Kinetics and mechanism of the nucleophilic substitution reaction of imidazole with bis(2,4,6-trichlorophenyl) oxalate and bis(2,4-dinitrophenyl) oxalate

Article abstract of DOI:10.1021/jo951627g

The kinetics of the imidazole-catalyzed decomposition of bis(2,4,6-trichlorophenyl) oxalate (TCPO) and bis(2,4-dinitrophenyl) oxalate (DNPO) was investigated by the stopped-flow technique. Pseudo-first-order rate constants were determined as a function imidazole concentration in the temperature range 6-45 °C by fitting the temporal changes in absorbance throughout the 245 to 345 nm wavelength range for TCPO and at 420 nm for DNPO. The reaction proceeds by release of two molecules of substituted phenol and formation of 1,1′-oxalyldiimidazole (ODI) for both esters. The identity of ODI was confirmed in the reaction of imidazole with TCPO by its UV absorbance spectrum and ¹³C-NMR spectrum. The reaction of imidazole with TCPO has a second-order dependence on imidazole concentration and an observed negative activation energy of -6.2 ± 0.3 kJ/mol, whereas the DNPO reaction has a first-order dependence on imidazole concentration and an observed positive activation energy of 12.0 ± 0.6 kJ/mol. The differences in the temperature dependence and order of the reaction with respect to imidazole for the two oxalate esters are explained by a shift in the rate-determining step from addition to the acyl group for DNPO to imidazole-catalyzed release of the phenol leaving group for TCPO. These kinetics results are useful in interpreting the initial reaction steps in peroxyoxalate chemiluminescence.

Full text of DOI:10.1021/jo951627g

J . Org. Chem. 1996, 61, 2657-2663
2657
Kin etics a n d Mech a n ism of th e Nu cleop h ilic Su bstitu tion Rea ction
of Im id a zole w ith Bis(2,4,6-tr ich lor op h en yl) Oxa la te a n d
Bis(2,4-d in itr op h en yl) Oxa la te
Andrew G. Hadd and J ohn W. Birks*
Department of Chemistry and Biochemistry and Cooperative Institute for Research in
Environmental Science (CIRES), University of Colorado, Boulder, Colorado 80309-0216
Received September 6, 1995X
The kinetics of the imidazole-catalyzed decomposition of bis(2,4,6-trichlorophenyl) oxalate (TCPO)
and bis(2,4-dinitrophenyl) oxalate (DNPO) was investigated by the stopped-flow technique. Pseudo-
first-order rate constants were determined as a function imidazole concentration in the temperature
range 6-45 °C by fitting the temporal changes in absorbance throughout the 245 to 345 nm
wavelength range for TCPO and at 420 nm for DNPO. The reaction proceeds by release of two
molecules of substituted phenol and formation of 1,1′-oxalyldiimidazole (ODI) for both esters. The
identity of ODI was confirmed in the reaction of imidazole with TCPO by its UV absorbance spectrum
and 13C-NMR spectrum. The reaction of imidazole with TCPO has a second-order dependence on
imidazole concentration and an observed negative activation energy of -6.2 ( 0.3 kJ /mol, whereas
the DNPO reaction has a first-order dependence on imidazole concentration and an observed positive
activation energy of 12.0 ( 0.6 kJ /mol. The differences in the temperature dependence and order
of the reaction with respect to imidazole for the two oxalate esters are explained by a shift in the
rate-determining step from addition to the acyl group for DNPO to imidazole-catalyzed release of
the phenol leaving group for TCPO. These kinetics results are useful in interpreting the initial
reaction steps in peroxyoxalate chemiluminescence.
In tr od u ction
shown to have a first- and second-order dependence on
the imidazole concentration, and this effect has been
attributed to concurrent “nucleophilic” and “general-base”
catalysis by imidazole.11 Additionally, Alvarez et al. have
suggested that the nature and concentration of the
catalyst affects the formation of different intermediates
in the PO-CL reaction.8 However, the catalytic role of
imidazole and its effect on the formation of intermediates
in the PO-CL reaction has not been well characterized.
In peroxyoxalate chemiluminescence (PO-CL), an aryl
oxalate ester reacts with hydrogen peroxide in the
presence of a fluorophore to generate light. The PO-CL
reaction has become a widely used analytical tool for
trace analysis,1,2 and in a number of cases a 10- to 100-
fold increase in sensitivity over conventional fluorescence
detection has been demonstrated.3-5 Several authors
have investigated the kinetics and mechanism of the PO-
CL reaction.6-11 Although several features of the mech-
anism have been explored, the key pathway(s) and nature
of the reactive intermediate(s) remain highly uncertain.
We are exploring the reaction kinetics and mechanism
of peroxyoxalate chemiluminescence in order to evaluate
the possibility of detecting single molecules. This paper
explores the use of the common PO-CL catalyst, imida-
zole, and its effect on the rate-determining steps of the
light-generating reaction.
The imidazole-catalyzed hydrolysis of esters bearing
good leaving groups is known to proceed by a nucleo-
philic-catalysis pathway.12-14 For example, the interme-
diate formation of N-acetylimidazole has been observed
for the imidazole-catalyzed hydrolysis of p-nitrophenyl
acetate.15 Recently, Neuvonen has examined the neutral
and imidazole-catalyzed hydrolysis of bis(4-nitrophenyl)
oxalate (4-NPO).16 The imidazole-catalyzed degradation
of 4-NPO was found to proceed by the successive release
of two 4-nitrophenyl groups with rapid formation of 1,1′-
oxalyldiimidazole (ODI). Neuvonen suggested that ODI
is the dominant intermediate in the PO-CL reaction.
The peroxyoxalate chemiluminescence reaction is ac-
celerated by the presence of a weak base, with imidazole
(ImH) having the greatest effect on the chemilumines-
cence yield and kinetics.10 The PO-CL reaction has been
ODI itself has been used for the chemiluminescence
detection of hydrogen peroxide in the presence of a
fluorophore.17 The uncatalyzed reaction of ODI with
hydrogen peroxide was found to have faster kinetics but
similar quantum yields when compared to the imidazole-
catalyzed reaction of bis(2,4,6-trichlorophenyl) oxalate
(TCPO) with hydrogen peroxide. In spite of these studies,
the intermediacy of ODI in peroxyoxalate chemilumines-
X Abstract published in Advance ACS Abstracts, April 1, 1996.
(1) Hadd, A. G.; Birks, J . W. in Peroxyoxalate Chemiluminescence:
Mechanism and Analytical Detection; Sievers, R. E., Ed.; J ohn Wiley
and Sons: New York, 1995; pp 209-239.
(2) Robards, K.; Worsfold, P. J . Anal. Chim. Acta 1992, 266, 147-
173.
(3) Kobayashi, S.; Imai, K. Anal. Chem. 1980, 52, 424.
(4) Sigvardson, K. W.; Birks, J . W. Anal. Chem. 1983, 55, 432-435.
(5) Mellbin, G.; Smith, B. E. F. J . Chromatogr. 1984, 312, 203.
(6) Rauhut, M. M. Acc. Chem. Res. 1969, 2, 80-87.
(7) Catherall, C. L.; Palmer, T. F.; Cundall, R. B. J . Chem. Soc.,
Faraday Trans. 2 1984, 80, 837-849.
(8) Alvarez, F. J .; Parekh, N. J .; Matuszewski, B.; Givens, R. S.;
Higuchi, T.; Schowen, R. L. J . Am. Chem. Soc. 1986, 108, 6435-6437.
(9) Milofsky, R. E.; Birks, J . W. J . Am. Chem. Soc. 1991, 113, 9715-
9723.
(10) Hanaoka, N.; Givens, R. S.; Schowen, R. L.; Kuwana, T. Anal.
Chem. 1988, 60, 2193-2197.
(11) Orlovic, M.; Schowen, R. L.; Givens, R. S.; Alvarez, F.; Ma-
tusweski, B.; Parekh, N. J . Org. Chem. 1989, 54, 3605.
(12) Bruice, T. C.; Schmir, G. L. J . Am. Chem. Soc. 1956, 76, 1663-
1667.
(13) Kirsch, J . F.; J encks, W. P. J . Am. Chem. Soc. 1984, 86, 833-
837.
(14) Neuvonen, H. J . Chem. Soc., Perkin Trans 2 1995, 951.
(15) Bender, M. L.; Turnquest, B. W. J . Am. Chem. Soc. 1957, 79,
1652-1655.
(16) Neuvonen, H. J . Chem. Soc. Perkin Trans. 2 1995, 945.
(17) Stigbrand, M.; Ponten, E.; Irgum, K. Anal. Chem. 1994, 66,
1766.
0022-3263/96/1961-2657$12.00/0 © 1996 American Chemical Society

2658 J . Org. Chem., Vol. 61, No. 8, 1996
Hadd and Birks
CDCl3. After 5 min, the solvent was partially evaporated
under vacuum. NMR spectra were recorded over a 2-h period.
cence has not been fully established. Since single mol-
ecule detection would require the production of high
concentrations of the reactive intermediate(s) responsible
for the energy transfer to the analyte molecule, clarifying
the identity and relative contributions of different inter-
mediates is important.
In addition to the formation of ODI as a possible
intermediate in the PO-CL reaction, the two oxalate
esters TCPO and DNPO undergo very different hydroly-
sis reactions. The hydrolysis of DNPO results in the
rapid decarbonylation and decarboxylation of the aryl
hydrogen oxalate without formation of oxalic acid.18-20
Although the much faster kinetics of DNPO and different
hydrolysis mechanism has been explored, further work
comparing common oxalate esters could provide ad-
ditional insight necessary for developing a general per-
oxyoxalate chemiluminescence mechanism.
We investigated the isolated reactions of TCPO and
DNPO with imidazole in order to better understand the
first of several key steps leading to chemiluminescence.
The kinetics of the reaction was studied using the
stopped-flow technique to monitor temporal changes in
the ultraviolet (UV) absorption spectrum. Reaction
conditions were chosen with imidazole in large excess
over each ester, and pseudo-first-order rate constants
were obtained as a function of imidazole concentration
and temperature. 13C-NMR was used to aid in identify-
ing the intermediate formed in the reaction of TCPO.
Resu lts
Effect of Im id a zole Con cen tr a tion on th e Sp ectr a
of 2,4,6-Tr ich lor op h en ol a n d 2,4-Din it r op h en ol.
Deprotonation of phenols results in a characteristic shift
in the absorbance spectrum to longer wavelengths. In
order to investigate the ability of imidazole to deprotonate
each phenol, absorbance spectra were determined in
acetonitrile as a function of added imidazole, as shown
in Figure 1. The spectra show only a small increase in
phenolate absorbance at 320 nm upon addition of imi-
dazole to TCP (Figure 1a). By comparison, in a 50/50 by
volume mixture of water and MeCN, there is a very large
increase in trichlorophenolate absorbance at 320 nm with
increasing imidazole concentration. In the case of DNP,
intense absorption bands grow in at 380 and 425 nm
upon addition of imidazole in neat MeCN (Figure 1b),
indicating a large degree of ionization. From these
results it is concluded that in acetonitrile the relative
+
pKa’s are DNP < ImH2 < TCP.
TCP O Rea ction Kin etics a n d Sp ectr a of Rea ction
P r od u cts a n d In ter m ed ia tes. The reaction of TCPO
with an excess of imidazole was monitored in the 245 to
345 nm wavelength range as a function of imidazole
concentration and temperature. An example of a three-
dimensional time, wavelength, and absorbance plot is
shown in Figure 2. Pseudo-first-order rate constants
were determined using the simplified reaction model
Exp er im en ta l Section
ka
9
Ch em ica ls. Bis(2,4,6-trichlorophenyl) oxalate, imidazole,
triethylamine, 2,4,6-trichlorophenol (TCP), bis(2,4-dinitrophen-
yl) oxalate, 2,4-dinitrophenol (DNP), and 1,1′-oxalyldiimidazole
(technical grade, g90% with principal impurity being imida-
zole) were purchased from Aldrich. Burdick and J ackson
HPLC-grade acetonitrile was used for all dilutions and experi-
ments. Chemicals and solvents were used without further
purification. Stock solutions of 0.10 M imidazole, 2.0 mM
TCPO, and 2.0 mM DNPO were prepared and stored in the
dark prior to dilution and analysis. Fresh DNPO solutions
were prepared every 6 h.
Kin etics Mea su r em en ts. A Hewlett-Packard 8452 diode
array spectrometer was used to determine the UV absorption
spectra of each of the reagents and possible products. Stopped-
flow measurements were made using an Applied Photophysics
DX-17MV stopped-flow spectrophotometer (Applied Photo-
physics, Leatherhead, UK) thermostated by a circulating water
bath. In a typical experiment, one syringe of the stopped-flow
instrument was filled with 0.20 mM TCPO in acetonitrile and
the other syringe filled with 5.0, 10.0, 15.0, or 20.0 mM imi-
dazole, also in acetonitrile. After 1:1 mixing of the reagents,
absorbance versus time data were collected in the range of 245
to 345 nm in 5-nm intervals over 1000 s. Three-dimensional
plots of absorbance, wavelength, and time were analyzed using
a global nonlinear regression fitting program, GLINT, avail-
able from Applied Photophysics. Each data fit utilized a
minimum of 1000 data points at each wavelength. The
kinetics of the reaction of DNPO (0.020 mM, final concentra-
tion) with imidazole (0.5 to 2.0 mM, final concentration) was
determined by following the dinitrophenol absorbance at 420
nm for 0.5 s. All reported rate constants are tabulated using
postmixing reagent concentrations.
A
8 B + 2P
kb
98 C
(1)
B
The experimental data fit this model well and were
used to calculate the spectra of the intermediate B and
phenol product P. The rate constants were derived by
monitoring the array of molar absorbtivity changes of the
calculated spectra over time. The rate constant ka follows
essentially the increase in the phenol absorbance, and
kb the decrease in the absorbance of intermediate B. The
calculated spectrum of P matched a standard spectrum
of 2,4,6-trichlorophenol well within experimental error
at each temperature and imidazole concentration. At-
tempts to model the reaction and follow the kinetics using
the stepwise addition of imidazole were unsuccessful.
Apparently, the second imidazole substitution is very fast
in comparison to the first substitution.
The derived first-order rate constants as a function of
imidazole concentration and temperature are given in
Table 1 for ka and Table II for kb. The rate constant ka
is strongly affected by the imidazole concentration. A
plot of the pseudo-first-order rate constant versus imi-
dazole concentration for a temperature of 24.7 °C is
provided in Figure 3. The rate constants were fit to a
second-order polynomial, resolving the pseudo-first-order
rate constant into zero-, first-, and second-order contribu-
tions according to the equation:
13C-Nu clea r Ma gn etic Reson a n ce Mea su r em en ts (13C-
NMR). A Varian VXR-300S nuclear magnetic resonance
spectrometer was used to obtain 13C-NMR spectra. Imidazole,
0.10 g, was added to 0.05 g of TCPO dissolved in 15 mL of
ka ) a + b[ImH] + c[ImH]2
(2)
The second-order term dominates the reaction kinetics,
and the reaction approximately follows the rate equa-
tion:
ka ) c[ImH]2
(3)
(18) J ennings, R. N.; Capomacchia, A. C. Anal. Chim. Acta 1988,
205, 207.
(19) Orosz, G.; Dudar, E. Anal. Chim. Acta 1991, 247, 141-147.
(20) Neuvonen, H. J . Chem. Soc., Perkin Trans. 2 1994, 89.
An Arrhenius plot of the third-order (first order in
TCPO, second order in ImH) rate coefficient, c (eq. 2), is

Kinetics and Mechanism of Nucleophilic Substitution
J . Org. Chem., Vol. 61, No. 8, 1996 2659
a
F igu r e 2. Example of an absorbance and wavelength versus
time plot obtained from monitoring the reaction of imidazole
with TCPO. Conditions: 5.0 mM ImH and 0.10 mM TCPO at
25 °C.
ences in absolute values between the intermediate and
ODI can be attributed to calculational errors and impuri-
ties in the ODI standard of up to 10% at each wavelength.
A
13C-NMR spectrum of the reaction mixture of TCPO
with an excess of imidazole resulted in the following
resonances that differed from 2,4,6-trichlorophenol and
imidazole: 117.6, 130.3, 139.4, 154.7, 163.3, 163.7 ppm.
The first four signals correspond to the 13C-NMR spec-
trum of an authentic sample of ODI. The remaining two
signals are probably due to subsequent decomposition
products formed in the slow second step of reaction 1 (B
f C).
b
The effect of temperature and imidazole concentration
on the decomposition rate of a commercial sample of ODI
are summarized in Table 3. ODI is known to decompose
into imidazole and carbon monoxide,17 but product iden-
tification was not performed in this study. Qualitatively,
the trend of increasing reaction rate with increasing
temperature and imidazole concentration for the rate
constants given in Tables 2 (kb for the reaction of TCPO
with ImH) and 3 agree. However, the rate constants in
Table 2 are generally twice as fast as for a commercial
sample of ODI. The two reaction systems differ in that
TCP is generated in the reaction of TCPO with ImH but
is not present in the ODI sample.
DNP O Rea ction Kin etics. Table 4 contains the
pseudo-first-order rate constants determined from the
appearance of 2,4-dinitrophenol at 420 nm for the reac-
tion of DNPO with imidazole. Rate constants recorded
in the table were determined using a fit to a single
exponential rise. The pseudo-first-order rate constants
increase linearly with increasing imidazole concentration
at each temperature, as seen in Figure 6. The slopes of
the pseudo-first-order rate constants as a function of
imidazole concentration were calculated using a linear
least-squares fit. The slope corresponds to the second-
order (first order in DNPO, first order in ImH) rate
constant, and an Arrhenius plot for the reaction is
provided as Figure 7. A positive activation energy for
the reaction of DNPO with imidazole of 12.0 ( 0.6 kJ /
mol is obtained.
F igu r e 1. Effect of imidazole concentration on the absorption
spectra of 2,4,6-trichlorophenol (a) and 2,4-dinitrophenol (b)
in acetonitrile.
given in Figure 4 and displays an observed negative
activation energy of -6.2 ( 0.3 kJ /mol.
The absorption spectrum of intermediate B (eq 1) was
calculated from temporal changes in the molar absorp-
tivities over the wavelength range 245 to 345 nm using
the GLINT program. This spectrum is compared with a
standard spectrum of 1,1′-oxalyldiimidazole in Figure 5.
The spectra agree extremely well in shape, and differ-
At 0.5 and 1.0 mM imidazole concentration, an im-
provement to the residuals was achieved using a biex-
ponential fit to the rise, which resolves the stepwise
release of both 2,4-dinitrophenolate ions. However, the

2660 J . Org. Chem., Vol. 61, No. 8, 1996
Hadd and Birks
Ta ble 1. P seu d o-F ir st Or d er Ra te Con sta n t, ka , a s a F u n ction of Im id a zole Con cen tr a tion a n d Tem p er a tu r e for TCP O.a
TCP O F in a l Con cen tr a tion of 0.10 m M
[ImH], mM
temp (°C)
2.5
5.0
7.5
10.0
5.8
15.1
24.7b
35.1
44.7
4.19 ( 0.05 × 10-2
6.19 ( 0.02 × 10-2
6.22 ( 0.01 × 10-2
6.07 ( 0.04 × 10-2
5.42 ( 0.01 × 10-2
0.191 ( 0.003
0.241 ( 0.001
0.201 ( 0.001
0.195 ( 0.002
0.172 ( 0.001
0.463 ( 0.003
0.511 ( 0.002
0.480 ( 0.003
0.455 ( 0.002
0.373 ( 0.004
0.909 ( 0.006
0.928 ( 0.001
0.893 ( 0.002
0.788 ( 0.006
0.725 ( 0.003
Rate constants in units of s-1
.
Additional rate constants shown in Figure 1: 3.5 mM ImH, 0.100 s-1, and 20.0 mM ImH, 3.58 s-1
,
a
b
at 24.7 °C.
Ta ble 2. P seu d o-F ir st-Or d er Ra te Con sta n t, kb, a s a
F u n ction of Im id a zole Con cen tr a tion a n d Tem p er a tu r e
for TCP O.a TCP O F in a l Con cen tr a tion of 0.10 m M
[ImH], mM
temp
(°C)
2.5
5.0
7.5
10.0
5.8 0.60 ( 0.01 0.81 ( 0.05
0.96 ( 0.05
0.91 ( 0.03
1.29 ( 0.01
1.50 ( 0.05
1.67 ( 0.01
15.1 0.52 ( 0.01 0.913 ( 0.001 1.18 ( 0.01
24.7 0.62 ( 0.01 0.782 ( 0.001 1.44 ( 0.03
35.1 0.78 ( 0.03 1.17 ( 0.01
44.7 1.40 ( 0.01 1.60 ( 0.01
1.29 ( 0.01
3.00 ( 0.004 3.20 ( 0.003
a
Rate constants in units of 10-3 s-1
.
two-exponential contribution could not be resolved for
trends within the entire imidazole concentration and
temperature range explored. The data presented in
Table 4 are thus an approximation of the kinetics of the
slowest step, i.e., the release of the second phenol.
Discu ssion
A general mechanism for the nucleophilic reaction of
imidazole with an oxalate ester is shown below, with the
understanding that the same details of the first phenol
substitution also apply to the second substitution.
F igu r e 3. Effect of imidazole concentration on the pseudo-
first-order rate constant (ka) for the reaction of TCPO with
imidazole. Conditions: Final concentrations of 0.10-mM TCPO
and 2.5 to 20.0 mM imidazole at 25 °C.
dazole-catalyzed decomposition of the tetrahedral inter-
mediate), the reaction is second-order in ImH:
(4)
rate ) k1[Ox-OAr][ImH]
k-1 , k2[ImH] (6)
k2[ImH] . k-1 (7)
rate ) k2K1[Ox-OAr][ImH]2
The mechanism begins with a pre-equilibrium step in
which ImH adds to one of the carbonyls to form a
zwitterionic, tetrahedral intermediate. In aprotic sol-
vents the expulsion of the neutral amine is favored over
expulsion of the negative phenoxide ion, making the
reverse of the addition reaction highly favored in the
absence of a base catalyst. A second imidazole molecule
may act as a general-base catalyst by accepting a proton
from the acylimidazolium ion and causing the release of
the phenoxide ion. Applying the steady-state approxima-
tion to the tetrahedral intermediate, the derived rate law
is given by
where Ox-OAr is the oxalate ester and K1 ) k1/k-1
.
The substituents and relative pKa’s of the leaving
groups will affect the relative rates of the three reaction
steps, as quantified by k1, k-1, and k2. An electron-
withdrawing substituent in the leaving group shifts the
equilibrium (k1/k-1) of reaction 4 to the right by promoting
addition of the amine,21 the inductive effect of DNP being
greater than that of TCP. The rate of decomposition of
the tetrahedral intermediate depends on the pKa of the
leaving group. DNP is much more acidic than TCP in
water (pKa’s of 4.08 and 7.42),22,23 dimethyl furan (pKa’s
of 6.4 and 12.4),24 dimethyl sulfoxide (pKa’s of 5.1 and
10.2),24 and propylene carbonate (pKa’s of 14.9 and 18.0).24
The imidizolium ion, ImH2+, has pKa’s intermediate
rate ) k1k2[Ox-OAr][ImH]2/(k-1 + k2[ImH]) (5)
Depending on the values of the rate constants k-1 and
k2, the reaction will be either first- or second-order in
imidazole concentration. For k-1 , k2[ImH] (i.e., rate-
limiting step is addition of ImH to the carbonyl), the
reaction is first-order in ImH, and for k-1 . k2[ImH] (i.e.,
a pre-equilibrium with the rate-limiting step being imi-
(21) Menger, F. M.; Smith, J . H. J . Am. Chem. Soc. 1972, 94,
3824.
(22) Lange’s Handbook of Chemistry, 14th ed.; Dean, J . A., Ed.,
McGraw-Hill: New York, 1992.
(23) Dictionary of Organic Compounds, 5th ed.; Buckingham, J ., Ed.,
Chapman and Hall: New York, 1982; Vol. 5.

Kinetics and Mechanism of Nucleophilic Substitution
J . Org. Chem., Vol. 61, No. 8, 1996 2661
Ta ble 3. Effect of Tem p er a tu r e a n d Im id a zole
Con cen tr a tion on th e Decom p osition Kin etics of a
Com m er cia l Sa m p le of ODI. Con d ition s: 0.10 m M ODI
w ith 2.5 m M Im H a t Ea ch Tem p er a tu r e, a n d a t 25 °C for
Va r ia tion of Im id a zole Con cen tr a tion
temp, °C
kb, 10-3 s-1
[ImH], mM
kb, 10-3 s-1
6.8
15.1
25.0
34.6
44.6
0.20 ( 0.02
0.41 ( 0.01
0.38 ( 0.03
0.45 ( 0.01
0.48 ( 0.02
2.5
5.0
10.0
15.0
0.38 ( 0.03
0.39 ( 0.02
0.59 ( 0.02
0.78 ( 0.01
Ta ble 4. P seu d o-F ir st-Or d er Ra te Con sta n ts a s a
F u n ction of Tem p er a tu r e a n d Im id a zole Con cen tr a tion
for DNP O Mon itor ed a t 420 n m .a Con d ition s: 0.02 m M
DNP O F in a l Con cen tr a tion
[ImH], mM
temp
(°C)
0.05
1.00
1.25
1.50
2.00
6.7 34.4 ( 0.1 68.8 ( 0.2 81.6 ( 0.3 95.0 ( 0.4 127 ( 1
15.0 40.8 ( 0.1 79.9 ( 0.3 95.2 ( 0.1 110 ( 1
139 ( 3
166 ( 2
199 ( 2
230 ( 5
25.1 43.9 ( 0.1 92.5 ( 0.5 110 ( 1
129 ( 2
152 ( 1
178 ( 4
35.0 52.0 ( 0.1 113 ( 0.2
44.6 56.8 ( 0.2 129( 0.1
132 ( 1
151 ( 2
a
Rate constants in units of s-1
.
F igu r e 4. Arrhenius plot for third-order rate coefficient c (eq
2) for the reaction of ImH with TCPO.
F igu r e 6. Effect of imidazole concentration on the pseudo-
first-order rate constant ka for the reaction of DNPO with ImH.
Conditions: Final concentrations of 0.020 mM DNPO and 0.5
to 2.0 mM imidazole at 25 °C.
different rate-determining steps for the two reactions
according to the proposed mechanism. In the case of
DNPO, reaction of the tetrahedral intermediate to form
products, k2[ImH], is fast compared to decomposition to
products, k-1, so that the addition reaction is the rate-
limiting step and the net reaction is first-order in ImH
(eq 6). For TCPO, the elimination reaction is slow
compared to decomposition of the tetrahedral intermedi-
ate, with the result that the reaction is second order in
ImH (eq 7). The key features of the reaction are
presented in Scheme 1. The same detailed mechanism
occurs for the second substitution as for the first, but the
nature of the first substituent affects the rate of the
second step, as discussed below.
F igu r e 5. Absorption spectrum of intermediate B (eq 2)
calculated by the kinetics fitting program GLINT compared
with a standard spectrum of 1,1′-oxalyldiimidazole.
between those of DNP and TCP in water (pKa of 7.0)22
and in DMSO (pKa of 6.3).23 Our spectroscopic results
+
place the pKa of ImH2 intermediate between those of
TCP and DNP; i.e., ImH can abstract a proton from DNP
but not from TCP. Thus, the relative ease of expulsion
of the leaving groups are 2,4-dinitrophenolate, followed
by imidazole, followed by 2,4,6-trichlorophenolate.
The differences in reaction order with respect to ImH
for the DNPO and TCPO reactions can be explained by
The catalytic effect of triethylamine, a non-nucleophilic
base, was used to help confirm the mechanism described
above. In Table 5, the effect of TEA on the pseudo-first-
order rate constants are given for the reactions of DNPO
(24) Acid-Base-Dissociation Constants in Dipolar Aprotic Solvents,
IUPAC Commission on Electroanalytical Chemistry, Chemical Data
Series No. 35, Blackwell Scientific Publications: Oxford, 1990.

2662 J . Org. Chem., Vol. 61, No. 8, 1996
Hadd and Birks
Ta ble 5. Effect of Tr ieth yla m in e Con cen tr a tion on th e
P seu d o-F ir st-Or d er Ra te Con sta n ts in th e Rea ction of
Im id a zole w ith TCP O a n d DNP O. Con d ition s: 0.10 m M
oxa la te ester , 5.0 m M Im id a zole
[TEA], mM
TCPO: ka, s-1
DNPO: ka, s-1
0
0.216 ( 0.005
2.23 ( 0.03
3.26 ( 0.04
5.71 ( 0.02
9.76 ( 0.05
100 ( 3
96.8 ( 0.5
103 ( 1
110 ( 3
118 ( 3
2.50
5.00
10.0
15.0
low or even negative observed activation energies because
of the combination of the temperature dependences of the
pre-equilibrium and rate-determining steps of the reac-
tion, as shown below:
o
o
o
K1 ) e-∆G /RT ) e∆S /R e-∆H /RT
(8)
k2 ) Ae-E /RT
(9)
a
kobs ) k2K1
(10)
o
+ ∆Ho)/RT
kobs ) Ae∆S /R e-(E
(11)
(12)
a
Ea,obs ) Ea + ∆Ho
F igu r e 7. Arrhenius plot for the second-order rate constant
of the reaction of DNPO with imidazole.
The apparent activation energy, Ea,obs, is the sum of
the enthalpy for formation of the imidazole adduct, which
is negative, and the activation energy for the elimination
of the phenol, which is positive. If the enthalpy is more
negative than the activation energy is positive, the
reaction will have an apparent negative activation en-
ergy, as was found for the reaction with TCPO.
Sch em e 1
In the case of DNPO, the addition of ImH (k1) is
expected to be more exothermic than for TCPO so that
the reverse reaction has a larger energy barrier and is
slower. Because DNP is a better leaving group than TCP,
the elimination reaction is expected to be faster for
DNPO, making the addition step rate-determining. For
DNPO, then, the activation energy reflects the barrier
to the addition reaction (eq 6). The proposed reaction
coordinate diagram given in Figure 8 qualitatively il-
lustrates the differences in the potential energy surfaces
for the reactions of TCPO and DNPO with ImH that
result in the observed reaction orders and activation
energies.
Since the proposed mechanism involves a pre-equilib-
rium step, experiments were carried out in which the
corresponding phenol was added to the reaction mixture
in order to determine whether a shift in the position of
the equilibrium would affect the reaction rate in the
predicted manner; i.e., in the simplest picture one would
expect no effect on the DNPO reaction and a decrease in
the rate of the TCPO reaction. For the addition of 0.04
mM DNP to 0.02 mM DNPO and 1.0 mM ImH, the
reaction rate decreased by approximately 20%. In the
reaction of 0.10 mM TCPO with 5.0 mM ImH, the
addition of 0.02 mM TCP increased the rate of reaction
by approximately 50%. The effect of DNP on the DNPO
reaction is quite small (as expected) considering that
twice as much DNP was added as DNPO. This result
does suggest, however, that the reaction may be partially
limited by the establishment of a pre-equilibrium. The
effect of TCP on the TCPO reaction gave the opposite
effect of that predicted and is probably due to the fact
that phenols form strong complexes with their phenox-
and TCPO with imidazole. The results are consistent
with the proposed rate-limiting steps of each reaction.
For the reaction of DNPO with imidazole, the effect of
TEA is quite small, consistent with the rate-limiting step
being addition of ImH to DNPO. For TCPO, however,
TEA has a very large effect on the measured rate
constant, consistent with TEA accepting a proton from
the ImH adduct of TCPO and promoting expulsion of the
trichlorophenolate anion (k2 in eq 4 is accelerated).
This proposed mechanism also explains the differences
in temperature dependence for the two reactions. Reac-
tions involving a pre-equilibrium step (eq 7) often have

Kinetics and Mechanism of Nucleophilic Substitution
J . Org. Chem., Vol. 61, No. 8, 1996 2663
zoates, depending on the nature of the leaving group, was
observed in a study by Menegheli et. al.29 The rate-
determining step for 2,4-dinitrophenyl p-nitrobenzoate
was addition of imidazole to the acyl carbonyl followed
by rapid release of the 2,4-dinitrophenolate ion, as
observed here for DNPO. In contrast, the rate-determin-
ing step for other benzoates was release of the corre-
sponding phenoxide, as found here for TCPO. A similar
dependence of the rate-limiting step on the pKa of the
leaving group also has been observed for other acyl-
transfer reactions.30,31 Kirsch and J encks noted that with
a good leaving group such a p-nitrophenyl acetate, the
uncatalyzed addition of imidazole dominates the kinet-
ics.31 With less acidic phenols, expulsion of the leaving
group becomes more difficult and requires general-base
catalysis to stabilize the zwitterionic intermediate. Neu-
vonen noted both uncatalyzed and general-base catalyzed
nucleophilic substitutions of imidazole in reactions with
4-nitrophenyl acetate, chloroacetate, and dichcloroac-
etate.16 Activation of the acetyl group shifts the reaction
from the catalyzed to the uncatalyzed addition of imida-
zole, again consistent with our observations for TCPO
and DNPO.
F igu r e 8. Qualitative reaction coordinate diagram for the first
substitutions of DNPO and TCPO with imidazole.
ides in aprotic solvents.25 Thus, the phenol can stabilize
the elimination of the phenoxide and speed the decom-
position of the tetrahedral intermediate, again consistent
with the proposed rate-limiting step in the TCPO reaction
with imidazole.
The observations reported here are not without pre-
cedent. A dominant second-order dependence on the
imidazole concentration has been observed for the imi-
dazole-catalyzed hydrolysis of monocarboxylic acid es-
ters.26,27 Negative energies of activation have been
reported for the aminolysis28 and imidazole-catalyzed
hydrolysis28 of esters in aprotic solvents. It was postu-
lated that imidazole acts as a nucleophile to form a
tetrahedral intermediate whose decomposition is cata-
lyzed by another molecule of imidazole by accepting a
proton from the acylimidazolium ion, analogous to our
Scheme 1.
The combination of nucleophilic and general-base
catalysis was confirmed for diaryl esters in a study of
the imidazole-catalyzed hydrolysis of bis(4-nitrophenyl)
oxalate, with formation of ODI having a dominant
second-order dependence on the imidazole concentra-
tion.16 Finally, a similar second-order dependence, with
or without a first-order dependence, on imidazole con-
centration was observed by Orlovic et al. in the imidazole-
catalyzed reaction of hydrogen peroxide with TCPO in
75%/25% MeCN-water.11 Milofsky and Birks observed
a second-order dependence on the imidazole concentra-
tion in the photoinitiated PO-CL reaction carried out in
ethyl acetate.9
A final consideration is the relative rates of the first
and second substitutions of ImH into TCPO and DNPO.
In the case of TCPO, the second substitution appears to
be much faster than the first, while for DNPO the second
substitution is as much as four times slower. This can
be explained if the order of activation by the acyl groups
toward substitution on the adjacent carbonyl is
DNP(CdO) > Im(CdO) > TCP(CdO)
(13)
Thus, substitution of ImH for one TCP in TCPO will
speed the second substitution, while substitution of ImH
for DNP in DNPO will slow the second substitution. This
inequality scheme also corresponds to the relative elec-
tron withdrawing strengths of the acyl groups.
The results from this study are helpful in explaining
the catalytic effects of imidazole on reactions of the two
oxalate esters most commonly used for peroxyoxalate
chemiluminescence. We are currently investigating the
reaction kinetics by observing the chemiluminescence
signals under similar reaction conditions, but with
hydrogen peroxide and a fluorophore added, in order to
further understand the dominant reaction pathways in
the PO-CL reaction.
Ack n ow led gm en t. The stopped-flow instrument
was provided by NSF grant BIR-9302748. We thank
Shelley Copley for several helpful discussions of this
work.
A difference in the rate-determining step for the
imidazole-catalyzed hydrolysis of substituted phenyl ben-
J O951627G
(25) Hadzi, D.; Novak, A.; Gordon, J . E. J . Phys. Chem. 1963, 67,
1118.
(26) Menger, F. M. J . Am. Chem. Soc. 1966, 88, 3081.
(27) Su, C.; Watson, J . W. J . Am. Chem. Soc. 1974, 96, 1854.
(28) Singh, T. D.; Taft, R. W. J . Am. Chem. Soc. 1975, 97, 3867.
(29) Menegheli, P.; Farah, J . P. S.; Seoud, O. A. E. Ber. Bunsenges.
Phys. Chem. 1991, 95, 1610-1615.
(30) Neuvonen, H. J . Chem. Soc., Perkin Trans. 2 1987, 159.
(31) Kirsch, J . F.; J encks, W. P. J . Am. Chem. Soc. 1984, 86, 837.