Halide effects in the hydrolysis reactions of (±)-7β,8α-dihydroxy- 9α, 10α-epoxy-7,8,9,10-tetrahydrobenzo-[α]pyrene

Article abstract of DOI:10.1021/tx9701743

Rates of reaction of (±)-7β,8α-dihydroxy-9α,10α-epoxy-7,8,9,10- tetrahydrobenzo[α]pyrene (DE-2) have been determined in 1:9 dioxane-water solutions containing 1.0 M KC1, 0.5 M KBr, and 0.1 M NaI over the pH range 4- 13. These pH-rate profiles are more complicated than those for reaction of DE-2 in 0.2 M NaC1O₄ solutions and are interpreted in part by mechanisms in which halide ion attacks the diol epoxide as a nucleophile at intermediate pH, resulting in the formation of a trans-halohydrin. Reaction of DE-2 in these halide solutions at pH < ca. 5 occurs by rate-limiting carbocation formation, followed by capture of the intermediate carbocation by halide ion. The relative magnitudes of the rate constants for reaction of the intermediate carbocation with halide ions are estimated from product studies. The halohydrins are unstable intermediates and react quickly in subsequent reactions to yield tetrols in a ratio different than that formed from reaction of the carbocation with solvent. Nucleophilic attacks of 1.0 M CI^- , 0.5 M Br^-, and 0.1 M I^- on DE-2 are the principal reactions in the pH range ca. 6-9, leading to intermediate trans-halohydrins that hydrolyze to tetrols. At pH ca. 9-11, halohydrin formed from attack of halide ion on DE-2 reverts back to epoxide, leading to a negative break in the pH-rate profile. The main product-forming reaction of DE-2 at pH 11.3 is the spontaneous reaction. At pH > 12, the rate of reaction of DE-2 increases due to a second- order reaction of HO^- with DE-2.

Full text of DOI:10.1021/tx9701743

630
Chem. Res. Toxicol. 1998, 11, 630-638
Ha lid e Effects in th e Hyd r olysis Rea ction s of
(()-7â,8r-Dih yd r oxy-9r,10r-ep oxy-7,8,9,10-tetr a h yd r oben zo-
[a ]p yr en e
Bin Lin,† Lanxuan Doan,† Haruhiko Yagi,‡ Donald M. J erina,‡ and
Dale L. Whalen*,†
Department of Chemistry and Biochemistry, University of Maryland, Baltimore County, Baltimore,
Maryland 21250, and Laboratory of Bioorganic Chemistry, NIDDK, National Institutes of Health,
Bethesda, Maryland 20892
Received September 23, 1997
Rates of reaction of (()-7â,8R-dihydroxy-9R,10R-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene
(DE-2) have been determined in 1:9 dioxane-water solutions containing 1.0 M KCl, 0.5 M
KBr, and 0.1 M NaI over the pH range 4-13. These pH-rate profiles are more complicated
than those for reaction of DE-2 in 0.2 M NaClO4 solutions and are interpreted in part by
mechanisms in which halide ion attacks the diol epoxide as a nucleophile at intermediate pH,
resulting in the formation of a trans-halohydrin. Reaction of DE-2 in these halide solutions
at pH < ca. 5 occurs by rate-limiting carbocation formation, followed by capture of the
intermediate carbocation by halide ion. The relative magnitudes of the rate constants for
reaction of the intermediate carbocation with halide ions are estimated from product studies.
The halohydrins are unstable intermediates and react quickly in subsequent reactions to yield
tetrols in a ratio different than that formed from reaction of the carbocation with solvent.
Nucleophilic attacks of 1.0 M Cl-, 0.5 M Br-, and 0.1 M I- on DE-2 are the principal reactions
in the pH range ca. 6-9, leading to intermediate trans-halohydrins that hydrolyze to tetrols.
At pH ca. 9-11, halohydrin formed from attack of halide ion on DE-2 reverts back to epoxide,
leading to a negative break in the pH-rate profile. The main product-forming reaction of
DE-2 at pH 11.3 is the spontaneous reaction. At pH > 12, the rate of reaction of DE-2 increases
due to a second-order reaction of HO- with DE-2.
ion and yields a mixture of tetrols that are the result of
ca. 6% cis and 94% trans hydration.2 This reaction
proceeds via an intermediate carbocation, and thus trans
attack of solvent on the carbocation is favored over cis
attack.2 At pH ca. 7-8, the reaction of DE-2 undergoes
change from hydronium ion-catalyzed hydrolysis to spon-
taneous hydrolysis, which yields ca. 44% cis-tetrol and
56% trans-tetrol (5, 7).
In tr od u ction
The environmental carcinogen benzo[a]pyrene is me-
tabolized in part to a mixture of diastereomeric, bay-
region 7,8-diol 9,10-epoxides (1), one with the benzylic
C-7 hydroxyl group cis to the epoxide group [(+)-DE-1,1
structure not shown] and the second with the benzylic
hydroxyl group trans to the epoxide group [(+)-DE-2].
(+)-DE-2 possesses strong carcinogenic activity and is
thought to be the metabolite responsible for the activity
of the parent hydrocarbon (2).
It has been reported that halide ions catalyze the
formation of tetrol product resulting from cis hydration
of DE-2 at pH 7 in 1:9 acetone-water (8). From the
observation that tetrol product ratios from hydrolysis of
DE-2 in solutions containing increasing chloride ion
concentrations change significantly, although there is
little accompanying change in reaction rate, it was
concluded that chloride ion reacts primarily with an
intermediate carbocation 3, formed in a rate-limiting
step, to yield chlorohydrin 5 (Scheme 1). The stereo-
chemistry of this chlorohydrin is reported to be trans (9).
Subsequent reaction of 5 to yield more cis-tetrol than is
formed in the absence of halide ion would then account
for the increased cis/trans hydration ratios. Direct SN2
attack of iodide and bromide ions, but not chloride ion,
The hydrolysis reactions of DE-1 and DE-2 have
received considerable attention (3-7). At pH < ca. 7 in
water, the hydrolysis of DE-2 is catalyzed by hydronium
† University of Maryland, Baltimore County.
‡ National Institutes of Health.
Abbreviations: DE-1, 7â,8R-dihydroxy-9â,10â-epoxy-7,8,9,10-tet-
1
2
The terms cis and trans are used to describe the relative stereo-
rahydrobenzo[a]pyrene; DE -2, 7â,8R-dihydroxy-9R,10R-epoxy-7,8,9,
10-tetrahydrobenzo[a]pyrene; HEPES, N-(2-hydroxyethyl)piperazine-
N′-2-ethanesulfonic acid; MOPSO, 3-(N-morpholino)-2-hydroxypropane-
sulfonic acid; CHES, 2-(N-cyclohexylamino)ethanesulfonic acid; HPLC,
high-pressure liquid chromatography.
chemistry of the C-8 and C-9 groups and also the stereochemistry of
addition of solvent or external nucleophiles to the epoxide group of
DE-2 such that the resulting hydroxyl group at C-9 and hydroxyl or
other group at C-10 in the product have cis and trans stereochemistries,
respectively.
S0893-228x(97)00174-4 CCC: $15.00 © 1998 American Chemical Society
Published on Web 05/23/1998

Halide Effects in Diol Epoxide Hydrolysis
Chem. Res. Toxicol., Vol. 11, No. 6, 1998 631
Ta ble 1. Su m m a r y of Hyd r olysis P r od u ct Yield s fr om
Sch em e 1
Rea ction of DE-2 in 0.2 M Na ClO4, 1:9 Dioxa n e-Wa ter a
yield of
pHb
cis-tetrol:trans-tetrol
tetrols (%)c
4.00 (4.08)
5.09 (5.73)
8.11 (8.15)d
10.01 (9.82)
11.18 (10.98)
12.0
6:94
6:94
44:56
42:58
36:64
37:63
100
100
86
72
57
53
a
All reactions were allowed to proceed at room temperature
b
for 15 h. The number in parentheses is the pH of the reaction
solution after 15 h. c The percent yield of tetrols from reaction of
DE-2 at pH > 8 is relative to the yield of tetrols formed from the
d
acid-catalyzed hydrolysis of DE-2 at pH < 5.5. HEPES buffer (2
× 10-3 M) was used to maintain pH.
results also show that the reaction of DE-2 at pH > ca.
7 yields halohydrins by a different pathway involving the
bimolecular attack of halide ion on DE-2. This second-
order reaction becomes the major reaction pathway for
reaction of DE-2 at pH ca. 7-10 when [Cl-] > ca. 0.3 M,
[Br-] > ca. 0.03 M, and [I-] > ca. 0.003 M. Mechanisms
of product formation from reaction of DE-2 as functions
of pH and halide ion concentrations are discussed.
at high halide concentrations was indicated by increases
in the rates of reaction of DE-2 in solutions containing
these halide ions.
Exp er im en ta l P r oced u r es
The reactions of DE-2 with dAMP (9), poly(A) (10), and
DNA (11) in solutions containing chloride ion also yield
greater amounts of cis adducts than are formed in the
absence of chloride ion. It was proposed that chloride
ion reacts with the carbocation formed from reaction of
DE-2 with H+ to yield a trans-chlorohydrin, followed by
SN2 attack of DNA on the trans-chlorohydrin to yield cis
adducts. The synthesis of the trans-chlorohydrin and its
reaction with deoxyadenosine to give higher yields of cis
adducts than are formed from reaction of DE-2 with
deoxyadenosine in the absence of chloride ion have
recently been reported (9).
Ma ter ia ls. (()-DE-2 was prepared by published procedures
(3, 16). (Ca u tion : This material is carcinogenic and should
be handled with caution.) Dioxane was distilled from sodium
prior to use. All other reagents were purchased from com-
mercial sources.
Kin etic P r oced u r es. For each kinetic run, approximately
5 µL of a stock solution of DE-2 in dioxane (ca. 1 mg/mL) was
added to 2.0 mL of reaction solution in the thermostated cell
compartment (25.0 ( 0.2 °C) of a UV-vis spectrophotometer.
Reactions were monitored at 348 nm, and pseudo-first-order rate
constants were calculated by nonlinear regression analysis of
the absorbance vs time data. For kinetic runs at intermediate
pH, approximately 10-3 M MES, HEPES, MOPSO, or CHES
buffer was used to maintain pH.
P r od u cts fr om Rea ction s of DE-2 in 1:9 (v/v) Dioxa n e-
Wa ter Solu tion s, 0.2 M Na ClO4. Aliquots (10.0 µL) of DE-2
in dioxane (ca. 0.5 mg/mL) were added to vials containing 2.0
mL of 0.2 M NaClO4 in 1:9 dioxane-water (v/v) whose pH had
been adjusted with either 0.2 M HClO4 in 1:9 dioxane-water
or 0.2 M NaOH in 1:9 dioxane-water. After swirling, the vials
were capped and allowed to stand at room temperature for 15
h (ca. 8 half-lives for the spontaneous reaction). An aliquot (10.0
µL) of 2-(1-naphthyl)ethanol in dioxane was then added to each
vial to serve as an HPLC standard. The pH of each reaction
solution was adjusted to ca. 5-8, and they were analyzed by
HPLC on a reverse-phase C18 column with 60% methanol-40%
water as eluting solvent. Products were monitored by UV
detection at 254 nm. The retention times for the trans-tetrol,
cis-tetrol, and 2-(1-naphthyl)ethanol standard are 8.7, 10.7, and
12.6 min, respectively. The results of these analyses are
summarized in Table 1 and are consistent with earlier product
studies of the hydrolysis of DE-2 in solutions containing only
0.1 M NaClO4, for which only tetrol ratios were determined (5).
Relative yields of tetrol products were calculated by comparing
the area of the tetrol HPLC peaks with that of the standard at
each pH.
The ratio of cis-tetrol:trans-tetrol from reaction of DE-2 at
pH 4.0 and 5.1, where the hydronium ion-catalyzed reaction
predominates, is ca. 6:94. HPLC analyses of the tetrol products
from reaction of DE-2 at pH 4.0 for 10-15 min or for 15 h at
room temperature give identical results. Solutions of tetrol
products showed no detectable change when allowed to stand
for >12 h at pH 2.1 and 12.0 and reanalyzed by HPLC, thus
indicating that no isomerization of the cis- and trans-tetrols had
occurred.
We have previously reported that rates and products
of hydrolysis of indene oxide (12), 9,10-phenanthrene
oxide (13), 7-methyl-substituted benz[a]anthracene 5,6-
oxides (14), styrene oxide, and several vinyl epoxides (15)
are dependent on chloride ion concentrations. These
results were interpreted in terms of a mechanism in
which chloride ion acts as an efficient nucleophile, both
toward reactant epoxides and toward intermediate car-
bocations. To avoid complications associated with the
chemical reactivity of chloride ion, sodium perchlorate,
a nonnucleophilic salt, was used instead of potassium
chloride or sodium chloride to maintain ionic strength
in a study of the hydrolysis reactions of DE-1 and DE-2
(5). We have also determined the rates of reaction of
DE-2 in solutions containing halide salts as a function
of the pH of the solution. Our interest in further
characterizing the kinetic parameters and mechanisms
of reaction of DE-2 in solutions containing halide salts
was stimulated in part by the recent reports on the
mechanisms of reaction of DE-2 in solutions containing
halide ions, in both the absence and presence of DNA
(8-11). We report here detailed rate-pH profiles and
product studies for the reaction of DE-2 in solutions
containing chloride, bromide, and iodide ions. These rate
and product studies confirm reports (8-11) that in the
reaction of DE-2 at sufficiently low pH, halide ions
effectively capture a carbocation intermediate to yield
halohydrins, which then react quickly to yield tetrol
products. Rate constants for the reaction of the carboca-
tion with chloride and bromide ions are estimated. Our

632 Chem. Res. Toxicol., Vol. 11, No. 6, 1998
Lin et al.
The rate profiles in Figure 1 for reaction of DE-2 in
1.0 M KCl, 0.5 M KBr, and 0.1 M NaI solutions are more
complicated than that for reaction of DE-2 in 0.2 M
NaClO4 solutions. These rate profiles show that DE-2
reacts at about the same rate at pH ca. 4-5 in solutions
of all four salts. At slightly higher pH, however, the
profiles for reaction of DE-2 in solutions containing halide
salts level out into plateaus. For example, the rate for
reaction of DE-2 in 1.0 M KCl solutions decreases with
increase of pH until the pH reaches ca. 6.5. The KCl
profile then levels out in a rate plateau, significantly
higher than the profile for 0.2 M NaClO4, that extends
to approximately pH 9. The rate of reaction of DE-2 then
decreases again with increase in pH until it is only
slightly greater than that in 0.2 M NaClO4 solution at
pH 11.3.
F igu r e 1. Plots of log kobsd vs pH for the reaction of DE-2 in
1:9 dioxane-water (v/v) solutions containing 0.1 M NaI, 0.5 M
KBr, 1.0 M KCl, and 0.2 M NaClO4, 25 °C. The curves through
the data points for NaI, KBr, and KCl are theoretical, based on
eq 5 and the kinetic parameters listed in Table 2. The curve for
NaClO4 is theoretical, based on eq 1 and the kinetic parameters
listed in Table 2.3 Concentrations of hydroxide at pH < 12 were
calculated from the equation [HO-] ) KW/[H+], with KW ) 10-14
M2, and were determined directly at pH > 12.
At pH 7, DE-2 reacts 5 times faster in 1.0 M KCl
solution than in 0.2 M NaClO4 solution; at pH 8, DE-2
is 10 times more reactive in 1.0 M KCl solution. The rate
plateaus for reaction of DE-2 in KBr and NaI solutions
are considerably higher than that for reaction in KCl
solution, even though the concentrations of bromide and
iodide ions are one-half and one-tenth, respectively, that
of KCl. At pH > ca. 12, the reactivity of DE-2 increases
due to a second-order reaction of DE-2 with hydroxide
ion.
P r od u cts fr om Rea ction of DE-2 in Solu tion s Con ta in -
in g Ha lid e Ion . Aliquots (10.0 µL) of DE-2 in dioxane (ca. 0.5
mg/mL) were added to vials containing 2.0 mL of 1:9 dioxane-
water (v/v) containing concentrations of NaCl, NaBr, and NaI
varying between 0 and 0.2 M. Ionic strength was kept constant
at 0.2 by addition of NaClO4. The initial pH of chloride,
bromide, and iodide solutions was adjusted with 0.2 M HClO4
in 1:9 dioxane-water to <4.4, <4.2, and <3.5, respectively. After
swirling, the vials were capped and allowed to stand at room
temperature for 20 min. An aliquot (10.0 µL) of 2-(1-naphthyl)-
ethanol in dioxane was then added to each vial to serve as an
HPLC standard. The pH of each reaction solution, which
generally had changed by <0.1 pH unit, was then adjusted to
ca. 5-8. Solutions were analyzed by HPLC on a reverse-phase
C18 column with 60% methanol-40% water as eluting solvent
(cf. Figure 3). The total yields of tetrols from reaction of DE-2
in solutions containing halide ion, based on comparison of tetrol
HPLC peaks with that of standard, were indistinguishable from
those in the absence of halide ion.
Product yields and shapes of the rate profiles for
reaction of DE-2 in solutions containing halide salts are
readily interpreted in terms of the mechanism outlined
in Scheme 2, in which halide ions not only capture a
carbocation intermediate 3 in the acid-catalyzed hydroly-
sis of DE-2 but also attack DE-2 as nucleophiles. As a
result, nucleophilic attack of halide on DE-2 becomes the
principal reaction leading to tetrol product in the pH
range ca. 6.5-10.0. In Scheme 2, kH is the second-order
rate constant for epoxide ring opening of DE-2 with
hydronium ion to yield carbocation 3, ko is the first-order
rate constant for the spontaneous reaction, k1 is the
second-order rate constant for nucleophilic attack of
halide ion on neutral epoxide, Ka is the dissociation
constant for deprotonation of halohydrin 7b, k2 is the
second-order rate constant for reaction of the carbocation
3 with halide ion, k3 is the first-order rate constant for
hydrolysis of halohydrin 7 by a route that does not
involve the free carbocation 3, and kOH is the second-order
rate constant for reaction of hydroxide ion with neutral
epoxide. In the mechanism of Scheme 2, it is assumed
that the spontaneous reaction of DE-2 does not proceed
to any significant extent via carbocation 3, although this
reaction pathway cannot be ruled out as a minor com-
ponent of ko. Profiles similar in shape to those for the
reaction of DE-2 in KCl, KBr, and NaI solutions have
been observed for the reactions of a number of other
epoxides in solutions containing KCl and have been
explained in terms of such a mechanism (12-15).
Hydrolysis of DE-2 at 1:9 dioxane-water with 1.0 M KCl at
pH 8.0, where nucleophilic attack of chloride ion on the epoxide
group to yield chlorohydrin is the principal reaction of DE-2,
yields tetrols resulting from 32-33% cis hydration and 67-68%
trans hydration. This ratio is very similar to that formed from
reaction of DE-2 in 0.2 M NaCl solution at pH 4.
Resu lts a n d Discu ssion
Plots of log kobsd for reaction of DE-2 in solutions
containing 0.2 M NaClO4, 1.0 M KCl, 0.5 M KBr, and
0.1 M NaI over the pH range 4-13 are provided in Figure
1. The rate data for reaction of DE-2 in 0.2 M NaClO4
solutions are accommodated by eq 1, where kH is the
kobsd ) kH[H+] + ko + kOH[HO-]
(1)
The profiles in Figure 1 for KBr, KCl, and NaI possess
a region below pH ca. 6 in which the reaction rate
increases with increase in hydronium ion concentration
(region A), a plateau region between pH ca. 7 and 9
(region B), a region between pH 9 and 11 (region C) where
the rate of reaction of DE-2 decreases until it reaches a
value similar to that for the spontaneous reaction rate
(labeled D in Figure 1), and a region at pH > 12 where
the rate of reaction of DE-2 increases with increasing
hydroxide ion concentration (region E). Mechanisms of
second-order rate constant for the acid-catalyzed reaction,
ko is the first-order rate constant for the spontaneous
reaction, and kOH is the second-order rate constant for
reaction of hydroxide ion with DE-2. These rate con-
stants are summarized in Table 2. The second-order
reaction of hydroxide ion with DE-2 was not observed in
an earlier study of the reactions of DE-2 in 0.1 M NaClO4
solutions (5) because rate data at pH > 11 were not
collected.

Halide Effects in Diol Epoxide Hydrolysis
Chem. Res. Toxicol., Vol. 11, No. 6, 1998 633
a
Ta ble 2. Su m m a r y of Ra te Con sta n ts for Rea ction of DE-2 in 0.1 M Na I, 0.5 M KBr , 1.0 M KCl, a n d 0.2 M Na ClO4
Solu tion s, 1:9 Dioxa n e-Wa ter (v/v), 25 °C
solution
rate constant
kH (M-1 s-1
ko (s-1
kOH (M-1 s-1
k1 (M-1 s-1
k-1Ka/k3 (M)
1.0 M KCl
0.5 M KBr
0.1 M NaI
)
(2.44 ( 0.12) × 103
(1.54 ( 0.11) × 10-4
(5.2 ( 0.5) × 10-3
(1.07 ( 0.06) × 10-3
(2.0 ( 0.3) × 10-10
(2.03 ( 0.17) × 103
(1.72 ( 0.27) × 103
)
(1.44 ( 0.19) × 10-4
0.9 × 10-5
b
)
)
(8.54 ( 0.76) × 10-3
(6.2 ( 1.2) × 10-10
(9.8 ( 0.6) × 10-2
(3.7 ( 0.4) × 10-9
Values of kH, ko, and kOH for reaction of DE-2 in 1:9 dioxane-water, 0.2 M NaClO4, are (1.83 ( 0.10) × 103 M-1 s-1, (1.03 ( 0.04) ×
a
10-4 s-1, and (1.5 ( 0.2) × 10-3 M-1 s-1, respectively. Rate constants for reaction of DE-2 at pH > 11 exhibited greater error than those
b
for reaction of DE-2 at lower pH, possibly due to the instability of ketone formed from the spontaneous reaction.
Sch em e 2
solutions at pH ca. 4 at about the same rate demonstrates
that a second-order reaction of DE-2 with halide ion at
this pH is small relative to the rate of acid-catalyzed ring
opening of DE-2.
Since the rate-limiting step of the hydrolysis reactions
of DE-2 at pH < ca. 4.5, in both the presence and absence
of halide ions, is the formation of carbocation 3 by
reaction of H+ with DE-2, the relative reactivities of
halide ions with 3 cannot be determined by kinetic
measurements. Instead the relative reactivities of halide
ions with 3 must be estimated by studying the products
of reaction of DE-2 as a function of halide ion concentra-
tion. Therefore, tetrol yields from the reaction of DE-2
at pH 3.5-4.5 as functions of varying concentrations of
halide ions were determined. In the absence of halide
ion, DE-2 reacts at this pH via the acid-catalyzed
pathway to yield 6% cis-tetrol and 94% trans-tetrol (5).
water (v/v) solutions at pH 5.00 ( 0.02 as functions of varying
F igu r e 2. Plots of kobsd for the reaction of DE-2 in 1:9 dioxane-
In solutions of increasing halide ion concentrations at this
pH, the reaction of DE-2 gives increasing yields of cis-
tetrol. Plots of the percent cis-tetrol formed from reaction
of DE-2 at pH ca. 4 as functions of halide ion concentra-
tions are given in Figure 3. Since there is no detectable
increase in the rate of reaction of DE-2 in chloride
solutions other than that expected of a normal salt effect,
the change in tetrol product ratio with increasing chloride
concentrations must be attributed to the capture of an
intermediate by chloride ion, after rate-limiting formation
of the intermediate, followed by a secondary reaction of
the chlorohydrin(s) to yield a tetrol mixture of different
composition. This mechanism is also assumed to hold
for the reaction of DE-2 in bromide and iodide solutions
at pH < 4.
NaClO4 and KCl concentrations, 25 °C. The slopes of the NaClO4
and KCl plots are (6.9 ( 1.0) × 10-3 and (5.5 ( 0.6) × 10-3 M-1
s-1, respectively.3
reaction of DE-2 in each rate region will be discussed
separately.
Region A. In this pH range the rate of reaction of
DE-2 is proportional to H+ concentration, and acid-
catalyzed hydrolysis of DE-2 is the predominant reaction.
The reactivity of DE-2 in 1.0 M KCl is only slightly
greater than that in 0.2 M NaClO4 solution. Plots of kobsd
for reaction of DE-2 in KCl and NaClO4 solutions of
varying salt concentrations at pH 5.0 are shown in Figure
2. The slopes of the two plots are very similar, and
therefore the small increase in the rate constant for
reaction of DE-2 in 1.0 M KCl solution compared to that
in 0.2 M NaClO4 solution must be due to a normal salt
effect. The fact that DE-2 reacts in KCl, KBr, and NaI
These results are readily accommodated by the kH[H+]
pathway for reaction of DE-2 in Scheme 2 to yield

634 Chem. Res. Toxicol., Vol. 11, No. 6, 1998
Lin et al.
reacts to form tetrol products via the free carbocation 3,
then there should be no change in the ratio of cis- and
trans-tetrols from reaction of DE-2 with increasing halide
ion concentrations. The increase in yield of cis-tetrol
product observed from reaction of DE-2 in solutions
containing halide ion must therefore be due to the
incursion of a new pathway other than the ks route for
tetrol product formation. The new pathway is labeled
k3 in Schemes 2 and 3.
From Scheme 3, the ratio of products formed from the
ks and k3 pathways is given by the ratio (ks[3])/(k3[7]).
However, without knowing the rate ratio k-2/k3 for
reaction of each halohydrin 7, the partitioning ratio k2/
ks for reaction of 3 cannot be determined from our product
data. However, a lower limit of this ratio can be
estimated by assuming (1) that carbocation 3 partitions
by reacting with solvent (ks) and by reacting with halide
ion (k2) to form halohydrin 7 and (2) that all of the
halohydrin 7 that is formed from the k2 reaction goes on
to tetrol product via the k3 pathway, e.g., k3[7] . ks[3].
From published data for the hydrolysis of trans-chloro-
hydrin 5 in NaCl solutions (9), the percent cis-tetrol
product formed as a function of NaCl concentration
increases somewhat more slowly than the percent cis-
tetrol product from reaction of DE-2 in the same NaCl
solutions at low salt concentrations, indicating that k-2
and k3 may be comparable in magnitude. However, the
equilibrium between 3 and 7 will shift toward 7 in
solutions with increasing halide concentrations, resulting
in lower concentrations of 3, and therefore the second
assumption may hold reasonably well for the reaction of
DE-2 in most of the halide solutions of Figure 3.
F igu r e 3. Plots of percent cis-tetrol in the mixture of cis- and
trans-tetrols from reaction of DE-2 at room temperature in
solutions of varying NaI (pH < 3.5), NaBr (pH < 4.2), and NaCl
(pH < 4.4) concentrations. Ionic strength was kept constant at
0.2 with NaClO4. The curves are theoretical, based on eq 4.
Values of (% cis)∞ are calculated to be 35.3 ( 1.3, 33.6 ( 0.9,
and 34.8 ( 0.2 for KCl, KBr, and KI, respectively. Values of
k2/ks are calculated to be 11.8 ( 1.4, 97 ( 17, and (3.4 ( 0.2) ×
102 M-1 for Cl-, Br-, and I-, respectively.3
Sch em e 3
With the assumption that all of the halohydrin 7
formed as an intermediate in the acid-catalyzed hydroly-
sis of DE-2 in halide solutions from the k2 step goes on
to tetrol product via the k3 pathway, then the mole
fraction of tetrol products formed by the ks route is given
by eq 2. The percent of cis-tetrol in the product mixture
ks
f(ks route) )
(2)
ks + k2[X-]
will be determined by the relative amounts of tetrol
product formed from the ks and k3 pathways and will vary
between 6%, when all of the tetrol products are formed
via the ks route in the absence of halide ion, and ca. 35%,
when all of the tetrol products are formed via the k3 route
at high halide concentrations. The mole fraction of tetrol
product formed by the ks route is given by eq 3. In eq 3,
carbocation 3 as the intermediate that is trapped by
halide ion. Since the carbocation-forming step (kH) is
rate-limiting, in both the presence and absence of halide
ion, it can be concluded that carbocation 3 is captured
by solvent (ks) faster than it returns to DE-2 and H+.
Thus, epoxide ring opening of DE-2 by H+ to yield 3 is
irreversible. At pH < ca. 5, most of the tetrol product
from reaction of DE-2 in 1.0 M KCl, 0.5 M KBr, and 0.1
M NaI solutions is formed from the intermediate halo-
hydrin, formed from capture of carbocation 3 by halide
ion (k2 pathway).
(% cis)∞ - (% cis)x
f(ks route) )
(3)
(% cis)∞ - (% cis)o
(% cis)x is the percent of cis-tetrol formed from reaction
of DE-2 in a solution of a given halide concentration, (%
cis)o is the percent of cis-tetrol formed in the absence of
halide ion (via the ks route), and (% cis)∞ is the percent
of cis-tetrol formed from reaction of DE-2 in solutions of
infinite halide concentration (via the k3 route). Combin-
ing eqs 2 and 3 provides eq 4, which relates the percent
In view of the fact that DE-2 reacts in 1.0 M KCl, 0.5
M KBr, and 0.1 M NaI solutions at pH < ca. 5 to yield
carbocation 3 mainly by the kH route, then the lowest
energy pathway for 3 to return to DE-2 under this
condition is by the reverse of the kH step, according to
the principle of microscopic reversibility (17). Thus,
hydroxide-induced ring closure of 7 (via its conjugate base
6) is precluded as a significant reaction pathway under
these conditions. Therefore, the mechanism of hydrolysis
of DE-2 in solutions at low pH containing halide ion is
given by Scheme 3. If carbocation 3 reacts with solvent
faster than it reacts with halide ion, or if halohydrin 7
(% cis)∞ - (% cis)o
1 + (k2/ks)[X-]
(% cis)x ) (% cis)∞ -
(4)
of cis-tetrol product to the concentration of halide ion.

Halide Effects in Diol Epoxide Hydrolysis
Chem. Res. Toxicol., Vol. 11, No. 6, 1998 635
Nonlinear fitting of the percent cis-tetrol vs [X-] to eq 4
yields a lower limit of the value for k2/ks and a value of
(% cis)∞, the calculated percent of cis-tetrol formed from
reaction of DE-2 in solution with infinite halide concen-
tration.3 Fitting of the data in Figure 3 provides values
(lower limits) for k2/ks of 11.8 ( 1.4, 97 ( 17, and (3.4 (
0.2) × 102 M-1 for reaction of DE-2 in chloride, bromide,
and iodide solutions, respectively. Calculated values of
(% cis)∞ are 35.3 ( 1.3, 33.6 ( 0.9, and 34.8 ( 0.2 as the
percent of cis-tetrol formed from reactions of chlorohy-
drin, bromohydrin, and iodohydrin, respectively, via the
k3 route.
From Figure 3, values of the percent cis-tetrol product
from reaction of DE-2 at pH 4 in solutions containing
[Cl-] > ca. 0.2 M, [Br-] > ca. 0.05 M, and [I-] > ca. 0.02
M are considerably closer to the extrapolated value at
infinite halide concentration than they are to the percent
cis-tetrol product from reaction of DE-2 in the absence
of halide ion (via reaction of 3 with solvent). Several
important conclusions can be drawn from these results.
First, in solutions containing Br- and I- ions in concen-
trations equal to or greater than those concentrations
listed above and Cl- > ca. 0.4-0.5 M, most of the tetrol
product from reaction of DE-2 at pH 4 is formed from
halohydrin via the k3 pathway, and not from the free
carbocation 3 via the ks pathway (k3[7] > ks[3]). If this
were not true, the percent cis-tetrol in the product
mixture from reaction of DE-2 would be much closer to
that observed for reaction of DE-2 in the absence of
halide ion, where the tetrols are formed by reaction of
solvent with 3. Second, carbocation 3 must react with
halide ion present in these concentrations faster than it
reacts with water (k2[X-] . ks). If this were not true,
the percent cis-tetrol in the product mixture would not
change as a function of halide ion concentration as
rapidly as it does.
Ra te Equ a tion . Although halohydrin 7 may not be
a steady-state intermediate in the hydrolysis of DE-2 at
pH < ca. 4, where the rate of the second-order reaction
of DE-2 with H+ will at some pH exceed the rate of the
first-order hydrolysis of 7, it is safe to assume that it will
be a steady-state intermediate in the hydrolysis of DE-2
at pH’s greater than that at which halide ion exerts a
rate effect by nucleophilic attack on DE-2 (pH > ca. 5
for I- and 6 for Cl-). The trans-chlorohydrin 5 is reported
to be considerably more reactive than DE-2 in aqueous
buffer solution (9), and the corresponding bromohydrin
and iodohydrin are expected to be much more reactive
than 5 due to the fact that bromide and iodide are better
leaving groups than chloride ion in solvolysis reactions.
For the reaction of DE-2 in halide solutions under the
conditions of Figure 1, as concluded in the foregoing
discussion, k2[X-] . ks and k3[7] > ks[3]. From these
conditions and the assumption that intermediates 6 and
7b are steady-state intermediates, the rate expression
for reaction of DE-2 is given by eq 5. Weighted nonlinear
values for kH, ko, kOH, k1, and k-1Ka/k3.3 A summary of
these parameters is provided in Table 2.
Region B. For halide solutions having pH values in
this range and containing sufficient concentrations of
halide ion, nucleophilic attack of halide ion on DE-2
becomes the predominant reaction. It is assumed that
the second-order, nucleophilic attack of halide ion occurs
to yield trans-oxyanion 6. In the pH range of region B,
the equilibrium between 6 and its conjugate acid, halo-
hydrin 7b, lies sufficiently in favor of 7b such that all of
6 formed from nucleophilic attack of X- on DE-2 proceeds
on to tetrol product via the k3 route. In this pH range
k1[X-] > kH[H+] and (k-1Ka)/(k3[H+]) , 1, and the first
term of eq 5 reduces to k1[X-]. For 1.0 M KCl, 0.5 M
KBr, and 0.1 M NaI, this kinetic term is greater than
the spontaneous rate constant and gives a rate plateau
in the pH-rate profile that is significantly higher than
that due to the spontaneous reaction. For example, the
rate constant for reaction of DE-2 in 1.0 M KCl solution
at pH 8.5 is 10 times larger than the spontaneous rate
constant for reaction of DE-2 in 0.2 M NaClO4 solution
at the same pH and ca. 7 times larger than that for
reaction of DE-2 at pH 11.2 in the same solvent. Thus,
approximately 90% of the reaction of DE-2 in 1.0 M KCl
solution at pH 8.5 goes by way of nucleophilic addition
of chloride ion to yield chlorohydrin 7b, followed by
solvolysis of chlorohydrin via the k3 route to yield tetrols.
In 0.5 M KBr and 0.1 M NaI solutions at this pH, an
even larger fraction of the reaction of DE-2 hydrolyzes
by this mechanism.
In 0.2 M NaClO4 solution, the mechanism of reaction
of DE-2 changes from acid-catalyzed hydrolysis to spon-
taneous hydrolysis in the pH range ca. 6.5-8.5. Acid-
catalyzed hydrolysis of DE-2 yields only about 6% cis-
tetrol, whereas the spontaneous hydrolysis of DE-2 yields
ca. 44% cis-tetrol (5). Thus, the yield of cis-tetrol
increases as the hydrolysis mechanism changes from
acid-catalyzed to spontaneous, in the absence of halide
ion. At pH 4.5, where the acid-catalyzed reaction pre-
dominates, there is an increase in yield of cis-tetrol with
increasing concentrations of chloride ion, as summarized
in Figure 3. However, at pH 8.5, where the spontaneous
reaction of DE-2 predominates, there is actually a slight
decrease in yield of cis-tetrol brought about by increasing
concentrations of chloride ion, from about 44% in the
absence of chloride ion to about 30% in 1.0 M KCl. Thus,
the presence of halide ion only increases the yields of cis-
tetrol from reaction of DE-2 in that pH range where the
acid-catalzyed hydrolysis of DE-2 predominates.
At pH 7 in the absence of halide ion, DE-2 reacts partly
by acid-catalyzed hydrolysis and partly by the spontane-
ous reaction. From the rate constants kH and ko (Table
2), it is calculated from eq 1 that 64% of DE-2 reacts at
pH 7.0 by the acid-catalyzed route and the remaining
36% by the spontaneous route. Therefore, added halide
ion both captures the intermediate carbocation 3 and
adds as a nucleophile to DE-2. At this pH, DE-2 reacts
5 times faster in 1.0 M KCl solution than in 0.2 M
NaClO4, and thus the rate enhancement due to nucleo-
philic attack of Cl- on DE-2 is approximately 8 times
greater than the rate of acid-catalyzed hydrolysis. There-
fore, at this pH 1.0 M chloride ion reacts mainly as a
nucleophile in attacking DE-2 and to a lesser extent as
a nucleophile in capturing the intermediate carbocation
3 from that portion of the reaction proceeding via acid-
catalyzed hydrolysis. The relative amount of DE-2 that
kH[H+] + k1[X-]
kobsd
)
+ ko + kOH[HO-]
(5)
k-1Ka
1 +
k3[H+]
least-squares fit of the rate data to eq
5
yields
3
Curve fitting of data and graphing were done with PRISM, a
program available from GraphPad Software Co.

636 Chem. Res. Toxicol., Vol. 11, No. 6, 1998
Lin et al.
reacts by each mechanism at pH 7 depends on the halide
ion concentration. Our observation that Cl- acts as an
efficient nucleophile in reacting with DE-2 at pH 7.0 in
1:9 dioxane-water solutions contrasts to an earlier report
that chloride ion does not exert a very large rate effect
on the reaction of DE-2 in 10% acetone-water solutions
at pH 7.0 containing 10 mM sodium cacodylate buffer
(8). This apparent discrepancy may be reconciled if the
differences in conditions between the present work and
earlier studies in aqueous acetone are taken into account.
Part of the difference may be caused by the presence of
10 mM cacodylate buffer in the aqueous acetone solution,
which, like phosphate (6, 7), should be an efficient general
acid catalyst that would give rise to a larger background
rate leading to carbocation 3. Acetone might also act
slightly different than dioxane as a cosolvent, resulting
in a slight change of the relative rates of acid-catalyzed
and spontaneous hydrolysis of DE-2 at pH 7.
Region s C a n d D. Sp on ta n eou s Rea ction of DE-
2. Above pH ca. 9.5, the value of (k-1Ka)/(k3[H+]) becomes
comparable to or greater than unity. This results in a
gradual lowering of the kinetic term in [X-] with increase
in pH until it becomes < ko at pH ca. 11.3. Thus, the
equilibrium (HX + DE-2 h 7) shifts gradually to the left
in favor of epoxide with increase in pH until ko[DE-2] >
k3[7] at pH 11.5, where kobsd becomes approximately equal
to ko. The larger values of kobsd for reaction of DE-2 at
pH 11.3 in 0.5 M KBr and 1.0 M KCl compared to that
in 0.2 M NaClO4 most likely reflect a salt effect on ko.
The spontaneous reaction of DE-2 (ko) is more favor-
able than either acid-catalyzed or hydroxide-catalzyed
reactions in the pH range ca. 7.5-11.5. Therefore, the
principal reactions of DE-2 in halide solutions with pH
8-10 are the spontaneous reaction (ko) and nucleophilic
addition of halide ion (k1). The relative amount of
reaction proceeding via each pathway will depend on the
concentration of halide ion.
The cis-tetrol:trans-tetrol ratio from reaction of DE-2
at pH 8-10 in 0.2 M NaClO4 solutions, where the
spontaneous reaction predominates, is approximately 44:
56. The yield of tetrol products from the spontaneous
reaction of DE-2 in this pH region, however, is found to
be only 74-86% of the yield of tetrol products from the
acid-catalyzed reaction at pH 4-5. It has been reported
that a ketone product [(()-7â,8R-dihydroxy-9-keto-7,8,9,
10-tetrahydrobenzo[a]pyrene] formed in the spontaneous
reaction of DE-1 is unstable in water solutions and
further reacts to form products that cannot be detected
by HPLC (5, 7). A possible explanation for the lower
yields of tetrol products from the spontaneous reaction
of DE-2 is that some of this same ketone (perhaps 10-
20%) is formed from this reaction, in addition to tetrol
products, but is not stable to reaction conditions. The
yield of tetrols from reaction of DE-2 at pH > 10 appears
to be reduced somewhat further, and the explanation for
this result is not clear.
Region E. In region E, kobsd increases with increase
in hydroxide ion concentration. For aryl- and vinyl-
substituted epoxides, an increase in rate at pH > 12 is
due to nucleophilic addition of HO- to the epoxide group,
leading to trans-diols (15). However, product studies of
the reaction of DE-2 in hydroxide solutions with [HO-]
> 0.01 M indicate that the yields of tetrols become lower
with increasing hydroxide concentrations. Although
there is an increase in the trans-tetrol:cis-tetrol product
ratio with increasing hydroxide concentration, there is
not a good rate-product correlation if it is assumed that
the second-order reaction of DE-2 and hydroxide ion is
solely due to nucleophilic addition of hydroxide leading
to trans-tetrol. Further work is necessary to clarify the
mechanism of reaction of DE-2 in highly basic solutions.
Ster eoch em istr y of Nu cleop h ile Atta ck on Ca r -
boca tion 3. Acid-catalyzed hydrolysis of DE-2 yields
94% trans-tetrol and only 6% cis-tetrol, and this observa-
tion has been rationalized by a mechanism in which there
is energetically favorable trans, axial attack of solvent
on the more stable conformation of carbocation 3 (18, 19).
Reaction of DE-2 at pH 4.7 in 1:9 dioxane-water
containing sodium azide yields a 1:4 mixture of azide
products arising from cis and trans addition of azide ion,
respectively, to 3 (20). Thus, reactions of 3 with water
and azide ion nucleophiles result in the formation of
minor yields of cis products. Therefore, it is very prob-
able that 3 also reacts with halide ion to give some cis-
halohydrin 7a as a minor product, which may not be
easily detected, along with the major trans-halohydrin
7b.
Mech a n ism of Hyd r olysis of Ha loh yd r in s 7. The
acid-catalyzed hydrolysis of DE-2 in solutions containing
sufficient concentrations of halide ion, e.g., 0.2 M Cl-
proceeds completely via intermediate halohydrin(s), which
lead to tetrol products by the k3 pathway. One possible
mechanism for this k3 pathway involves ionization of 7
to a carbocation-halide ion pair, followed by reaction of
solvent with this ion-pair intermediate. A second pos-
sible mechanism involves the ionization of 7 to a car-
bocation that is conformationally different from that
formed in the reaction of DE-2 by the kH route, followed
by reaction of this carbocation with solvent at a rate that
is comparable to or greater than that for its conforma-
tional isomerization. This mechanism is attractive be-
cause it would account for the fact that the ratios of cis-
and trans-tetrol products from reactions of all three
halohydrins via the k3 pathway are almost the same.
In solvolysis reactions leading to relatively stable
carbocations such as 3, ionization occurs first to an
intimate ion pair, then to a solvent-separated ion pair,
and finally to fully separated ions, 3 + X- (21). Tetrol
product can arise from reaction of solvent with each
intermediate along the reaction pathway. Since solvoly-
sis studies of halohydrin 7 in solutions free of external
halide salts have not yet been carried out, the amount of
tetrol product formed from 7 via a freely solvated ion 3
under such conditions is not known. Even if k-2 is
comparable to or greater than k3, capture of 3 by halide
ion in halide solutions lowers the steady-state concentra-
tions of 3 in the reaction of DE-2. Consequently, if there
are other product-forming pathways for reaction of 7, e.g.,
k3, then the relative amount of tetrol product from
reaction of 7 via a freely solvated ion 3 (ks) in solutions
containing halide salts will decrease with increasing
halide concentrations. If hydrolysis of 7 occurs by an ion-
pair mechanism, the relative amount of tetrol forming
at the earlier ion-pair stage in the reaction of 7 will
increase with increasing halide concentration. Reaction
of solvent-separated ion pairs with solvent would also
account for the fact that both inversion and retention at
carbon occurs. More work is needed to clarify the
mechanisms of reaction of these halohydrins.
,
Rela tive Rea ctivities of Ha lid e Ion s w ith DE-2
a n d w ith Ca r boca tion 3. From the k1 values listed in
Table 1, the relative reactivities of Cl-, Br-, and I- toward

Halide Effects in Diol Epoxide Hydrolysis
Chem. Res. Toxicol., Vol. 11, No. 6, 1998 637
bimolecular addition to DE-2 are 1:8:96, respectively.
This order of nucleophilicity for halide ions in water
solution is observed in SN2 reactions (22). In the reaction
of halide ion with carbocation 3, the order of nucleophi-
licity is the same, e.g., I- > Br- > Cl-. However, the
relative reactivities for reaction of Cl-, Br-, and I- with
3 are 1:8:28, respectively. Thus, Br- is ca. 8 times more
reactive than Cl- toward reaction with both DE-2 and
carbocation 3. However, I- is 96 times more reactive
than Cl- toward DE-2 but only 28 times more reactive
than Cl- in reaction with 3. One possible explanation
for this reduced reactivity of I- with 3 relative to that of
Br- is that I- reacts with 3 at the diffusional limit and
this rate constant cannot be exceeded.
reaction of DE-2 in 1:9 dioxane-water containing 1.0 M
KCl, 0.5 M KBr, and 0.1 M NaI in the pH range 4-13
are determined.
Ack n ow led gm en t. This work was funded in part by
a Special Research Initiative Support Award from the
UMBC Designated Research Initiative Fund. Helpful
discussions with Dr. J ane Sayer (National Institutes of
Health) are greatly appreciated.
Refer en ces
(1) Thakker, D. R., Levin, W., Wood, A. W., Conney, A. H., Yagi, H.,
and J erina, D. M. (1984) Stereoselective biotransformation of
polycyclical aromatic hydrocarbons to ultimate carcinogens. In
Drug StereochemistrysAnalytical Methods and Pharmacology
(Wainer, I. W., and Drayer, D. E., Eds.) pp 271-296, Marcel
Dekker, New York.
(2) J erina, D. M., Yagi, H., Thakker, D. R., Sayer, J . M., van Bladeren,
P. J ., Lehr, R. E., Whalen, D. L., Levin, W., Chang, R. L., Wood,
A. W., and Conney, A. H. (1984) Identification of the ultimate
carcinogenic metabolites of the polycyclic aromatic hydrocar-
bons: Bay-region (R,S)-diol-(S,R)-epoxides. In Foreign Compound
Metabolism (Caldwell, J ., and Paulson, G. D., Eds.) pp 257-266,
Taylor and Francis Ltd., London.
Carbocation 3 is also trapped in aqueous solution by
azide ion, and the kaz/ks ratio is observed to be 2.5-3.0
× 102 M-1 (23). The kaz/ks ratios for activation-limited
-
attack of N3 on stable carbocations (24, 25) are orders
of magnitude greater than that observed for reaction of
-
3 with N3 (23). Consequently, it has been argued by
J encks and co-workers that low values of kaz/ks for
reaction of certain carbocations indicate that the reaction
of N3- with the carbocation occurs at the diffusional limit,
whereas reaction of the carbocation with solvent (ks) is
activation-limited (26). The kaz/ks ratio for reaction of 3
with I- is larger than that for reaction of 3 with N3-, so
if N3- reacts with 3 at the diffusional limit, then I- most
likely does also. If the diffusion-limited rate constant is
assumed to be 5 × 109 M-1 s-1 (26) and kaz assigned this
value, then k2 for reaction of halide ions with 3 is
calculated to be g(2.4 × 108) M-1 s-1 for Cl-, g(1.8 ×
109) M-1 s-1 for Br-, and g(6 × 109) M-1 s-1 for I-.
(3) Yagi, H., Thakker, D. R., Hernandez, O., Koreeda, M., and J erina,
D. M. (1977) Synthesis and reactions of the highly mutagenic 7,8-
diol 9,10-epoxides of the carcinogen benzo[a]pyrene. J . Am. Chem.
Soc. 99, 1604-1611.
(4) Keller, J . W., Heidelberger, C., Beland, F. A., and Harvey, R. G.
(1976) Hydrolysis of syn- and anti-benzo[a]pyrene diol epoxides:
stereochemistry, kinetics and the effect of an intramolecular
hydrogen bond on the rate of syn-diol epoxide solvolysis. J . Am.
Chem. Soc. 98, 8276-8277.
(5) Whalen, D. L., Montemarano, J . A., Thakker, D. R., Yagi, H., and
J erina, D. M. (1977) Changes of mechanisms and product
distributions in the hydrolysis of benzo[a]pyrene-7,8-diol 9,10-
epoxide metabolites induced by changes in pH. J . Am. Chem. Soc.
99, 5522-5524.
(6) Whalen, D. L., Ross, A. M., Montemarano, J . A., Thakker, D. R.,
Yagi, H., and J erina, D. M. (1979) General acid catalysis in the
hydrolysis of benzo[a]pyrene 7,8-diol 9,10-epoxides. J . Am. Chem.
Soc. 101, 5086-5088.
Su m m a r y
(7) Islam, N. B., Gupta, S. C., Yagi, H., J erina, D. M., and Whalen,
D. L. (1990) Trapping of a carbocationic intermediate in the
spontaneous hydrolysis reaction of 7â,8R-dihydroxy-9â,10â-
epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene: mechanism of the spon-
taneous and general acid-catalyzed hydrolysis reaction of bay-
region benzo[a]pyrene 7,8-diol9,10-epoxides. J . Am. Chem. Soc.
112, 6363-6369.
(8) Wolfe, A. R., and Meehan, T. (1994) Halide-catalyzed cis product
formation in the hydrolysis of anti-benzo[a]pyrene diol epoxide
and its alkylation of poly(A). Chem. Res. Toxicol. 7, 110-119.
(9) Meehan, T., Wolfe, A. R., Negrete, G. R., and Song, Q. (1997)
Benzo[a]pyrene diol epoxide-DNA cis adduct formation through
a trans chlorohydrin intermediate. Proc. Natl. Acad. Sci. U.S.A.
94, 1749-1754.
(10) Wolfe, A. R., Song, Q., and Meehan, T. (1996) The effect of
temperature on the stereochemistry of adducts and hydrolysis
products derived from anti-benzo[a]pyrene diol epoxide in the
presence and absence of halide ions. Polycyclic Aromat. Hydro-
carbons 10, 203-210.
(11) Wolfe, A. R., Yamamoto, J ., and Meehan, T. (1994) Chloride ions
catalyze the formation of cis adducts in the binding of anti-benzo-
[a]pyrene diol epoxide to nucleic acids. Proc. Natl. Acad. Sci.
U.S.A. 91, 1371-1375.
(12) Whalen, D. L., and Ross, A. M. (1976) Specific effects of chloride
ion on epoxide hydrolysis. The pH-dependence of the rates and
mechanisms for the hydrolysis of indene oxide. J . Am. Chem. Soc.
198, 7859-7861.
(13) Whalen, D. L., Ross, A. M., Dansette, P. M., and J erina, D. M.
(1977) Specific effects of chloride ion in the hydrolysis of a
K-region arene oxide. J . Am. Chem. Soc. 99, 5672-5676.
(14) Nashed, N. T., Balani, S. K., Loncharich, R. J ., Sayer, J . M.,
Shipley, D. Y., Mohan, R. S., Whalen, D. L., and J erina, D. M.
(1991) Solvolysis of K-region arene oxides: substituent effects on
reactions of benz[a]anthracene 5,6-oxide. J . Am. Chem. Soc. 113,
3910-3919.
(15) Ross, A. M., Pohl, T. M., Piazza, K., Thomas, M., Fox, B., and
Whalen, D. L. (1982) Vinyl epoxide hydrolysis reactions. J . Am.
Chem. Soc. 104, 1658-1665.
(16) Yagi, H., Akagi, H., Thakker, D. R., Mah, H. D., Koreeda, M.,
and J erina, D. M. (1977) Absolute stereochemistry of the highly
At pH < ca. 5 in 1.0 M KCl, 0.5 M KBr, and 0.1 M NaI
solutions, DE-2 reacts with hydronium ion in a rate-
determining step to yield carbocation 3, which is ef-
ficiently trapped by halide ions to yield halohydrins.
Relative reactivities of the halide ions with 3 are I-
>
Br- > Cl-, with I- reacting at or near the diffusional limit
with 3. In solutions with these halide concentrations,
the halohydrin intermediates hydrolyze mainly by a route
that does not involve carbocation 3, but rather they
hydrolyze by another mechanism possibly involving
attack of solvent on an ion pair or involving a carbocation
intermediate that is conformationally different from that
formed from reaction of DE-2 and H+. Hydrolysis of the
halohydrin by this pathway yields ca. 35% cis-tetrol and
65% trans-tetrol, a mixture enriched in cis-tetrol com-
pared to that from reaction of solvent with 3. When DE-2
is hydrolyzed in solutions at intermediate pH containing
sufficient halide ion concentrations (e.g., 1.0 M KCl, 0.5
M KBr, and 0.1 M NaI), nucleophilic attack of halide ion
on DE-2 becomes the principal reaction. This reaction
leads to a trans-halohydrin intermediate, which hydro-
lyzes to form a tetrol mixture containing slightly less
tetrol than formed from the spontaneous reaction of DE-2
in the absence of halide ion. Thus, the presence of halide
ion only increases the yields of cis-tetrol from reaction of
DE-2 in that pH range where the acid-catalzyed hydroly-
sis of DE-2 predominates. At pH greater than ca. 8.5-
9.5, the pH-dependent equilibrium DE-2 + H+ + X-
h
7b shifts to the left, resulting in a decrease in the rate of
reaction of DE-2, until at pH ca. 11.2 the rate is equal to
that of the spontaneous reaction. Kinetic parameters for

638 Chem. Res. Toxicol., Vol. 11, No. 6, 1998
Lin et al.
mutagenic 7,8-diol 9,10-epoxides derived from the potent carcino-
gen trans-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene. J . Am. Chem.
Soc. 99, 2358-2359.
(21) Winstein, S., Klinedinst, P. E., J r., and Robinson, G. C. (1961)
Salt effects and ion pairs in solvolysis and related reactions. XXI.
Acetolysis, bromide exchange and the special salt effect. J . Am.
Chem. Soc. 83, 4986-4989.
(22) Swain, C. G., and Scott, C. B. (1953) Quantitative correlation of
relative rates. Comparison of hydroxide ion with other nucleo-
philic reagents toward alkyl halides, esters, epoxides and aryl
halides. J . Am. Chem. Soc. 75, 141-147.
(23) Lin, B., Islam, N., Friedman, S., Yagi, H., J erina, D. M., and
Whalen, D. L. (1998) Change in rate-limiting step in general acid-
catalyzed benzo[a]pyrene diol epoxide hydrolysis. J . Am. Chem.
Soc. 120, 4327-4333.
(17) Carey, F. A., and Sundberg, R. J . (1984) Advanced Organic
Chemistry Part A: Structure and Mechanism, p 176, Plenum
Press, New York.
(18) Gillilan, R. E., Pohl, T. M., and Whalen, D. L. (1982) Substituent
effects on rates and product distributions in the hydrolysis
reactions of naphthalene tetrahydro epoxides. Models for tetrahy-
dro epoxides of polycyclic aromatic hydrocarbons. J . Am. Chem.
Soc. 104, 4481-4482.
(24) Bunton, C. A., and Huang, S. K. (1972) Reactions of the tri-p-
anisylmethyl carbonium ion with nucleophiles. J . Am. Chem. Soc.
94, 3536-3544.
(25) Ritchie, C. D. (1975) Cation-anion combination reactions. XIII.
Correlation of the reactions of nucleophiles with esters. J . Am.
Chem. Soc. 97, 1170-1179.
(26) Richard, J . P., Rothenberg, M. E., and J encks, W. P. (1984)
Formation and stability of ring-substituted 1-phenylethyl car-
bocations. J . Am. Chem. Soc. 106, 1361-1372.
(19) Sayer, J . M., Yagi, H., Lilverton, J . V., Friedman, S. L., Whalen,
D. L., and J erina, D. M. (1982) Conformational effects in the
hydrolyses of rigid benzylic epoxides: implications for diol ep-
oxides of polycyclic hydrocarbons. J . Am. Chem. Soc. 104, 1972-
1978.
(20) Lin, B., Doan, L., Yagi, H., J erina, D. M., and Whalen, D.
Quantitative aspects of the effects of halide and azide ions on the
acid-catalyzed hydrolysis of (()-7â,8R-Dihydroxy-9R,10R-epoxy-
7,8,9,10-tetrahydrobenzo[a]pyrene. Polycyclic Aromat. Hydrocar-
bons, submitted for publication.
TX9701743