6621
J. Am. Chem. SOC.1989, 111, 6621-6627
Homolytic and Heterolytic Scission of Organic Hydroperoxides by (meso-Tetraphenylporphinato)iron(111) and Its Relation to Olefin Epoxidation Regine Labeque and Lawrence J. Marnett* Contribution from the Department of Chemistry, Wayne State University, Detroit. Michigan 48202. Received October 7, 1988
Abstract Reaction of 1O-hydroperoxyoctadeca-8,12-dienoicacid (10-00H-18:2) with chloro(meso-tetraphenylprphinato)iron(III) (Fe3+-TPP) in CH2Cl2produces two products identified as 10-oxodec-8-enoicacid (lo-oxo-10:1) and lO-oxmtadeca-8,12-dienoic acid (10-oxo-18:2). These compounds are derived from an alkoxyl radical intermediate generated by homolytic cleavage of 10-00H-18:2. Only a trace of the heterolytic cleavage product 10-hydroxycctadeca-8,12-dienoicacid (10-OH-18:2) is observed. Inclusion of imidazole alters the product profile so that 10-OH-18:2 is the major product (62%). Reaction of 10-00H-18:2 or t-BuOOH (0.05 M) with Fe3+-TPP (0.001 M) and cis-stilbene ( 5 M) produces trans-stilbene oxide as the only oxidation product (55% based on hydroperoxide). Oxidation in the presence of imidazole (0.3 M) produces approximately equal amounts of cis-stilbeneoxide (22%) and trans-stilbene oxide (23%). Butylated hydroxytoluene inhibits formation of both epoxides from cis-stilbene whereas p-methoxyanisoleselectively inhibits formation of cis-stilbeneoxide. The data indicate that two separate oxidizing agents are produced in the reaction of hydroperoxides with Fe3+-TPP. In the absence of imidazole, Fe3'-TPP cleaves 1O-OOH- 18:2 and t-BuOOH homolytically to alkoxyl radicals and Fe4+=0.TPP. Both species oxidize the hydroperoxide to peroxyl radicals that epoxidize cis-stilbene nonstereospecifically. In the presence of imidazole, Fe3+-TPP also cleaves the hydroperoxides heterolytically to alcohol and Fe4+=0.TPP", which epoxidizes cis-stilbene stereospecifically. The results reconcile apparently conflicting observations in the literature and suggest that the pathway of hydroperoxide cleavage by heme complexes and heme proteins is a sensitive function of the environment of the heme.
contrast to complexes such as 1, complexes analogous to 2 do not appear to catalyze stereospecific olefin epoxidation.6 Recently, Traylor and Xu reported that hydroperoxides are oxidized to peroxyl radicals during reactions with heme complexes and suggested hydroperoxide scavenging of 1 competes with its reaction with olefins (eq 3).' They offered a unifying hypothesis that
Studies of the reactions of metalloporphyrins with iodosylbenzene (PhIO), peracids, and hydroperoxides have provided important insights into the nature of the oxidizing agents produced from hemeproteins such as peroxidases, catalase, and cytochrome P-450. The consensus of these studies is that PhIO and peracids transfer an oxygen atom with two oxidizing equivalents (the result of heterolytic cleavage) to form a high valent iron-oxo complex (eq 1) capable of epoxidizing o1efins.l In the case of ferric I I
Fe3+ + PhIO (RC03H)
-
*'Ae4+=0 + Phi (RCO,H)
I
1+ ROOH42
I I
-
L+=o + RO* + I
H+
(2)
2
porphyrins, the iron-oxo complex (1) contains a ferry1 iron and a radical cation of the porphyrin.l8v2 These complexes are electronically analogous to the initial oxidizing agents (compounds I) produced during the catalytic turnover of peroxidase^.^ Although PhIO and peracids are useful mechanistic tools, they are of limited biological relevance because they do not occur in plant or animal metabolism. In contrast, hydrogen peroxide and fatty acid hydroperoxides are widely distributed in the plant and animal kingdom, and they support a variety of oxidations by hemeproteins. Studies of their reactions with heme complexes and hemeproteins can provide direct information about the nature of the oxidants in many biological oxidations and highlight differences in mechanism induced by the protein component of hemeproteins. Despite intensive investigationof heme-hydroperoxide reactions, a clear mechanistic consensus has not evolved. Several studies of hydroperoxide-dependent oxidations by model ferric porphyrins suggest the heme group reacts with hydroperoxides by a different mechanism than it reacts with PhIO and pera ~ i d s . l ~ v ~Hemes 3~ appear to cleave the hydroperoxide bond homolytically to produce an alkoxyl radical and a ferryl-oxo complex (2) that contains only one of the oxidizing equivalents of the hydroperoxide (eq 2). The ferryl-oxo complex (2) is electronically analogous to compound I1 of p e r o x i d a ~ e s . ~ ~InJ ~ - ~ *Address correspondence to L.J.M. at Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, TN 37232.
0002-7863/89/1511-6621$01.50/0
Hf
(3)
heme-hydroperoxide reactions proceed by heterolytic scission analogous to their peracid counterparts but that secondary reaction with hydroperoxide consumes 1. One can estimate the extent of heterolytic hydroperoxide cleavage using stereospecific olefin epoxidation to detect 1.28 The absence of molecules that react with ferryl-oxo complexes 2 in a diagnostic fashion limits one's ability to quantitate homolytic
(1)
1
Fe% + R m H
+ ROO'+
(1) (a) Groves, J. T.; Nemo, T. E.; Myers, R. S. J . Am. Chem. SOC.1979, 101, 1032-1033. (b) Groves, J. T.; Nemo, T. E. J . Am. Chem. SOC.1983, 105, 5786-5791. (c) Lee, W. A.; Bruice, T. C. J . Am. Chem. SOC.1985, 107, 513-514. (d) Lee, W. A.; Yuan, L.-C.; Bruice, T. C. J . Am. Chem. SOC.1988, 110, 4277-4283. (e) Collman, J. P.; Kodadek, T.; Brauman, J. I . J . Am. Chem. SOC.1986, 108, 2588-2594. (2) (a) Groves, J. T.; Watanabe, Y. J . Am. Chem. SOC.1986, 108, 507-508. (b) Groves, J. T.; Hauslater, R. C.; Nakamura, M.; Nemo, T. E.; Evans, B. J. J . Am. Chem. SOC.1981, 103, 2884-2886. (c) Balch, A. L.; Latos-Grazynski, L.; Renner, M. W. J . A m . Chem. SOC. 1985, 107, 2983-3002. (3) (a) Dunford, H. B.; Stillman, J. S. Coord. Chem. Reu. 1976, 19, 187-251. (b) Roberts, J. E.; Hoffman, 9. M.; Rutter, R.; Hager, L. P. J. Biol. Chem. 1981, 256, 21 18-2121. (c) Dolphin, D.; Forman, A,; Borg, D. C.; Fajer, J.; Felton, R. H. Proc. Natl. Acad. Sci. U.S.A.1971,68, 614-618. (d) La Mar, G. N.; deRopp, J. S. J . A m . Chem. SOC.1980, 102, 395-397. (e) Cytochrome P-450: Structure, Mechanism and Biochemistry; Ortiz de Montellano, P. R., Ed.; Plenum: New York, 1986. (f) Guengerich, F. E.; Macdonald, T. C. Acc. Chem. Res. 1984, 17, 9-16. (4) (a) Mansuy, D.; Bartoli, J.-F.; Momenteau, M. Tetrahedron Lett. 1983, 23, 2781-2784. (b) Lindsay-Smith, J. R.; Mortimer, D. N. J . Chem. SOC.,Perkin Tram. 2 1986, 1743-1749. (c) Koola, J. D.; Kochi, J. K. J . Org. Chem. 1987,52,4545-4553. (d) Mansuy, D. Pure. Appl. Chem. 1987.59, 759-770. ( 5 ) Balch, A. L.; Chan, Y.-W.; Cheng, R.-J.; La Mar, G. N.; Latos-Grazynski, L.; Renner, M. W. J . Am. Chem. Soc. 1984, 106, 7779-7785. (6) (a) Mansuy, D.; Battioni, P.; Renaud, J. P. J . Chem. Soc., Chem. Commun. 1984, 1255-1257. (b) Chin, D.; La Mar, G. N.; Balch, A. L. J . A m . Chem. Soc. 1980, 102, 5945-5947. (7) Traylor, T. G.; Xu, F. J . Am. Chem. SOC.1987, 109, 6201-6202.
0 1989 American Chemical Society
6622 J . A m . Chem. SOC.,Vol. 111, No. 17, 1989
Labeque and Marnett
Scheme I
CSHI,,
10-OH-18:2
hydroperoxide scission. This highlights t h e need for novel approaches t h a t provide a complete picture of t h e chemistry of hydroperoxide cleavage by metal complexes and metalloproteins. We recently described a fatty acid hydroperoxide, 10-hydroperoxyoctadeca-8,12-dienoicacid ( 1 0 - 0 0 H - 1 8 : 2 ) , t h a t is capable of simultaneously differentiating homolytic and heterolytic scission of t h e hydroperoxide bond (Scheme I).8a Heterolytic scission produces lO-hydroxyoctadeca-8,12-dienoicacid (1 0-OH- 18:2) whereas homolytic scission produces a n alkoxyl radical t h a t undergoes @-scissiont o 10-oxodec-8-enoic acid (10-oxo-10: 1) or oxidation t o 10-oxooctadeca-8,12-dienoicacid (1O-oxo-18:2). 1 0 - 0 0 H - 1 8 : 2 is readily synthesized from linoleic acid, can be produced with radioactive label for sensitive product detection, a n d exhibits solubility properties t h a t enable it to be used in aqueous or organic solvents. We have used 1 0 - 0 0 H - 1 8 : 2 to demonstrate t h a t ferric bleomycin catalyzes homolytic hydroperoxide cleavage and prostaglandin H synthase catalyzes heterolytic hydroperoxide ~ l e a v a g e . ~ The importance of defining t h e mechanism of hydroperoxide reduction by model heme complexes prompted us to investigate t h e reaction of 1O-OOH-18:2 with chloro(mes0-tetraphenylporphinato)iron(III) (Fe3+-TPP) in CH2C12. We have also conducted experiments to relate t h e pathway of hydroperoxide cleavage t o t h e oxidants produced t h a t epoxidize olefins. Our results provide strong evidence for homolytic scission of the peroxide bond and explain discrepancies between previous studies that employed olefin epoxidation to infer the identity of the oxidant produced in heme-hydroperoxide reactions.
Experimental Procedures Materials. cis-Stilbene, trans-stilbene, Fe3+-TPP, tert-butyl hydroperoxide (t-BuOOH, 90%), and iodosobenzene diacetate were purchased from Aldrich (Milwaukee, WI) and butylated hydroxytoluene was from Sigma (St. Louis, MO). Phenol (Fisher) was distilled prior to use. Silica gel was from Merck (Rahway, NJ), and 3% OV-17 on 100/120 Chromosorb Q was obtained from Anspec (Ann Arbor, MI). [1-'4C]1000H-18:2 was synthesized as previously described.8bPhIO was prepared according to the method described by Lucas and Kennedy.Io p-Methoxythioanisole was a gift from Patrick PI&, Wayne State University. Solvents used for chromatography were HPLC grade. Other chemicals were reagent grade. (8) (a) Labeque, R.; Marnett, L. J. J . Am. Chem. SOC.1987, 109, 2828-2829. (b) Labeque, R.; Marnett, L. J. Biochemisfry 1988, 27, 7060-7070. (9) (a) Padbury, G.;Sligar, S. C.; Labeque, R.; Marnett, L. J. Biochemisfry 1988, 27, 7846-7852. (b) Marnett, L. J.; Chen, Y.-N. P.; Maddipati, K. R.; PIE, P.; Labeque, R. J . B i d . Chem. 1988, 263, 16532-16535. ( I O ) Lucas, H. J.; Kennedy, E. R. Org. Synrh. 1955, 3, 482-483.
Purification of cis-Stilbene. Commercially available cis-stilbene was contaminated with 2-3% trans-stilbene. Therefore cis-stilbene was purified by medium-pressure liquid chromatography on a silica gel column eluted with hexane. The fractions eluting from the column were assayed by gas chromatography (GC) using a program that was isothermal at 80 'C for 3 min followed by a linear gradient to 240 OC at 8 'C/min. The retention times of cis- and trans-stilbene were 11.5 and 14.1 min, respectively. Fractions of cis-stilbene containing less than 0.2% transstilbene were combined and concentrated. The purified cis-stilbene was 99.8% pure. Instrumentation. HPLC was performed with an LDC pump connected to an Alltech Partisil 10 column (10-pm silica, 4.6 X 250 mm). Radioactivity eluting from the column was quantitated in a continuous fashion with a Radiomatic Flo-One H P radioactivity flow detector using Scinti-Verse LC, premixed scintillant, from Fisher. G C analysis was performed on a Varian Model 3700 with flame ionization detection using a 2-ft column packed with 3% OV-17 on Chromosorb Q. Product quantitation was performed by integrating the GC signal with a Hewlett Packard H P 3380A integrator. Reaction of 10-00H-18:2 with Fe3+-TPP. Fe3*-TPP (100 pM) was dissolved in CH2C12. Reactions were initiated by addition of 5 mM [ 1-I4C]10-00H-18:2 (final concentration) and allowed to continue at room temperature for 30 min with stirring. An aliquot of the reaction mixture was analyzed by normal-phase HPLC (hexane/tetrahydrofuran/acetic acid:895/100/5). Imidazole, or phenol, when present, was added to the solution of Fe3+-TPP 2 min prior to the addition of 1000H-18:2. cis-StilbeneOxidation. cis-Stilbene (3-5 M) and Fe'+-TPP (1 mM) were dissolved in CH2C12,and hydroperoxide (t-BuOOH or 1 0 - 0 0 H 18:2) was added to initiate the reaction. The PhIO oxidation of cisstilbene was performed as follows: cis-stilbene (250 mM) and Fe3+-TPP (7 mM) were dissolved in CH2CI2,and PhIO was slowly added. The reaction mixtures were assayed by GC using a linear gradient from 80 to 240 O C at 3 "C/min. The retention times of cis-stilbene, cis-stilbene oxide, trans-stilbene, and trans-stilbene oxide were 12.7, 16.5, 19.3, and 20.5 min, respectively. Results Products of the Reaction of 10-00H-18:2 with Fe3+-TPP. W h e n 1 0 - 0 0 H - 1 8 : 2 ( 5 m M ) was reacted with Fe3+-TPP (100
wM) in CH2C12,two products resulted. These compounds were identified as 1O-oxo-18:2 and 10-oxo-10:1. They exhibited identical chromatographic a n d mass spectral properties as reported previously.8b lO-Oxo-18:2 a n d 10-oxo- 10: 1 a r e derived from a n alkoxyl radical formed by homolytic scission of 1O-OOH-18:2 by Fe3+-TPP. Only trace amounts of the heterolytic scission product, 10-OH-18:2, were detected. T h e time course of reaction of 100 0 H - 1 8 : 2 with Fe3+-TPP showed complete consumption of the hydroperoxide after 20 min ( d a t a not shown). T h e ratio of t h e yields of 1o-oxo-18:2 to 10-oxo-10:1 was constant with time. NO
J . Am. Chem. Soc., Vol. 111, No. 1 7 , 1989 6623
Scission of Organic Hydroperoxides
I00xo-1~1
Table I. Reaction of l0-Hydroperoxy-8,12-Octadecadienoic Acid
with Fe’+-TPP 10-OXO-1 8 ~ 2 , 10-0x0- IO: 1, 10-OH-18:2, 7% % %
* *
