Tropane alkaloids from Darlingia darlingiana

Article abstract of DOI:10.1016/S0031-9422(99)00263-0

A new γ-pyranotropane, darlingine N-oxide, was isolated from the bark and leaves of Darlingia darlingiana, along with the known compound, darlingine. 1D-NOESY NMR experiments indicated that the N-methyl groups of both alkaloids were orientated towards the pyran ring.

Full text of DOI:10.1016/S0031-9422(99)00263-0

Phytochemistry 52 (1999) 529±531
Tropane alkaloids from Darlingia darlingiana
a
Peter L. Katavic , Mark S. Butler *, Ronald J. Quinn , Paul I. Forster ,
a,
a
b
b
Gordon P. Guymer
a
Queensland Pharmaceutical Research Institute, Grith University, Nathan, 4111 QLD, Australia
Queensland Herbarium, Brisbane Botanic Gardens, Mt. Coot-tha Rd., Toowong, 4066 QLD, Australia
b
Received 9 February 1999; received in revised form 27 April 1999; accepted 27 April 1999
Abstract
A new g-pyranotropane, darlingine N-oxide, was isolated from the bark and leaves of Darlingia darlingiana, along with the
known compound, darlingine. 1D-NOESY NMR experiments indicated that the N-methyl groups of both alkaloids were
orientated towards the pyran ring. # 1999 Elsevier Science Ltd. All rights reserved.
Keywords: Darlingia darlingiana; Proteaceae; Bark; Leaves; Tropane alkaloids; Darlingine; Darlingine N-oxide
1. Introduction
(17 mg). The leaves contained 1 and 2 in a similar
ratio to the bark.
The only proteaceous plants known to produce alka-
The major alkaloid 1 was identi®ed as the known
compound (+)-darlingine (1). Darlingine (1) has been
previously isolated from D. darlingiana (Bick et al.,
1979b), as well as D. ferruginea (Bick, Gillard & Leow,
1979c) and B. montana (Bick et al., 1979a). In 1995
Majewski and Lazny established the absolute stereo-
chemistry of 1 by an enantiomeric synthesis of (� )-dar-
loids are Agastachys odorata, Bellendena montana,
Darlingia darlingiana, D. ferruginea, Knightia deplan-
chei and K. strobilina (Lounasmaa & Tamminen,
1
(
1
993). A small group of these alkaloids, bellendine
Bick, Bremner & Gillard, 1971; Motherwell et al.,
971), isobellendine (Bick, Gillard & Leow, 1979a),
darlingine (1) (Bick, Gillard & Leow, 1979b) and stro-
biline (Lounasmaa, Pusset & Sevenet, 1980), contain a
1
13
lingine. Some of the H and C NMR resonances of 1
reported by Bick et al. (1979c) have been reassigned
using 2D NMR experiments and are given in the ex-
perimental section.
Â
unique g-pyranotropane skeleton. In this report, we
describe the isolation of a new g-pyranotropane alka-
loid, darlingine N-oxide (2), found in addition to dar-
lingine (1) in the bark and leaves of D. darlingiana (F.
Muell.) L.A.S. Johnson.
The spectroscopic data of the minor alkaloid 2 was
similar to that of 1, which suggested these compounds
to be closely related.
A molecular formula of
2. Results and discussion
An alkaloid extract from the bark (100 g) of D. dar-
lingiana was separated by counter current chromatog-
raphy [solvent system CHCl ±MeOH±H O (13:7:8)] to
3
2
give darlingine (1) (74 mg) and darlingine N-oxide (2)
*
Corresponding author.
0031-9422/99/$ - see front matter # 1999 Elsevier Science Ltd. All rights reserved.
PII: S0031-9422(99)00263-0

5
30
P.L. Katavic et al. / Phytochemistry 52 (1999) 529±531
Table 1
6
NMR data of darlingine N-oxide (2) (DMSO-d )
13
1
Position
C
H
gCOSY
gHMBC
2
3
4
5
6
161.6
119.2
174.6
121.4
69.0
4.18 d (6.0)
1.65 m
2.75 m
H-7b
C-4, C-5, C-7, C-8, C-9, C-11, N-CH
C-5, C-6, C-8, C-9
3
7
7
8
8
9
1
1
1
2
3
a
b
a
b
32.9
H-7b, H-8a, b
H-6, H-7a, H-8a, b
H-7a, b, H-8b
H-7a, b, H-8a, H-9
H-8b, H-10b
H-10b
C-5, C-6, C-8, C-9
C-6, C-7, C-9, C-10
C-6, C-7, C-9, C-10
28.5
1.71 m
2.52 m
71.6
35.3
3.65 dd (5.4, 6.0)
2.82 d (18.6)
3.24 dd (5.4, 18.6)
C-6, C-7, C-8, C-10, C-11, N-CH
C-5, C-8, C-9, C-11
3
0a
0b
1
H-9, H-10a
C-5, C-8, C-9, C-11
158.1
17.5
9.4
-CH
-CH
3
2.26 s
1.82 s
3.07 s
C-2, C-3
C-2, C-3, C-4
C-6, C-9
3
N±CH
3
50.1
C H NO , determined by accurate mass measure-
17
HSQC and HMBC) were recorded using standard
pulse sequences. 1D-NOESY as described by Stott,
Stonehouse, Keeler, Hwang and Shaka (1995). Positive
ESI-MS: Fisons VG Platform II. High resolution posi-
tive ESI-MS: Bruker BioAPEX 47e. Counter current:
Sanki Centrifugal Partition Chromatograph Model
LLB-M.
1
3
3
+
ment of the [M+H] mass ion peak (m/z 236.1276) in
the positive ESI mass spectrum, indicated that 2 had
an extra oxygen atom compared to darlingine (1).
Examination of the 1D and 2D NMR data (Table 1)
established that 2 had the same gross structure as 1
and the extra oxygen atom in 2 could only be present
as an N-oxide. This was con®rmed by a zinc mediated
reduction of 2 to give (+)-darlingine.
3
.1. Plant material
1D-NOESY NMR experiments were used to estab-
lish the orientation of the N-methyl groups in darlin-
gine N-oxide (2) and darlingine (1). Selective
refocusing of the N-methyl group (d 3.07) of darlingine
N-oxide (2) gave enhancements to H-6 (d 4.18), H-9 (d
Bark and leaves of D. darlingiana [(F. Muell.)
L.A.S. Johnson] were collected in November 1995
from Timber Reserve 165 in North Queensland,
Australia. D. darlingiana is a ditypic genus endemic to
the `wet tropics' and grows as a canopy tree in com-
plex notophyll vineforest on basalt or granite derived
soils. A voucher specimen (PIF 18157) of this species
is lodged at the Queensland Herbarium.
3.65) and H-10b (d 3.24). No enhancements were
observed to either H-7b or H-8b, which indicated that
the N-methyl group was orientated towards the pyran
ring. Enhancements to H-6 (d 4.14), H-9 (d 3.42) and
H-10b (d 2.96) and none to H-7b or H-8b upon selec-
tive refocusing of the N-methyl group (d 2.30) of dar-
lingine (1) indicated the N-methyl group was also
orientated towards the pyran ring. This orientation is
the opposite to the way the N-methyl group is depicted
in literature for the other g-pyranotropanes, apart
from a paper on the X-ray crystal structure of bellen-
dine (Motherwell et al., 1971).
3
.2. Extraction and isolation
The dried and ground bark (100 g) of D. darlingiana
was extracted sequentially with CH Cl , MeOH and
2
2
H O. The CH Cl and MeOH extracts were combined
2
2
2
and partitioned between 2 M H SO and CHCl . The
4
2
3
acid layer was basi®ed to pH 10 with 25% NH OH
4
and partitioned with CHCl . The CHCl3 layer (110
3
mg) was separated by counter current chromatography
using the solvent system CHCl ±MeOH±H O (13:7:8)
with the MeOH±H O layer as the stationary phase to
3. Experimental
3
2
2
Optical rotations: JASCO P-1020 polarimeter.
13
give darlingine (1) and a 1:15 mixture of 1 and 2. The
counter current separation was repeated on the mix-
ture to give darlingine (1) (overall yield 75 mg, 0.075%
dry weight) and darlingine N-oxide (2) (17 mg, 0.017%
dry weight). An identical scheme was used to isolate
alkaloids 1 and 2 from the leaves.
1
NMR: Varian INOVA H NMR (600 MHz) and
C
NMR (150 MHz): CDCl and DMSO-d were used as
3
6
solvents with chemical shifts calculated relative to the
1
13
1
13
solvent peaks H d 7.26, C d 77.0 and H d 2.49,
C
d 39.5, respectively. 2D NMR data (gradient COSY,

P.L. Katavic et al. / Phytochemistry 52 (1999) 529±531
531
3
.3. Darlingine (1)
i®ed by C18 column chromatography to give darlin-
gine (1) (2.5 mg), which was identical in all respects to
the natural product.
2
6
26
[
a ] +51.78 (CHCl ; c 1.00); [a]D +46.48 (MeOH;
D
3
c 1.00) [lit. (Majewski & Lazny, 1995) +51.58 (MeOH;
c 1.02)]; positive ESI-MS m/z 220 [M+H] ; H NMR
+
1
(
CDCl ) d 1.48 (1H, m, H-8b), 1.78 (1H, m, H-7a),
3
Acknowledgements
1
2
2
1
4
9
.90 (3H, s, 3-CH ), 2.07 (1H, d, J = 18.6 Hz, H-10a),
3
.17 (1H, m, H-7b), 2.19 (1H, m, H-8a), 2.21 (3H, s,
-CH ), 2.30 (3H, s, N-CH ), 2.96 (1H, dd, J = 5.4,
We would like to thank Mr. Rick Willis (Australian
Institute of Marine Science, Townsville) for the high
resolution mass measurement and Dr. Greg Pierens
3
3
8.6 Hz, H-10b), 3.42 (1H, dd, J = 5.4, 6.0 Hz, H-9),
1
3
.14 (1H, d, J = 6.0 Hz, H-6); C NMR (CDCl ): d
3
(
QPRI, Grith University) for assistance with NMR.
.7 (q, 3-CH ), 17.6 (q, 2-CH ), 29.2 (t, C-8), 33.1 (t,
3
3
C-10), 33.4 (t, C-7), 36.7 (q, N-CH ), 55.5 (d, C-6),
3
5
1
7.8 (d, C-9), 120.2 (s, C-3), 123.5 (s, C-5), 159.3 (s, C-
1), 160.6 (s, C-2), 177.1 (s, C-4); COSY and HMBC
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3.4. Darlingine N-oxide (2)
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2
7
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D
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� 1
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Â
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+
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3
.5. Reduction of darlingine N-oxide (2)
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alkaline with 25% NH OH and extracted with
4
CH Cl . The CH Cl -soluble material was further pur-
2
2
2
2