Laser flash photolysis kinetic studies of α-methoxy-β-phosphatoxyalkyl radical heterolysis reactions: A method for alkoxyalkyl radical cation detection [12]

Full text of DOI:10.1021/ja000763m

6128
J. Am. Chem. Soc. 2000, 122, 6128-6129
Laser Flash Photolysis Kinetic Studies of
r-Methoxy-â-phosphatoxyalkyl Radical Heterolysis
Reactions: A Method for Alkoxyalkyl Radical Cation
Detection
Martin Newcomb,*,† Neil Miranda,† Xianhai Huang,‡ and
David Crich*,‡
Department of Chemistry, Wayne State UniVersity
Detroit, Michigan 48202
Figure 1. (A, B) Signal decay from Chl* (510 nm) and growth from
(Ph3N)+• (650 nm) following 355 nm irradiation of acetonitrile solutions
containing 1 × 10-3 M chloranil and 1 × 10-4 M Ph3N with (A) no
added dihydropyran and (B) 1 × 10-3 M dihydropyran present. The
triphenylaminium cation radical absorbs at 510 nm; this results in a
residual signal at 510 nm in panel A and signal growth at 510 nm
following depletion of Chl* in panel B. (C) Absorbance at 650 nm
following irradiation of a TFE-ACN (5:95, v:v) solution containing PTOC
ester 1b and 1 × 10-4 M Ph3N.
Department of Chemistry, UniVersity of Illinois at Chicago
Chicago, Illinois 60607
ReceiVed March 2, 2000
Nucleotide C4′ radicals are implicated in the cleavage of DNA
by antitumor antibiotics such as bleomycin.1 Under anaerobic
conditions, model C4′ radicals are thought to undergo initial
heterolysis to give a radical cation and a phosphate anion.2 Rapid
proton transfer may then occur to give an allyl radical and
phosphoric acid,2,3 or the radical cation may be trapped by
nucleophiles such as water. Alternatively, a direct pathway to the
allyl radical involving concerted [1,3]-elimination of phosphoric
acid was found computationally by Zipse.4 Schulte-Frohlinde and
co-workers followed the rates of dialkylphosphoric acid formation
from â-phosphatoxyalkyl radicals,5 and Giese and co-workers
observed an allyl radical product from reaction of an R-alkoxy-
â-phosphatoxyalkyl radical by ESR spectroscopy.6 Evidence of
heterolysis of C4′ DNA radicals has been reported by Giese’s
group,7 and Giese, Rist and co-workers observed CIDNP effects
from photolysis of a precursor to an R-alkoxy-â-phosphatoxyalkyl
radical.8
decay period, but the signal for the aminium cation radical
continued to grow after Chl* was depleted (Figure 1B). These
results show that the DHP radical cation produced in acetonitrile
eventually oxidized Ph3N (Scheme 1).
Scheme 1
A serious obstacle for direct studies of reactions of C4′ DNA
radical models is the absence of a prominent UV chromophore
in both the R-alkoxy-â-phosphatoxyalkyl radicals and the products
of heterolysis. We report a probe technique that detects enol ether
radical cations, thus signaling a heterolysis reaction, and permits
kinetic studies. In laser flash photolysis (LFP) studies, we employ
triarylamines as reporters that are oxidized by enol ether radical
cations to triarylaminium cation radicals that are readily detected.
Preliminary LFP studies demonstrated that the method was
viable. Irradiation of chloranil with 355 nm light gives triplet
chloranil (Chl*) which is a powerful oxidant and relatively long-
lived. When Chl* was produced in an acetonitrile (ACN) solution
containing Ph3N, the rate of decay of the signals from Chl* was
comparable to that of growth of the (Ph3N)+• signals indicating
an uncomplicated electron-transfer process (Figure 1A). When
the reaction was repeated with dihydropyran (DHP) present, the
rate of Chl* decay was accelerated due to its reaction with both
DHP and Ph3N. Some (Ph3N)+• was produced during the Chl*
Similar results were found in acetonitrile with (p-BrC6H4)3N
and with both amines in the highly polar solvent 2,2,2-trifluoro-
ethanol (TFE). In aqueous acetonitrile solutions, however, the
Ar3N reporter method cannot be used. Triplet chloranil oxidation
of Ar3N was observed in aqueous ACN solutions as above, but
reactions conducted with added DHP showed no growth of the
(Ph3N)+• signal after Chl* decay was complete. These results
indicate that the DHP radical cation reacted with water rapidly
in a reaction that eventually consumed this oxidant.
Reactions of R-methoxy-â-phosphatoxyalkyl radicals were
studied by the use of the Ar3N reporter method. We used PTOC
esters9 as radical precursors in LFP studies. PTOC esters are
cleaved by 355 nm laser irradiation and have been employed in
studies of â-aryl-â-phosphatoxyalkyl radicals.3,10 The synthetic
sequence to the PTOC esters is described in the Supporting
Information.
The sequence of reactions following 355 nm laser irradiation
of PTOC esters 1 is shown in Scheme 2. Photolyses gave the
pyridine-2-thiyl radical (2) and acyloxyl radicals that rapidly
decarboxylated to give the desired radicals 3. The tert-butyl groups
in radicals 3 preclude proton transfer to give an allyl radical and
phosphoric acid. Radicals 3 reacted by heterolysis to give the
enol ether radical cation 4 and phosphate anion, and radical cation
4 reacted with Ar3N to give the detectable triarylaminium cation
radicals. A control reaction with a simple alkyl radical generated
by photolysis of a PTOC ester resulted in no signal formation
† Wayne State University.
‡ University of Illinois at Chicago.
(1) Stubbe, J.; Kozarich, J. W.; Wu, W.; Vanderwall, D. E. Acc. Chem.
Res. 1996, 29, 322-330.
(2) Beckwith, A. L. J.; Crich, D.; Duggan, P. J.; Yao, Q. W. Chem. ReV.
1997, 97, 3273-3312.
(3) Newcomb, M.; Horner, J. H.; Whitted, P. O.; Crich, D.; Huang, X.;
Yao, Q. W.; Zipse, H. J. Am. Chem. Soc. 1999, 121, 10685-10694.
(4) Zipse, H. J. Am. Chem. Soc. 1997, 119, 2889-2893.
(5) Behrens, G.; Koltzenburg, G.; Ritter, A.; Schulte-Frohlinde, D. Int. J.
Radiat. Biol. 1978, 33, 163-171; Koltzenburg, G.; Behrens, G.; Schulte-
Frohlinde, D. J. Am. Chem. Soc. 1982, 104, 7311-7312; Behrens, G.;
Koltzenburg, G.; Schulte-Frohlinde, D. Z. Naturforsch. C 1982, 37, 1205-
1227.
(9) The acronym PTOC is for pyridine-2-thione-N-oxycarbonyl. See:
Barton, D. H. R.; Crich, D.; Motherwell. W. B. Tetrahedron 1985, 41, 3901-
3924.
(10) Whitted, P. O.; Horner, J. H.; Newcomb, M.; Huang, X.; Crich, D.
Org. Lett. 1999, 1, 153-156. Choi, S. Y.; Crich, D.; Horner, J. H.; Huang,
X.; Newcomb, M.; Whitted, P. O. Tetrahedron 1999, 55, 3317-3326; Choi,
S. Y.; Crich, D.; Horner, J. H.; Huang, X.; Martinez, F. N.; Newcomb, M.;
Wink, D. J.; Yao, Q. W. J. Am. Chem. Soc. 1998, 120, 211-212.
(6) Peukert, S.; Batra, R.; Giese, B. Tetrahedron Lett. 1997, 38, 3507-
3510.
(7) Meggers, E.; Dussy, A.; Scha¨fer, T.; Giese, B. Chem. Eur. J. 2000, 6,
485-492 and references therein.
(8) Gugger, A.; Batra, R.; Rzadek, P.; Rist, G.; Giese, B. J. Am. Chem.
Soc. 1997, 119, 8740-8741.
10.1021/ja000763m CCC: $19.00 © 2000 American Chemical Society
Published on Web 06/08/2000

Communications to the Editor
J. Am. Chem. Soc., Vol. 122, No. 25, 2000 6129
Scheme 2
Scheme 3
In our studies, recombination of the free ions cannot compete
with reaction of radical cation 4 with Ar3N. The measured rate
constants for fragmentation of radicals 3 are the products of rate
constants and equilibrium constants shown in Scheme 3.
We make the reasonable assumption that the diffusive escape
rate constants (kesc) are equal for the diphenyl- and diethylphos-
phate-containing SSIPs in a given solvent. If we also assume that
the equilibrium constant Ksol is the same for both phosphates in
a common solvent, then the faster reaction of radical 3a compared
to that of 3b would reflect either the difference in the equilibrium
constants for the neutral radicals and the CIPs or the difference
in rate constants of the heterolysis reactions that produce the CIPs.
It is noteworthy that, in reactions of â-aryl-â-phosphatoxyalkyl
radicals apparently involving heterolysis and collapse of the CIP,10
[1,2]-migrations of the diphenylphosphate group are about 100
times faster than those of the diethylphosphate group.13,14
This work demonstrates that the Ar3N reporter method can be
applied in LFP studies of DNA radical models as an indicator of
heterolysis and for kinetic measurements, with the caveat that it
cannot be used with aqueous solutions. The observed rate
constants for fragmentations are the products of rate constants
and equilibrium constants (Scheme 3), and factoring out the rate
constants for heterolyses that produce the CIPs will require
additional information. Intramolecular variants of the method
which are under development could address these points and
expand the dynamic range of the technique.
Table 1. Rate Constants for Fragmentations of Radicals 3a
c
radical
solventb
kfrag (s-1
)
kox (M-1 s-1
)
3a
ACNd
(8 ( 1) × 106
(9 ( 1) × 106
>2 × 107
1.2 × 1010
1.2 × 1010
1.7 × 1010
1.7 × 1010
1.6 × 1010
0.7 × 1010
ACNe
5% TFE
ACN
4.5% TFE
5% TFEf
90% TFE
3b
no reaction observed
(2.0 ( 0.2) × 106
(2.7 ( 0.2) × 106
>1 × 107
a Reactions at (23 ( 2) °C conducted with 1-2 × 10-4 M Ph3N
present unless noted. b ACN ) acetonitrile, TFE ) 2,2,2-trifluoro-
ethanol; the volume-% of TFE is listed for mixtures of TFE and ACN.
c Second-order rate constant for electron transfer determined by dividing
observed pseudo-first-order trapping rate constant (ktrap) by amine
concentration. d Average results for reactions with 1 × 10-4, 5 × 10-4
,
and 1 × 10-3 M Ph3N. e (p-BrC6H4)3N used for detection. f Average
results for 1 × 10-4 and 2.5 × 10-4 M Ph3N.
from Ph3N, showing that neither the pyridine-2-thiyl radical (2)
nor the alkyl radical oxidized the amine.
Acknowledgment. This work was supported by grants from the
National Institutes of Health (GM56511 to M.N. and CA60500 to D.C.).
We are grateful to Professor J. H. Horner for experimental assistance in
LFP studies and to Professor M. Schmittel for a helpful discussion.
Figure 1C shows a kinetic trace for formation of (Ph3N)+• from
reaction of 3b; the shape clearly indicates a convolution of rate
constants. Individual rate constants were extracted by fitting the
kinetic traces to the expression for consecutive first-order reac-
tions. The results are listed in Table 1. A precise rate constant
for diphenylphosphate cleavage from 3a could be determined in
ACN. Ph3N and (p-BrC6H4)3N gave the same results, apparently
because oxidations of both of the amines by radical cation 4 are
diffusion-controlled. In solutions containing TFE and ACN,
phosphate fragmentation from 3a was too fast to determine the
rate constants. No signal was observed from reaction of radical
3b in ACN, apparently because this fragmentation was too slow,
but kinetics could be measured in 5% TFE in ACN solution. The
solvent polarity effects are as expected for heterolysis reac-
tions.11,12
Supporting Information Available: Synthetic methods for prepara-
tion of PTOC esters 1 (PDF). This material is available free of charge
via the Internet at http://pubs.acs.org.
JA000763M
(13) The only kinetic data for phosphate fragmentation with which we can
compare our results is for diethylphosphate cleavage from radical 5 in MeOH
to give radical cation 6 where competition kinetic experiments gave a rate
constant of >3 × 109 s-1 at 25 °C.8 Whereas our method reports on the
formation of diffusively free radical cations, the formation of MeOH-trapped
products from radical cation 6 probably occurs in the SSIP and possibly in
the CIP. This conclusion is based on stereoselective alcohol trapping of the
radical cations produced from the diasteromeric radicals 7 where stereochem-
ical information could be maintained in the CIP but would be lost for
diffusively free species.14 Thus, the large difference in the heterolysis rate
constants for 3b and 5 probably reflects equilibrium constants and the rate
constant for diffusive escape from the SSIP that are in the kinetics for 3b but
not in those for 5.
A detailed view of fragmentation of the â-phosphate radicals
3 is shown in Scheme 3. The heterolysis produces a contact ion
pair (CIP) that can collapse back to radical 3 or solvate to give
a solvent-separated ion pair (SSIP). The SSIP equilibrates with
the CIP and also further solvates to give diffusively free species.
(11) The ET(30) solvent polarity values (see ref 12) are as follows: ACN,
45.6; TFE, 59.8; 4.5% TFE in ACN, 55.6 (measured); methanol, 55.4.
(12) Reichardt, C. Chem. ReV. 1994, 94, 2319-2358.
(14) Crich, D.; Gastaldi, S. Tetrahedron Lett. 1998, 39, 9377-9380.