one electron
oxidation
•(+)
H●
●
CO2R
1
P-450
(
two electron
H●
oxidation)
80% retention
oxygen-18
3
O
O
isomerase
CO2R
7
8
CO2R
oxygen
rebound
(a)
(b)
disproportionation
OH
FeIV–OH
FeIV–OH
FeIII H2O
•(+)
•
•
●H
●H
●
●
H
●H
4
5
oxygen
FeIII
HO
●H
rebound
CO2R
Scheme 4
dehydrogenase
2
●
prior to reduction by a dehydrogenase. Finally, an additional
mechanism would involve a two-electron oxidation of 1 to
generate carbocation 3, as illustrated in the brackets in Scheme
6
Scheme 3
3
. Such a process has some precedent with the accepted
mechanisms of thromboxane and prostacyclin synthases,
1
5
ions. The levels of incorporation were determined by selective
ion monitoring (SIM) and the values in Table 1 are corrected for
natural isotope abundances. There was no significant incorpora-
tion (M + 2 = 0 ± 0.3) into the M + 2 ions and these are not
included in Table 1. Of particular significance for this study is
the M + 1/M + 3 ratio in the molecular ions of both 1 and 2,
which is independent of total incorporation (12–17%). The
extracted littorine 1 on each of the four days had an M + 1/M +
where carbocation intermediates are implied, and in some
recent mechanistic studies16 on P-450 systems, where carboca-
tions gave rise to side products during the hydroxylation of
cyclopropane substrates. As illustrated in Scheme 3, rearrange-
ment to carbocation 4 followed by a direct collapse to aldehyde
6 prior to reduction would not require any loss of oxygen-18.
Some exchange then at the aldehyde level is also consistent with
the experimental result. A fuller discussion of the various
mechanistic possibilities is given elsewhere.17
3
1
= 1.26 ± 0.02, whereas the hyoscyamine had a value of M +
/M + 3 = 2.1 ± 0.07. This change of ratio represents a loss of
We thank the CIBA foundation for an ACE award and the
EPSRC for a Studentship (C. W. W.).
ca. 25–29% of the M + 3 abundance in hyoscyamine relative to
littorine 1 and a corresponding enrichment of the M + 1
abundance of hyoscyamine 2 relative to littorine in going from
Notes and References
1
to 2. Thus there is an exchange of ca. 25–29% of oxygen-18
†
E-mail: david.o’hagan@durham.ac.uk
for oxygen-16 in going from littorine 1 to hyoscyamine 2.
Our working hypothesis for the rearrangement of 1 and 2
involves two enzymatic activities, an iron–oxo isomerase and a
1
R. Robinson, Proceedings of the University of Durham Philosophical
Society, 1927–1932, 8, 14.
dehydrogenase, as illustrated in Scheme 3. This hypothesis
2 R. Robinson, Structural Relations of Natural Products, Clarendon
Press, Oxford, 1955.
3 E. Leete, N. Kowanko and R. A. Newmark, J. Am. Chem. Soc., 1975, 97,
developed from ideas of Sankawa,1
1,12
who rationalised a
number of isomerisation reactions in terms of iron–oxo enzyme
processes. Three mechanistic scenarios emerge in the light of
this result. In general, iron–oxo processes are considered to
6
826.
4
5
6
N. C. J. Chesters, D. O’Hagan and R. J. Robins, J. Chem. Soc., Chem.
Commun., 1995, 127.
R. J. Robins, P. Bachmann and J. G. Woolley, J. Chem. Soc., Perkin
Trans. 1, 1994, 615.
IV
involve radicals and it can be envisaged that a Fe –O· species
9
abstracts hydrogen (the 3A-pro-R hydrogen) to generate a
substrate radical 3. After rearrangement, the product radical 4 is
E. Leete, Can. J. Chem., 1987, 65, 226; E. Leete, J. Am. Chem. Soc.,
quenched in a classical manner by delivery of an hydroxyl
radical from Fe –OH (oxygen rebound) to generate hydrate 5
1
984, 106, 7271.
IV
7 M. Ansarin and J. G. Woolley, J. Chem. Soc., Perkin Trans. 1, 1995,
followed by collapse of the hydrate to give aldehyde 6. The high
retention of oxygen-18 found experimentally is compatible with
this process if (i), a partially stereospecific collapse of the
hydrate occurs under enzymatic control, or (ii) a fully
stereospecific process removes the unlabelled oxygen, but there
is some exchange at the aldehyde prior to reduction by a
dehydrogenase. A non-stereospecific process would of course
result in 50% loss of oxygen-18 and this is not observed.
A similarly high retention of oxygen (80%) was observed
previously13 in the oxidation of 7 to 8 in a P-450 mediated
487.
8 J. R e´ tey and D. Arigoni, Experientia, 1966, 22, 783.
9
N. C. J. E. Chesters, K. Walker, D. O’Hagan and H. G. Floss, J. Am.
Chem. Soc., 1996, 118, 925.
1
1
1
0 R. J. Robins, N. C. J. E. Chestrs, D. O’Hagan, A. J. Parr, N. J. Walton
and J. G. Woolley, J. Chem. Soc., Perkin Trans. 1, 1995, 481.
1 M. F. Hashim, T. Hakamatsuka, Y. Ebizuka and U. Sankawa, FEBS
Lett., 1990, 271, 219.
2 T. Hakamatsuka, M. F. Hashim, Y. Ebizuka and U. Sankawa,
Tetrahedron, 1991, 47, 5969.
13 M. Akhtar, M. R. Calder, D. L. Corina and J. N. Wright, Biochem. J.,
1982, 201, 569.
14 M. Akhtar and J. N. Wright, Nat. Prod. Rep., 1981, 8, 527.
process operating during oestrogen biosynthesis. It was con-
cluded1
3,14
in that case that the observation is most consistent
1
1
1
5 V. Ullrich and R. Brugger, Angew. Chem., Int. Ed., Engl., 1994, 33,
with a disproportionation process [Scheme 4(a)] involving
Fe OH and the product radical, rather than the more common
oxygen rebound [Scheme 4(b)] process. It is poignant that a
similar level of oxygen-18 retention is found in this study and
such a disproportionation must remain under consideration. The
partial loss of isotope can then be attributed to some exchange
of the carbonyl oxygen of aldehyde 6 with the aqueous medium
1
911.
IV
6 M. Newcomb, M.-H. Le Tadic-Biadatti, D. L. Chestney, E. S. Roberts
and P. F. Hollenberg, J. Chem. Soc., 1995, 117, 12091.
7 D. O’Hagan and R. J. Robins, Chem. Soc. Rev., 1998, in the press.
Received in Cambridge, UK, 2nd February, 1998; Revised manuscript
received 2nd March 1998; 8/01722K
1046
Chem. Commun., 1998