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Short Communication Open Access
Ipvelutine, 7β-Acetoxy-2α-(tigloyloxy)tropane,
an Unusual Tropane Alkaloid from
Ipomoea velutina
R. BR. (Convolvulaceae)
Sonja Christina OTT 1, Kristina JENETT-SIEMS 1, Karsten SIEMS 2,
Frank MÜLLER 3, Monika HILKER 3, Eckart EICH * 1
1 Institut für Pharmazie (Pharmazeutische Biologie), Freie Universität Berlin, Königin-Luise-Str. 2-4, D-14195
Berlin, Germany.
2 AnalytiCon Discovery, Hermannswerder Haus 17, D-14473 Potsdam, Germany.
3 Institut für Biologie (Angewandte Zoologie/Ökologie der Tiere), Freie Universität Berlin, Haderslebener-
Str. 9, D-12163 Berlin, Germany.
* Corresponding author. E-mails: kjsiems@zedat.fu-berlin.de (K. Jenett-Siems), eckeich@zedat.fu-berlin.de
(E. Eich)
Sci Pharm. 2013; 81: 543–548 doi:10.3797/scipharm.1303-13
Published: June 4th 2013 Received: March 19th 2013
Accepted: June 4th 2013
This article is available from: http://dx.doi.org/10.3797/scipharm.1303-13
© Ott et al.; licensee Österreichische Apotheker-Verlagsgesellschaft m. b. H., Vienna, Austria.
This is an Open Access article distributed under the terms of the Creative Commons Attribution License
(http://creativecommons.org/licenses/by/3.0/), which permits unrestricted use, distribution, and reproduction
in any medium, provided the original work is properly cited.
Abstract
Convolvulaceae provide a rich source of tropane alkaloids, however, 2-substi-
tuted tropanes have been described for only few species of this taxon. In this
note, 2,7-diesters such as ipvelutine [7β-acetoxy-2α-(tigloyloxy)tropane] isolated
from the vegetative parts of the Australian Ipomoea velutina R. BR. are
described as a new group of tropane diesters.
Keywords
Ipomoea velutina • Convolvulaceae • Ipvelutine • 7β-Acetoxy-2α-tigloyloxytropane •
2,7-Disubstituted Tropanes • Structure Elucidation
Introduction
During our continuous studies on secondary metabolites of the Convolvulaceae, this plant
family has been shown to produce a plethora of tropane alkaloids, especially 3-tropanols
and their esters (e. g. [1, 2]), as well as some 3,6-disubstituted tropanes [3] or the
polyhydroxylated calystegines [4]. This underlines the chemotaxonomic relationship with
their sister family Solanaceae where the biosynthetic pathway of tropane alkaloids is well
investigated. The main route leads to two stereoisomeric 3-hydroxytropanes, namely
544 S. C. Ott et al.:
Sci Pharm. 2013; 81: 543–548
3α-tropanol (basic component of the well-known atropine and other esters), and
3β-tropanol which is also precursor of the calystegines. 2-Substituted tropane alkaloids
could only be found as a by-product in the Solanaceae [5]. Accordingly, amongst the
tropane alkaloids of the Convolvulaceae 2-substituted ones are extremely rare, too, and
could only be detected in some Calystegia, Erycibe, and Ipomoea species [6].
Results and Discussion
In the alkaloidal screening of Convolvulaceae via GC-MS analysis the basic extracts of the
Australian Ipomoea velutina R. BR. revealed the presence of several unknown substances.
In the basic extract of the vegetative parts seven unknown nitrogen-containing compounds
were detected: one main alkaloid and six minor ones (0.7–18.7% of the main alkaloid by
integration of the corresponding GC-MS peaks). The molecular formula of the main
compound (1) is consistent with C15H23NO4 (m/z 281).
The 1H-NMR (Table 1) in combination with HSQC and HMBC experiments showed two
acylic residues: a C5-acid containing a double bond, namely tiglic acid, as well as acetic
acid. Both were confirmed by fragmentation ions in the EIMS as products of α-cleavage
neighbouring the ester carbonyls: m/z 83 (C4H7−CO+; HRMS: [C5H7O]+ as 83.04959,
calcd. 83.04969) and m/z 43 (CH3−CO+).
Tab. 1. 1H- and 13C-NMR data of ipvelutine (in MeOD)
atom
1H-NMR
(in MeOD)
13C-NMR*
(in MeOD)
1
3.55
br d
3.2 Hz
72.9
2a
5.02
ddd
2.2 Hz; 5.9 Hz; 11.3 Hz
68.9
3e
3a
1.98
1.49
m
dtd
6.4 Hz; 12.1 Hz; 12.8 Hz
22.8
4a
4e
1.89
1.68
m
ddd
2.3 Hz; 6.7 Hz; 13.7 Hz
27.5
5
3.82
br t
5.2 Hz
64.7
6n
6x
2.36
2.27
dd
ddd
8.0 Hz; 14.6 Hz
3.5 Hz; 6.3 Hz; 14.7 Hz
37.8
7n
4.61
dd
3.4 Hz; 7.9 Hz
70.8
N−CH3
2.91
s
40.9
1'
167.8
2'
129.0
3'
6.96
dq
1.2 Hz; 6.9 Hz
139.3
CH3−4'
1.83
d
7.1 Hz
11.9
CH3−5'
1.84
d
0.9 Hz
14.1
1''
176.7
CH3−2''
1.93
s
21.9
*…taken from HSQC/HMBC.
The HSQC spectrum revealed a characteristically downfield shifted N−CH3 (δC 40.9, δH
2.91) as well as three methylene signals (δC 37.8, 27.5, and 22.8) and four methine groups
Ipvelutine, an Unusual Tropane Alkaloid from Ipomoea velutina R. BR. (Convolvulaceae) 545
Sci Pharm. 2013; 81: 543–548
(δC 72.9, 70.8, 68.9, and 64.7). From the 1H-1H-COSY, the complete coupling sequence
could be deduced. As a result, 1 (Fig. 1) could be identified as a 2,7-disubstituted tropane.
The substitution pattern of the tropane diester was derived from the mass spectrometric
data on the basis of the specific mass fragmentation in bridge-substituted tropanes. The
most important fragment is [M − X−COO−CH=CH2]+ after expulsion of the ethylene bridge
C-6−C-7 including its substituent; this allows a prediction of the substituents' positions in
3,6/7-disubstituted tropanes [7, 8]. Regarding 1, there are two possible key ions: in case of
acetylation in position 7 m/z 195 or in case of acetylation in position 2 m/z 155. As there is
only a veritable peak at m/z 195, 1 has to be acetylated in position 7 of the tropane.
The relative stereochemistry of 1 was deduced from characteristic coupling constants: H-7
showed a doublet-doublet with coupling constants of 3.4 Hz and 7.9 Hz that can also be
observed in the 7β-substituted schizanthines C-E [9]. This corresponds with the
experience that, for steric reasons, bridge substituents usually are exo-orientated. H-2
showed a trans-diaxial coupling constant J = 10 Hz which is – according to [10] and [11] –
specific for α-orientated substituents at C-2. These conclusions were also confirmed by
NOE measurements: H-2 (δH 5.02) showed correlations to H-1 (δH 3.55), to the equatorial
H-3e (δH 1.98) and to the axial H-4a (δH 1.89) which is only possible if H-4a and H-2 are
both axial [11]. H-7 (δH 4.61) was correlated to H-1 (δH 3.55) and – only enabled by its
endo-position – to the axial H-3a (δH 1.49) and H-6n (δH 2.36).
Thus, 1 (ipvelutine) was identified as 7β-acetoxy-2α-(tigloyloxy)tropane.
Fig. 1. Structure of ipvelutine [7β-acetoxy-2α-(tigloyloxy)tropane], main alkaloid from
the vegetative parts of Ipomoea velutina R. BR.
In the vegetative parts and/or roots, eight minor compounds related to ipvelutine could be
detected by GC-MS analysis. They were identified by their fragmentation patterns;
characteristic base peaks of those 2,7-disubstituted tropanes are m/z 95 and m/z 82 or
m/z 81 together with a prominent peak at m/z 156, and of their nortropane derivatives m/z
125 and m/z 81 including a half-maximal peak at m/z 108. An additional result of the
systematic GC-MS screening is the detection of ipvelutine (appearing as deacetylated
derivative in GC-MS analysis) in vegetative parts of Convolvulus graminetinus, C.
sagitattus, and Ipomoea abrupta. Both Convolvulus species afforded similar structures, as
well, and, additionally, the corresponding nortropanes in the roots. Ipvelutine-related
substances were also found in Ipomoea asarifolia and I. plebeia. The mass fragmentation
patterns obtained by GC-MS analysis show that these variations include differences in the
stereostructure at C-2 or/and C-7, alternation of the position of the substituents,
methylbutyric and hydroxymethylbutyric acid as diverging acyl components, change of the
546 S. C. Ott et al.:
Sci Pharm. 2013; 81: 543–548
bridge substituents' position from C-7 to C-6 and a hydroxy group as additional substituent
(for details see [12]).
2,7-Dihydroxynortropane showing the same substitution pattern as ipvelutine is also
synthesized by root cultures of Calystegia sepium (Solanaceae). Incorporation
experiments with 15N-labelled 3-tropanone revealed that, unless 2,7-dihydroxynortropane
derives the regular tropane alkaloid pathway, it is not an intermediate in calystegine
biosynthesis, but can be seen as a by-product [5].
From the pharmacological point of view, the finding of ipvelutine and derivatives is of
interest since they show structural similarity to bao gong teng A [13] obtained from the
vegetative parts of Erycibe obtusifolia (Convolvulaceae). Bao gong teng A is characterized
by strong miotic properties and therefore used as an antiglaucoma agent in medicinal
products. This pharmacological effect is contradictory to that of atropine/hyoscyamine
having significance as a mydriatic in ophthalmology and being one of the most commonly
used tropanes of natural origin.
Experimental
General procedures
1H-NMR and 1H-1H-COSY spectra were obtained on a Bruker AMX 400 MHz, HSQC and
HMBC spectra on a Bruker DRX 500 MHz (TMS as internal standard). EIMS and HR-
EIMS were recorded on a Varian MAT 711 (80 eV), FABMS on a Varian MAT CH5DF. The
GC-MS system consisted of a Fisons GC 8060 coupled to a quadrupole mass spectro-
meter Fisons MD 800c.
Plant material
Roots and vegetative parts of Ipomoea velutina R. BR. grown from seeds collected in the
wild at Florence Falls, Litchfield National Park, Northern Territory/Australia, were
harvested in the greenhouse of the Institut für Pharmazie, Freie Universität Berlin. A
voucher specimen is deposited at the herbarium of the Berlin-Dahlem Botanical Garden –
Botanical Museum (BGBM), Freie Universität Berlin, Germany.
Extraction and isolation of ipvelutine
235 g dried and ground vegetative parts of Ipomoea velutina were extracted 4 h with 3 L
MeOH three times and once with a mixture of 2.4 L MeOH and 600 mL 2% aqueous
tartaric acid. After evaporation of the MeOH (50°C i. V.), the residue was redissolved in
600 mL 2% aqueous tartaric acid and extracted with petrol ether, CH2Cl2, and EtOAc,
respectively (3 x 500 mL each). Then, the aqueous layer was alkalinized (pH 10) with
aqueous NH3 (25%) and extracted with 4 x 500 mL CH2Cl2. The united alkaline CH2Cl2
fractions gave 172 mg crude alkaloid fraction which was dissolved in 50 mL 2% aqueous
tartaric acid again and extracted with petrol ether, CH2Cl2, and EtOAc (3 x 50 mL each).
After addition of aqueous NH3 (pH 10), the aqueous layer was extracted with 4 x 50 mL
CH2Cl2. After drying over Na2SO4 and evaporation of CH2Cl2 (40°C i. V.), the alkaline
fractions were united and 10 mg ipvelutine were gained (81% purity according to NMR
spectra).
Ipvelutine, an Unusual Tropane Alkaloid from Ipomoea velutina R. BR. (Convolvulaceae) 547
Sci Pharm. 2013; 81: 543–548
7β-Acetoxy-2α-(tigloyloxy)tropane [(1S,2S,5R,7R)-7-(acetyloxy)-8-methyl-8-aza-
bicyclo[3.2.1]oct-2-yl (2E)-2-methylbut-2-enoate, ipvelutine, 1]
Yellow oil. 1H-NMR (400 MHz, MeOD): see Table 1. 13C-NMR (100.6 MHz, MeOD): see
Table 1. MS (EI, 80 eV, 110°C): m/z (%) = 281 (2) [M]+, 239 (83), 195 (7), 156 (100), 142
(60), 140 (35), 112 (11), 98 (46), 96 (84), 95 (91), 94 (50), 85 (41), 84 (31), 83 (27), 55
(22), 43 (20). (+)-FAB MS (80 eV): m/z = 282 [M+H]+. HR MS (80 eV): m/z = 281.16256
(calcd. 281.16271 for C15H23NO4), 239.15283 (calcd. 239.15214 for C13H21NO3),
156.10254 (calcd. 156.10245 for C8H14NO2+), 142.08678 (calcd. 142.08681 for
C7H12NO2+), 140.10749 (calcd. 140.10754 for C8H14NO+), 98.062524 (calcd. 98.06059 for
C5H8NO+), 95.072728 (calcd. 95.073499 for C6H9N).
GC-MS analysis
Ground plant parts (50 g) were extracted three times with 500 mL MeOH (80%). After
evaporation the residue was dissolved in 2% aqueous tartaric acid and extracted with
petrol ether, CH2Cl2, and EtOAc. The aqueous layer was alkalinized and extracted with
CH2Cl2. To purify the extracts obtained, this procedure was repeated with corresponding
smaller amounts of the solvents. The resulting extracts were subjected to GC-MS analysis.
Samples were injected at 240°C (split 1:20) and separated on a DB-1 column (0.32 mm x
30 m, J&W Scientific, California) by raising temperature from 70°C to 300°C at 6°C/min.
Helium was used as carrier gas. Retention indices (RI): Kovats indices [14] were
calculated in relation to a set of co-injected hydrocarbons.
Acknowledgement
The authors are indebted to Ms. U. Ostwald (FU Berlin) for providing the FABMS and
HRMS spectra.
Authors’ Statement
Competing Interests
The authors declare no conflict of interest.
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