A novel substitution process for the preparation of alkoxy-, aryloxy-, and acyloxy-substituted 1,2-dioxetanes

Full text of DOI:10.1021/ja962245a

J. Am. Chem. Soc. 1997, 119, 245-246
245
Scheme 1
A Novel Substitution Process for the Preparation of
Alkoxy-, Aryloxy-, and Acyloxy-Substituted
1,2-Dioxetanes
Hashem Akhavan-Tafti,* Robert A. Eickholt,
Zahra Arghavani, and A. Paul Schaap
Lumigen, Inc., 24485 West Ten Mile Rd.
Southfield, Michigan 48034
Scheme 2
ReceiVed July 2, 1996
Stabilized 1,2-dioxetanes which are enzymatically triggered
to undergo chemiluminescent decomposition are widely used
in numerous applications including immunoassays, gene expres-
sion studies, Western blotting, Southern blotting, DNA sequenc-
ing, and the identification of infectious agents.1 The dioxetane
4-methoxy-4-(3-phosphoryloxyphenyl)spiro[1,2-dioxetane-3,2′-
tricyclo[3.3.1.13,7]decane], disodium salt, and analogous deriva-
tives are chemiluminescent substrates for alkaline phosphatase.
Common structural features of these dioxetanes include an
alkoxy group, a spiroadamantyl group, and a protected aryl oxide
group.
electrophile at sulfur converts the thioalkyl or thioaryl substituent
into a good leaving group which ionizes to a benzylic carbo-
cation. It seems likely that the cationic center is also stabilized
by overlap with a lone pair on oxygen. No products resulting
from carbocationic rearrangements were observed, although the
presence of unidentified minor reaction products prevents us
from definitively excluding their formation. The lack of
significant amounts of rearranged products can be attributed to
the thermodynamic stability of the adamantyl ring system. An
SN2 mechanism would appear unlikely on steric grounds.
Recently, the nucleophilic substitution of a chloride leaving
group on an ozonide has been reported.6 Fluoride-, methoxy-,
and acetoxy-substituted ozonides accompanied by ring-opened
products were formed by an SN1 reaction. It is particularly
significant in the present case that the additional ring strain of
the four-membered ring does not preclude substitution in favor
of ring fragmentation as is the case with epoxides.7
The preparation of these alkoxy-substituted dioxetanes has
generally been accomplished by photooxygenation of the
corresponding vinyl ether.2 Other methods for preparing
dioxetanes with alkoxy substituents from the corresponding vinyl
ethers are known, e.g. electron-transfer oxidation with oxygen
and triarylaminium cation radical salts,3 oxidation by Cr(VI)
or Mo(VI) oxide diperoxides,4 and oxidation with triethylsilyl
hydrotrioxide.5 Nevertheless, there is no general method for
synthesizing dioxetanes incorporating other types of groups in
place of the alkoxy moiety. Synthetic methods are needed to
prepare triggerable dioxetanes bearing groups which would be
adversely affected during the oxidation of the vinyl ether or
1
which would deactivate the vinyl ether toward O2. We now
Vinyl sulfides I (R1 ) C2H5, CH2CF3, or p-C6H4F) can be
prepared by TiCl4-initiated reaction of the corresponding aryl
adamantyl ketone with the requisite mercaptan.8 We have
discovered that vinyl sulfides can also be formed by coupling
adamantanone and a thioester with a low-valent titanium reagent
prepared from TiCl3 and LiAlH4 in the presence of Et3N in THF.
The latter reaction achieves the direct formation of both carbon-
carbon bonds of the vinyl sulfide. Although the formation of
vinyl sulfides by reduction of vinyl sulfones with LiAlH4-
TiCl4,9 and the reductive elimination of â-halo sulfoxides with
report a general method for the preparation of a variety of
alkoxy-, aryloxy-, and acyloxy-substituted dioxetanes from a
common dioxetane intermediate. This process does not require
the preparation of each individual vinyl ether precursor.
The key feature of this method is the replacement of a thiol
group SR1 of an alkylthio- or arylthio-substituted dioxetane by
one of various oxygen nucleophiles (Scheme 1). Oxidation of
vinyl sulfide I with 1O2 at temperatures between 0 and -78 °C
produces the sulfur-substituted dioxetane II. Treatment of II
with a stoichiometric amount of a Lewis acid oxidant such as
N-chlorosuccinimide (NCS), Hg(OAc)2, or H2O2/I2 and an
excess of a nucleophilic oxygen compound leads to formation
of oxygen-substituted dioxetanes III in moderate yields.
The direct replacement of a group on the dioxetane ring with
another group is without precedent in dioxetane chemistry.
Further, no reaction involving the direct introduction of sub-
stituents on a pre-formed dioxetane ring has been reported. This
novel reaction sequence is best explained by an SN1 mechanism
involving a dioxetane carbocation (Scheme 2). Reaction of the
10
Zn-TiCl4 have been reported, neither of these methods
involves the direct formation of the sulfur-substituted C-C
double bond from two carbonyl compounds.
Photooxygenation of vinyl sulfide I in the presence of either
methylene blue or Rose Bengal is readily monitored by thin
layer chromatography (TLC) or 1H NMR spectroscopy by
observing the disappearance of the vinyl sulfide. Additionally,
heating a small portion of the reaction solution leads to easily
detectable chemiluminescence indicating formation of the sulfur-
substituted dioxetane. Treatment of sulfur-substituted dioxe-
tanes II with F- in DMSO produces yellow or orange-red
chemiluminescence. Sulfur-substituted dioxetanes of relatively
lower thermal stability (R1 ) C2H5) are not isolated at this point
but instead reacted at -78 °C with a compound containing a
hydroxyl group or its salt. Sulfur-substituted dioxetanes of
higher thermal stability (R1 ) CH2CF3 and p-C6H4F) can be
isolated first or reacted at 0 to -78 °C. A Lewis acid oxidant
(1) (a) Schaap, A. P. Photochem. Photobiol. 1988, 47S, 50S. (b) Schaap,
A. P.; Akhavan, H.; Romano, L. J. Clin. Chem. 1989, 35, 1863. (c) Beck,
S.; Ko¨ster, H. Anal. Chem. 1990, 62, 2258. (d) Legris, F.; Martel-Pelletier,
J.; Pelletier, J.-P.; Colman, R.; Adam, A. J. Immun. Meth. 1994, 168, 111.
(e) Bronstein, I.; Olesen, C. E. M.; Martin, C. S.; Schneider, G.; Edwards,
B.; Sparks, A.; Voyta, J. C. Bioluminescence and Chemiluminescence
Fundamentals and Applied Aspects; Campbell, A. K., Kricka, L. J., Stanley,
P. E., Eds.; Wiley: Chichester, 1994; pp 269-272.
(2) Schaap, A. P.; Handley, R. S.; Giri, B. P. Tetrahedron Lett. 1987,
935.
(3) (a) Curci, R.; Lopez, L.; Troisi, L.; Rashid, S. M. K.; Schaap, A. P.
Tetrahedron Lett. 1987, 28, 5319. (b) Lopez, L.; Troisi, L.; Mele, G.
Tetrahedron Lett. 1991, 32, 117.
(6) Griesbaum, K.; Schlindwein, K. J. Org. Chem. 1995, 60, 8062.
(7) Keul, H.; Pfeffer, B.; Griesbaum, K. Chem. Ber. 1984, 117, 2193.
(8) Mukaiyama, T.; Saigo, K. Chem. Lett. 1973, 479.
(9) Akgun, E.; Mahmood, K.; Mathis, C. A. J. Chem. Soc., Chem.
Commun. 1994, 6, 761.
(10) Retrakul, V.; Poochaivatananon, P. Tetrahedron Lett. 1983, 24(5),
531.
(4) Curci, R.; Lopez, L.; Troisi, L.; Rashid, S. M. K.; Schaap, A. P.
Tetrahedron Lett. 1988, 29, 3145.
(5) Posner, G. H.; Webb, K. S.; Nelson, W. M.; Kishimoto, T.; Seliger,
H. H. J. Org. Chem. 1989, 54, 3252.
S0002-7863(96)02245-7 CCC: $14.00 © 1997 American Chemical Society

246 J. Am. Chem. Soc., Vol. 119, No. 1, 1997
Table 1. Dioxetanes III Prepared (X ) COt-Bu)
Communications to the Editor
phosphate and other derivatives which can be used in enzyme-
linked assays.
dioxetane
R2
oxidant
yield (%)
The surprising stability of thioalkyl- and thioaryl-substituted
dioxetanes II also deserves comment in view of the known
thermal instability of other sulfur-substituted dioxetanes.11 The
most stable sulfur-substituted dioxetanes previously reported are
derived from 4,5-dialkyl-2,3-dihydrothiophene and decompose
with a half-life of a few minutes at room temperature.12 Even
incorporation of a spiroadamantyl group in other sulfur-
containing dioxetanes has been insufficient to achieve a
significant degree of stabilization. Two such dioxetanes bearing
one and two sulfur substituents, respectively, on the dioxetane
ring have been reported to rapidly and completely decompose
on attempted isolation at room temperature.11a In contrast,
sulfur-substituted dioxetanes used in the present study can be
manipulated at temperatures of -10 to 25 °C. Dioxetanes
bearing a (p-fluorophenyl)thio or (trifluoroethyl)thio group can
be chromatographically purified at room temperature. Reaction
of each of the sulfur-substituted dioxetanes with the triggering
agent 0.1 M n-Bu4NF in DMSO cleaves the O-X bond in
seconds and induces a chemiluminescent decomposition at rates
several orders of magnitude more rapid than thermal decom-
position. The progress of this reaction can be monitored by
eye in a darkened room. In our experience, the reagent n-Bu4-
NF/DMSO is both a strong base and a powerful nucleophile
and cleaves various types of phenol protecting groups including
carboxylic esters, glycosides, and phosphate esters as well as
silyl ethers.
1
2
3
4
5
CH3
NCS
NCS
NCS
NCS
52
21
19
56
52
CH2CF3
CH2CHdCH2
C6H5
COCH3
Hg(OAc)2
(1-1.5 equiv based on I) is added to the sulfur-substituted
dioxetane and the solution maintained at 0 to -78 °C for 10-
30 min. The oxygen nucleophile (typically, 1-1.5 equiv based
on I) is then added at low temperature and the reaction mixture
warmed to room temperature. Alternately, the oxygen nucleo-
phile may be present during the photooxygenation. Dioxetanes
III were purified by silica gel chromatography.
The generality of the substitution process appears to be limited
only by the availability of the oxygen nucleophile. Pivalate-
protected dioxetanes 1-5 which have been prepared from the
corresponding pivalate-protected ethylthio vinyl sulfide via the
corresponding sulfur-substituted dioxetane 6 (II, X ) COt-Bu,
R1 ) C2H5) illustrate the scope of the reaction as shown in
Table 1. Yields reported are after isolation and chromatographic
purification and are unoptimized. Numerous other dioxetanes
have been prepared, the details of which will be reported in a
full paper. These include dioxetanes III in which X ) H and
R2 ) alkyl, aryl, acetyl, allyl, substituted alkyl, and polyha-
loalkyl groups. In general, substitution reactions using sulfur-
substituted dioxetanes derived from vinyl sulfides where X )
COt-Bu were somewhat slower than those where the phenol
was unprotected. The rates of substitution were improved by
using a larger excess of the oxygen nucleophiles in those cases.
Since the oxygen-containing group OR2 is not introduced until
after formation of the dioxetane ring, it is possible to prepare
dioxetanes which would be problematic to prepare by photo-
oxygenation of the vinyl ether precursor. This process is
amenable to the preparation of dioxetanes with R2 groups such
A more detailed study of the thermal stability and triggered
decomposition of the sulfur-substituted dioxetanes, including
the unexpected stability of a (trifluoroethyl)thio-substituted
dioxetane in alkaline amine buffers, will be presented in a
subsequent publication. The full scope of the generality of this
nucleophilic substitution method with regard to the electrophilic
reagent, other oxygen nucleophiles, and stabilizing groups is
being explored and will be detailed in a full report of this work.
1
as alkenes and dienes which themselves react with O2 (diox-
Acknowledgment. We thank Dr. Richard S. Handley for assistance
in the preparation of this manuscript and for helpful discussions.
etane 3) or groups which could interfere with the photooxy-
genation either by quenching of singlet oxygen or by quenching
or bleaching the photosensitizer. It is also amenable to the
preparation of dioxetanes with groups which make the vinyl
ether electron-deficient and therefore less reactive or unreactive
to singlet oxygen. An aryloxy-substituted dioxetane 4 and an
acyloxy-substituted dioxetane 5 have been prepared for the first
time by this reaction.
The stable 1,2-dioxetanes 1-5 have long half-lives at room
temperature, but can be triggered by an activating agent to
decompose rapidly with half-lives ranging from seconds to a
few minutes depending on the reaction medium. These diox-
etanes are useful in their own right and as precursors to the
Supporting Information Available: Synthetic procedures and
analytical data (7 pages). See any current masthead page for ordering
and Internet access instructions.
JA962245A
(11) (a) Adam, W.; Encarnacion, L. A. Chem. Ber. 1982, 115(7), 2592.
(b) Ando, W.; Watanabe, K.; Migita, T. J. Chem. Soc., Chem. Commun.
1975, (24), 961. (c) Geller, G.; Foote, C. S.; Pechmann, D. B. Tetrahedron
Lett. 1983, 673. (d) Handley, R. S.; Stern, A. J.; Schaap, A. P. Tetrahedron
Lett. 1985, 26, 3183.
(12) (a) Adam, W.; Griesbeck, A.; Gollnick, K.; Knutzen-Mies, K. J.
Org. Chem. 1988, 53, 1492. (b) Gollnick, K.; Knutzen-Mies, K. J. Org.
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