Chem. Res. Toxicol. 1993,6, 351-355
35 1
Reactions of the Vitamin E Model Compound 2,2,5,7,8-Pentamethylchroman-6-01with Peroxyl Radicals Daniel C. Liebler,*tt Jeanne A. Burr,? Shigenobu Matsumoto,l and Mitsuyoshi Matsuo’ Department of Pharmacology and Toxicology, College of Pharmacy, University of Arizona, Tucson, Arizona 85721, and Tokyo Metropolitan Institute of Gerontology, 35-2 Sakaecho, Itabashi-ku, Tokyo 173, Japan Received January 4, 1993
The vitamin E model compound 2,2,5,7,8-pentamethylchroman-6-01 (lb) was oxidized by peroxyl radicals generated by thermolysis of 2,2’-azobis(2,4-dimethylvaleronitrile) in oxygenated solvents. Oxidation of l b yielded 4a,5-epoxy-4a,5-dihydro-8a-hydroperoxy-2,2,5,7,8-pentamethylchroman-6(8aH)-one(5b), 7,8-epoxy-7,8-dihydro-8a-hydroperoxy-2,2,5,7,8-pentamethylchroman-6(8aH)-one (6b), 8a-[(2,4-dimethyl-l-nitrilopent-2-yl)dioxyl-2,2,5,7,8-pentamethylchroman-6(8aH)-one (3b),and a 5,5’-spirodimer product (7b). In otherwise identicalreactions, yields of chromanone 3b and epoxides 5b/6b increased with increasing solvent polarity, whereas the yield of spirodimer 7b decreased. Deuterium substitution at Csainhibited oxidation of l b to spirodimer 7b and favored the formation of a novel 5,7’-spirodimer 11. The results demonstrate that the reaction medium controls the balance between competing reactions of chromanoxyl radical 2b and are consistent with the formation of epoxides and 8a-substituted tocopherones as the predominant products of a-tocopherol oxidation in biological membranes.
Introduction The biologicalantioxidant vitamin E [principally (RRR)a-tocopherol; la, Chart I] is a principal scavenger of peroxyl radicals in biological membranes (1). Antioxidant reactions of la appear to play a critical role in preventing oxidative injury by toxic and carcinogenic chemicals (2). Like other phenols, la traps radicals by hydrogen atom transfer from the chromanol ring system to form a chromanoxyl radical 2a (3). Although chromanoxyl radicals such as 2a may be reduced back to the corresponding chromanols by other reductants ( 4 - 9 , competing reactions may convert the chromanoxyl radicals to other products. In the first, peroxyl radicals add to chromanoxyl radical 2a para to the phenoxy oxygen to form 8a-(alkyldioxy)chromanones such as 3a (8-10) in a reaction analogous to that observed for other antioxidant phenoxy1radicals (11). In another reaction, oxygen addition to the 8a-position of 2a followed by hydrogen abstraction forms 8a-hydroperoxychromanone 4a (10, 12). In a third reaction, peroxyl radicals oxidize 2a to epoxychromanones 5a/6a (10, 12). Depending on reaction conditions, epoxides are thought to be formed either by further oxidation of hydroperoxide 4a (12) or by an initial epoxidation of 2a, followed by oxygen addition and hydrogen abstraction (10). However, the mechanism of epoxide formation has not been unambiguously established in any system. The fourth pathway yields 5,5’-spirodimer 7a and trimer products (13),which may be formed via an intermediate quinone methide 8a (14, 15). A notable feature of chromanol oxidations is that the distribution of products varies dramatically with reaction conditions. For example, peroxyl radicals derived from thermolysis of 2,2’-azobis(2,4-dimethylvaleronitrile) (AMUniversity of Arizona.
* Tokyo Metropolitan Institute of Gerontology. +
0893-228x/93/2706-0351$04.00/0
Chart I
1
5
2
6
9
W
O
3
I 7
4
8
11
VNI1 in benzene oxidized la to spirodimer 7a and 8a(alky1dioxy)chromanone 3a (13). In acetonitrile, 8a-(alky1dioxy)chromanone la, epoxides 5a/6a, and hydroperoxide 4a were formed, but spirodimer 7a was not (IO). Oxidation of la by tert-butylperoxyl radicals in tert-butyl hydroperoxide yielded epoxide 5a, but 8a-(alky1dioxy)chromanones and dimers were not formed ( 1 2 ) . Since these studies employed similar reaction conditions and involved structurally similar peroxyl radicals, differences in reaction solvent may have accounted for differences in product distribution. We felt that a systematic study of the relationship between reaction solvent propAbbreviations: AMVN, 2,2’-azobis(2,4-dimethylvaleronitrile); ‘HCOSY, proton homonuclear correlation spectroscopy.
0 1993 American Chemical Society
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erties and product distribution might reconcile apparently contradictory reports of vitamin E antioxidant chemistry. Here we report the effects of reaction solvent on the oxidation of the vitamin E model compound 2,2,5,7,8pentamethylchroman-6-01(1b) by AMVN-derivedperoxyl radicals.
Experimental Procedures General. Compound l b and [5a-CD3]-lbwere synthesized as described previously (16, 17). AMVN was from Wako Pure Chemical Industries, Ltd. (Tokyo, Japan). CD3CN was from E. Merck (Darmstadt, Germany). Solvents were of the highest available purity and were obtained from standard commercial sources. Isopropyl ether (Wako) was purified before use by passage through an activated alumina column to remove antioxidants added as stabilizers. HPLC analyses of l b and its oxidation products were done with a Hewlett-Packard Model 1090 liquid chromatograph (Hewlett-Packard GmbH, Waldbronn, Germany) equipped with a Model 1040 diode-array detector and controlled with a H P 300 (Series 9000) computer and H P Chem-Station software. Preparative reverse-phase HPLC was done with a Waters Radial Compression Module System (WatersAssociates, Milford, MA) equipped with a NovaPak 25 X 100 cm cartridge. 'H- and 13C-NMR spectra were obtained with a Varian VXR-400s (Varian Associates, Palo Alto, CA) instrument operating a t 399.95 MHz for 'H and 100.58 MHz for I3C. Mass spectra were obtained with a Finnegan MAT-90 instrument (Finnegan MAT, Bremen, Germany) equipped with a Micro PDP-11/73 computer (U.S. Design, Palo Alto, CA) and a Finnegan ICIS data system. Samples were introduced by direct probe insertion. Oxidation of lb. Compound l b (10 mg, 45.5 pmol) and AMVN (40 mg, 161 pmol) were heated in the dark a t 60 "C in 1mL of solvent (see Table I) in a glass screw-cap tube. Samples (10 pL) were removed with a glass syringe and added to 1 mL of CH3CNin an autosampler vial. Compound l b and its oxidation products were separated by reverse-phase HPLC on a Hypersil ODS, 5-pm, 4.6 X 100 mm analytical column (Hewlett Packard) eluted with 80% CH30H/20% H20 a t a flow rate of 1mL min-'. After a 5 min of isocratic elution, nonpolar products were eluted with alinear gradient to 90% CH30H/10% HzOin 5 min, followed by isocratic elution at that composition for 10 min. Compound l b was detected a t by UVabsorbanceat 292 nm; allother products were detected at 255 nm. Calibration curves for each of the products were constructed with authentic product standards. 8a-(Alkyldioxy)chromanones3b. Reverse-phase HPLC of l b oxidation products yielded two peaks corresponding to 8a(alky1dioxy)chromanones 3b. The two peaks were isolated by preparative reverse-phase HPLC with a mobile phase of 85 % CH30H/15% H 2 0 at a flow rate of 7 mL min-l. Peak I: UV (HPLC mobile phase) X, 242 nm; MS m/z 361 (M+) (1.41, 329 (11.8),235 (23.7), 219 (100); MS m/z 361.2249 (calcd for C21H31NO4 361.2253); 1H-NMR (CDC13)6 0.81 (d, 3 H), 0.89 (d, 3 H), 0.98 (d, 3 H), 1.02 (d, 3 H), 1.15 (s, 3 H), 1.39 (s, 3 H), 1.54 (s, 3 H), 1.58 (m, 2 H), 1.67 (s, 3 H), 1.70 (m, 1 H), 1.75 (m, 1 H), 1.82 (m, 1 H), 1.84 (s, 3 H), 1.85 (m, 1 H), 1.85 (s, 3 H), 1.99 (s, 3 H), 2.15 (m, 1 H), 2.60 (m, 1 H) ppm. Peak 11: UV (HPLC mobile phase) A, 242 nm; MS m/z 361 (M+)(5.7), 329 ( 7 3 , 2 3 5 (29.4),219 (100);MS m/z 361.2237 (calcd for C21H31N04 361.2253); 'H-NMR (CDC13) 6 0.76 (d, 3 H), 0.849 (d, 3 H), 1.14 ( ~ , H), 3 1.40 (s, 3 H), 1.49 (m, 2 H), 1.53 (s, 3 H), 1.58 (s, 3 H), 1.66 (m, 1 H), 1.83 (s, 3 H), 1.86 (s, 3 H), 1.89 (s, 3 H), 2.22 (s, 1 H), 2.61 (m, 1 H), 2.22 (m, 1 H) ppm. Configurations a t Cas and CZ,(of the AMVN-derived alkyl chain) for products in each peak were tentatively assigned by comparison with spectra of four diastereomers of 8a-(alkyldioxy)tocopherones3a. Compound 3b diastereomers in peak l exhibited C4methylene resonances a t higher field (6 2.15 and 2.60) and were tentatively assigned the configuration 8a(S),2'(S) and 8a(R),2'(R). Compound 3b diastereomers in peak 2 exhibited C4methylene resonances a t lower
Liebler e t al. field ( 6 2.61 and 2.82) and were tentatively assigned the configuration 8a(R),2'(S) and 8a(S),2'(R). The rationale for configurational assignments based on NMR shift data has been described previously (9,lO). Two sets of resonances were detected for some of the alkyl protons of the 8a-(alkyldioxy) moiety for each product peak. 'H-NMR and 'H-homonuclear correlation (IH-COSY) spectra and a summary of 'H shift assignments are provided as supplementary material. 13C-NMR Analysis of AMVN-Dependent Oxidation of [13C]-lb. Compound l b (5 mg, 22.8 pmol) and AMVN (20 mg, 80.5 pmol) were dissolved in 0.5 mL of oxygen-saturated CD3CN in a pressure-resistant 5-mm NMR tube and heated in the NMR probe a t 60 OC. 'H-decoupled 13C-NMRspectra were acquired after 15, 30, 60, and 120 min of reaction. A standard 13Cpulse sequence was used to acquire 256 scans a t each time point. At 120 min, 10 pL of the reaction mixture was analyzed by reversephase HPLC as described above. To simulate reverse-phase HPLC analysis of the products, the remaining reaction mixture was applied to an octadecylsilyl Sep-Pak cartridge (Waters Associates, Milford, MA) prewetted with CH3CN. The column was washed with 2 mL of CH30H, and the eluents werecombined, evaporated in vacuo, and redissolved in CD3CN for 13C-NMR analysis as described above. Oxidation of [lia-CDJ-lb. Compound [5a-CD3]-lb was oxidized with AMVN in hexane as described above. Yield of spirodimer 7b was measured by HPLC (see above), except that a mobile phase of CH3OH/HzO (85:15 v/v) a t 1.5 mL min-' was used to resolve dimers 7b and 11. Reverse-phase HPLC of reaction products afforded spirodimer 11-d6: MS m/z 441.2930 (calcd for C28H31D504 441.2927); UV-vis, MS, and HPLC elution of 11-d5were identical to those of 1 1 - d ~formed by K3Fe(CN)6 oxidation of [5a-CD~]-lb(see below). Potassium Hexacyanoferrate(II1) Oxidation of l b a n d [5a-CDJ-lb. Compound l b o r [5-CD31-lb (560mg) wasdissolved in hexane (70 mL) and shaken in a separatory funnel with a solution of K3Fe(CN)e(3.26 g) in 0.2 N potassium hydroxide (32 mL) for 10 min. The hexane phase was separated and washed twice with water, dried over Na2S04,and evaporated to dryness. The yellow mixture of 5,5'-spirodimer 7b and 5,7'-spirodimer 11 was first purified by silica gel column chromatography, and the two dimers then were resolved by reverse-phase HPLC (see above). Compound 11 (from oxidation of unlabeled lb): UV (CH3CN) A,, 301 (c 4900), 345 ( t 2000), 420s (t 700) nm; IR (KBr) Xmax 1670, 16505, 1595 cm-'; MS m/z 436 (M+), 421, 380, 324, 218, 203, 175, 164; MS m/z 436.2597 (calcd for C2&& 436.2614); 'H-NMR (CDC13) 6 (CH3) 2.06, 2.00, 1.95, 1.83, 1.29, 1.26, 1.25, 1.24 ppm; 13C-NMR (CDC13) 6 202.6 (s), 145.9 (s), 145.5 (s), 144.8 (s), 143.0 (s), 127.1 (s), 120.3 (s), 119.6 (s), 117.9 (s), 117.7 (s), 115.6 (s), 81.0 (s), 73.8 (s), 72.3 (s), 33.1 (t), 32.4 (t), 28.4 (t),27.4 (q), 26.5 (q), 26.4 (q), 26.3 (q), 21.0 (t),18.9 (t),17.8 (t), 13.9 (q), 11.2 (q), 11.0 (q), 10.8 (4) ppm. Compound 11-d5 (from oxidation of [5a-CD3]-lb): MS m/z 441 (M+), 426, 385, 329, 221, 205, 177; MS m/z 441.2893 (calcd for C28H31D504 441.2927);'H-NMR (CDC13)resonance a t 2.06 ppm missing; 13CNMR (CDC13)resonances a t 28.4 (t) and 11.0 (9) ppm missing.
Results and Discussion Compound l b was incubated at 60 "C in cyclohexane, benzene, isopropyl ether, ethyl acetate, tetrahydrofuran, or acetonitrile with AMVN, which generates peroxyl radicals by thermolysis and oxygen addition: RN=NR
-
2R
+ N,
2R' + 0, 2 R 0 0 ' (2) Residual l b and its oxidation products were analyzed by reverse-phase HPLC with diode-array UV detection. Protic solvents (e.g., alcohols) were not used for these studies, as solvent incorporation into products can occur, which complicates product distribution (9). Oxidation of
Chem. Res. Toxicol., Vol. 6, No. 3, 1993 353
Oxidation of a Vitamin E Model Compound 2
50
50
I E 40
E- 40
< 30
u KJ 0
0
L
30
L
a 20
a 20 L
L
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0
-
n '0
n
7
0
0
60
120
10
180
time, min
0 0
30
60
90
120
time, min
Figure 1. Time course of consumption of l b ( 0 )and formation a t 60 "C in of oxidation products 3b (A),4b/5b (B), and 7b (0) cyclohexane (panel A) and acetonitrile (panel B). Reaction mixtures contained 45.5 mM l b and 161 mM AMVN. Table I. Effect of Reaction Solvent on Yields of Oxidation Products in AMVN-Dependent Oxidation of lb product yield, 7% of lb consume@ solvent €b 5bI6b 3b 7b cyclohexane benzene isopropyl ether ethyl acetate tetrahydrofuran acetonitrile
2.015 2.274 3.88 6.02 7.58 35.94
8.2 f 4.5 8.4 f 2.6 18.2 f 2.5 17.8f 3.8 20.6 f 3.7 20.2 f 2.5
11.1f 2.6 24.2f0.3 42.7 f 1.0 57.95 8.9 62.9 f 7.0 54.2 f 5.9
23.85 3.2 15.6 f 2.3 17.6 f 2.2 4.9f 1.2 1.4 f 0.8 ndc
Product yields were determined after 30 min of reaction at 60
"C. * Solvent dielectric constant (28). Not detected.
l b yielded 8a-(alky1dioxy)chromanones (3b), epoxytocopherone 5b, and 5,5'-spirodimer 7b in yields that varied with the reaction solvent (see below). Identities of compounds 5b and 7b were confirmed by coelution with authentic standards (12) and by diode-array scanning of their UV spectra, whereas the previously uncharacterized compound 3b was characterized by diode-array UV scanning, mass spectrometry, and NMR spectroscopy. Compound 3b eluted as two peaks, each of which contained 2 diastereomers. A peak that was incompletely resolved from epoxytocopherone 5b and had an identical UV spectrum was tentatively identified as epoxytocopherone 6b. The two epoxide peaks were quantified together for measurements of product distribution. Time course studies of l b oxidation and product formation (Figure 1) indicated that l b was oxidized at approximately the same rate in all solvents studied except cyclohexane, in which l b was oxidized more slowly. Accumulation of products 3b and 5b in all solvents was linear for up to 60 min and then leveled off as l b oxidation exceeded 80%. In contrast, levels of 5,5'-spirodimer 7b peaked at 60 min and then declined thereafter. The concomitant appearance of other product peaks near 7b suggested that 7b may have been further oxidized during the reaction. Distributions of l b oxidation products at 30 min are summarized in Table I. Since both l b depletion and product formation were linear in this time range, relative product amounts should reflect relative rates of formation. The formation of epoxides 5b/6b and chromanones 3b increased with increasing solvent dielectric constant E , whereas spirodimer 7b formation decreased. Epoxides and chromanones increased from about 8% and 11%, respectively, of total products in cyclohexane to about 20% and 52 % , respectively, of total products in acetonitrile. The distribution of l b oxidation products in acetonitrile was virtually identical to that for oxidation of la under similar conditions (IO). It is evident from the
product yields in Table I that other products of the reactions were not measured. Products of l b oxidation that were observed, but not quantified, were 8a-hydroperoxychromanone 4b, quinone 9, and dihydroxy dimer 10. Tentative identification of these other products was based on comparisons of their chromatographic characteristics and diode-array UV spectra with those of authentic standards (18). Trimers formed from lb, which would be expected in systems where spirodimer 7b is formed, would not have eluted in the HPLC system we employed. Table I also indicates that the yield of chromanone 3b increased with increasing solvent dielectric constant t (except for acetonitrile). In contrast, spirodimer 7b yield was maximal (about 23%) in cyclohexane, the least polar solvent, whereas epoxide 5b/6b yield was maximal in acetonitrile, the most polar solvent. Plots of epoxide yield and spirodimer yield versus log E for the solvents (excluding acetonitrile) were log-linear and displayed a reciprocal relationship (data not shown). The average sum of epoxide and spirodimer yields in all of the solvents (including acetonitrile) was approximately constant at 26 f 6 %. Thus, epoxides 5b/6b and spirodimer 7b appeared to account for a fixed fraction of the products. On the basis of this finding, we postulated that epoxides 5b/6b might be formed via a reaction intermediate that yields either the epoxides or 5,5'-spirodimer 7b depending on the reaction solvent. Because an unstable intermediate might not survive chromatographic analysis, we attempted to detect intermediates in the oxidation of [5a-l3CH3I-lb by 13C-NMR. A proton-decoupled scan of the reaction mixture after 15 min at 60 "C revealed the resonances of [5aJ3CH31-lb at 12 ppm, in addition to a group of resonances centered at 10.8 ppm, which are attributed to the four diastereomers of chromanone [5a-13CH31-3b(9), and another resonance at 12.7 ppm attributed to epoxide [5a-13CH3]-5b(12) (Figure 2A). Other signals at 20-30 ppm are due to AMVN and its decomposition products. The [5a-13CH31-lbresonance declined with time, whereas those for 3b, 5b, and 6b (15.3 ppm) increased (Figure 2BC). HPLC analysis at 120 min revealed a product distribution identical to that at 120 min in incubations with unlabeled l b (see above). None of the resonances was due to a labile intermediate, since passage of the reaction mixture through an octadecylsilyl silica solidphase extraction column did not affect the 13C-NMR spectrum of the reaction mixture (Figure 2D). These results indicate that intermediates in the oxidation of l b to epoxides 5b/6b were too unstable to accumulate to levels detectable by 13C-NMR. Additional studies will be required to establish the mechanism of epoxide formation. One possible route to spirodimer 7b is by hydrogen abstraction from chromanoxyl radical 2b, to form quinone methide 8b (Scheme I). In this case, a deuterium isotope effect would be expected for oxidation of [5a-CD3]-lbto 7b. Indeed, AMVN-dependent oxidation of unlabeled l b produced 5,5'-spirodimer 7b in about &fold higher yield than did oxidation of [5a-CD31-lb under the same conditions (Figure 3). This isotope effect suggests that a hydrogen abstraction from 2b forms quinone methide 8b. These data do not indicate whether the hydrogenabstracting species is a peroxyl radical, an alkoxy1radical (see below), or another chromanoxyl radical 2b (i.e., a disproportionation reaction of 2b). In oxidations of [5a-CD31-lb, a new product was observed, which eluted on reverse-phase HPLC columns
354
Chem. Res. Toxicol., Vol. 6, No. 3, 1993
Liebler et al.
7 0
E C
T
601
401 0 ' 0
1
20
40
60
time, min Figure 3. Formation of product 7b with AMVN-initiated oxidation of unlabeled l b ( 0 )and [5a-CD3]-lb (w) in hexane.
C
I
D
PPm
Figure 2. 13C-NMRanalysis of AMVN dependent oxidation of [5-13C-methy1]-lbinCD3CNat6OOC. Proton-decoupledspectra were recorded a t 15 min (A), 60 min (B), and 120 min (C). After 120min, the reaction mixture was passed through an octadecylsilyl solid-phase extraction column and then reanalyzed (D).
Scheme I
,
i
ll'ds
just after 7b and was identified as a novel 5,7'-spirodimer 1 ld5. This previously unreported dimer evidently is formed from the 5- and 7-quinone methides, which arise from competing hydrogen abstractions from the 5- and 7-positions of chromanoxyl radical 2b (Scheme I). Deuterium substitution thus slows the formation of [5a-CDzl8b and instead directs hydrogen abstraction to C,a, which yields the corresponding 7a-quinone methide. Interestingly, potassium hexacyanoferrate(II1) oxidation of lb, a
convenient method for preparing 5,5'-spirodimer 7b (19), also yielded a 5,7'-spirodimer 11. Moreover, potassium hexacyanoferrate(II1) oxidation of [5a-CD&lb yielded the deuterated analog I I-&. Mass spectra of 11 and 1 l-d5 from potassium hexacyanoferrate(II1) oxidation and of 1 1 - d ~from AMVN oxidation all exhibited identical fragmentation patterns and 'H- and 13C-NMRspectra, except for differences expected from deuterium substitution. Our results suggest that differences in the fate of la/ lb in peroxyl radical-dependent oxidations may be due largely to solvent effects on the reactions of chromanoxyl radical 2b. Increased solvent polarity increases the yield of 8a(alky1dioxy)chromanones 3b and epoxides, whereas decreased solvent polarity instead favors formation of spirodimer 7b and related dimer and trimer products. This pattern of solvent effect would reconcile all previously reported product distributions in peroxyl radical oxidations of l a (9, 10, 12,20-22). Solvent polarity also may dictate product distribution by influencing the types of oxidizing radicals formed from AMVN. Horswill and Ingold (23) have suggested that nonpolar solvents enhance the formation of alkoxyl radicals from azo compounds. Alkoxy1radicals display a marked preference for hydrogen abstraction and may be the principal oxidants responsible for forming quinone methide 8a/b and the spiro dimers. In support of this hypothesis, spirodimer 7a/b and other products of quinone methide 8a/b are principal products of la/b oxidation by alkoxyl (15,24) and benzoyloxyl(25) radicals. The formation of products through other alkoxyl radical dependent reactions may account for the shortfall in product recovery observed in nonpolar solvents (Table I). In lipid bilayers, epoxides and 8a-substituted tocopherones are the principal products of l a oxidation by peroxyl radicals, whereas dimers have not been observed (21, 26). Nevertheless, Draper and colleagues isolated dimer and trimer oxidation products from samples of rodent liver (27). These observations suggest that oxidation of l a may occur in diverse enough reaction environments to yield both dimers and epoxides. Alternatively, the formation of dimers and trimers may reflect l a oxidation by alkoxyl radicals even in relatively polar reaction environments, as in the oxidation of la to 5,5'spirodimer 7a by benzoyloxyl radicals in acetonitrile (25). Further studies of the oxidation of la in biological membranes may help to resolve these questions.
Acknowledgment. We thank Dr. Graham Burton for helpful suggestions and Mr. Peter F. Baker for mass spectral analyses. This work was supported in part by USPHS Grant CA47943.
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Oxidation of a Vitamin E Model Compound Supplementary Material Available: Figures 4-8 depicting
1H-NMR, 1H-COSY spectra, and 1H resonance assignments for compound 3b, peaks 1and 2 ( 5 pages). Ordering information is given on any current masthead page.
References (1) Machlin,L. J. (1991)VitaminE. InHandbookofuitamins (Machlin,
L. J., Ed.) 2nd ed., pp 99-144, Marcel Dekker, New York. (2) . , Chow. C. K. (1991) Vitamin E and oxidative stress. Free
Radical
~ i o lk. e d . ii, 215-232. (3) Burton. G. W.. and Innold. K. U. (1986) Vitamin E: amlication of the principles of physical organic chemistry to the expioiation of its structure and function. Acc. Chem. Res. 19, 194-201. (4) Packer, J. E., Slater, T. F., and Willson, R. L. (1979) Direct observation of a free radical interaction between vitamin E and vitamin C. Nature 278,737-738. (5) Mehlhorn, R. J., Sumida, S., and Packer, L. (1989) Tocopheroxyl radical persistence and tocopherol consumption in liposomes and in vitamin E-enriched rat liver mitochondria and microsomes. J.Biol. Chem. 264, 13448-13452. (6) Niki, E., Tsuchiya, J., Tanimura, R., and Kamiya, Y. (1982) Regeneration of vitamin E from alpha-chromanoxyl radical by glutathione and vitamin C. Chem. Lett., 789-792. (7) Kalayanaraman, B., Darley-Usmar, V. M., Wood, J., Joseph, J., and Parthasarathy,S. (1992)Synergisticinteraction betweenthe probucol phenoxy1radical and ascorbic acid in inhibiting the oxidation of low density lipoprotein. J. Biol. Chem. 267, 6789-6795. (8) Winterle, J.,Dulin,D.,andMill,T. (1984)Productaandstoichiometry of reaction of vitamin E with alkylperoxy radicals. J. Org. Chem. 49,491-495. (9) Yamauchi, R., Mataui, T., Satake, Y., Kato, K., and Ueno, Y. (1989) Reaction products of alpha-tocopherol with a free radical initiator, 2,2’-azobis(2,4-dimethylvaleronitrile).Lipids 24, 204-209. (10) Liebler, D. C., Baker, P. F., and Kaysen, K. L. (1990) Oxidation of vitamin E: Evidence for competingautoxidation and peroxyl radical trapping reactions of the tocopheroxyl radical. J. Am. Chem. SOC. 112,6995-7000. (11) Howard, J. A. (1973) Homogenous Liquid Phase Autoxidations. In Free Radicals (Kochi, J. K., Ed.) Vol2., pp 3-62, Wiley, New York. (12) Matsuo, M., Matsumoto, S., IItaka, Y., and Niki, E. (1989)Radical scavengingreactions of vitamin E and its model compound, 2,2,5,7,8pentamethylchroman-6-01,in a tert-butylperoxylradical generating system. J. Am. Chem. SOC.111, 7179-7185. (13) Yamauchi, R., Matsui, T., Kato, K., and Ueno, Y. (1989) Reaction of alpha-tocopherol with 2,2’-azobis(2,4-dimethylvaleronitrile)in benzene. Agric. Biol. Chem. 12, 3257-3262. (14) Skinner, W. A., and Parkhurst, R. M. (1966) Oxidation products of vitamin E and its model, 6-hydroxy-2,2,5,7,8-pentamethylchroman.
VIII. Oxidation with benzyl peroxide. J. Org. Chem. 31, 12481251.
(15) Suarna, C., Craig, D. C., Cross, K. J., and Southwell-Keely, P. T. (1988) Oxidations of vitamin E (alpha-tocopherol) and ita model compound 2,2,5,7,8-pentamethyI-6-hydroxychroman. A new dimer. J. Org. Chem. 53, 1281-1284. (16) Ozawa, T., Hanaki, A., Matsumoto, S., and Matauo, M. (1978) Electron spin resonance studies of radicals obtained by the reaction of a-tocopherol and its model compound with superoxide ion. Biochim. Biophys. Acta 531, 72-78. (17) Nilsson, J. L. G., Sievertsson, H., and Selander, H. (1968)Synthesis of methyl substituted 6-hydroxychromans, model compounds of tocopherols. Acta. Chem. Scand. 22, 316C-3170. (18) Matsumoto, S., Matsuo, M., and Iitaka, Y. (1987) Reactions of a hydroperoxide, 8a-hydroperoxy-2,2,5,7,8-pentamethylchroman6(8aH)-one,derived from a vitamin E modelcompound dimerization and x-ray crystal structure. J. Chem. Res. Suppl.,58-59. (19) Skinner, W. A,, and Alaupovic, P. (1963) Vitamin E oxidation with alkaline ferricyanide. Science 140, 803-805. (20) Yamauchi, R., Kato, K., and Ueno, Y. (1988)Formation of trimers of alpha-tocopherol and its model compound, 2,2,5,7,8-pentamethylchroman-6-01,in autoxidizing methyl linoleate. Lipids 23,779783. (21) Liebler, D. C., Kaysen, K. L., and Burr, J. A. (1991) Peroxyl radical trapping and autoxidation reactions of alpha-tocopherol in lipid bilayers. Chem. Res. Toxicol. 4, 89-93. (22) Yamauchi, R., Matsui, T., Kato, K., and Ueno, Y. (1990) Reaction products of a-tocopherol with methyl linoleate-peroxyl radicals. Lipids 25, 152-158. (23) Horswill, E. C., and Ingold, K. U. (1966) The oxidation of phenols. I. The oxidation of 2,6-di-t-butyl-4-methylphenol, 2,6-di-t-butylphenol, and 2,6-dimethylphenol with peroxy radicals. Can. J. Chem. 44, 263-268. (24) Sumarno, M., Atkinson, E., Suarna, C., Saunders, J. K., Cole, E. R., and Southwell-Keely, P. T. (1987) Solvent influence on model oxidations of alpha-tocopherol. Biochim.Biophys. Acta 920, 247250. (25) Goodhue, C. T., and Risley, H. A. (1965)Reactions of vitamin E with peroxides. 11. Reaction of benzoyl peroxide with d-alpha-tocopherol in alcohols. Biochemistry 4, 854-858. (26) Liebler, D. C., and Burr, J. A. (1992) Oxidation of vitamin E during iron-catalyzed lipid peroxidation: evidence for electron-transfer reactions of the tocopheroxyl radical. Biochemistry 31,8278-8284. (27) Draper, H. H., Csallany, A. S., and Chiu, M. (1966) Isolation of a trimer of alpha-tocopherol from mammalian liver. Lipids 2,47-54. (28) Weast, R. C., Ed. (1976)Handbook of Chemistry and Physics, 56th ed., pp E55-E57, CRC Press, Cleveland.
