pyoverdin,9 melanin,10 and styelsamine11 and in the hydroxy-
kynurenine-mediated cross-linking of proteins in the mam-
malian eye lens.12
Scheme 1
Several experiments were carried out that support this
mechanism and differentiate it from alternatives. When
diaminobenzene 10 is treated with ribose phosphate under
the same conditions used to oxidize 3, DMB (yields of
6-10%) is also formed in an oxygen-dependent reaction.
This supports the intermediacy of 6 (Scheme 3). In addition,
imine 5 may be intermediates and that it may be possible to
find relevant nonenzymatic chemistry for the conversion of
3 to 4. Here, we demonstrate the validity of this analysis
and propose that the conversion of 3 to DMB can occur to
a large extent by a facile nonenzymatic oxidative cascade
(Scheme 2).
Scheme 3
Scheme 2
when this reaction is carried out with D-[13C-1]ribose, NMR
analysis of the resulting DMB demonstrated that the 13C was
localized at the C-2 position. This labeling pattern is identical
to the biosynthetic labeling pattern [1] suggesting that the
biomimetic and the biosynthetic conversion of 6 to DMB
are occurring by the same mechanism.
HPLC analysis of the 10 + 11 reaction mixture (Scheme
3) identified DMB and an additional compound as the major
reaction products (Figure 1). Structure 12 was assigned to
DMB (4) was formed when a synthesized sample of 3
was treated with catalytic amounts of potassium ferricyanide,
a mild oxidizing agent known to oxidize amino phenols to
the corresponding quinone imines. This supports the proposal
that DMB can be formed by the nonenzymatic oxidation of
3.8 Aerobic oxidation of 3 under physiological conditions
(pH 7, 37 °C) also gave DMB. However, the reaction was
approximately 30 times slower than the ferricyanide-mediated
oxidation. DMB was not formed after 24 h in argon-purged
buffer, demonstrating that its formation requires oxygen.
Having established the feasibility of converting 3 to 4,
we next turned our attention to the mechanism of this
complex oxidation process. Our current mechanistic proposal
is outlined in Scheme 2. In this proposal, oxidation of the
electron-rich diamine 3 to the bisimine 5 followed by a
tautomerization gives 6. Cyclization to give 7 followed by a
second two-electron oxidation gives 8. Aromatization of 8
by extrusion of 9 gives 4. Analogous oxidative cascades, in
which electron-rich benzenes undergo complex reactions,
have been proposed in the biosynthesis of actinomycin,8
Figure 1. HPLC analysis of the reaction shown in Scheme 3.
this compound on the basis of its MS, UV, and NMR spectra.
Compound 12 was stable and did not undergo conversion
to DMB (Scheme 4). The detection of 12 provides support
(9) Dorrestein, P. C.; Poole K.; Begley T. P. Org. Lett. 2003, 5, 2215-
2218.
(10) Fatibello-Filho O.; Vieira, I. C. Analyst 1997, 122, 345-350.
(11) Skyler, D.; Heathcock, C. H. Org. Lett. 2001, 3, 4323-4324.
(12) Aquilina, J. A.; Carver, J. A.; Truscott, R. J. W. Biochemistry 2000,
39, 16176-16184.
(7) Singh, M. P.; Sasmal S.; Lu, W.; Chatterjee, M. N. Synthesis 2000,
10, 1380-1390.
(8) Barry, C.; Nayar, P.; Begley, T. P. Biochemistry 1989, 28, 6323.
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Org. Lett., Vol. 5, No. 13, 2003