavenger needs to out-compete the target molecules and CO2 and/or be able to efficiently scavenge CO3·¯ . We have proposed that complexes such as MnIIITM-2-PyP can efficiently inhibit peroxynitrite-mediated oxidations even in the presence of CO2 (13Ferrer-Sueta G. Batinic-Haberle I. Spasojevic I. Fridovich I. Radi R. Chem. Res. Toxicol. 1999; 12: 442-449Crossref PubMed Scopus (157) Google Scholar). Moreover, the reaction of MnIIITM-2-PyP with ONOO– in the presence of CO2 produced more oxidation of the metal complex than expected based on simple competition kinetics (13Ferrer-Sueta G. Batinic-Haberle I. Spasojevic I. Fridovich I. Radi R. Chem. Res. Toxicol. 1999; 12: 442-449Crossref PubMed Scopus (157) Google Scholar), suggesting a probable reaction of the complex with CO3·¯ . The possible reaction of Mn(III) porphyrins with CO3·¯ has also been proposed recently (26Zhang H. Joseph J. Gurney M. Becker D. Kalyanaraman B. J. Biol. Chem. 2002; 277: 1013-1020Abstract Full Text Full Text PDF PubMed Scopus (97) Google Scholar) in experiments related to the effect of bicarbonate on the peroxidase activity of Cu,Zn superoxide dismutase. In aerated aqueous solution, the stable oxidation state of manganese porphyrins used in this study is Mn(III). However, given the low oxygen tension inside the cell, cellular components, like low molecular weight reductants (27Spasojevic I. Batinic-Haberle I. Fridovich I. Nitric Oxide. 2000; 4: 526-533Crossref PubMed Scopus (69) Google Scholar) and probably some dehydrogenases (28Faulkner K.M. Liochev S.I. Fridovich I. J. Biol. Chem. 1994; 269: 23471-23476Abstract Full Text PDF PubMed Google Scholar), can produce Mn(II) porphyrins and maintain them in the reduced state. Mn(II) porphyrin chemistry has been studied since the seventies (29Hambright P. Chock P.B. Inorg. Chem. 1974; 13: 3029-3031Crossref Scopus (26) Google Scholar, 30Duncan I.A. Harriman A. Porter G. J. Chem. Soc. Faraday Trans. II. 1980; 76: 1415Crossref Scopus (17) Google Scholar, 31Pasternack R.F. Banth A. Pasternack J.M. Johnson C.S. J. Inorg. Biochem. 1981; 15: 261-267Crossref PubMed Scopus (67) Google Scholar), but only recently has its biochemistry begun to be explored (27Spasojevic I. Batinic-Haberle I. Fridovich I. Nitric Oxide. 2000; 4: 526-533Crossref PubMed Scopus (69) Google Scholar). Mn(II) porphyrins may have a number of advantages over Mn(III) porphyrins with regard to their scavenging and antioxidant activity. For instance, they could rapidly scavenge oxidizing radicals such as CO3·¯ and yield innocuous products, or they could reduce strong oxidants like ONOO– or H2O2 via a two-electron transfer reaction without producing any secondary radicals. This latter reaction is particularly important if Mn(II) can be regenerated at the expense of readily available biological reductants (14Ferrer-Sueta G. Quijano C. Alvarez B. Radi R. Methods Enzymol. 2002; 349: 23-37Crossref PubMed Scopus (72) Google Scholar). Carbonate radical anion is long known to radiation chemists (32Weeks J.L. Rabani J. J. Phys. Chem. 1966; 70: 2100-2106Crossref Scopus (190) Google Scholar, 33Behar D. Czapski G. Duchovny I. J. Phys. Chem. 1970; 74: 2206-2210Crossref Scopus (165) Google Scholar) but has only recently drawn attention of biochemists because of its formation from the reaction of ONOO– with CO2 (16Beckman J.S. Beckman T.W. Chen J. Marshall P.A. Freeman B.A. Proc. Natl. Acad. Sci. U. S. A. 1990; 87: 1620-1624Crossref PubMed Scopus (6718) Google Scholar, 17Radi R. Peluffo G. Alvarez M.N. Naviliat M. Cayota A. Free Radic. Biol. Med. 2001; 30: 463-488Crossref PubMed Scopus (675) Google Scholar, 18Radi R. Cosgrove T.P. Beckman J.S. Freeman B.A. Biochem. J. 1993; 290: 51-57Crossref PubMed Scopus (348) Google Scholar, 19Lymar S.V. Hurst J.K. J. Am. Chem. Soc. 1995; 117: 8867-8868Crossref Scopus (495) Google Scholar, 20Denicola A. Freeman B.A. Trujillo M. Radi R. Arch. Biochem. Biophys. 1996; 333: 49-58Crossref PubMed Scopus (508) Google Scholar, 21Bonini M.G. Radi R. Ferrer-Sueta G. Ferreira A.M. Augusto O. J. Biol. Chem. 1999; 274: 10802-10806Abstract Full Text Full Text PDF PubMed Scopus (281) Google Scholar) and the effect of bicarbonate on the peroxidase activity of Cu,Zn superoxide dismutase (34Liochev S.I. Fridovich I. Free Radic. Biol. Med. 1999; 27: 1444-1447Crossref PubMed Scopus (54) Google Scholar). Carbonate radical anion is the conjugate base of a strong acid (pK a < 0) (35Czapski G. Lymar S. Schwarz H. J. Phys. Chem. A. 1999; 103: 3447-3450Crossref Scopus (102) Google Scholar) and a strong oxidant (36Huie R. Clifton C. Neta P. Radiat. Phys. Chem. 1991; 38: 477-481Google Scholar) with a characteristic spectrum in the visible (ϵ600 = 1860 m–1cm–1) (32Weeks J.L. Rabani J. J. Phys. Chem. 1966; 70: 2100-2106Crossref Scopus (190) Google Scholar, 33Behar D. Czapski G. Duchovny I. J. Phys. Chem. 1970; 74: 2206-2210Crossref Scopus (165) Google Scholar). Nevertheless, its reactivity is somewhat selective, and its potential targets in biological systems include sulfur-containing and aromatic amino acids (37Adams G.E. Aldrich J.E. Bisby R.H. Cundall R.B. Redpath J.L. Willson R.L. Radiat. Res. 1972; 49: 278-289Crossref PubMed Scopus (216) Google Scholar, 38Chen S.N. Hoffman M.Z. Radiat. Res. 1973; 56: 40-47Crossref PubMed Scopus (158) Google Scholar). Manganese porphyrins display linear free energy relationships between ligand or complex properties, e.g. pK a of pyrrolic nitrogens, E½ (Mn(III)/Mn(II)), and rate constants, such as the catalytic rate constant of O2·¯ dismutation (5Batinic-Haberle I. Spasojevic I. Hambright P. Benov L. Crumbliss A.L. Fridovich I. Inorg. Chem. 1999; 38: 4011-4022Crossref Scopus (241) Google Scholar, 39Batinic-Haberle I. Spasojevic I. Stevens R.D. Hambright P. Fridovich I. J. Chem. Soc. Dalton Trans. 2002; : 2689-2696Crossref Scopus (124) Google Scholar). These linear free energy relationships break down if N-alkylpyridinium substituted porphyrins contain alkyl groups longer than ethyl, which has been ascribed to steric and solvation differences in the metal surroundings (39Batinic-Haberle I. Spasojevic I. Stevens R.D. Hambright P. Fridovich I. J. Chem. Soc. Dalton Trans. 2002; : 2689-2696Crossref Scopus (124) Google Scholar). The complexes possess two axially coordinated water molecules, and they can undergo up to two ionization steps in the alkaline pH range. Data in the literature with respect to these ionization reactions are diverse, both in the methods used and in the results. For instance, MnIIITM-4-PyP displays a single pK a of 10 according to 1H NMR (40Yushmanov V.E. Imasato H. Tominaga T.T. Tabak M. J. Inorg. Biochem. 1996; 61: 233-250Crossref PubMed Scopus (57) Google Scholar) but two pK as at 10.9 and 12.3 (41Chen F. Cheng S. Yu C. Liu M. Su Y.O. J. Electroanal. Chem. 1999; 474: 52-59Crossref Scopus (39) Google Scholar) or at 8.0 and 10.6 (42Harriman A. Porter G. J. Chem. Soc. Faraday Trans. II. 1979; 75: 1532Crossref Google Scholar) by spectrophotometric titration. In what follows, we examine the reduction of ONOO– by Mn(III) porphyrins and the reaction of both Mn(II) and Mn(III) porphyrins with CO3·¯ . We also use spectrophotometric titration to determine the relevant pK a values. Chemicals—Mn(III) porphyrins used in this work are listed in Table I along with their electric charge at pH 7. MnIIITCPP and MnIIITSPP were purchased from MidCentury Chemicals, Chicago, IL, and the other porphyrins were synthesized as described previously (39Batinic-Haberle I. Spasojevic I. Stevens R.D. Hambright P. Fridovich I. J. Chem. Soc. Dalton Trans. 2002; : 2689-2696Crossref Scopus (124) Google Scholar). Peroxynitrite was synthesized from hydrogen peroxide and sodium nitrite in acidic solution (43Radi R. Beckman J.S. Bush K.M. Freeman B.A. J. Biol. Chem. 1991; 266: 4244-4250Abstract Full Text PDF PubMed Google Scholar). All other chemicals were commercial. Mn(II) porphyrins were prepared by the reduction of Mn(III) porphyrins with equimolar dithionite in N2O-saturated solutions containing 0.1 m carbonate at pH 10.4. Dithionite solution was prepared in helium-saturated solutions containing 0.1 m carbonate at pH 10.4, and its concentration was assessed immediately prior to its use by reduction of Fe(CN)63– (ϵ418 = 1012 m–1cm–1) (44Scaife C.W.J. Wilkins R.G. Inorg. Chem. 1980; 19: 3244-3247Crossref Scopus (45) Google Scholar).Table IStructure, abbreviation, and electric charge at pH 7 of the Mn(III) complexes used in this work Methods—Stopped-flow kinetic measurements were carried out using an SX-17MV Stopped-Flow from Applied Photophysics coupled with a 1-cm-long mixing cell. Briefly, Mn(III) porphyrins (0.8 to 6 μm) in acid phosphate solution were mixed in a 1:1 ratio with ONOO– in 10-fold or greater excess, dissolved in a known concentration of NaOH. The reaction was monitored by the change in absorbance in the Soret band of the porphyrin, and the plots were fitted to a simple exponential function. All experiments were carried out at 37 °C. The pH was measured at the outlet of the stopped flow. γ-Radiolysis experiments were carried out with a 137Cs source (Radiation Machinery Corporation Parsippany, NJ). The dose rate (9.8 gray/min) was determined using the Fricke dosimeter (1 mm FeSO4 in 0.8 n H2SO4) based on G(FeIII) = 15.6 and ϵ302(FeIII) = 2200 m–1cm–1. Pulse radiolysis experiments were carried out with a 5-MeV Varian 7715 linear accelerator (0.2–1.5 μs electron pulses, 200 mA current). All measurements were made at room temperature in a 1-cm spectrosil cell using three light passes (optical path length 3.1 cm). A 150-watt xenon-mercury lamp was used as the light source. The detection system included a Bausch & Lomb grating monochromator model D330/D331 Mk.11 and a Hamamatsu R920 photomultiplier. The signal was transferred through a Sony/Tektronix 390AD programmable digitizer to a micro PDP-I 1/24 computer, which operated the whole pulse radiolysis system. Generation of CO3·¯ —The radical was generated upon irradiation of N2O-saturated (∼25 mm) aqueous solutions containing 0.5 m carbonate at pH ≥ 8.5 via the reactions shown below in Equations 2, 3, 4, 5 (the species radiation yields are given in parentheses in Equation 2). H2O→eaq-(2.6),·OH(2.7),H·(0.6),H3O+(2.6),H2O2(0.72)(Eq. 2) eaq-+N2O→N2+OH-+·OHk3=9.1×109M-1S-1(45)(Eq. 3) ·OH+CO32-→CO3·¯+OH-k4a=3.9×108M-1S-1(45)(Eq. 4) ·OH+HCO3-→CO3·¯+H2Ok4b=8.5×106M-1S-1(45)(Eq. 5) The pulse intensity was set to produce between 2 and 4 μmCO3·¯ , and manganese porphyrin concentration ranged between 15 and 60 μm. Ionization of Axial Water of Mn(III) Porphyrins—Solutions containing 10 μm Mn(III) porphyrin and 1 m K2CO3 at pH ≥ 13 were mixed in a 1:1 ratio with HCl solutions of varying concentrations to yield pH 7.5–13.2. The mixture was made directly in the spectrophotometer cell using a RX2000 rapid mixing accessory from Applied Photophysics. Spectra were recorded from 350 to 600 nm at each pH in a Cary 50 spectrophotometer. Spectral data were analyzed using Microcal Origin software. MnIIITSPP was dissolved in 0.1 m K2CO3, because the complex precipitates in more concentrated solutions. Reaction of Mn(III) Porphyrins with Peroxynitrite—The reaction of Mn(III) porphyrins with excess of peroxynitrite was studied by stopped flow. The observed pseudo-first order rate constants were plotted versus peroxynitrite concentration at each pH to obtain the second order constant (k ox), which increases upon increasing the pH. Fig. 1 shows plots of k ox versus pH for four representative complexes. Given that the pK a of ONOOH is 6.6 ± 0.1 (46Kissner R. Nauser T. Bugnon P. Lye P.G. Koppenol W.H. Chem. Res. Toxicol. 1997; 10: 1285-1292Crossref PubMed Scopus (567) Google Scholar, 47Merényi G. Lind J. Goldstein S. Czapski G. J. Phys. Chem. 1999; 103: 5685-5691Crossref Scopus (96) Google Scholar), our results demonstrate, as reported previously for several porphyrin complexes (13Ferrer-Sueta G. Batinic-Haberle I. Spasojevic I. Fridovich I. Radi R. Chem. Res. Toxicol. 1999; 12: 442-449Crossref PubMed Scopus (157) Google Scholar), that Mn(III) porphyrins react faster with ONOO– than with ONOOH. 2The electric charge of the complexes in the equations only considers the formal charge of the metal ion and the atoms directly bound to it, thus Mn(III) porphyrin is MnIIIP+, Mn(II) porphyrin is MnIIP, and O = Mn(IV) porphyrin is O = MnIVP. MmIIIP++ONOO-→O=MnIVP+·NO2(Eq. 6) MmIIIP++ONOOH→O=MnIVP+·NO2+H+(Eq. 7) ONOOH⇆ONOO-+H+pKa(Eq. 8) Hence, the effective second-order rate constant of the reaction of Mn(III) porphyrin with peroxynitrite (k ox) is pH-dependent and is given by Equation 9, where k 6 and k 7 are the rate constants of the reactions shown in Equations 6 and 7, respectively. kox=(k6Ka+k7[H+])/(Ka+H+)(Eq. 9) Table II summarizes the values of k 6 and pK a obtained for all ten complexes. The pH profiles of all cationic complexes fit Equation 9 assuming that k 7 was very small or zero. In the case of MnIIITSPP, a value of k 7 < 105m–1s–1 can be put forward whereas MnIIITCPP did not display any significant variation in k ox in the pH range from 6.4 to 7.5. It is important to remember that a value of 105m–1s–1 is at the low end of the rate constants measurable by the method used herein. The pK a values of ONOOH obtained in the case of all cationic complexes fall below the literature value of 6.6 although it is higher for the anionic MnIIITSPP (Table II). This variation can be explained if the derived pK a values belong to the outer-sphere complex between ONOO– and Mn(III) porphyrin, and the value deviates from 6.6 because of the relative stabilization of ONOO– by the electric charge on the Mn(III) porphyrin.Table IISecond order rate constants for the reaction shown in Equation 6 and apparent pKa values for peroxynitrite obtained from fitting the data of kox to Equation 9, at 37 ± 0.1 °CComplexk6pKaM-1S-1MnIII TM-2-PyP1.9 ± 0.1 × 107aFrom Ref 13.6.2 ± 0.1MnIII TE-2-PyP3.4 ± 0.1 × 1076.4 ± 0.1MnIII TnPr-2-PyP1.4 ± 0.01 × 1076.1 ± 0.1MnIII TnBu-2-PyP1.3 ± 0.03 × 1076.2 ± 0.1MnIII TnHex-2-PyP1.3 ± 0.04 × 1076.3 ± 0.1MnIII TnOct-2-PyP1.4 ± 0.05 × 1076.1 ± 0.1MnIII TM-3-PyP4.2 ± 0.1 × 106aFrom Ref 13.6.1 ± 0.1MnIII TM-4-PyP4.3 ± 0.2 × 106aFrom Ref 13.6.3 ± 0.1MnIII TSPP3.4 ± 0.6 × 1057.4 ± 0.3MnIII TCPP1.0 ± 0.1 × 105NDa From Ref 13Ferrer-Sueta G. Batinic-Haberle I. Spasojevic I. Fridovich I. Radi R. Chem. Res. Toxicol. 1999; 12: 442-449Crossref PubMed Scopus (157) Google Scholar. Open table in a new tab Ionization of Axial Water Molecules— Fig. 2A displays the spectral changes experienced by MnIIITM-2-PyP upon the change in pH from 8.8 to 12.8. No isosbestic point was detectable in the region from 300 to 600 nm, implying that more than one equilibrium is involved. Panel B shows the spectrophotometric titration curve at 454 nm, which is the λmax for this complex at acidic and neutral pH. The plot displays a minimum and two inflection points, which is consistent with two ionization steps (pK a1 and pK a2) shown below in Equations 10 and 11, respectively. H2O-MnIIIP+⇆HO-MnIIIP+H+(Eq. 10) HO-MnIIIP⇆O=MnIIIP-+H+(Eq. 11) The spectral data were fitted to Equation 12, shown below, where H2A+ represents H2O-MnIIIP+, HA represents OH-MnIIIP, and A– represents O = MnIIIP–. Abs=AbsH2A+[H+]2+AbsHAKa1[H+]+AbsA-Ka1Ka2[H+]2+Ka1[H+]+Ka1Ka2(Eq. 12) Spectral data at ten significant wavelengths were fitted simultaneously to four variable parameters: two wavelength-dependent, namely AbsHA, AbsA–, and two wavelength-independent, namely K a1 and K a2. The parameter AbsH2A+ was obtained from the mean experimental value at the lowest pH. MnIIITCPP and MnIIITSPP showed a simpler behavior within the pH range studied, which was consistent with only one ionization equilibrium; nevertheless, reported data suggest another pK a below 14 (48Carnieri N. Harriman A. Porter G. J. Chem. Soc. Dalton Trans. 1982; : 931Crossref Scopus (67) Google Scholar). The results are summarized in Table III.Table IIIAxial water pKa1 and pKa2 obtained by fitting the spectrophotometric titration data to Equation 12ComplexpKa1pKa2MnIII TM-2-PyP10.511.4MnIII TE-2-PyP10.811.1MnIII TnPr-2-PyP11.211.2MnIII TnBu-2-PyP11.011.2MnIII TnHex-2-PyP11.011.1MnIIITnOct-2-PyP10.711.0MnIIITM-3-PyP11.513.2aThese values fall beyond the pH range studied.MnIIITM-4-PyP11.613.4aThese values fall beyond the pH range studied.MnIIITSPPbObtained in 0.05 M carbonate buffer.12.3>13aThese values fall beyond the pH range studied.MnIIITCPPbObtained in 0.05 M carbonate buffer.12.6>13aThese values fall beyond the pH range studied.a These values fall beyond the pH range studied.b Obtained in 0.05 M carbonate buffer. Open table in a new tab Spectral Changes upon Oxidation and Reduction of Mn(III) Porphyrins—Oxidation and reduction of Mn(III) porphyrin were carried out by γ-radiolysis to assess the spectral changes at 500–650 nm. O = Mn(IV) porphyrin was produced via the reaction shown in Equation 13 in N2O-saturated solutions containing 0.5 m carbonate at pH 10.5. MnIIIP++CO3·¯+H2O→O=MnIVP+CO32-+2H+(Eq. 13) Mn(II) porphyrin was generated through the reaction shown in Equation 14, shown below, in N2O-saturated solutions containing 0.1 m 2-propanol and 50 mm phosphate at pH 7 as described previously (49Morehouse K.M. Neta P. J. Phys. Chem. 1984; 88: 1575-1579Crossref Scopus (26) Google Scholar). MnIIIP++[(CH3)2COH]·→MnIIP+(CH3)2CO+H+(Eq. 14) Difference spectra were calculated as oxidized minus reduced complex. The difference spectra are exemplified in Fig. 3 for MnIIITM-2-PyP. In all cases the changes in absorbance associated with the Mn(III) to Mn(IV) transition is about four times larger than that for the Mn(II) to Mn(III) transition. Kinetics of the Oxidation of Mn(III) Porphyrins by CO3·¯ —The reaction of 2–4 μmCO3·¯ with 15–60 μm Mn(III) porphyrin was studied at pH 8.5–13 by pulse radiolysis. The reaction could not be studied at pH < 8.5, because the rate of carbonate oxidation by ·OH (Equations 4 and 5) decrease substantially with the decrease in the pH, i.e. pKa(HCO3–/CO32–) = 10.2. The reaction was followed at 570–575 nm for MnIIIT-(alkyl)-2(3,4)-PyP5+ and at 595 nm for MnIIITCPP3– and MnIIITSPP3–. The changes in the absorbance obeyed first-order kinetics, and k obs was linearly dependent on the porphyrin concentration (Fig. 4) and increased upon increasing the pH as shown in Fig. 5 for four representative complexes. The pH profiles of k 13 were fitted to Equation 15, below, and the results are presented in Table IV. k13=(kbasicKaapp+kacid[H+])/(Kaapp+H+)(Eq. 15) Fig. 5The effect of pH on k 13 for four representative complexes: MnIIITM-4-PyP (squares), MnIIITM-2-PyP (circles), MnIIITSPP (triangles), and MnIIITCPP (inverted triangles). The increment of k 13 with pH correlates with deprotonation of water molecules axially coordinated to the manganese. Solid lines represent the best fits to Equation 15. All experiments were carried out in N2O-saturated solutions containing 0.5 m carbonate buffer.View Large Image Figure ViewerDownload Hi-res image Download (PPT)Table IVRate constants for the oxidation of manganese porphyins by CO3·¯ at room temperatureComplexMnIIIP++CO3·¯MnIIP++CO3·¯(k16)kacidkbasicpK aapp108 M-1S-1109 M-1S-1MnTM-2-PyP2.7 ± 0.154 ± 410.82.1 ± 0.04MnTE-2-PyP3.1 ± 0.239 ± 311.02.4 ± 0.8MnTnPr-2-PyP3.7 ± 0.241 ± 911.12.0 ± 0.4MnTnBu-2-PyP3.6 ± 0.253 ± 911.13.4 ± 0.7MnTnHex-2-PyP3.3 ± 0.335 ± 110.72.4 ± 1.0MnTnOct-2-PyP3.2 ± 0.232 ± 110.53.7 ± 0.5MnTM-3-PyP3.0 ± 0.243 ± 111.55.4 ± 0.8MnTM-4-PyP3.0 ± 0.452 ± 111.94.0 ± 0.6MnTCPP12.0 ± 0.328aValues generated by extrapolation using pK a1 from Table III.12.6bThe values of k acid, k basic; and pK aapp were obtained by fitting the observed rate constants of the reaction in Equation 13, Equation 14, Equation 15. Values of k 16 were obtained at pH 10.4.3.5 ± 0.4MnTSPP5.3 ± 0.112aValues generated by extrapolation using pK a1 from Table III.12.3bThe values of k acid, k basic; and pK aapp were obtained by fitting the observed rate constants of the reaction in Equation 13, Equation 14, Equation 15. Values of k 16 were obtained at pH 10.4.1.3 ± 0.5a Values generated by extrapolation using pK a1 from Table III.b The values of k acid, k basic; and pK aapp were obtained by fitting the observed rate constants of the reaction in Equation 13, Equation 14, Equation 15. Values of k 16 were obtained at pH 10.4. Open table in a new tab Oxidation of Mn(II) Porphyrins by CO3·¯ —The reaction of 2–4 μmCO3·¯ with 16–45 μm Mn(II) porphyrin, shown below in Equation 16, was studied at pH 10.4 by pulse radiolys
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