Synthesis and antioxidant profile of all-rac-α-selenotocopherol

Article abstract of DOI:10.1021/jo052133e

all-rac-α-Selenotocopherol (6c) has been synthesized in 11 steps in 6.6% total yield. Key steps include chloromethylation to approach the persubstituted aromatic 9b and cyclization of alcohol precursor 10 by radical homolytic substitution at selenium to f

Full text of DOI:10.1021/jo052133e

Synthesis and Antioxidant Profile of all-rac-r-Selenotocopherol
David Shanks,† Riccardo Amorati,‡ Maria Grazia Fumo,‡ Gian Franco Pedulli,‡
Luca Valgimigli,*,‡ and Lars Engman*,†
Department of Chemistry, Organic Chemistry, Box 599, Uppsala UniVersity,
SE-751 24 Uppsala, Sweden, and Dipartimento di Chimica Organica “A. Mangini”, UniVersita` di
Bologna, Via S. Giacomo 11, 40126 Bologna, Italy
lars.engman@kemi.uu.se
ReceiVed October 12, 2005
all-rac-R-Selenotocopherol (6c) has been synthesized in 11 steps in 6.6% total yield. Key steps include
chloromethylation to approach the persubstituted aromatic 9b and cyclization of alcohol precursor 10 by
radical homolytic substitution at selenium to form the selenotocopherol heterocycle. Determination of
the OH bond dissociation enthalpy (BDE) of 6c by electron paramagnetic resonance (EPR) equilibration
techniques gave a value of 78.1 ( 0.3 kcal mol-1, approximately 1 kcal mol-1 higher than that of
R-tocopherol. Kinetic studies performed by measuring oxygen uptake of the induced oxidation of styrene
in the presence of an antioxidant showed that selenotocopherol (6c) was a slightly poorer inhibitor than
R-tocopherol, in agreement with the BDE values. In contrast to simpler selenotocopherol analogues, 6c
was not regenerable in the presence of a stoichiometric coreductant in a two-phase lipid peroxidation
model.
Introduction
Oxidative stress is a contributing factor in many of the so-
called “diseases of aging” which include, for example, condi-
tions such as atherosclerosis and Alzheimer’s disease.1 Our
bodies have an endogenic arsenal of enzymatic and nonenzy-
matic antioxidants to combat oxidative degradation, but because
it is apparent that these defenses are sometimes breached, there
is a current interest in the development and application of new
synthetic antioxidants for “antioxidant therapy”. In cell mem-
branes, the most important antioxidant is Vitamin E, which is
a collective term for a series of structurally related chromanols
named tocopherols (1a-d), differing only in the number and
position of the aromatic methyl groups, where R-tocopherol is
the most abundant.2
In the development of new antioxidants to bolster our defense
against oxidative stress, the R-tocopherol structure is therefore
a natural starting point. The possibilities for variation of the
structure are numerous. Two decades ago, Ingold and co-
workers discovered that decreasing the size of the nonaromatic
ring from six- to five-membered (2) resulted in a significant
increase in antioxidant activity, presumably due to a better lp
SOMO overlap in the phenoxyl radical.3 The side chain of
R-tocopherol has been varied to provide water-soluble analogues
† Uppsala University.
‡ Universita` di Bologna.
(1) Finkel, F.; Holbrook, N. J. Nature 2000, 408, 239.
(2) Burton, G. W.; Ingold, K. U. Acc. Chem. Res. 1986, 19, 194.
(3) Burton, G. W.; Doba, T.; Gabe, E. J.; Hughes, L.; Lee, F. L.; Prasad,
L.; Ingold, K. U. J. Am. Chem. Soc. 1985, 107, 7053.
10.1021/jo052133e CCC: $33.50 © 2006 American Chemical Society
Published on Web 12/27/2005
J. Org. Chem. 2006, 71, 1033-1038
1033

Shanks et al.
with altered pharmacokinetic properties (3).4 Recently, variations
in the phenolic ring, as exemplified by pyrimidin-5-ol 4 and
pyridin-3-ol 5, were found to produce antioxidants with
improved air stability and hydrogen-atom donating capacity.5,6
SCHEME 1
The nature of the heteroatom in the fused heterocyclic ring
is another target for investigation. The aforementioned pyridinol-
and pyrimidinol-based antioxidants use amine substituents to
improve potency. Ingold has prepared the racemic sulfur
analogue of R-tocopherol (6b), which proved to be less effective
as a hydrogen-atom donor than the parent tocopherol (6a).7
Within our group, a series of simple chalcogen-substituted
analogues (7a-d) were tested, and the selenium-substituted
analogue (7c) seemed particularly promising, being catalytically
regenerated by a stoichiometric coreductant.8 We have also
previously synthesized a selenium-substituted R-tocopherol
analogue lacking only aromatic methyl groups (8), although this
compound was never tested for antioxidant activity.9 To further
assess the true potential of selenium-substituted antioxidants,
we have now synthesized the full selenium analogue of racemic
R-tocopherol (6c) and investigated its antioxidant activity.
SCHEME 2a
a Conditions: (a) Pb(OAc)4, CH2Cl2, quantitative; (b) NaCN, MeOH,
reflux, 47%; (c) MeI, K2CO3, acetone, quantitative; (d) KOH, H2O, ethylene
glycol, 140 °C, 90%; (e) LiAlH4, THF, reflux, 97%; (f) PBr3, Et2O, 97%;
(g) Br2, CH2Cl2, quantitative.
aromatic 9 could be made, it was expected that radical precursor
10 could be approached by the same means as those previously
reported for the synthesis of analogue 8.9
Our initial approach toward structure 9 from phenol 11
involved oxidation to the ortho-quinol acetates 12 (Scheme 2).12
The resulting mixture of regioisomers was used directly for
conjugate addition/rearomatization to give benzonitrile 13 in
moderate yields.13 This was converted to benzyl bromide 9a in
high yield via O-methylation to give 14, via nitrile hydrolysis
to give 15, via reduction of the carboxylic acid to give 16, via
benzylic substitution to give 17, and, finally, via aromatic
bromination.
A more expedient route, giving benzyl chloride 9b in three
steps, entailed bromination of phenol 11 to give 18 and
methylation to give 19, followed by microwave-assisted chlo-
romethylation of the product anisole (Scheme 3). Subsequent
synthesis toward ketone 24 proceeded as expected, in good
yields, following procedures developed for the synthesis of 8.9
Thus, the hexa-substituted aromatic was subjected to benzylic
substitution with the enolate of tert-butyl acetoacetate to give
Results and Discussion
Synthesis. Homolytic substitution at selenium is now a well-
established tool for the construction of selenacycles. Within our
group, this technique has been applied to the synthesis of simple
R-tocopherol analogues,9 and Schiesser has recently used it for
the construction of selenacycle-fused pyridines10 and selenap-
enams.11 This was therefore the method of choice for synthesiz-
ing selenotocopherol (6c) (Scheme 1). If the persubstituted
(4) Bolkenius, F. N.; Grisar, J. M.; De Jong, W. Free Radical Res.
Commun. 1991, 14, 363.
(5) Pratt, D. A.; DiLabio, G. A.; Brigati, G.; Pedulli, G. F.; Valgimigli,
L.; J. Am. Chem. Soc. 2001, 123, 4625.
(6) Wijtmans, M.; Pratt, D. A.; Valgimigli, L.; DiLabio, G. A.; Pedulli,
G. F.; Porter, N. A. Angew. Chem. Int. Ed. 2003, 42, 4370.
(7) Zahalka, H. A.; Robillard, B.; Hughes, L.; Lusztyk, J.; Burton, G.
W.; Janzen, E. G.; Kotake, Y.; Ingold, K. U. J. Org. Chem. 1988, 53, 3739.
(8) Malmstro¨m, J.; Jonsson, M.; Cotgreave, I. A.; Hammarstro¨m, L.;
Sjo¨din, M.; Engman, L. J. Am. Chem. Soc. 2001, 123, 3434.
(9) Al-Maharik, N.; Engman, L.; Malmstro¨m, J.; Schiesser, C. H. J. Org.
Chem. 2001, 66, 6286.
(11) Carland, M. W.; Martin, R. L.; Schiesser, C. H. Org. Biomol. Chem.
2004, 2, 2612.
(12) Quinkert, G.; Du¨rner, G.; Kleiner, E.; Adam, F.; Haupt, E.; Leibfritz,
D. Chem. Ber. 1980, 113, 2227.
(10) Fenner, T.; Schiesser, C. H. Molecules 2004, 9, 472.
(13) Leitich, J.; Wessely, F. Monatsh. Chem. 1964, 95, 116.
1034 J. Org. Chem., Vol. 71, No. 3, 2006

Antioxidant Profile of all-rac-R-Selenotocopherol
SCHEME 3a
FIGURE 1. Experimental (top) and computer-simulated (bottom)
spectra obtained by irradiating compound 6c in benzene containing 10%
v/v di-tert-butyl peroxide at 298 K.
OH Bond Dissociation Enthalpy (BDE). To measure the
OH BDE value for selenotocopherol (6c), we used the EPR
radical equilibration technique. Of the various methods used
for the determination of bond strengths, this one seems at present
to guarantee the best accuracy.14-17 For this purpose, we
measured the equilibrium constant, Ke, for the hydrogen-atom
transfer among a reference phenol (ArOH), in this case BHA
(BDE ) 77.2 kcal mol-1),14,18 the selenotocopherol (6c,
SeTocOH), and the corresponding phenoxyl radicals (eq 1)
generated under continuous photolysis in deoxygenated benzene
at room temperature (25 °C).
a Conditions: (a) Br2, CH2Cl2, quantitative; (b) MeI, K2CO3, acetone,
quantitative; (c) (CH2O)n, HCl, AcOH, 154 °C, 57%; (d) tert-butyl
acetoacetate, NaH, THF, 78%; (e) HCl, H2O, reflux, 98%; (f) ethylene
glycol, p-TsOH, benzene, reflux, quantitative; (g) (i) t-BuLi, THF, -78
°C, (ii) Se, -78 °C to room temperature, (iii) BnBr, room temperature,
66% overall; (h) HCl, H2O, THF, room temperature, 98% (i) Mg, 4,8,12-
trimethyltridecyl bromide, THF, reflux, 47%; (j) (i) oxalyl chloride, toluene,
(ii) 2-mercaptopyridine N-oxide, DMAP, toluene, reflux, 49%; (k) BBr3,
CH2Cl2, -78 °C, quantitative.
SeTocOH + ArO• h SeTocO• + ArOH
(1)
In the calculation of Ke, the equilibrium constant for eq 1, the
initial concentrations of SeTocOH and ArOH were used, and
the relative radical concentrations were determined by means
of EPR spectroscopy. The BDE for selenotocopherol (6c) was
calculated, in the assumption that the entropic term can be
neglected,15 by means of eq 2 from Ke and the BDE value of
the reference phenol.
20, decarboxylative hydrolysis to give 21, protection as the
glycol ketal to give 22, lithiation, selenium insertion and
benzylation to give 23, and, finally, deprotection to give 24.
Grignard addition to ketone 24 proved troublesome, and
significant quantities of starting material and aldol-type byprod-
ucts were isolated in addition to alcohol 10. It was found that
addition of borontrifluoride etherate suppressed formation of
the aldol byproducts, although the yield remained moderate.
Cyclization according to the Barton-Crich protocol formed
selenochroman 25, which was smoothly deprotected using BBr3
to provide selenotocopherol (6c).
Electron Paramagnetic Resonance (EPR) Spectra of the
Free Radical Obtained from Selenotocopherol (6c). Radicals
were produced at room temperature inside the cavity of an EPR
spectrometer by reacting selenotocopherol (6c) with alkoxyl
radicals generated photolytically from di-tert-butyl peroxide in
deoxygenated benzene solutions. By photolyzing solutions of
6c, an intense spectrum centered at g ) 2.0101 was observed,
as shown in Figure 1 together with its computer simulation.
Hyperfine splitting constants of 4.94 G (3H), 5.68 G (3H), 1.75
G (3H), and 1.37 G (2H) were extracted. These spectral pa-
rameters are consistent with the structure of the selenotocoph-
eroxyl radical.
BDE(SeTocO-H) = BDE(ArO-H) - RT ln(Ke) (2)
From these measurements, repeated under different light intensi-
ties to check the constancy of Ke, we obtained a BDE value of
78.1 ( 0.3 kcal mol-1 for selenotocopherol (6c). The BDE of
(14) Lucarini, M.; Pedrielli, P.; Pedulli, G. F.; Cabiddu, S.; Fattuoni, C.
J. Org. Chem. 1996, 61, 9259-9263.
(15) Brigati, G.; Lucarini, M.; Mugnaini, V.; Pedulli, G. F. J. Org. Chem.
2002, 67, 4828-4832.
(16) Pratt, A. D.; DiLabio, G. A.; Valgimigli, L.; Pedulli, G. F.; Ingold,
K. U. J. Am. Chem. Soc. 2002, 124, 11085-11092.
(17) Valgimigli, L.; Brigati, G.; Pedulli, G. F.; DiLabio, G.; Mastragos-
tino, M.; Arbizzani, C.; Pratt, D. A. Chem.-Eur. J. 2003, 9, 4997-5010.
(18) All the BDE values determined in benzene solution by means of
the EPR radical equilibration technique,14 based on the O-H BDE of 2,4,6-
tri-tert-butylphenol determined many years earlier by Mahoney et al.19 using
calorimetric measurements, must be downscaled by 1.1 kcal mol-1 because
of the revision of the heat of formation of E-azobenzene.20
(19) Mahoney, L. R.; Ferris, F. C.; DaRooge, M. A. J. Am. Chem. Soc.
1969, 91, 3883.
(20) Mulder, P.; Korth, H.-G.; Pratt, D. A.; DiLabio, G. A.; Valgimigli,
L.; Pedulli, G. F.; Ingold, K. U. J. Phys. Chem. A 2005, 109, 2647-2655.
J. Org. Chem, Vol. 71, No. 3, 2006 1035

Shanks et al.
(9)
∆[O2]t ) -kp/kinh [AH] ln(1 - t/τ)
It transpires that selenotocopherol has an inhibition rate,
kinh ) 1.2 × 106 M-1 s-1, and a stoichiometric coefficient, n )
1.9, similar to those of R-tocopherol, which has kinh ) 3.2 ×
106 M-1 s-1 and n ) 2.0. Thus, the selenotocopherol (6c) is
only slightly less effective than its oxygen-containing analogue
R-tocopherol, in agreement with the slightly higher value of
the OH BDE. This, and the fact that the stoichiometric
coefficients are nearly identical, implies that both antioxidants
act by the same mechanism.
Evaluation of the Regenerability of Selenotocopherol (6c).
To evaluate whether selenotocopherol (6c) was regenerable by
a stoichiometric coreductant, it was tested in a two-phase lipid
peroxidation model similar to that previously described.8 In brief,
a thermostated vial is charged with a chlorobenzene solution
of linoleic acid. First, the antioxidant to be tested is added,
followed by an aqueous solution of the coreductant (N-acetyl
cysteine). The mixture is stirred and allowed to come to
equilibrium for 30 min before oxidation is induced by addition
of a solution of an azo initiator. The oxidation is followed by
sampling the organic phase every 10 min, separating the
components by high-pressure liquid chromatography (HPLC),
and monitoring the formation of conjugated dienes at 234 nm.
In the absence of NAC, selenotocopherol had an inhibition
period of 58 min, compared to R-tocopherol (6a), which had a
tinh ) 80 min. We expected, in analogy to selenide 7c, that the
inhibition period of selenotocopherol would be significantly
increased in the presence of NAC, but to our surprise, no
difference was observed. Hence, although the simple seleno-
tocopherol analogue 7c is catalytically regenerable, the full
selenotocopherol (6c) is not. At the moment, we can only
speculate as to the origin of this difference in reactivity. Possible
reasons include steric hindrance at the redox-active site or a
changeover in mechanism from electron transfer to hydrogen-
atom transfer with increasing substitution of the selenide.
Methyl-protected selenotocopherol 25 was also tested and was
shown to have no inhibiting effect whatsoever, demonstrating
the necessity of the phenolic hydrogen for activity. In analogy
to selenide 7c, selenotocopherol is not expected to possess any
hydroperoxide-decomposing activity. Further studies into the
antioxidant and regeneration mechanisms are underway in our
laboratories.
FIGURE 2. Oxygen consumption recorded during the thermally
initiated autoxidation of styrene (4.7 M) in air-saturated chlorobenzene
at 30 °C, in the presence of AMVN (5 × 10-3 M) as the initiator and
selenotocopherol (6c) or R-tocopherol (6a), at 5.0 × 10-6 M.
R-tocopherol is valued at 77.15 kcal mol-1 by the same
technique,18 and thus, it can be seen that the BDE in the
selenium-containing analogue is approximately 0.95 kcal mol-1
higher than in R-tocopherol. This is in good agreement with
the calculated difference of 0.94 kcal mol-1 21
.
Kinetic Measurements. Kinetic measurements with peroxyl
radicals have been performed by studying the inhibited autoxi-
dation of a standard oxidizable substrate (styrene) at 303 K in
chlorobenzene, initiated by 2,2′-azobis(2,4-dimethylvaleronitrile)
(AMVN), in the presence of selenotocopherol (6c) or R-tocoph-
erol (R-TOH) as the reference antioxidant (Figure 2).
The autoxidation was followed by monitoring the oxygen
consumption in an oxygen-uptake apparatus built in our
laboratory and based on a Validyne DP15 differential pressure
transducer, which has been previously described.22 The observed
kinetics are in accord with equations 3-8.
Ri
9
initiator
R• + O2 f ROO•
8 R•
(3)
(4)
kp
9
ROO• + RH
8 ROOH + R•
(5)
(6)
2kt
ROO• + ROO• 98 products
Experimental Section
kinh
ROO• + ArOH 8 ROOH + ArO•
(7)
(8)
Synthesis. A. 3-Cyano-2,5,6-trimethylphenol (13). To a solu-
tion of o-quinol acetates 1212 (1:1 mixture, 2 g, 10.3 mmol) in
methanol (20 mL) was added sodium cyanide (1.11 g, 22.7 mmol).
The mixture was stirred at room temperature for 15 min and then
heated at reflux for 40 min. Water (30 mL) was added, and the
methanol was removed on the rotary evaporator. The aqueous phase
was extracted repeatedly with EtOAc, and the combined organic
phase was washed with saturated brine, dried over MgSO4, and
evaporated to give a crude brown residue. This was recrystallized
from CCl4 to give the title compound as a white solid (780 mg,
ROO• + ArO• f products
Compound 6c showed a neat inhibition period, τ, in styrene,
whose length provided the stoichiometric coefficient, i.e., the
number of peroxyl radicals trapped by one molecule of
antioxidant, n ) Riτ/[AH], where Ri is the initiation rate
measured in a preliminary set of experiments using R-tocopherol
and [AH] is the initial concentration of antioxidant. Integration
of the oxygen consumption trace affords the rate of reaction
with peroxyl radicals, kinh, provided kp is known (eq 9).
1
47%): mp 132-133 °C; H NMR δ 7.02 (1H, s), 4.96 (1H, s),
2.40 (3H, s), 2.26 (3H, s), 2.20 (3H, s); 13C NMR δ 152.5, 136.6,
128.2, 126.0, 124.5, 118.8, 110.5, 20.1, 14.5, 12.6. Anal. Calcd
for C10H11NO: C, 74.51; H, 6.88. Found: C, 74.32; H, 6.87.
B. 3-Cyano-2,5,6-trimethylanisole (14). A mixture of phenol
13 (2.26 g, 14.1 mmol), MeI (10.04 g, 70.5 mmol), and K2CO3
(19.55 g, 141.0 mmol) in acetone (70 mL) was stirred at room
temperature for 22 h. The product slurry was filtered, and the solvent
was removed in vacuo. The residue was partitioned between water
(21) Calculations performed at the (RO)B3LYP/LANL2DZdp//(U)B3LYP/
LANL2DZ level. Shanks, D.; Frisell, H.; Ottosson, H.; Engman, L. Org.
Biomol. Chem. 2006, accepted for publication.
(22) Amorati, R.; Pedulli, G. F.; Valgimigli, L.; Attanasi, O. A.;
Filippone, P.; Fiorucci, C.; Saladino, R. J. Chem. Soc., Perkin Trans. 2
2001, 2142-2146.
1036 J. Org. Chem., Vol. 71, No. 3, 2006

Antioxidant Profile of all-rac-R-Selenotocopherol
and dichloromethane, and the aqueous phase was extracted with
dichloromethane. The combined organic phase was washed with
brine, dried over MgSO4, and evaporated to give the title compound
acetic acid (3.5 mL) was heated at 154 °C for 30 min in a
microwave reactor. The product was precipitated with water, and
the supernatant was decanted. The crude solid was dissolved in
ether, washed with water and saturated brine, and dried over
MgSO4, and the solvent was removed in vacuo to give a crude
brown solid. This was purified by column chromatography (pentane/
dichloromethane 9:1) to give the title compound (554 mg, 57%)
1
(2.46 g, quantitive) as a beige solid: mp 59-60 °C; H NMR δ
7.17 (1H, s), 3.69 (3H, s), 2.43 (3H, s), 2.24 (3H, s), 2.23 (3H, s);
13C NMR δ 157.3, 137.2, 136.2, 132.7, 129.2, 118.6, 111.1, 60.4,
20.0, 14.6, 13.1. Anal. Calcd for C11H13NO: C, 75.40; H, 7.48.
Found: C, 75.22; H, 7.47.
1
as a white crystalline solid: mp 101-102 °C; H NMR δ 4.87
(2H, s), 3.66 (3H, s), 2.41 (3H, s), 2.40 (3H, s), 2.28 (3H, s); 13C
NMR δ 156.4, 136.3, 133.9, 132.4, 130.0, 123.9, 60.5, 45.7, 21.0,
14.3, 12.9. Anal. Calcd for C11H14BrClO: C, 47.60; H, 5.08.
Found: C, 47.75; H, 5.02.
C. 3-Methoxy-2,4,5-trimethylbenzoic Acid (15). A mixture of
nitrile 14 (1.0 g, 5.7 mmol), KOH (960 mg, 17.1 mmol), water
(1.4 mL), and ethylene glycol (11 mL) was heated (bath temp of
140 °C) for 7 h. The solution was then cooled to room temperature
and treated with aqueous HCl (24 mL, 1 M). The precipitated solid
was collected by filtration and dried under vacuum over P2O5,
giving the title compound (1.0 g, 90%) as a white powder: mp
159-160 °C; 1H NMR δ 11.70 (1H, br s), 7.65 (1H, s), 3.69 (3H,
s), 2.55 (3H, s), 2.28 (3H, s), 2.26 (3H, s); 13C NMR δ 173.2, 157.7,
135.9, 135.7, 131.8, 128.4, 127.1, 60.4, 20.1, 13.8, 13.2. Anal. Calcd
for C11H14O3: C, 68.02; H, 7.27. Found: C, 67.88; H, 7.23.
D. 3-Methoxy-2,4,5-trimethylbenzyl Alcohol (16). To a solution
of the acid 15 (981 mg, 5.05 mmol) in dry THF (11 mL) at 0 °C
under nitrogen was added LiAlH4 (383 mg, 10.1 mmol) portionwise.
After complete addition, the mixture was allowed to come to room
temperature for 30 min and then heated at reflux for 20 h. Excess
LiAlH4 was quenched by careful addition of aqueous HCl (3 M)
at 0 °C, and the aqueous phase was extracted with Et2O. The
combined organic phase was washed with brine, dried over MgSO4,
and evaporated to give the title compound (880 mg, 97%) as a
white powder: mp 87-88 °C; 1H NMR δ 6.94 (1H, s), 4.63 (2H,
s), 3.68 (3H, s), 2.27 (3H, s), 2.25 (3H, s), 2.20 (3H, s), 1.51 (1H,
br s); 13C NMR δ 157.2, 137.4, 135.7, 129.4, 126.7, 125.4, 64.0,
60.3, 20.1, 12.6, 11.5. Anal. Calcd for C11H16O2: C, 73.30; H, 8.95.
Found: C, 73.42; H, 8.99.
E. 3-Methoxy-2,4,5-trimethylbenzyl Bromide (17). Neat PBr3
(710 mg, 2.6 mmol) was added dropwise to a solution of alcohol
16 (860 mg, 4.77 mmol) in Et2O (10 mL). The reaction mixture
was stirred at room temperature for 4.5 h, before quenching with
water. The organic phase was diluted with Et2O, washed with water
and brine, dried over MgSO4, and evaporated to give the title
compound (1.13 g, 97%) as a pale-blue powder: mp 70-71 °C;
1H NMR δ 6.93 (1H, s), 4.49 (2H, s), 3.68 (3H, s), 2.32 (3H, s),
2.23 (3H, s), 2.20 (3H, s); in good agreement with previously
published data;23 13C NMR δ 157.4, 136.0, 134.3, 130.9, 127.9,
127.2, 60.4, 33.3, 20.0, 12.8, 11.7. Anal. Calcd for C11H15BrO: C,
54.34; H, 6.22. Found: C, 54.23; H, 6.16.
I. tert-Butyl-2-(2-bromo-5-methoxy-3,4,6-trimethylbenzyl) Ac-
etoacetate (20). Neat tert-butyl acetoactetate (475 mg, 3 mmol)
was added dropwise to a stirred slurry of NaH (60% in mineral
oil, 120 mg, 3 mmol) in dry THF (4 mL) at 0 °C under nitrogen.
The mixture was allowed to come to room temperature over 30
min, and the solution became homogeneous. The benzyl chloride
9b (278 mg, 1 mmol) dissolved in THF (3 mL) was added by
syringe, and the mixture was heated at reflux for 15 h. The reaction
was then allowed to cool to room temperature and was quenched
by addition of saturated aqueous NH4Cl. The aqueous phase was
extracted with ether, and the combined organic phase was washed
with brine, dried over MgSO4, and evaporated to give crude product.
Excess tert-butyl acetoactetate was removed by Kugelrohr distil-
lation under vacuum, and the residue was subjected to column
chromatography (pentane/Et2O 85:15) to give the title compound
as a colorless viscous oil (311 mg, 78%): 1H NMR δ 3.93 (1H, t,
J ) 7.3 Hz), 3.60 (3H, s), 3.39 (1H, dd, J ) 14.5, 7.3 Hz), 3.32
(1H, dd, J ) 14.5, 7.3 Hz), 2.37 (3H, s), 2.27 (3H, s), 2.25 (3H, s),
2.17 (3H, s), 1.37 (9H, s); 13C NMR δ 203.4, 168.9, 156.3, 135.8,
135.4, 129.8, 129.6, 124.2, 82.1, 60.3, 59.6, 31.9, 30.0, 28.0, 21.0,
14.1, 13.8. Anal. Calcd for C19H27BrO4: C, 57.15; H, 6.82.
Found: C, 57.12; H, 6.95.
J. 4-(2-Bromo-5-methoxy-3,4,6-trimethylphenyl)-2-butanone
(21). A solution of acetoacetate 20 (5.0 g, 12.5 mmol) in aqueous
HCl (9 M, 80 mL) was heated at reflux for 15 h. The mixture was
then cooled to room temperature and extracted with diethyl ether.
The organic phase was washed with water and saturated brine and
then dried over MgSO4 to give the title compound (3.65 g, 98%)
1
as a white solid: mp 84-85 °C; H NMR δ 3.63 (3H, s), 3.09
(2H, m), 2.64 (2H, m), 2.38 (3H, s), 2.26 (3H, s), 2.25 (3H, s),
2.19 (3H, s); 13C NMR δ 208.3, 156.3, 137.5, 135.8, 129.4, 128.3,
123.7, 60.4, 42.5, 30.1, 28.7, 21.0, 14.1, 13.2. Anal. Calcd for
C14H19BrO2: C, 56.20; H, 6.40. Found: C, 56.31; H, 6.43.
K. 2-[2-(2-Bromo-5-methoxy-3,4,6-trimethylphenyl)ethyl]-2-
methyl-1,3-dioxolane (22). A mixture of ketone 21 (910 mg, 3.0
mmol), p-toluenesulfonic acid (60 mg, 0.3 mmol), and ethylene
glycol (380 mg, 6.1 mmol) in benzene (15 mL) was heated at reflux
for 13 h. The solution was then cooled to room temperature, diluted
with diethyl ether, washed with saturated aqueous NaHCO3 and
brine, dried over MgSO4, and evaporated to give the title compound
as a colorless oil (1.04 g, quantitive): 1H NMR δ 4.04 (4H, s),
3.67 (3H, s), 2.96 (2H, m), 2.41 (3H, s), 2.33 (3H, s), 2.28 (3H, s),
1.86 (2H, m), 1.46 (3H, s); 13C NMR δ 156.2, 138.6, 135.6, 128.9,
128.2, 123.8, 110.0, 65.0, 60.4, 37.5, 29.3, 23.9, 21.0, 14.1, 13.1.
L. 2-[2-(2-Benzylselenenyl-5-methoxy-3,4,6-trimethylphenyl)-
ethyl]-2-methyl-1,3-dioxolane (23). To a solution of the arylbro-
mide 22 (2.74 g, 8.0 mmol) in dry THF (80 mL) at -78 °C under
nitrogen was added t-BuLi (8.4 mL, 8.0 mmol). The mixture was
stirred at -78 °C for 5 min, and then finely ground selenium (662
mg, 8.4 mmol) was added in one portion. The mixture was allowed
to approach room temperature for 15 min until all selenium had
dissolved. Benzyl bromide (1.14 mL, 9.6 mmol) was added, and
the reaction mixture was stirred at room temperature for a further
3 h and then quenched by addition of saturated aqueous NH4Cl.
The aqueous phase was extracted with ether, and the combined
organic phase was washed with brine, dried over MgSO4, and
evaporated to give crude product. This was purified by flash
F. 2-Bromo-5-methoxy-3,4,6-trimethylbenzyl Bromide (9a).
Bromine (750 mg, 4.7 mmol) dissolved in dichloromethane (8 mL)
was added dropwise to a solution of aromatic 17 (1.09 g, 4.5 mmol)
in dichloromethane (10 mL), and the reaction mixture was stirred
for 4.5 h at room temperature. Excess bromine was destroyed by
addition of saturated aqueous Na2S2O3 (20 mL). The aqueous phase
was extracted with dichloromethane, and the combined organic
phase was washed with brine, dried over MgSO4, and evaporated
to give the title compound (1.44 g, quantitative) as a white
1
powder: mp 103-104 °C; H NMR δ 4.77 (2H, s), 3.66 (3H, s),
2.39 (6H, s), 2.27 (3H, s); 13C NMR δ 156.4, 136.4, 133.9, 132.3,
129.9, 123.9, 60.5, 33.9, 21.0, 14.3, 12.9. Anal. Calcd for C11H14-
Br2O: C, 41.03; H, 4.38. Found: C, 41.05; H, 4.31.
G. 4-Bromo-2,3,6-trimethylanisole (19). Anisole 19 was pre-
pared by the same method as that used for anisole 14, using phenol
1
188 as starting material: quantitative yield; H NMR δ 7.28 (1H,
s), 3.70 (3H, s), 2.36 (3H, s), 2.29 (3H, s), 2.27 (3H, s); 13C NMR
δ 156.3, 135.2, 131.9, 131.7, 130.1, 120.0, 60.2, 20.0, 16.0, 13.8.
H. 2-Bromo-5-methoxy-3,4,6-trimethylbenzyl Chloride (9b).
A mixture of anisole 19 (800 mg, 3.5 mmol), paraformaldehyde
(524 mg, 17.5 mmol), concentrated HCl (37%, 7 mL), and glacial
(23) Gruter, G. -J. M.; Akkerman, O. S.; Bickelhaupt, F. J. Org. Chem.
1994, 59, 4473.
J. Org. Chem, Vol. 71, No. 3, 2006 1037

Shanks et al.
chromatography (pentane/EtOAc 90:10) to give the title compound
as a yellow solid (2.29 g, 66%): mp 37-39 °C; H NMR δ 7.17
(3H, m), 7.03 (2H, m), 3.98 (2H, m), 3.95 (2H, m), 3.81 (2H, s),
3.66 (3H, s), 2.91 (2H, m), 2.48 (3H, s), 2.26 (3H, s), 2.22 (3H,s),
1.69 (2H, m), 1.38 (3H, s); 13C NMR δ 157.8, 144.5, 141.3, 139.3,
128.9, 128.4, 128.0, 127.9, 127.1, 126.8, 110.1, 65.0, 60.3, 39.2,
33.0, 30.0, 23.8, 21.7, 14.1, 13.1.
2.28 (3H, s), 2.23 (3H, s), 2.23 (3H, s), 1.93 (2H, m), 1.84-1.03
(24H, several peaks), 0.92 (12H, several peaks); 13C NMR δ (some
characteristic peaks) 149.6, 133.6, 133.0, 122.0, 120.6, 119.8, 45.1.
Anal. Calcd for C29H50OSe: C, 70.56; H, 10.21. Found: C, 70.30;
H, 10.04.
EPR and Thermochemical Measurements. Deoxygenated
benzene solutions containing the phenols (0.01-0.001 M) and di-
tert-butyl peroxide (10% v/v) were sealed under nitrogen in a
suprasil quartz EPR tube. The sample was inserted at room
temperature in the cavity of an EPR spectrometer and photolyzed
with the unfiltered light from a 500 W high-pressure mercury lamp.
The temperature was controlled with a standard variable temperature
accessory and was monitored before and after each run with a
copper-constantan thermocouple.
1
M. 2-(2-Benzylselenenyl-5-methoxy-3,4,6-trimethylphenyl)-
ethyl Methyl Ketone (24). A mixture of ketal 23 (2.22 g, 5.1
mmol), THF (25 mL), and aqueous HCl (25 mL, 3 M) was stirred
for 15 h at room temperature. The aqueous phase was extracted
with diethyl ether, and the combined organic phase was washed
with brine, dried over MgSO4, and evaporated to give the title
1
compound (1.96 g, 98%) as a yellow solid: mp 80-81 °C; H
NMR δ 7.17 (3H, m), 6.99 (2H, m), 3.81 (2H, s), 3.66 (3H, s),
3.07 (2H, m), 2.49 (3H, s), 2.38 (2H, m), 2.23 (3H, s), 2.21 (3H,
s), 2.10 (3H, s); 13C NMR δ 208.7, 157.8, 143.4, 141.4, 139.3,
128.9, 128.5, 128.4, 127.7, 127.1, 126.9, 60.4, 44.2, 33.0, 30.0,
29.2, 21.7, 14.1, 13.2. Anal. Calcd for C21H26O2Se: C, 64.77; H,
6.73. Found: C, 64.52; H, 6.63.
The EPR spectra were recorded on a spectrometer equipped with
a microwave frequency counter for the determination of the
g-factors, which were corrected with respect to that of the perylene
radical cation in concentrated H2SO4 (g ) 2.00258).
When using mixtures of BHA and selenotocopherol (6c), the
molar ratio of the two equilibrating radicals was obtained from the
EPR spectra and used to determine the equilibrium constant, Ke.
Spectra were recorded a few seconds after starting to irradiate to
avoid significant consumption of the phenols during the course of
the experiment.
Relative radical concentrations were determined either by double
integration of the signals due to the two equilibrating species or
by comparison of the digitized experimental spectra with computer-
simulated ones, as previously described.14
Kinetic Measurements. The rate constants for the reaction of
the antioxidants with peroxyl radicals have been measured by
following the autoxidation of pure styrene (4.7 M) in chlorobenzene,
at 303 K using AMVN (5 × 10-3 M) as an initiator . The
antioxidant concentration was kept constant for all measurements
(5.0 × 10-6 M) to compare more easily their behavior. Initiation
rates, Ri, were determined for each condition in preliminary ex-
periments by the inhibitor method using R-tocopherol as the
reference antioxidant: Ri ) 2[R-TOH]/τ.3
HPLC Peroxidation Assay. An HPLC equipped with an
autoinjector, a thermostated sample rack, a 5 µL injection loop, a
photodiode array detector, and a relay-controlled stirring plate was
used for the peroxidation studies. In a typical experiment, linoleic
acid in chlorobenzene (7.5 mL, 36.2 mM) was stirred at 1000 rpm,
42 °C, in a 20 mL vial. To this solution was added the antioxidant
in ethanol (107 µL, 3.0 mM) by micropipet, followed by an aqueous
solution of NAC (8.0 mL, 1.0 mM). An injection routine, whereby
every 10 min 5 µL samples of the lower organic phase were
separated by a silica column (4 µ, 150 × 4.6 mm) eluted with
hexane/ethanol mixtures, was commenced. Stirring (controlled by
relay from the HPLC) was stopped 30 s prior to sampling to allow
phase separation and immediately resumed thereafter. Prior to the
fourth injection, a thermostated solution of AMVN in chlorobenzene
(0.5 mL, 22.4 mM) was added to initiate oxidation. Formation of
conjugated dienes was monitored at 234 nm. The inhibition period
(tinh) was determined by the least-squares method as the point where
the uninhibited oxidation phase was projected to intersect the initial
peroxide concentration.
N. 1-(2-Benzylselenenyl-5-methoxy-3,4,6-trimethylphenyl)-
3,7,11,15-tetramethyl-3-hexadecanol (10). To a slurry of mag-
nesium (48 mg, 2 mmol) in dry THF (3 mL) heated at reflux under
nitrogen was slowly added 4,8,12-trimethyltridecyl bromide (458
mg, 1.5 mmol) dissolved in dry THF (2 mL). The reaction mixture
was heated at reflux for 5 h and then cooled to -78 °C. First, BF3‚
Et2O (114 µL, 0.9 mmol) was added, followed by ketone 24 (300
mg, 0.77 mmol) in dry THF (5 mL). The mixture was stirred at
-78 °C for 15 min and then brought to room temperature and stirred
overnight. The reaction was quenched by addition of saturated
aqueous NH4Cl. The aqueous phase was extracted with ether, and
the combined organic phase was washed with brine, dried over
MgSO4, and evaporated to give the crude product mixture, which
was separated by column chromatography (pentane/Et2O 85:15) to
give the title compound as a pale viscous oil (227 mg, 48%) and
recovered starting material (140 mg, 47%), as well as traces of
aldol product: 1H NMR δ 7.17 (3H, m), 7.02 (2H, m), 3.81 (2H,
s), 3.67 (3H, s), 2.84 (2H, m), 2.50 (3H, s), 2.26 (3H, s), 2.53 (3H,
s), 1.65-1.03 (27H, m), 0.86 (12H, m); 13C NMR δ (some
characteristic peaks) 157.8, 144.8, 141.3, 139.3, 128.9, 128.5, 128.0,
127.8, 127.0, 126.9, 73.3. Anal. Calcd for C37H60O2Se: C, 72.16;
H, 9.82. Found: C, 71.94; H, 9.93.
O. 6-Methoxy-2,5,7,8-tetramethyl-2-(4,8,12-trimethyltridecyl)-
selenochromane (25). A solution of alcohol 10 (230 mg, 0.37
mmol) and oxalyl chloride (0.4 mL) in dry toluene (2.2 mL) was
stirred at room temperature for 15 h. All volatile components were
removed under vacuum, and the residue was dissolved in toluene
(1.5 mL). This solution was transferred by canulla to a slurry of
the sodium salt of 2-mercaptopyridine N-oxide (67 mg, 0.45 mmol)
and DMAP (4.5 mg, 0.04 mg) and heated at reflux in toluene (3
mL). The solvent was then removed in vacuo, and the residue was
purified by column chromatography (pentane/dichloromethane 80:
20) to give the title compound as a colorless oil (93 mg, 49%): 1H
NMR δ 3.65 (3H, s), 2.83 (1H, ddd, J ) 16.9, 7.4, 4.1 Hz), 2.75
(1H, ddd, J ) 16.9, 8.9, 4.3 Hz), 2.23 (6H, s), 2.21 (3H, s), 1.92
(2H, m), 1.84-1.03 (24H, several peaks), 0.86 (12H, several peaks);
13C NMR δ (some characteristic peaks) 154.5, 133.6, 133.4, 127.7,
127.1, 126.1, 60.5. Anal. Calcd for C30H52OSe: C, 70.97; H, 10.32.
Found: C, 71.13; H, 10.48.
Acknowledgment. We thank the Swedish Research Council,
the University of Bologna, and MIUR (Rome; research project
“Free Radical Processes in Chemistry and Biology: Funda-
mental Aspects and Applications in Environment and Material
Sciences”) for financial support.
P. 2,5,7,8-Tetramethyl-2-(4,8,12-trimethyltridecyl)selenochro-
man-6-ol (6c). BBr3 (394 µL, 1.0 M in CH2Cl2, 0.39 mmol) was
added dropwise to a solution of the selenide 25 in dichloromethane
(2.4 mL) at -78 °C under nitrogen. The mixture was brought to
room temperature and stirred for 15 h. The reaction was quenched
by addition of water. The aqueous phase was extracted with
dichloromethane, and the combined organic phase was washed with
brine, dried over MgSO4, and evaporated to give the title compound
(97 mg, quantitive) as a pale viscous oil: 1H NMR δ 2.88 (1H,
ddd, J ) 16.9, 7.4, 4.1 Hz), 2.78 (1H, ddd, J ) 16.9, 8.9, 4.3 Hz),
Supporting Information Available: General synthetic details
and 1H spectra of compounds 19, 22, and 23. This material is
available free of charge via the Internet at http://pubs.acs.org.
JO052133E
1038 J. Org. Chem., Vol. 71, No. 3, 2006