Aporphine and bisaporphine alkaloids from Aristolochia lagesiana var. intermedia

Article abstract of DOI:10.1016/j.phytochem.2009.11.010

Corydines, isocorydines, and analogous aporphine alkaloids were isolated from the leaves of Aristolochia lagesiana var. intermedia, together with three bisaporphine salts (lagesianines B-D). Their structures were determined by chemical derivatizations and spectroscopic analyses. Lagesianines B and C are the first examples of N-CH₂-N′ and C-2-O-C-1′ linked dimeric aporphine alkaloids, respectively, while the monomeric units of lagesianine D, which has a carbon skeleton, are linked through C-7-C-5′ via an ethane-1,2-diol group (C-7-CHOHCHOH-C-5′).

Full text of DOI:10.1016/j.phytochem.2009.11.010

Phytochemistry 71 (2010) 469–478
Contents lists available at ScienceDirect
Phytochemistry
journal homepage: www.elsevier.com/locate/phytochem
Aporphine and bisaporphine alkaloids from Aristolochia lagesiana var. intermedia
a
a
a
b
Marcia L.R. Ferreira , Inara C. de Pascoli , Isabele R. Nascimento , Julio Zukerman-Schpector ,
Lucia M.X. Lopes
a
a,
*
Instituto de Química, São Paulo State University, UNESP, C.P. 355, 14801-970 Araraquara, SP, Brazil
Departamento de Química, São Carlos Federal University, UFSCar, Via Washington Luiz km 235, C.P. 676, 13565-905 São Carlos, SP, Brazil
b
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 11 August 2009
Received in revised form 2 November 2009
Available online 28 December 2009
Corydines, isocorydines, and analogous aporphine alkaloids were isolated from the leaves of Aristolochia
lagesiana var. intermedia, together with three bisaporphine salts (lagesianines B–D). Their structures were
determined by chemical derivatizations and spectroscopic analyses. Lagesianines B and C are the first
0
0
examples of N–CH
2
–N and C-2–O–C-1 linked dimeric aporphine alkaloids, respectively, while the mono-
0
meric units of lagesianine D, which has a carbon skeleton, are linked through C-7–C-5 via an ethane-1,2-
diol group (C-7–CHOHCHOH–C-5 ).
Keywords:
0
Aristolochia lagesiana var. intermedia
Aristolochiaceae
Alkaloids
Ó 2009 Elsevier Ltd. All rights reserved.
Aporphine
Bisaporphine
Alkaloid dimer
Lagesianines
Corydine
Isocorydine
Corydine N
a-oxide
1
. Introduction
Aristolochiaceae species contain three small but characteristic
plied to the mesothorax of third-instar Anticarsia gemmatalis (vel-
vetbean caterpillar) larvae, increased the duration of the larval
phase and decreased both the pupae weight and the larval viability
(Vieira et al., 2009). As part of our continuing studies on the Aris-
tolochiaceae family, in this paper we report the isolation and struc-
tural elucidation of aporphine alkaloids from A. lagesiana.
groups of nitrogenated compounds: 4,5-dioxoaporphines, aristo-
lactams, and aristolochic acids, which have a phenanthrene chro-
mophore moiety in their structures (Kumar et al., 2003; Lopes
et al., 2001; Nascimento and Lopes, 2003; Tsurata et al., 2002). Bra-
zilian Aristolochia species have been shown to contain compounds
belonging to these groups as well as other classes of alkaloids, such
as aporphine (Lopes et al., 2001), tetrahydroisoquinoline (Francisco
et al., 2003; Lopes et al., 2001), benzylisoquinoline (Lemos et al.,
2
. Results and discussion
Compounds 1–13 (Fig. 1) were isolated by chromatography and
partition procedures from the ethanol extracts of the leaves, and
analyzed by spectrometric methods (IR, UV, NMR, and MS).
Compound 1 was identified as (+)-isocorydine based on an anal-
1
1
993; Lopes et al., 1995) and bisbenzylisoquinoline (Cortes et al.,
987), 8-benzylberbine (Lopes, 1992; Lopes and Humpfer, 1997),
and 13-oxidodibenzo[a,g]-quinolizidinium (Watanabe and Lopes,
995).
Previous studies on the roots of Aristolochia lagesiana Ule var.
1
13
ysis of its MS, UV, IR, H and C NMR spectra (Guinaudeau et al.,
975). With the assistance of gHMBC, gHMQC, and COSY experi-
1
1
0
0
ments, the chemical shifts of all hydrogen and carbon atoms in
the molecule were assigned (Tables 1 and 2). The equatorial orien-
intermedia Hoehne led to the isolation of 8,8 -cis-cubebin, 8,8 -
trans-cubebin, and 8,8 -trans-bicubebin A (Pascoli et al., 2006).
The essential oil from the roots has a predominance of sesquiter-
penes (ꢀ30%) and oxygenated sesquiterpenes (ꢀ50%), with eu-
desm-7(11)-en-1-ol being the most abundant compound
0
tation for H-6a
the coupling between this hydrogen and H-5
the magnitude of the coupling constant between H-6a
13.2 Hz) indicated that they are trans-diaxial and in an anti-peri-
planar position on the C ring.
Considering all these data as well as the positive signal of the
optical activity observed for 1 ([ +220), a 6aS configuration
and two P, out of four, conformations could be proposed for 1 (b
a
in relation to the B ring was established based on
(1.3 Hz). In addition,
and H-7b
a
a
(
(Francisco et al., 2008). Extracts of A. lagesiana, when topically ap-
a]
D
*
Corresponding author. Tel.: +55 16 3301 6663; fax: +55 16 3301 6692.
E-mail address: lopesxl@iq.unesp.br (L.M.X. Lopes).
0
031-9422/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.phytochem.2009.11.010

4
70
M.L.R. Ferreira et al. / Phytochemistry 71 (2010) 469–478
4
4
R
3
-
H CO
2
3a
H CO
Cl
3
5
3
A
B
+
4
1
b
R
R
1
1a
N
1
N
H
1
1
R3
R O
2
6a
7a
R O
2
3
C
1a
H
R O
R O
7
D
2
8
H CO 10
3
H CO
3
1
2
3
4
1
3
4
R
R
R
R
R
R
R
R
1
2
4
6
9
CH3
CH3
CH3
H
H
H
H
βCH3
αCH3
H
H
3
7
8
CH3
H
H
CH3
CH3
H
H
H
CH3
CH3
H
H
H
CH3
CH3 βCH3
αCH3
H
CH3
H
OH
-
-
H CO
H CO
Cl
Cl
OCH3
3
3
+
-
+
+
N
H
O
N
N
HO
H CO
OCH3
OH
3
H H H H
H CO
HO
3
H CO
H CO
OCH3
3
3
5
10
3
4
H CO
H CO
3
3
5
+
H
-
N
Cl
NH
OH
-
H
O
H CO
HO
1
6a
7
+
3
Cl
3'
N
2''
4'
OCH3 HO
OH
1''
OCH3
5
'
OH HN
H CO
H CO
6a'
1' OCH3
OH
3
3
9
7
'
OCH3
2HCl
OCH3
9'
11
12
Fig. 1. Chemical structures of isolated compounds 1–12.
Table 1
1
3
H NMR spectroscopic data for compounds 1 and 2 (CDCl , 500 MHz, J in Hz).
H
1
2
1H d
gCOSY
1H d
2D-gNOESY
3
6.63 br s
6.74 s
3
H-4b, H-4a, OCH -2
4
4
5
5
6
7
7
8
9
b
a
b
2.64 ddt (16.5, 3.5, 1.3)
H-4
H-4b, H-5b, H-5
H-4 , H-4b, H-5
a
, H-5b
3.59–3.66 m
H-3
H-3
3.11 ddddd (16.5, 12.1, 5.5, 1.3, 1.0)
2.41 ddd (12.1, 11.3, 3.5)
2.94 ddd (11.3, 5.5, 1.3)
2.80 ddt (13.2, 3.5, 1.3)
2.37 ddd (13.2, 13.0, 1.0)
2.97 dd (13.0, 3.5)
6.76 dd (8.0, 1.0)
6.79 d (8.0)
a
a
a
a
2.94 dd (17.5, 3.5)
4.03–4.10 m
3.77–3.84 m
3.70–3.77 m
3.54–3.61 m
3.17 dd (13.0, 3.5)
6.86 dd (8.3, 1.0)
6.89 d (8.3)
3.09–3.13 br s
8.67 br s
a
a
H-5b, H-4
a, H-6a
aa
H-7b, H-7
a, H-5
H-7a
b
a
H-6a
a
, H-7a
H-7b, H-6a
a
H-6a
a
, H-8
H-7a
OCH
3
-10
N–CH
OH-11
3
2.46 br s
8.76 br s
OCH
3
-1, N–CH
3
OCH
OCH
OCH
3
3
3
-1
-2
-10
3.64 s
3.84 s
3.85 s
3.73 s
3.93 s
3.92 s
OH-11, N-CH
H-3
H-9
3
and d, Fig. 2). Furthermore, ROESY experiments showed spatial
interaction between N–CH and both H-5, which is consistent with
an N–CH group in an equatorial position and a P (b) conformation.
The MS, UV, and IR spectra of compound 2 were very similar to
3
well-resolved signals, including that of the N–CH hydrogens (Ta-
3
bles 1 and 2). In addition, DEPT 135° and 90° experiments did
not show signals for all the carbons expected for an isocorydine
structure. Nevertheless, gHMBC experiments showed correlations
between hydrogens and carbons or nitrogen (Fig. 3, Table 2), which
3
1
13
those of 1. However, the H and C NMR spectra of 2 did not have

M.L.R. Ferreira et al. / Phytochemistry 71 (2010) 469–478
471
Table 2
1
3C NMR spectroscopic data for compounds 1 and 2 (CDCl
3
, d).
C
1
2
13C (d)a
gHMBC
13C (d) a
gHMBC
1
1
1
2
3
3
4
5
6
7
7
8
9
1
1
1
142.2 s
125.9 s
129.3 s
151.3 s
111.1 d
129.9 s
29.4 t
52.7 t
62.9 d
35.9 t
130.2 s
118.9 d
110.9 d
149.5 s
144.0 s
120.2 s
43.9 q
62.1 q
55.9 q
56.2 q
OCH
3
-1, H-3
143.2 s
OCH
OCH
H-3
H-8
3
-1, H-3
a
b
126.2 br s
122.1 br s
153.1 s
H-7b, H-3
OCH
3
-2, H-3
3
-2, H-3
111.0 d
126.2 br s
26.2 br t
52.8 br t
63.3 br d
33.2 br t
126.9 br s
119.7 d
111.7 d
150.3 s
a
H-5
H-5
a
a
, H-3
3
N-CH , H-4a
H-7b, N-CH
H-8
H-7b, H-8
a
a
3
, H-5a, H-7a
H-7
a
0
1
1a
OCH
3
-10, H-8, OH-11
OCH
H-9, OH-11
H-7
3
-10, H-8, OH-11
H-9, OH-11
H-7b, H-8, OH-11
H-5a
144.3 s
119.3 br s
42.4 br q
62.3 q
56.0 q
56.2 q
a
N–CH
3
OCH
OCH
OCH
3
3
3
-1
-2
-10
a
13
C NMR data were assigned with assistance of DEPT 135° and gHMQC experiments.
were compatible with the basic structure of the isocorydine. The
NOE enhanced coupled (gated decoupling) 13C NMR spectrum
showed the multiplicities of all carbons, with most of them as
broad signals, which suggested that conformational equilibrium
had a 6aS configuration. Therefore, alkaloids 1 and 2 are diastereo-
mers, with opposite configurations at the N-6 stereocenter. Inter-
estingly, when analyzed by X-ray crystallography, crystals from 2
showed the same absolute configuration as previously reported
for 1 (Ribar et al., 1992). Aporphine alkaloids, in which chirotopic
stereogenic nitrogens can afford two N-methyl free base diastereo-
mers, are expected to be at the fast exchange limit (FEL) on the
NMR time-scale for interconversion due to facile nitrogen inver-
sion at room temperature (Glaser and Bernstein, 1991). Compound
2 should readily undergone conversion under higher temperature
during crystallization procedures. Hence, 1 and 2 should crystallize
when they reach the most stable conformation and configuration
(P).
was taking place in CDCl
3
solution, but not on the NMR time-scale.
In addition, repetitive H NMR experiments showed that the chem-
ical shifts of the aliphatic hydrogens changed. The N–CH hydro-
1
3
gens clearly shifted from d 2.56 to 3.13. In some cases, a double
signal could be observed, which were confirmed to be singlets by
2
D–J-resolved experiments. To confirm that 1 and 2 have the same
basic structure, both compounds were subjected to transforma-
tions with hydrochloric acid and methyl iodide to yield the same
corresponding salt derivatives (3 and 14). Generally, aporphine
1
alkaloids as either free or protonated bases show d
and 27 for C-4 in conformations M (a), P (b), M (c), and P (d),
respectively, and very similar d N–CH
(d ꢀ42), except for that
in the P (d) conformation (d ꢀ32) (Glaser and Bernstein, 1991)
Fig. 2). Based on a comparative analysis of these data with those
of 2, a P (d)-to-M (a) conversion can be proposed for 2 in solution.
C
ꢀ25, 29, 22,
Based on a comparison of its spectrometric data (MS, IR, UV, H
and 1 C NMR) to those reported in the literature (Chang et al.,
2001; Rasoanaivo et al., 1998) compound 4 was identified to be
(+)-norisocorydine. Compounds 5 and 6 were identified to be (+)-
3
C
3
(
corydine Na-oxide and (+)-corydine, respectively, based on a de-
tailed analysis of their spectroscopic data, as described for 1 and
The observed optical activity for 2 ([
a]
D
+235) indicated that 2 also
2, comparison of these data with those reported in the literature
H
H
H
R
3
R'
R
8
+
N
H
7
4
H
+
N
6
a
H H
H
H H
5
H
H
R'
R
R'
R
R'
H
(
S S
6 ,6a ,
M
)
a
H or OH
or H
R S
(6 ,6a ,
P
)
b
H or OH
or H
H
H
H
R
R
H
+
H
N
+
N
H H
H H
or H
R'
H
R'
H
R
R'
R
R'
(
R S
6 ,6a ,
M
)
c
H or OH
or H
S S
(6 ,6a ,
P
)
d
H or OH
Fig. 2. M and P B-ring chair-type conformations.

4
72
M.L.R. Ferreira et al. / Phytochemistry 71 (2010) 469–478
pling constants between the aliphatic hydrogens (Table 3). Consid-
H CO
3
ering that compound 9 follows the rule for all simple aporphines,
e.g., C-6aR aporphines are levorotatory while their S analogous
are dextrorotatory (Ringdahl et al., 1981), as observed for (+)-glau-
fidine and (+)-epiglaufidine (Guinaudeau and Shamma, 1985), the
N
H
H CO
3
HO
absolute configuration of
4R,6S,6aS,M. This compound (9) is thus referred to as lagesianine
9 could be established as being
H CO
3
A hereafter to avoid further confusion.
Fig. 3. Selected gHMBC correlations (C ? H, N
H) for compound 2.
The 1H and 13C NMR spectra of compound 10 were similar to
those of norisocorydine (4) (Tables 4 and 5). In addition, signals
for a methylene group (d
gens (d 9.87, 10.73) were observed. The HRMS spectra showed
quasi-molecular 741.2369
H C
3.64, 3.57; d 70.6) and two acid hydro-
(
Guinaudeau et al., 1983; Shafiee and Jafarabadi, 1998), and by an
analysis of the spectrometric data of their hydrochloride deriva-
tives 7 and 8.
H
+
ions
[M+KꢁHCl]
at
m/z
(
C
39
H
43ClN
2 8
O + K). These data suggested that 10 was an aporphine
The HRMS spectrum of compound 9 showed quasi-molecular
1
1
ions [M+H]+ at m/z 358.1652 (C20
than in 1. The H and C NMR spectra of 9 (Table 3) were very sim-
ilar to those of 1 (Tables 1 and 2). The main differences being the
dimer. A H– H COSY experiment showed coupling between H-6
(dH 9.87) and H-5 (dH 3.77) and H-6a (d 4.04), whereas H-6 (dH
1
1
additional methylene hydrogens (2H-1 ), 2H-4, and 2H-7, permit-
ting assignment of the chemical shifts of hydrogens and carbons
for one of the units. Thus, the two aporphinoid units should be
linked through the nitrogen atoms via a methylene group. The frag-
ment ions at m/z 326, 362 [326+HCl], 342, and 463 [440+Na] ob-
served in both HRMS and LRMS are consistent with this linkage
H24NO
5
), which is 16 u higher
0
1
13
H
0
0.73) showed coupling to H-6a (d
D-TOCSY experiments showed coupling between H-6a and the
H
4.47). In addition, selective
signals for a carbinolic methylene at C-4 (d
downfield chemical shifts observed for N–CH
7.09). The presence of an hydroxyl group in 9 was confirmed
H
C
4.75, d 64.0) and
0
0
3
H
(d 3.04) and H-3
(d
H
+
by the base peak [MꢁH
2 4
O+H] at m/z 340.1549 (C20H22NO ) in
the HRMS spectrum. Moreover, from the gHMBC spectrum show-
ing a correlation between H-4 and C-3 and the fact that the NOESY
experiments showed interaction between H-4 and H-3, it was pos-
sible to establish that the OH group was linked to C-4. These data
led us to propose the same basic structure for 9 as was previously
determined for crabbine (Yang et al., 1993). This was further sup-
ported by 1D-TOCSY experiments where couplings between H-5eq
3
(Fig. 4). Considering that, except for the N–CH hydrogens, the pro-
tonation of isocorydine did not significantly affect the chemical
shifts of the carbons and hydrogens, it can be proposed that the dif-
ferences between the chemical shifts of the corresponding carbons
0
0
and hydrogens on the B/B and C/C rings of both units of 10 are due
(d 3.78) and H-5ax (d 3.34), H-4 (d 4.75), and H-6a (d 3.68) were ob-
1
to different conformations and/or configurations at the N-6 and N-
served. However, a comparison of the H NMR spectroscopic data
for both compounds showed that they were not identical. Analys-
ing the spatial correlations, as established by NOESY experiments,
and considering the four possible conformations for 9, a M confor-
mation (a, Fig. 2) was determined for this alkaloid based on the
0
6
4
stereocenters. Based on the chemical shifts of C-4 (d 25.5) and C-
(d 30.2) and on the magnitude of the coupling constants between
0
N–H and H-6a, H-5, and H-1’’ (br q, J = 10.5 Hz), as well as those be-
0
0
0
tween N–H and H-6a , H-5 , and H-1’’ (br d J = 10.5 Hz), an M (a)
conformation could be assigned to one unit and a P (d) conforma-
tion to the other. These considerations are consistent with the ob-
spatial interactions between H-5ax and H-6a, N–CH
5
3
(d 3.04), H-
eq, and H-4eq (d 4.75), as well as on the magnitude of the cou-
0
served up-field chemical shifts of C-5 and C-5 in relation to those
0
of isocorydines due to a
c
-effect of N–H and N–H. Consequently,
this new dimer was named lagesianine B.
Table 3
1
1
H and 13C NMR spectroscopic data for compound 9.
The H NMR spectrum of 11 was very similar to that of 3. Essen-
tially, the only difference between them is the proportion of meth-
oxyl hydrogens (Table 6). The former showed signals for three
methoxyl groups at d ꢀ3.86 and one at d ꢀ3.68, whereas 3 showed
two and one methoxyl groups at the respective spectral regions.
9
H
1H (d)a,b
C
13C (d) b
gHMBC
1
1
1
2
3
3
4
5
144.4 c
125.5 s
123.3 s
153.6 s
112.1 d
129.6 s
s
3
OCH -1, H-3
a
b
The HRMS spectra of 11 showed the quasi-molecular ions at m/z
H-3
+
7
11.2204 ([C38
H
40
N
2
O
7
.HCl+K] ), and ions at m/z 683.2068
3
OCH -2, H-3
+
+
(
[C36
H
36
N
2
O
7
ꢂHCl+K] ) and 551.2048 ([C34
31 7
H O ] ), which may
3
7.09 s
H-3, H-4
H-5e
a
have arisen from retro Diels–Alder rearrangements involving the
4
5
5
6
7
7
8
9
e
4.75 br s
64.0 br d H-3
60.1 br t NCH
0
B and B rings. Under negative ion conditions, the HRMS spectra
a
e
a
a
e
3.34 dd (13.0, 3.5)
3.78 br dd (13.0, 1.4)
3.68 dd (13.5, 3.5)
3.28 br t (13.5)
3.08 dd (13.5, 3.5)
6.77 d (8.5)
3
3
basically showed two intense peaks at m/z 659.2077 ([M–
ꢁ
ꢁ
OHꢁOCH
3
ꢁH] ) and 631.1841 ([MꢁOHꢁOCH
3
ꢁCH CH
2 2
ꢁH] ),
6a
7
7a
8
63.3 br d NCH
32.9 t
126.6 s
119.7 d
111.7 d
H-8
H-9, H-7a
H-7e
and their corresponding ions with two, three, and four u higher
showed intensities compatible with two chlorines. Based on these
data, a dimeric aporphine structure was proposed for alkaloid 11,
6.80 d (8.5)
9
0
in which the two monomers are linked through C-2–O–C-1 . This
1
1
1
0
1
1a
150.2 s
OCH
H-9
H-8
3
-10, H-9, H-8
144.2 c
new bisaporphine alkaloid was, in turn, named lagesianine C.
s
119.3 s
42.0 br q
Compound 12 was suggested to be an aporphine dimer, since its
1
13
NCH
3
3.04 br s
3.69 s
3.88 s
NCH
OCH
OCH
OCH
10
3
3
3
3
H and C NMR, gHMQC, and DEPT 90° and 135° spectra showed
OCH
OCH
OCH
3
-1
3
-2
3
-
-1
-2
-
62.3 q
0
0
signals for two sets of aromatic rings (A/A and D/D ) with the same
d
56.1
q
q
56.2 d
substitution pattern as observed for 10, resonances for four meth-
3.83 s
0
0
1
0
ylene carbons at d 25.6 (C-4), 29.7 (C-4 ), 41.1 (C-5), and 34.9 (C-7 ),
0
OH-11
8.52 s
and six methyne carbons at d 55.8 (C-6a), 56.3 (C-7), 62.4 (C-5 ),
53.4 (C-6a ), and 68.5 and 70.6 (C-1 , C-2 ) (Tables 6 and 7). Thus,
0
00
00
a
Multiplicities were determined with the assistance of 1H– H COSY experiments,
J in Hz.
1
one methylene carbon must be replaced by two methyne carbons
b
0
Recorded in CDCl
3
, 11.7 T.
in each isocorydine unit. The correlations between C-1b (d
C
c,d
Assignments may be interchangeable within the same column.
0
0
0
1
26.2) and H-3 (d
H
6.69), H-4 (d
H
2.70), and H-7 (d
H
3.15), as well

M.L.R. Ferreira et al. / Phytochemistry 71 (2010) 469–478
473
Table 4
1
H NMR spectroscopic data for compounds 4 and 10 (CDCl
3
, 500 MHz, J in Hz).
4
10
H
3
1H (d)
gCOSY
1H (d)
gCOSY
H
1H (d)
gCOSY
0
6.66 s
6.64 s
3
6.64 s
3.57 m
0
0
0
0
4
a
3.35 ddd (17.5, 11.7, H-4b, H-5b, H- 3.57 m
H-4b, H-5b
4a
H-4b , H-5a , Y-5b
5
.0)
5a
H-4a
H-4a, H-5b
0
0
0
0
4
5
5
b
a
b
2.80 dd (17.5, 3.8)
3.59 dd (12.3, 5.0)
2.88 br dd (17.0, 3.5) H-4a, H-5b
3.77 m
4b
5a
5b
2.81 m
3.64 m
3.20 m
H-4a , H-5a
0
0
0
H-5b
0
0
0
3.02 ddd (12.3, 11.7, H-5a, H-4a
.8)
3.12 m
N-H, H-4b
H-4b , H-5a
3
a
0
0
6
a
3.85
H-7a, H-7b
4.04 ddd (13.0, 10.5, H-7a, H-7b, N-H 6a
.5)
3.28 br dd (13.0, 3.5) H-6a, H-7b
4.47 br dd (13.0, 3.0) H-7a , H-7b
3
0
0
0
0
7
7
8
9
a
b
3.07 dd (13.0, 4.0)
2.90 dd (13.5, 13.0)
6.76 d (8.0)
6.79 d (8.0)
-
H-6a
H-6a
7a
7b
2.93 br dd (13.0, 3.5) H-6a , H-7b
0
3.13 t (13.0)
6.78 m
H-6a, H-7a
2.49 t (13.0)
6.75 m
6.78 m
H-6a , H-7a
0
0
8
9
H-9
H-8
6.78 m
0
N–H
N–CH
9.87 br q (10.5)
3.64 m
H-6a, H-5b
N–CH –N
2
N –H
10.73 br d (10.5)
0
0
2
–N
3
.57 m
0
H-5a
0
OH-11
8.67 s
3.64 s
3.84 s
OH-11
8.71 s
3.66 s
3.84 s
0
0
OCH
OCH
3
-1
-2, OCH
3.66 s
3.84 s
OCH
OCH
3
-1
-2 , OCH
3
3
-
3
3
-
0
1
0
10
a
Multiplicity obscured by methoxyl signals.
Table 5
1
3C NMR spectroscopic data for compounds 4 and 10 (CDCl
3
, d).
4
10
C
13C (d) a
gHMBC
13C (d) a
gHMBC
H-3, OCH
C
13C (d) a
142.8 s
119.6 s
128.2 s
151.3 s
111.2 d
129.2 s
30.2 t
gHMBC
0
0
0
1
1
1
2
3
3
4
5
6
7
7
8
9
1
1
1
142.8 s
126.0 s
128.3 s
152.5 s
111.2 d
129.2 s
26.8 t
41.6 t
53.6 d
36.0 t
127.7 s
119.5 d
111.5 d
149.9 s
144.4 s
119.7 s
OCH
3
-1, H-3
143.1 s
119.4 s c
122.8 s
153.0 s
111.2 d
130.0 s c
25.5 t
41.0 t
53.32 d
34.8 t
3
-1
1
H-3 , OCH
3
-1
0
0
c
a
b
1a
0
0
H-5a
H-3, H-7b
1b
H-3 , H-7b
0
OCH
H-3
3
-2, H-3
OCH
3
-2, H-3
2
OCH
3
-2
0
3
0
0
a
3a
H-4b
H-3
0
H-3
4
0
5
43.8 t
0
0
a
a
H-7a, H-7b, H-5a
H-7b
H-8
H-7b, H-9
H-7b
6a
53.28 d
36.4 t
H-7b
H-8
0
0
7
126.4 s c
119.9 d
111.6 d
150.1 s
144.4 s
122.4 s
70.6 t
7a
0
126.2 s
119.6 d
111.6 d
149.7 s
144.1 s
b,c
H-7b
0
0
H-9, H-7b, H-7a
H-7b, H-7a
0
0
8
H-7a , H-7b
0
9
0
0
0
0
0
0
0
1
1a
OCH
H-9
H-8
3
-10, H-8, H-9
H-8, OCH
3
-10, OH-11
10
11
11a
H-8 , OCH
3
-10 , OH-11
0
H-9, OH-11
H-7a, OH-11
H-9 , OH-11
0
127.2 s b
0
0
H-7a , H-7b , Y-8
0
0
N–CH
OCH
OCH
2
N
H-5a
62.0 q c
56.0 q, 56.2 q
OCH
OCH
-1
0
0
62.2 q c
56.0 q, 56.2 q
3
-1
-2, OCH
61.9 q
3
0
3
3
-10
3
-2 , OCH
3
-10
a
b
c
The 13C NMR data were assigned with the assistance of DEPT 90° and 135°, gHMQC, and gHSQC experiments.
Assignments may be interchangeable within the same column.
Assignments may be interchangeable within the same line.
0
0
as between C-6a (d
C
53.4) and 2H-7 , as observed in gHMBC exper-
from the migration of an hydrogen of an OH group and simulta-
0
0
0
00
iments, allowed us to assign the chemical shifts for one of the units
I). Moreover, COSY experiments showed coupling between H-6a
4.05) and 2H-7 (d
.70) and H-4a (d
unit (II) was determined by the correlation between C-1b (d
22.8) and H-3 (d 6.68) and H-4 (d 3.13), as evidenced by gHMBC
experiments, and by the coupling between 2H-4 (d 3.13 and 2.92)
3.58 and 3.80). These data suggested that the linkage
between the two units should involve C-7 and C-5 . The HRMS
spectrum showed ions at m/z 781.2356, 747.2690, and 713.3029,
which correspond to the formulae C40
and C40 10, respectively. Under positive ion conditions, the
ESI-MS–MS of the ions at m/z 747.27 gave rise to a series of spectra
showing ions at m/z 328.15 (base peak), which could have arisen
neous cleavage of the bonds C-7–C-1 and/or C-5 –C-2 , and at
m/z 362.12 (327 + Cl), which arise from these bond-breaking pro-
cesses. Based on these data, formula 12a was proposed for the ions
at m/z 747.27 (Fig. 5). Moreover, these spectra showed ions at m/z
342.17 and 376.13 (341 + Cl), which suggested the same types of
0
(
(
2
0
0
d
H
H
3.15 and 3.32) and between H-4b (d
2.92) and H-5 (d
H
0
0
H
H
4.57). Similarly, the other
C
0
0
00
1
H
H
fragmentation from ions containing a CH
stead of CHOH. This methylene and an
have arisen from enols produced by dehydration of the diol. Based
on all these data, the molecular formula C40
2
group at C-1 or C-2 in-
H
a-carbonyl group could
and 2H-5 (d
H
0
H
44
N
2
O
10ꢂ2HCl could
be deduced for 12. The magnitude of the coupling constants of
H-6a (br d, J = 13.0 Hz) suggested a trans-di-axial orientation on
the C ring for H-6a and H-7 and a P conformation for unit I, in
which the substituent at C-7 is in an equatorial position, whereas
2 2 2 10
H43Cl N O10, C40H44ClN O ,
45 2
H N O
0
the multiplicity and coupling constants of H-6 (dddd, J = 13.0,

4
74
M.L.R. Ferreira et al. / Phytochemistry 71 (2010) 469–478
by avoiding the traditional acid/base procedures used for the isola-
H CO
OCH3
3
tion and purification of aporphine alkaloids. Lagesianine C is the
R R'
+
+
0
first example of a C-2–O–C-1 linked dimeric aporphine alkaloid,
N
N
+
H CO
OCH3
OH
3
+
K
whereas lagesianine D has a new carbon skeleton, in which the
H
HO
0
monomeric units are linked through C-7–C-5 via an ethane-1,2-
440
0
-
diol group (C-7–CHOHCHOH–C-5 ). Bisaporphine alkaloids linked
Cl
H CO
OCH3
0
0
3
through N–N via a carbonyl group (N–CO–N ) have been isolated
from Hernandia nymphaeifolia (Hernandiaceae) (Chen et al.,
R
H
R'
1
996a,b), but this is the first report of such linkage via a methylene
0
1
1
0a
0b
group (N–CH
N ), as in lagesianine B.
m/z 741
2
H
°
3
. Experimental
3
.1. General experimental procedures
H CO
OCH3
3
H
+
+
1
13
NH
N
One-dimensional ( H, C, DEPT, NOE enhanced coupled (gated
H CO
OCH3
OH
3
13
1
1
decoupling) C, and gNOESY) and two-dimensional ( H– H gCOSY,
gNOESY, gHMQC, H– C and H– N gHMBC, J-resolved) NMR
HO
1
13
1
15
HCl
experiments were performed on a Varian INOVA 500 spectrometer
H CO
OCH3
1
13
15
3
(
11.7 T) at 500 MHz ( H), 126 MHz ( C), and 51 MHz ( N), using a
deuterated solvent (CDCl ) (P 99.9% D) as an internal standard for
C NMR chemical shifts, and residual solvent as an internal stan-
3
m/z 362
m/z 342
13
1
15
dard for H NMR, and d are reported relative to TMS. N gHMBC
spectra were calibrated using nitromethane as an external stan-
dard and by setting the chemical shift to 381.9 so that the 15N d
are reported relative to liquid ammonia. Mass spectra (ESI-MS)
were obtained on a Fisons Platform II, and flow injection into the
electrospray source was used for LC–ESI-MS. High-resolution mass
spectra (HRMS) were obtained on a Bruker Daltonics ultrOTOFq
(ESI-TOFMS). IR spectra were obtained on a Perkin–Elmer 1600
FT-IR spectrometer using KBr discs. Optical rotations were mea-
sured on a Perkin–Elmer 341-LC polarimeter. Ultraviolet (UV)
absorptions were measured on a Perkin–Elmer UV–vis Lambda
14P diode array spectrophotometer.
Fig. 4. Key ions obtained in HR-ESI-MS of 10.
9
.0, 2.0, and 1.0 Hz), which were calculated by spectral simulations
using the FOMSC3 program (FOMSC3, 2004), suggested an M con-
0
formation for unit II, in which H-6a is in a trans-di-axial orienta-
0
0
tion in relation to one of N-H and H-7 (Fig. 2). The spatial
0
0
correlations between H-5 and 2H-4 and H-7 establish an equato-
rial position for H-5 and are consistent with this suggestion. The
downfield chemical shift observed for of C-4 in relation to C-4
0
0
(D
d = 4.1) could be explained by the b effect of the substituent at
C-5 . Hence, the conformation of this new compound was tenta-
0
HPLC analyses were performed using a Shimadzu liquid chro-
matograph (SPD-10 Avp) equipped with UV–vis and 341-LC polar-
imeter detectors. The columns were RP-18 (Varian, C18, with a
tively determined to be I-P and II-M, and it was named lagesianine
D.
particle size of 5
l
m, 250 ꢃ 4.6 mm for analytical analysis, and
2.1. Concluding remarks
250 ꢃ 20 mm for semi-preparative analysis), and chromatograms
were acquired at 336 and 254 nm. TLC: silica gel 60 PF254. Melting
points were recorded on a Microquímica MQAPF-302 melting point
apparatus and are uncorrected.
The isolation of a significant number of natural alkaloids in their
salt forms from A. lagesiana is noteworthy. This was only possible
Table 6
1
H NMR spectroscopic data for compounds 3, 11, and 12 (CDCl
3
, 500 MHz, J in Hz).
3
12
11
H
3
1H (d)a
1H (d)a
H
1H (d)a
H
1H (d)a
0
0
6.71 s
6.68 s
3
6.69 s
2.92 m
2.70 ddd (15.5, 2.5, 2.0)
4.57 ddd (13.0, 4.0, 2.0)
3, 3
4a, 4a
4b, 4b
5a, 5a
5b, 5b
6a, 6a
7a, 7a
7b, 7b
8, 8
9, 9
6.68 s
3.90 m
2.85 dd (17.5, 3.5)
3.59 m
0
0
0
0
0
0
0
0
0
0
0
0
0
0
4
4
5
5
6
7
7
8
9
1
a
3.90 ddd (17.5, 12.6, 5.5)
2.85 dd (17.5, 3.5)
3.65 dd (12.6, 5.5)
3.22 dddd (12.6, 12.6, 10.0, 3.5)
3.78 m
3.14 dd (13.5, 3.2)
3.30 dd (13.5, 13.0)
6.82 d (8.0)
3.13 m
2.92 m
3.80 m
3.58 m
4.71 br d (13.0)
3.85 m
4a
4b
5a
5b
6a
7a
b
a
b
a
a
b
3.08 m
b
4.05 br t (13.0)
3.15 t (13.0)
3.32 dd (13.0, 3.5)
6.80 m
6.80 m
3.58 m
3.11 dd (13.5, 3.5)
3.32 dd (13.0, 13.5)
6.82 d (8.5)
7b
0
0
00
0
0
6.80 m
6.80 m
3.68 m
8
9
2
6.86 d (8.0)
6.78 d (8.5)
0
0
0
0
0
N–CH
N–H
OH-11
OCH
OCH
OCH
3
3.04 d (5.0)
12.75 br m
8.67 s
3.69 s
3.90 s
N–CH
N–H
OH-11
OCH
OCH
OCH
N–CH
N–H, N –H
OH-11, OH-11
OCH
OCH
OCH
3
, N –CH
3.01 d (5.0)
3
3
0
0
0
b
9.90 br s
8.70 s
3.65 s
3.86 s
3.86 s
10.90 dddd (13.0, 9.0, 2.0, 1.0)
0
0
8.70 s
3.68 s
3.86 s
3.86 s
8.63 s
3.68 s
3.86 s
3.86 s
0
0
3
3
3
-1
-2
-10
3
3
3
-1
-2
-10
3
3
3
-1
-2
0
0
0
3.89 s
-10, OCH
3
-10
a
Multiplicities were determined with the assistance of 1H– H COSY experiments.
Signals not observed.
1
b

M.L.R. Ferreira et al. / Phytochemistry 71 (2010) 469–478
475
Table 7
1
3C NMR spectroscopic data for compounds and 3 and 12 (CDCl
3
, d).
3
12
C
13C (d)a
gHMBC
OCH -1, H-3
13C (d)a
gHMBC
OCH -1, H-3
C
13C (d)a
gHMBC
0
0
0
-1
1
1
1
2
3
3
4
5
6
7
7
8
9
1
1
1
143.1 s
126.1 s
122.1 s
152.9 s
111.0 d
127.0 s
26.1 t
52.3 t
62.9 d
33.1 t
126.5 s
119.6 d
111.6 d
150.1 s
144.2 s
119.3 s
3
143.2 s
127.1 s
122.8 s
153.0 s
111.3 d
127.1 s
25.6 t
41.1 t
55.8 d
56.3 d
129.2 s
119.9 d
111.7 d
150.1 s
144.4 s
119.4 s
68.5 d
3
1
143.1 s
H-3 , OCH
3
0
0
d
a
b
1a
0
0
0
H-3, H-7b
OCH -2, H-3
H-3, H-7a
1b
126.2 s
153.0 s
111.3 d
129.7 s
29.7 t
62.4 d
53.4 d
34.9 t
H-3 , H-4b , H-7a
0
0
0
3
OCH
3
-2, H-3
2
OCH
3
-2 , H-3
0
3
0
0
a
H-5a
H-3
3a
H-4b
H-3
0
0
H-3
4
0
N–CH
3
H-4a
H-9
5
0
0
0
a
a
H-5a, H-7b, N–CH
H-8
H-9, H-7b
H-7b
3
6a
H-7a , H-7b
0
0
7
H-8
0
126.4 s b
119.9 d
111.7 d
150.1 s
144.4 s
122.2 sb
70.6 d
7a
H-9
0
8
0
9
0
0
0
0
0
0
0
1
OCH
H-9
H-8
3
-10, H-8, H-9
3
H-8, OCH -10, OH-11
H-9
10
11
H-8 , OCH
3
-10 , OH-11
0
H-9 , OH-11
0
0
1a
H-8
11a
H-7b
0
0
00
1
2
N–CH
3
42.2 q
62.2 q
62.0 q e
56.1 q
56.1 q
OCH
OCH
OCH
-1
-2
0
0
62.2 q
56.0 q
56.2 q
e
c
c
OCH
OCH
OCH
3
3
3
-1
-2
-10
3
3
3
b
55.9 q
56.1 q b
-10
0
a
The 13C NMR data were assigned with the assistance of DEPT 90° and 135° and gHMQC experiments.
Assignments may be interchangeable within the same column.
Signals not observed.
b,c
d
e
Assignments may be interchangeable within the same line.
3.2. Plant materials
of conc. HCl to obtain the NMR spectroscopic data for 3 and
7
+ 8, respectively.
The plant materials were collected in Ituiutaba, MG, Brazil,
Compounds 1 (8.0 mg) and 2 (8.0 mg) were dissolved in acetone
in February 2003, and identified as A. lagesiana Ule var. interme-
dia Hoehne by Dr. Condorcet Aranha and Dr. Lindolpho Cappel-
lari Júnior. A voucher specimen (ESA 88885 02/13/2003) was
deposited at the herbarium of the Escola Superior de Agricultur-
a, Luiz de Queiroz (ESALQ), Piracicaba, SP, Brazil. The material
was separated according to the plant parts, dried (ꢀ45 °C),
and ground.
3
and individually subjected to methylation with CH I (200 lL) by
the usual procedures to give 14 (5.0 and 4.3 mg, respectively).
3.4. (6R,6aS,P)-Isocorydine (1)
Yellow solid; m.p. 182–185 °C (MeOH) [lit. (Guinaudeau et al.,
2
D
5
1975) 185–186 °C]; ½
a
ꢄ
þ 220 (c 1.30, CHCl ) [lit. (Ringdahl
3
MeOH
max
et al., 1981) +196° (c 1.0, CHCl
4.85), 269 (4.32), 303 (3.89); IR
599, 1461, 1262; for H and C NMR (CDCl ), see Tables 1 and
3
, respectively; ESI-MS (probe) 40 V, m/z (rel. int.): 341 [M] (20),
3
)]; UV k
nm (log e): 219
KBr
max
ꢁ1
(
m
cm : 3202, 2927, 2848,
3.3. Extraction, isolation, and chemical transformation
1
13
1
2
3
+
The plant material from the leaves (236.2 g) was extracted
exhaustively at room temperature with hexane, acetone, and EtOH
+
40 [MꢁH] (68), 326 (100), 310 (10), 298 (7).
(
ꢀ1.3 L for each solvent), successively, and the residues were ex-
3
.5. (6S,6aS,M)-Isocorydine (2)
tracted with EtOH (ꢀ 1.3 L for each solvent), in a soxhlet apparatus.
The extracts were individually concentrated.
White crystals; m.p. 201–205 °C (MeOH) [lit. (Guinaudeau et al.,
The initial crude ethanol extract (6.6 g) was fractionated by CC
2
D
5
1
1
975) 185 °C]; ½
a
ꢄ
þ 235 (c 1.50, CHCl
3
) [lit. (Ringdahl et al.,
nm (log e): 219 (4.85), 269
(
silica gel 60 H, 180.0 g, CHCl
give 32 fractions. Fraction 9 was subjected to PTLC (CHCl
NH OH 94:5:1) followed by semi-prep. HPLC (MeOH–H
give 1 (14.1 mg), 2 (208.0 mg), 4 (15.3 mg), and 6 (17.3 mg). Frac-
tion 10 after PTLC (CHCl –MeOH–NH OH 79:20:1) gave 5 (8.9 mg).
Fraction 17 was subjected to semi-prep. HPLC (CH CN–MeOH–H
:1:8) to give 13 (33.0 mg).
A portion (7.9 g) of the crude ethanolic extract (14.5 g) obtained
for soxhlet extraction was subjected to partitioning in AcOEt and
O. The aqueous fraction was concentrated (3.02 g) and fraction-
ated over Sephadex LH-20 (MeOH–H O gradient) to give 23 frac-
tions. Fractions 3 (1024.6 mg) and 4 (644.1 mg) were combined
and subjected to CC (silica gel 60 H, 50.0 g, CHCl –MeOH gradient,
3
–MeOH, gradient 5–100% MeOH) to
–MeOH–
O 3:2) to
MeOH
max
981) +196° (c 1.0, CHCl
3
)]; UV k
3
KBr
ꢁ1
(
4.32), 303 (3.89); IR
m
cm : 3202, 2927, 2848, 1599, 1461,
4
2
max
1
13
1
262; for H and C NMR (CDCl
3
), see Tables 1 and 2, respectively;
1
5
N NMR (50.7 MHz, CDCl
3
): d 42.1 (N-6); MS-EI (probe) 70 eV, m/z
3
4
+
rel. int.): 341 [M] (42), 326 (100).
3
2
O
1
3.6. (6S,6aS,M)-Isocorydine hydrochloride (3)
Yellow solid; m.p. 258–260 °C (MeOH); for 1H and 13C NMR
H
2
3
(CDCl ), see Tables 6 and 7, respectively; ESI-MS (probe)+50 V, m/
2
+
z (rel. int.): 376 [MꢁH] (7), 342 (47), 311 (50), 296 (55), 280
(60), 279 (100).
3
5
–100% MeOH) to give 40 fractions (ꢀ50 mL each). Fraction 10
gave 9 (34.5 mg). Fraction 5 by PTLC (CHCl
3
–MeOH, 19:1) gave 2
3.7. (6aS,M)-(+)-Norisocorydine (4)
(
(
(
35.5 mg), 10 (10.8 mg), and 6 (33.3 mg). Fraction 6 by PTLC
CHCl –MeOH, 19:1) gave (11.5 mg), 7 + 8 (11.6 mg), 11
1.5 mg), and 12 (10.4 mg).
Compounds 2 (25.0 mg) and 6 (15.1 mg) were dissolved in
CDCl and individually subjected to protonation using one drop
3
5
Brown solid; m.p. 185–187 °C (MeOH) [lit. (Chang et al., 2001)
2
5
202–204 °C]; ½
+158.5° (c 0.10, EtOH)]; H NMR (500 MHz, CDCl
dd, J = 17.5, 3.8 Hz, H-4b), 2.90 (1H, dd, J = 13.5, 13.0 Hz, H-7b),
aꢄ
þ 178 (c 0.20, CHCl
3
) [lit. (Chang et al., 2001)
D
1
3
): d 2.80 (1H,
3

4
76
M.L.R. Ferreira et al. / Phytochemistry 71 (2010) 469–478
[
M]+
H CO
-
H CO
3
3
Cl
2
NH
NH
2
H CO
+
H CO
+
3
OH
3
OH
HO
OCH3
HO
OCH3
+
OH
N
OH
HN
H CO
OCH3
OH
H CO
OCH3
OH
3
H
3
12a m/z 747
m/z 713
OCH3
OCH3
H CO
3
NH
2
H CO
HO
+
H CO
-
3
3
Cl
o
+NH
2
H CO
3
O
H
H CO
HO
OCH3
3
m/z 342
+
HN
H CO
OCH3
OH
3
H CO
3
NH
2
OCH3
H CO
HO
+
3
-
Cl
+
H CO
3
m/z 376
H CO
3
-
Cl
NH
2
H CO
+
3
H CO
3
HO
OCH3
NH
+
2
-
H CO
HO
+
O
HN
3
H CO
OCH3
OH
3
Cl
+
H CO
3
m/z 362
OCH3
Fig. 5. Key ions obtained in HR-ESI-MS of 12.
3
4
.02 (1H, ddd, J = 12.3, 11.7, 3.8 Hz, H-5b), 3.07 (1H, dd, J = 13.0,
.0 Hz, H-7a), 3.35 (1H, ddd, J = 17.5, 11.7, 5.0 Hz, H-4a), 3.59 (1H,
-1), 3.84 (6H, s, OCH
-10), 3.85 (1H, m, H-6a), 6.66 (1H, s, H-3), 6.76 (1H, d,
3 3
H-5), 3.85 (6H, s, OCH -2, OCH -10), 4.07 (1H, dd, J = 12.5, 6.0 Hz,
H-5), 4.47 (1H, dd, J = 13.0, 3.5 Hz, H-6a), 6.64 (1H, s, H-3), 6.86
(1H, d, J = 8.0 Hz, H-9), 7.12 (1H, dd, J = 8.0, 1.0 Hz, H-8), 8.77 (1H,
dd, J = 12.3, 5.0 Hz, H-5a), 3.66 (3H, s, OCH
, OCH
3
3
-
1
3
2
3
s, OH); C NMR (126 MHz, CDCl
3
): d 27.0 (t, C-4), 29.8 (t, C-7),
-10), 62.1 (q, OCH -11),
13
J = 8.0 Hz, H-8), 6.79 (1H, d, J = 8.0 Hz, H-9); C NMR data were
48.3 (q, NCH ), 56.1 (q, OCH -2, OCH
3
3
3
3
consistent with those previously reported (Rasoanaivo et al.,
65.6 (t, C-5), 74.5 (d, C-6a), 110.5 (d, C-3), 111.9 (d, C-9), 119.3 (s,
C-1b), 122.5 (s, C-3a), 125.6 (s, C-1a), 126.1 (d, C-8), 143.9 (s, C-
+
1
998); MS-EI (probe) 70 eV, m/z rel. int.): 327 [M] (100).
1
2
), 144.0 (s, C-11), 151.1 (s, C-2), 152.3 (s, C-10); ESI-MS (probe)
030 V, m/z rel. int.): 358 [M+H] (100).
+
3.8. (6S,6aS,M)-(+)-Corydine-Na-oxide (5)
Brown solid; 1H NMR (500 MHz, CDCl
): d 2.63 (1H, ddd,
J = 13.0, 12.5, 1.0 Hz, H-7), 3.03 (1H, dd, J = 18.0, 6.0 Hz, H-4), 3.14
3.9. (6R,6aS,P)-(+)-Corydine (6)
3
(
(
3H, s, NCH
3
), 3.24 (1H, br ddd, J = 18.0, 12.5, 6.0 Hz, H-4), 3.53
-11), 3.82 (1H, m,
Brown solid; m.p. 154–158 °C (MeOH) [lit. (Shafiee and
2
5
1H, dd, J = 12.5, 3.5 Hz, H-7), 3.66 (3H, s, OCH
3
Jafarabadi, 1998) m.p. 149–150 °C]; ½
aꢄ
þ 266 (c 0.36, CHCl
3
)
D

M.L.R. Ferreira et al. / Phytochemistry 71 (2010) 469–478
477
[
(
2
2
1
lit. (Shafiee and Jafarabadi, 1998) +202 (c 0.20, EtOH)]); 1H NMR
500 MHz, CDCl ): d 2.33 (1H, ddd, J = 13.7, 12.8, 1.0 Hz, H-7b),
.43 (1H, ddd, J = 12.0, 11.2, 3.9 Hz, H-5b), 2.46 (3H, s, NCH ),
.59 (1H, dd, J = 16.0, 3.9 Hz, H-4b), 2.85 (1H, ddd, J = 13.7, 3.5,
.3 Hz, H-6a), 2.93 (1H, ddd, J = 11.2, 6.0, 1.3 Hz, H-5a), 2.96 (1H,
culated for C37
(30).
H
37Cl
2
N
2
O
5
= 659.2088), 631.1789 (100), 603.1508
3
3
3.14. Lagesianine C (11)
2
5
1
dd, J = 12.8, 3.5 Hz, H-7a), 3.08 (1H, br ddd, J = 16.0, 12.0, 6.0 Hz,
H-4a), 3.66 (3H, s, OCH -11), 3.83 (3H, s, OCH -10), 3.84 (3H, s,
OCH -2), 6.61 (1H, s, H-3), 6.80 (1H, d, J = 8.0 Hz, H-9), 7.01 (1H,
dd, J = 8.0, 1.0 Hz, H-8), 8.61 (1H, s, OH); C NMR (126 MHz,
CDCl ): d 29.1 (t, C-4), 35.6 (t, C-7), 44.0 (q, NCH ), 52.8 (t, C-5),
6.0 (q, OCH -2), 56.0 (q, OCH -10), 62.0 (q, OCH -11), 62.8 (d, C-
Brown solid; ½
aꢄ
þ 38 (c 0.01, CHCl
3
); H NMR (CDCl
3
) see Ta-
D
3
3
ble 6; HR-ESI-MS (probe) 4500 V, m/z (rel. int.): 711.2204
+
3
[
6
5
MꢁHCl+K] (100) (calculated for C38
40 2 7
H N O
ꢂHCl + K = 711.2241),
83.2068 (70), 639.1022 (78), 619.2219 (75), 551.2048 (94),
07.1769 (89), 463.1502 (80); HR-ESI-MS (probe) 4500 V, m/z
= 659.2079),
13
3
3
5
6
1
1
3
3
3
(
rel. int.): 659.2077 (70) (calculated for C37
H37Cl
2
N
2
O
5
a), 110.8 (d, C-9), 111.3 (d, C-3), 119.3 (s, C-11a), 124.0 (s, C-3a),
24.3 (d, C-8), 126.5 (s, C-1a), 128.2 (s, C-1b), 130.9 (s, C-7a),
6
31.1841 (100) (calculated for C35
H33Cl
2
N
2
O
5
= 631.1766).
1
5
42.3 (s, C-1), 143.8 (s, C-11), 149.1 (s, C-2), 151.8 (s, C-10);
): d 46.8 (N-6); MS-EI (probe) 70 eV, m/z
rel. int.): 342 [M+H] (100).
N
3.15. Lagesianine D (12)
NMR (50.7 MHz, CDCl
3
+
34
D
Brown solid; m.p. 233–234 °C (MeOH); ½
aꢄ
þ 219 (c 0.02,
1
13
CHCl
3
); H and C NMR (CDCl
3
) see Tables 6 and 7, respectively;
+H]+
10 = 781.2296), 747.2690
44ClN 10 = 747.2684),
13.3029 [Mꢁ2HCl+H] (45) (calculated for
13.3074), 376.1323 (35), 362.1164 (57), 342.1676 (62), 328.1502
3
.10. (6S,6aS,M)-Corydine hydrochloride (7)
HR-ESI-MS (probe) 4500 V, m/z (rel. int.): 781.2356 [Mꢁ2H
2
(
[
7
7
48) (calculated for
40 2 2
C H43Cl N O
Brown solid; 1H NMR (500 MHz, CDCl
): d 2.81 (1H, br dd,
), 3.16 (1H, m,
H-7a), 3.24 (1H, m, H-7b), 3.68 (3H, m, H-5, H-6a), 3.69 (3H, s,
OCH -11), 3.85 (1H, m, H-4a), 3.86 (6H, s, OCH -2, OCH -10), 6.66
1H, s, H-3), 6.86 (1H, d, J = 8.0 Hz, H-9), 7.06 (1H, d, J = 8.0 Hz, H-
+
3
MꢁHClꢁH] (48) (calculated for
C
40
H
2
O
+
J = 17.5, 3.0 Hz, H-4b), 3.02 (3H, d, J = 3.0 Hz, NCH
3
40 45 2 10
C H N O =
3
3
3
(
100).
(
8
1
3
); C NMR (126 MHz, CDCl
3
): d 25.8 (t, C-4), 33.0 (t, C-7), 42.6
-2, OCH -10), 62.1 (q, OCH
3.16. Glycerol (13)
(q, NCH
3
), 52.5 (t, C-5), 56.1 (q, OCH
3
3
3
-
1
1
1
1), 63.1 (d, C-6a), 110.8 (d, C-3), 111.8 (d, C-9), 118.7 (s, C-3a),
20.9 (s, C-11a), 121.2 (s, C-1b), 124.9 (d, C-8), 125.6 (s, C-1a),
1H and 13C NMR data were consistent with those previously re-
ported (Pouchert, 1992).
27.2 (s, C-7a), 144.1 (s, C-11), 144.2 (s, C-1), 150.8 (s, C-2), 152.6
+
(s, C-10); ESI-MS (probe) 2030 V, m/z (rel. int.): 340 [MꢁHClꢁH]
3
.17. N-Methylisocorydine iodide (Menisperine, 14)
(
100).
Brown solid; m.p. 218–222 °C (MeOH) [lit. (Dwuma-Badu et al.,
980) 222–224 °C]; H and C NMR data were consistent with
3.11. (6R,6aS,M)-Corydine hydrochloride (8)
1
13
1
those previously reported (Dwuma-Badu et al., 1980; Marsaioli
et al., 1979).
Brown solid; 1H NMR (500 MHz, CDCl
): d 2.64 (1H, t,
3
J = 13.0 Hz, H-7b), 2.92 (1H, m, H-4a), 2.72 (3H, d, J = 4.5 Hz,
NCH ), 3.86 (1H, m, H-4a), 3.16 (1H, m, H-5b), 3.43 (1H, m, H-5a),
.44 (1H, ddd, J = 13.0, 3.5, 3.0 Hz, H-6a), 3.24 (1H, m, H-7a), 3.65
3H, s, OCH -11), 3.86 (6H, s, OCH -2, OCH -10), 6.66 (1H, s, H-3),
.87 (1H, d, J = 7.5 Hz, H-9), 7.10 (1H, d, J = 7.5 Hz, H-8); C NMR
): d 21.8 (t, C-4), 32.0 (t, C-7), 32.9 (q, NCH ),
9.5 (t, C-5), 56.1 (q, OCH -2, OCH -10), 61.9 (q, OCH -11), 58.1
3
Acknowledgements
4
(
6
(
4
(
3
3
3
1
3
The authors thank the Fundação de Amparo à Pesquisa do Esta-
do de São Paulo (FAPESP) and Conselho Nacional de Desenvolvi-
mento Científico e Tecnológico (CNPq) for financial support, and
CNPq for the fellowships to I.C. de Pascoli, J. Zukerman-Schpector
and L.M.X. Lopes. We also thank Dr. Condorcet Aranha and Dr. Lin-
dolpho Capellari Júnior for plant identification, and Dr. Norberto P.
Lopes for HRMS spectra.
126 MHz, CDCl
3
3
3
3
3
d, C-6a), 110.5 (d, C-3), 111.9 (d, C-9), 119.8 (s, C-11a), 120.0 (s,
C-3a), 120.6 (s, C-1b), 125.5 (d, C-8), 125.6 (s, C-1a), 126.7 (s, C-
a), 143.8 (s, C-11), 144.2 (s, C-1), 151.1 (s, C-2), 152.6 (s, C-10);
ESI-MS (probe) 2030 V, m/z (rel. int.): 340 [MꢁHClꢁH] (100).
7
+
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)
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[
[
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D
3
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Brown solid; m.p. 168–170 °C (MeOH); ½
a
ꢄ
þ 22 (c 0.04,
), see Tables 4 and 5, respec-
tively; HR-ESI-MS (probe) 4500 V, m/z (rel. int.): 741.2369
1
13
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3
+
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M+KꢁHCl] (25) (calculated for C39
H
43ClN
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2
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O
8
+ K = 741.2346),
+ K = 713.2033),
7
6
5
3
4
13.2069 (50) (calculated for
C
37
H
2
8
97.2317 (50), 669.1795 (51), 653.2030 (68), 639.2210 (75),
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ꢁ
500 V, m/z (rel. int.): 659.2073 [MꢁOHꢁOCH
3
ꢁOCH
3
]
(70), (cal-

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