Biosynthesis of tetraponerine-6. Evidence that two different pathways are
operating in the biosynthesis of the two tetraponerine skeletons
C. Devijver,a J. C. Braekman,a D. Dalozea and J. M. Pasteelsb
a Laboratory of Bio-organic Chemistry, Faculty of Sciences, CP160/07, University of Brussels, Av. F. D. Roosevelt, 50, B-1050
Brussels, Belgium
b Laboratory of Animal and Cellular Biology, Faculty of Sciences, CP 160/12, University of Brussels, Av. F. D. Roosevelt, 50,
B-1050 Brussels, Belgium
Evidence is presented showing that tetraponerine-6 2a from
Tetraponera sp. ants is biosynthesized from two molecules of
putrescine and a seven-carbon moiety coming from an eight-
carbon polyacetate chain through decarboxylation, in con-
trast with tetraponerine-8 1a, which derives from one
putrescine unit and a twelve-carbon polyacetate precursor.
be submitted to chemical degradation. Thus, 1000 Tetraponera
sp. ants were starved for three days, then fed with a 1 mol dm23
sucrose solution containing [1,4-14C]putrescine dihydrochlor-
ide (specific activity: 114 Ci mol21; total activity: 50 mCi).
They were killed ten days later by dropping them into methanol.
Using the already described2 extraction and purification
scheme, four of the tetraponerines (T-8 1a, T-4 1b, T-6 2a and
T-5 2c) could be isolated in pure state. All were radioactive.
Their specific activity and specific incorporation rate (SIR) are
reported in Table 1 (experiment 2), together with the values
measured from the first incorporation experiment5 for T-8
(experiment 1). Table 1 shows that the specific incorporation
rates of the four alkaloids isolated from experiment 2 are of the
same order of magnitude as that of T-8 1a from the first
experiment.5 A closer inspection, however, shows that, in
experiment 2, the specific incorporation rates of T-6 2a and T-5
2c are nearly twice that of T-8. Compound T-6 2a (6.50 mg),
which in this experiment was found to be the major alkaloid of
the venom, was then diluted with racemic synthetic4 material
(23.91 mg) as carrier, and divided into two parts, each of which
was separately submitted to a degradation scheme analogous to
that devised for T-85 (Scheme 2). This procedure allowed us to
split the molecule into three moieties:‡ N-methylpyrrolidine 9,
comprising C-4 to C-7, N-tosyl-p-bromophenacylprolinate 10,
comprising C-1 to C-3, C-9 and C-10, and p-bromophen-
acylhexanoate 11, comprising C-8 and C-11 to C-15 of T-6
(Scheme 2). The specific activities and relative specific
activities (RSA) of compounds 9 to 11, and of the two
The tetraponerines are a group of eight tricyclic alkaloids
isolated from the poison gland of the Neo Guinean ant
Tetraponera sp.1 Structural and synthetic work from our
laboratories2–4 showed that these compounds are distributed
into two families, which are based either on the 5-alkyldecahy-
dropyrido[1,2-c]pyrrolo[1A,2A-a]pyrimidine† skeleton 1 (tetra-
ponerines-3, -4, -7 and -8) or on the 5-alkyldecahydrodipyr-
rolo[1,2-a:1A,2A-c]pyrimidine† skeleton 2 (tetraponerines-1, -2,
-5 and -6) (Fig. 1). In each family, the compounds differ from
each other by the length of the alkyl chain (propyl or pentyl) and
(or) by the configuration of the carbon atom to which it is
attached.
Incorporation experiments with Tetraponera sp. ants using
sodium [1-14C]- and [2-14C]-acetate, l -[U-14C]glutamic acid,
l -[U-14C]ornithine hydrochloride, and [1,4-14C]putrescine di-
hydrochloride, followed by chemical degradation of radioactive
tetraponerine-8 (T-8) 1a, the major alkaloid of the venom,
allowed us to propose a biosynthetic pathway for this alkaloid.5
It was found that acetate is a precursor of the entire carbon
skeleton of T-8, but it is incorporated through two different
pathways: the pyrrolidine ring of T-8 comes from acetate by its
conversion, via the tricarboxylic acid cycle, into l -glutamic acid
3, l -ornithine 4 and putrescine 5, whereas the twelve remaining
carbon atoms appear to be derived from a polyacetate precursor,
for which a compound such as 6 is a likely candidate (Scheme
1).
+
H3N
–
HO2C
CO2
H2N
–
CO2
+
NH3
3
4
The presence of alkaloids based on two different, albeit
closely related, skeletons in one and the same organism is
intriguing, particularly with respect to their biosynthesis. We
now report our results from biosynthetic studies conducted on
tetraponerine-6 (T-6) 2a, which show that the two tetraponerine
skeletons 1 and 2 are assembled via two different pathways.
We had to repeat the incorporation experiments with
[1,4-14C]putrescine dihydrochloride, since the sample of T-6 2a
isolated from the first experiment5 decomposed before it could
H2N
H2N
+
1a
O
O
H
6
5
Scheme 1 Biogenetic scheme proposed for tetraponerine-8 1a
Table 1 Incorporation of [1,4-14C] putrescine (50 mCi, 114 Ci mol21) into
H
H
N
H
H
N
the tetraponerines
1
4
10
8
11
9
R
R
1
Experi-
ment
Amount/
mg
TAa/
mCi
102 SAb/
102 SIRc
N
H
N
H
3
Compound
Ci mol21 (%)
6
5
1
2
2
2
2
1a (T-8)
1a (T-8)
2a (T-6)
1b (T-4)
2c (T-5)
6.4
1.01
0.68
1.44
0.12
0.05
4.0
3.0
5.1
3.3
4.9
3.7
2.6
4.5
2.9
4.3
1a 9β-H; R = C5H11 (T-8)
b 9β-H; R = C3H7 (T-4)
c 9α-H; R = C5H11 (T-7)
d 9α-H; R = C3H7 (T-3)
2a 9β-H; R = C5H11 (T-6)
5.70
6.70
0.81
0.22
b 9β-H; R = C3H7
c 9α-H; R = C5H11
d 9α-H; R = C3H7
(T-2)
(T-5)
(T-1)
Fig. 1 Structures of the eight tetraponerines
a Total activity. b Specific activity. c Specific incorporation rate.
Chem. Commun., 1997
661
Devijver
