Litebamine N-homologues: Preparation and anti-acetylcholinesterase activity

Article abstract of DOI:10.1021/np970298f

Litebamine N-homologues were easily prepared from laurolitsine, generally via three reaction steps (N-alkylation, solvolysis with 1 M NH₄OAc under reflux, and the Mannich reaction) in more than 80% overall yield. Among the prepared compounds, N-propyl-, N-isobutyl-, and N-isopropylnorlitebamines exhibited moderate antiacetylcholinesterase activity (IC₅₀ ca. 7.0 μM), while the corresponding N-metho salt of N-propylnorlitebamine showed potent activity (IC₅₀ 2.70 μM).

Full text of DOI:10.1021/np970298f

4
6
J . Nat. Prod. 1998, 61, 46-50
Liteba m in e N-Hom ologu es: P r ep a r a tion a n d An ti-Acetylch olin ester a se Activity
†
‡
,†
Chi-Ming Chiou, J aw-J ou Kang, and Shoei-Sheng Lee*
School of Pharmacy, and Institute of Toxicology, College of Medicine, National Taiwan University, 1 J en-Ai Road, Section 1,
Taipei 100, Taiwan, Republic of China
X
Received J une 12, 1997
Litebamine N-homologues were easily prepared from laurolitsine, generally via three reaction
steps (N-alkylation, solvolysis with 1 M NH OAc under reflux, and the Mannich reaction) in
4
more than 80% overall yield. Among the prepared compounds, N-propyl-, N-isobutyl-, and
N-isopropylnorlitebamines exhibited moderate antiacetylcholinesterase activity (IC50 ca. 7.0
µM), while the corresponding N-metho salt of N-propylnorlitebamine showed potent activity
(IC50 2.70 µM).
Litebamine (11) is a novel phenanthrene alkaloid
isolated from the wood of Litsea cubeba.1 It possesses
antiplatelet aggregation activity through the inhibition
of thromboxane B2 formation induced by arachidonic
2
acid in washed rabbit platelets (15 µM, 77% inhibition).
Recently, it was demonstrated to possess activity against
acetylcholinesterase (AChE), with an IC50 value of 22.0
3
µM. The latter biological activity stimulated our inter-
est to study the potential of this type of isoquinoline
alkaloid as an anti-AChE agent. The following de-
scribes our effort in the preparation of litebamine
N-homologues (13-19) and their inhibitory effect on
AChE.
Resu lts a n d Discu ssion
A three-step preparation of litebamine (11) from
boldine (1) via a biogenetic approach has been developed
in our laboratory.4 The method for preparing the key
intermediate secoboldine (1a ) was modified recently to
a facile one-pot reaction via solvolysis of 1 with 1 M NH4-
OAc in EtOH-H2O under reflux, which afforded 1a in
high yield (>90%).5 This modification has increased the
total yield of litebamine (11) up to 80% from boldine
N-allylation and subsequent Hoffman-like degradation
reaction can occur if more reagent is added. These two
successive reactions gave N,N-diallylsecolaurolitsine,
which is almost TLC-inseparable from the desired
(1), a significant improvement compared with the 23%
+
product 7. The diallylseco derivative displays a [M]
obtained after alkylation with iodoacetic acid, followed
at m/z 393 and a base peak at m/z 110, corresponding
to fragment ion A (Figure 1), with R ) allyl and H
6
by a novel rearrangement. Based on these surveys, we
used the solvolysis method to prepare the key interme-
diates, the N-alkylsecolaurolitsines (3a -10a ). These
alkylsecoaporphines were prepared from laurolitsine (2),
which is an abundant aporphine alkaloid in Phoebe
1
replaced by another allyl group. The H-NMR spectra
of 3-8 showed N-alkyl, N-allyl, or N-benzyl signals in
addition to that of laurolitsine, and their EIMS spectra
displayed the common major peak at m/z 284 (fragment
ion B, Figure 1), obtained probably from a retro Diels-
formosana and was readily obtained as crude base (ca.
7
2
5% purity) upon alkalization of the acidic extract.
9
Alder fragmentation process, supporting their struc-
N-Alkyllaurolitsines (3-6) and N-benzyllaurolitsine
8) were prepared from crude 2 via direct reaction with
tures. Solvolysis of these 2-hydroxyaporphines with 1
M NH4OAc in EtOH-H2O (1:1) solution under a reflux
condition overnight gave the corresponding N-alkylseco-
laurolitsines (3a -6a ), N-allylsecolaurolitsine, (7a ) and
N-benzylsecolaurolitsine (8a ) in more than 90% yield
at a 300-mg level. These phenanthrenes showed char-
acteristic UV absorption maxima at 263.0, 279.6, 304.5,
(
the corresponding halide under reflux in alkaline condi-
8
tions. N-Allyllaurolitsine (7), however, could be pre-
pared easily only with suitable molar ratio (ca. 1: 0.72)
of the reactant to reagent (allyl bromide). Because of
the highly reactive property of allyl bromide, exhaustive
1
and 318.4 nm; H-NMR signals for H-9 and H-10 as an
*
To whom correspondence should be addressed. Phone: (886) 2-391-
AB system at δ ca. 7.45 and 7.56 (J AB ) 9.4 Hz);1
and EIMS spectra displaying the base peak (fragment
ion A) at m/z 58 + 14 n (n ) 0 from 3a ; n ) 1 from 4a ;
0,11
6
127. FAX: (886) 2-391-9098. E-mail: shoeilee@ha.mc.ntu.edu.tw.
†
School of Pharmacy.
Institute of Toxicology.
Abstract published in Advance ACS Abstracts, December 15, 1997.
‡
X
S0163-3864(97)00298-X CCC: $15.00
© 1998 American Chemical Society and American Society of Pharmacognosy
Published on Web 01/23/1998

Litebamine N-Homologues
J ournal of Natural Products, 1998, Vol. 61, No. 1 47
Å) in the litebamine N-homologues is close to that of
the corresponding atoms in acetylcholine of the extended
1
4
conformation (ca. 3.815 Å).
This might explain the
higher anti-AChE activity of these compounds. The
repulsion between the N-substituent and the aromatic
nucleus in secoaporphines 3a -6a and 9a , resulting in
3
an N and O distance unsuitable for binding to the
active site of AChE, might account for their low anti-
AChE activity. This study also indicated that the C3
or C4 N-substitution in the N-alkylated norlitebamines
possessed the optimal anti-AChE activity; however, a
planar C3 N-substitution (i.e., allyl group) gave a lower
such activity (15.80 µM for 17 vs 7.21 µM for 14). The
quaternary compound 20, obtained from a treatment of
F igu r e 1. Most significant fragment ions in the mass spectra
of 1a -10a (ions A and B) and 11-19 (ion C).
n ) 2 from 5a and 6a ), 70 (from 7a ), and 120 (from 8a ),
from a â-cleavage fragmentation.10
Mannich reaction of these secondary bases (3a -8a )
1
4 with methyl iodide, was about two and half times
4
with HCHO under acetate buffer conditions (pH 5.76)
more potent than 14, indicating the increased affinity
of the quaternary ammonium to the binding site of
AChE. This study discloses a part of the structure-
activity relationship of litebamine derivatives toward
AChE and will be of value for the development of anti-
AChE agents of isoquinoline-type skeleton.
gave the target products 13-18 in near quantitative
1
yield. The H-NMR spectra of these products showed
the two-proton singlet of H-2a at δ ca. 3.54, and the
EIMS spectra showed a common fragment ion (C) at
m/z 296 (Figure 1) from a retro Diels-Alder fragmenta-
tion process, thus confirming the structures of these
products.
Exp er im en ta l Section
N-Isopropyllaurolitsine (9) could not be obtained by
reacting laurolitsine with isopropyl bromide, probably
due to the steric effect of the isopropyl group to C-7 of
the aporphine moiety. To overcome this steric problem,
the key intermediate for the Mannich reaction, N-
Gen er a l Exp er im en ta l P r oced u r es. The physical
data of the prepared compounds were obtained from the
following instruments: Fisher-J ohns melting point ap-
paratus (uncorrected); Perkin-Elmer 1760-X IR FT
spectrometer; Hitachi 150-20 UV; J EOL J MX-HX110
+
i
isopropylsecolaurolitsine (9a ), [M] at m/z 355, N- Pr at
δ 1.13 (6H, d, J ) 6.3 Hz) and 2.93 (1 H, septet, J ) 6.3
Hz), was prepared by reductive N-alkylation of seco-
laurolitsine (2a ) (Me2CO-NaBH4) (89.8% yield). Com-
(HREIMS) and Finnigan TSQ-700 (EIMS) mass spec-
trometers; Bruker AC-80 and AMX-400 spectrometers
using solvent peak as reference standard.
P r ep a r a t ion of N-Alk ylla u r olit sin es (3-6) a n d
N-Ben zylla u r olitsin e (8). The preparation of N-
ethyllaurolitsine (3) is given as an example. The
mixture of crude laurolitsine (2) (6.0 g, ca. 25% purity),
DMF (60 mL), K2CO3 (2.82 g), and EtI (2.30 mL, 22.4
mmol) was stirred at 50-55 °C for 6 h under nitrogen.
The resulting suspension, after removal of the organic
solvent under reduced pressure, was diluted with H2O
+
pound 2a , [M] at m/z 313, was prepared from N-acetyl-
laurolitsine (10), which was obtained from N-acetylation
of 2 with Ac2O, by acid solvolysis (concd HCl-MeOH )
3
:4, reflux, 1 d, 87.0% yield).12 During the preparation
1
of 2a , N-acetylsecolaurolitsine (10a ), H NMR δ 1.92 (s,
3
H, NHAc), was also isolated from a reaction with
shorter reaction time (1 h). This suggests that 10a may
be the intermediate of 2a , and the mechanism of the
acid solvolysis could be drawn as shown in Figure 2,
suggesting the Hofmann-like ring opening, facilitated
by the aid of an acyl group.
(300 mL) and adjusted to pH 8.0 with 25% ammonia
water. The mixture was partitioned against CHCl3 (200
mL × 3). The combined CHCl3 layers were dried
(anhydrous Na2SO4) and evaporated. The residue was
Under experimental conditions similar to those used
for the preparation of 13-18, 9a gave 19. Along with
purified by Si gel chromatography (1-3% MeOH in
1
CHCl3) to give pure 3 (1.21 g): mp 175-177 °C; H-
1
the characteristic H-NMR signals of litebamine-like
NMR (CDCl3) δ 7.86 (1H, s, H-11), 6.81 (1H, s, H-8),
compounds, the EIMS spectrum of 19 showed the same
major fragment ion (C) at m/z 296 as the other litebam-
ine N-homologues (Figure 1), and ion A at m/z 72
confirming the assigned structure.
6
1
.61 (1H, s, H-3), 3.89 (3H, s, 10-OMe), 3.56 (3H, s,
-OMe), 3.06 (2H, m, N-CH2CH3), 1.11 (3H, t, J ) 7.2
+
Hz, N-CH2CH3); EIMS (70 eV) m/z 341 [M] (100), 326
(66), 284 (26).
This practical method not only gives a high yield of
N-alkylnorlitebamines (>80% in the last two steps) but
also simplifies the workup procedures inasmuch as a
sole product was obtained in each step. In addition, the
purification of the last two-step products, by recrystal-
lization or washing, was very facile.
Under the similar experimental conditions, crude 2
(6.20 g) with 1-bromopropane (2.62 mL, 30.5 mmol)
yielded 4 (1.45 g), crude 2 (5.10 g) with butyl bromide
(2.70 mL, 25.9 mmol) yielded 5 (1.23 g), crude 2 (5.30
g) with isobutyl bromide (2.70 mL, 28.5 mmol) yielded
6 (1.25 g), and crude 2 (2.00 g) with benzyl chloride (0.4
mL, 1.54 mmol) yielded 8 (580 mg).
The anti-AChE activity of these compounds was
1
3
1
evaluated by the colorimetric method.
The results
N-P r op ylla u r olitsin e (4): mp 162-166 °C; H-NMR
indicated that the litebamine N-homologues (13-19)
were more active than their respective secoaporphine
intermediates (3a -6a , and 9a ) (Table 1). Particularly,
those having a bulky group at the nitrogen showed large
differences in their inhibitory activity, such as 6.80 µM
(CDCl3) δ 7.86 (1H, s, H-11), 6.82 (1H, s, H-8), 6.61 (1H,
s, H-3), 3.89 (3H, s, 10-OMe), 3.56 (3H, s, 1-OMe), 3.20
(1H, dd, J ) 13.6, 3.9 Hz, H-7R), 3.12 (1H, ddd, J )
11.4, 5.9, 1.5 Hz, H-5â), 3.01 (1H, ddd, J ) 17.0, 11.4,
5.9 Hz, H-4â), 2.94 (1H, dd, J ) 13.6, 3.9 Hz, H-6a),
2.87 (1H, ddd, J ) 13.0, 9.8, 6.4 Hz) and 2.40 (1H, m)
(N-CH2C2H5), 2.62 (1H, br dd, J ) 17.0, 3.8 Hz, H-4R),
(
19) vs 72.02 µM (9a ). Molecular modeling studies
3
revealed that the distance between N and O (ca. 4.288

4
8
J ournal of Natural Products, 1998, Vol. 61, No. 1
Chiou et al.
F igu r e 2. Proposed mechanism of acid solvolysis of N-acylaporphine 10.
Ta ble 1. Inhibitory Effect of Secolaurolitsines (3a -6a and 9a ),
Litebamine N-Homologues (11, 13-20) on Electric Eel AChE
ammonia water. The mixture was partitioned against
CHCl3 (200 mL × 3). The combined CHCl3 layers were
dried (Na2SO4), and after removal of the solvent, the
extract was purified by Si gel chromatography (1%
MeOH in CHCl3) to give pure 7 (825 mg, 2.34 mmol):
secoaporphine
intermediates
IC50
(µM)
litebamine
N-homologues
IC50
a
(µM)a
1
1
22.00
14.45
7.21
7.79
6.51
3
4
5
6
a
a
a
a
27.22
107.34
110.67
61.99
13
14
15
16
1
mp 175-180 °C; H-NMR (CDCl3) δ 7.86 (1H, s, H-11),
6
1
.81 (1H, s, H-8), 6.62 (1H, s, H-3), 5.96 (1H, ddt, J )
7.2, 10.1, 6.6 Hz, N-CH2CHdCH2), 5.26 (1H, br d, J )
1
1
19
7
8
15.80
10.67
6.80
17.2 Hz, N-CH2CHdCHEHZ), 5.19 (1H, br d, J ) 10.1
Hz, N-CH2CHdCHEHZ), 3.90 (3H, s, 10-OMe), 3.56 (3H,
s, 1-OMe), 3.05 (2H, br d, J ) 6.6 Hz, N-CH2CHdCH2);
9
a
72.02
2
0
2.79
+
EIMS (70 eV) m/z 353 [M] (100), 338 (72), 326 (46),
a
The IC50 values were calculated from the dose-response curve
of at least four concentrations of each tested compound that gave
0-90% inhibition of the AChE activity.
2
84 (21), 163 (21), 72 (13).
P r ep a r a tion of N-Eth yl- (3a ), N-P r op yl- (4a ),
1
N-Bu tyl- (5a ), N-Isobu tyl- (6a ), N-Allyl- (7a ), a n d
N-Ben zylsecola u r olitsin es (8a ). The preparation of
N-benzylsecolaurolitsine (8a ) is given as an example.
The mixture of 8 (350 mg, 0.86 mmol), EtOH (10 mL),
and 1 M NH4OAc(aq) (10 mL) was heated under reflux
overnight and cooled to room temperature. The pure
crystalline product 8a (265 mg, 75.7% yield) was col-
2
.52 (1H, dd, J ) 13.6, 13.6 Hz, H-7â), 2.43 (1H, ddd, J
11.4, 11.4, 3.8 Hz, H-5R), 1.57 (2H, m, N-CH2CH2-
)
CH3), 0.94 (3H, t, J ) 7.4 Hz, N-C2H4CH3); COSY-45
data, δ 3.20 to δ 2.94 and 2.52; δ 3.12 to δ 3.01, 2.62,
and 2.43; δ 3.01 to δ 3.12, 2.62, and 2.43; δ 2.62 to δ
3
.12, 3.01, and 2.43; δ 2.94 to δ 3.20 and 2.52; δ 2.87 to
1
lected by suction. 8a : mp 88-92 °C; H-NMR (CDCl3)
δ 9.12 (1H, s, H-5), 7.70 (1H, d, J ) 9.1 Hz, H-10), 7.39
δ 2.40 and 1.57; δ 2.40, to δ 2.87 and 1.57, δ 1.57 to δ
2
3
+
.87, 2.40 and 0.94; EIMS (70 eV) m/z 355 [M] (100),
(
1H, d, J ) 9.1 Hz, H-9), 7.26 (1H, s, H-8), 7.12 (1H, s,
40 (51), 326 (46), 284 (21), 163 (21), 72 (13).
H-2), 4.04 (3H, s, 6-OMe), 3.84 (3H, s, 4-OMe), 3.83 (2H,
1
N-Bu tylla u r olitsin e (5): mp 98-100 °C; H-NMR
s, N-CH C H ), 3.24 (2H, m, H-11), 3.03 (2H, m, H-12);
2
6
5
+
δ 7.86 (1H, s, H-11), 6.82 (1H, s, H-8), 6.62 (1H, s, H-3),
.91 (3H, s, 10-OMe), 3.56 (3H, s, 1-OMe), 2.88 (1H, m)
EIMS (20 eV) m/z 403 [M] (12), 284 (100), 269 (9), 120
3
(54), 91 (14).
and 2.43 (1H, m) (N-CH2C3H7), 1.53 (2H, m, N-CH2-
Under a similar experimental condition, 3 (500 mg,
.47 mmol) yielded 3a (450 mg, 90.0% yield), 4 (500 mg,
CH2C2H5), 1.38 (2H, m, N-C2H4CH2CH3), 0.94 (3H, t, J
1
+
)
7.2 Hz, N-C3H6CH3); EIMS (70 eV) m/z 369 [M]
1.41 mmol) yielded 4a (484 mg, 96.8% yield), 5 (500 mg,
1.36 mmol) yielded 5a (483 mg, 96.6% yield), 6 (500 mg,
1.36 mmol) yielded 6a (475 mg, 95.0% yield), and 7 (500
mg, 1.42 mmol) yielded 7a (490 mg, 98.0% yield).
Melting points of 3a -7a are as follows: 154-156 °C
(100), 354 (51), 338 (19), 326 (60), 297 (14), 284 (36).
1
N-Isobu tylla u r olitsin e (6): mp 84-86 °C; H-NMR
CDCl3) δ 7.86 (1H, s, H-11), 6.82 (1H, s, H-8), 6.62 (1H,
(
s, H-3), 3.90 (3H, s, 10-OMe), 3.55 (3H, s, 1-OMe), 2.88
(
(
3a ), 164-168 °C (4a ), 148-150 °C (5a ), 106-110 °C
6a ), and 165-170 °C (7a ). For the other physical and
(1H, m) and 2.48 (1H, m) (N-CH2iC3H7), 1.85 [1H, m,
N-CH2CH(CH3)2], 0.93 [6H, d, J ) 7.2 Hz, N-CH2CH-
1
5
+
spectral data ( H-NMR and MS), see Lee et al.
(
CH3)2]; EIMS (70 eV) m/z 369 [M] (56), 326 (100), 297
P r ep a r a tion of Secola u r olitsin e (2a ). The mix-
ture of crude 2 (10.0 g), DMF (60 mL), and Ac2O (2 mL)
was stirred at room-temperature overnight in a sealed
tube, and the solvent was evaporated under reduced
pressure to give an amorphous residue that was recrys-
tallized from MeOH (50 mL) to give pure N-acetyllau-
(
19), 284 (6), 277 (28), 263 (27), 163 (12).
1
N-Ben zylla u r olitsin e (8): mp 96-98 °C; H-NMR
CDCl3) δ 7.88 (1H, s, H-11), 7.33 (5H, m, N-CH2C6H5),
.83 (1H, s, H-8), 6.62 (1H, s, H-3), 4.30 (1H, d) and 3.32
1H, d) (J AX ) 13.7 Hz, N-CH2C6H5), 3.91 (3H, s, 10-
(
6
(
+
OMe), 3.57 (3H, s, 1-OMe); EIMS (70 eV) m/z 403 [M]
1
rolitsine (10) (3.60 g): mp 272-274 °C; H-NMR (MeOH-
d4) δ 8.06 (1H, s, H-11), 6.77 (s) and 6.72 (s) (1H, H-8),
(100), 388 (46), 372 (16), 324 (28), 284 (39), 269 (16),
1
20 (16), 91 (70).
6
.61 (1H, s, H-3), 3.89 (3H, s, 10-OMe), 3.58 (3H, s,
P r ep a r a tion of N-Allylla u r olitsin e (7). The mix-
ture of crude laurolitsine (2) (10.0 g, ca. 25% purity, ca.
mmol), DMF (40 mL), KHCO3 (2.82 g), and allyl
bromide (0.5 mL, 5.78 mmol) was stirred at 50-55 °C
for 6 h under nitrogen. The resulting suspension, after
removal of organic solvent under reduced pressure, was
diluted with H2O (300 mL) and adjusted to pH 8.0 with
1-OMe), 2.20 (s) and 2.19 (s) (3H, NAc); EIMS (20 eV)
m/z 355 [M] (88), 296 (74), 283 (100), 269 (42), 240 (18).
+
8
The solution of 10 (500 mg, 1.41 mmol), MeOH (8 mL),
and 37% HCl (6 mL) was stirred under reflux for 24 h
in a sealed tube. The cooled solution, after removal of
organic solvent under reduced pressure, yielded a
brownish solid residue that was washed with H2O to

Litebamine N-Homologues
J ournal of Natural Products, 1998, Vol. 61, No. 1 49
+
give pure secolaurolitsine hydrochloride (2a .HCl) (430
mg, 87.3% yield): mp 236-238 °C; H-NMR (MeOH-
d4) δ 9.12 (1H, s, H-5), 7.76 (1H, d, J ) 9.1 Hz, H-10),
eV) m/z 354 (22), 353 [M] (100), 338 (22), 296 (68), 281
1
(16), 140 (15); HREIMS m/z 353.1617 (calcd for C21H23-
NO4, 353.1627).
7
.53 (1H, d, J ) 9.1 Hz, H-9), 7.20 (1H, s, H-8), 7.13
Under a similar experimental condition, 4a (300 mg,
0.85 mmol) yielded 14 (284 mg, 91.6% yield), 5a (300
mg, 0.81 mmol) yielded 15 (282.4 mg, 91.0% yield), 6a
(300 mg, 0.81 mmol) yielded 16 (280 mg, 90.4% yield),
7a (400 mg, 1.13 mmol) yielded 17 (385 mg, 93.1%
yield), 8a (51.6 mg, 128 µmol) yielded 18 (50.5 mg, 95.0%
yield), and 9a (51.6 mg, 145 µmol) yielded 19 (44.1 mg,
82.9% yield).
(1H, s, H-2), 4.05 (3H, s, 6-OMe), 3.84 (3H, s, 4-OMe),
3
1
2
3
.37 (2H, m, H-11), 3.24 (2H, m, H-12); mp of 2a 166-
+
70 °C; EIMS of 2a (20 eV) m/z 313 [M] (95), 284 (100),
69 (44); HREIMS m/z 313.1324 (calcd for C18H19NO4,
13.1314).
An experiment with a shorter reaction time (1 h) than
that of 10 gave an additional product, N-acetylsecolau-
1
rolitsine (10a ): Rf 0.34 (10% MeOH-CHCl3); mp 219-
N-P r op yln or liteba m in e (14): mp 175-177 °C; H-
1
2
21 °C; H-NMR (MeOH-d4) δ 9.11 (1H, s, H-5), 7.79
NMR (MeOH-d4) δ 9.12 (1H, s, H-5), 7.59 (1H, d, J )
9.1 Hz, H-10), 7.56 (1H, d, J ) 9.1 Hz, H-9), 7.17 (1H,
s, H-8), 4.02 (3H, s, 6-OMe), 3.76 (3H, s, 4-OMe), 3.78
(2H, s, H-2a), 3.15 (2H, m, H-11), 2.96 (2H, m, H-12),
2.59 (2H, t, J ) 7.4 Hz, N-CH2C2H5), 1.53 (2H, m,
N-CH2CH2CH3), 0.94 (3H, t, J ) 7.4 Hz, N-CH2-
(
7
3
1H, d, J ) 9.1 Hz, H-10), 7.43 (1H, d, J ) 9.1 Hz, H-9),
.18 (1H, s, H-8), 7.05 (1H, s, H-2), 4.06 (3H, s, 6-OMe),
.84 (3H, s, 4-OMe), 3.45 (2H, m, H-11), 3.18 (2H, m,
+
H-12), 1.92 (3H, s, NAc); EIMS (20 eV) m/z 355 [M]
(100), 296 (78), 283 (35), 263 (12); HREIMS m/z 355.1437
+
(calcd for C20H21NO5, 355.1420). A treatment of 10 (100
CH2CH3); EIMS (20 eV) m/z 368 (23), 367 [M] (97), 338
mg, 0.28 mmol) with 50% H2SO4 (5 mL) in a sealed tube
at 80 °C for 3.5 h also yielded 10a (50 mg, 50%) after
neutralization of the reaction mixture, which was puri-
fied through an Amberlite XAD-2 column, washed with
H2O, and eluted with MeOH.
(100), 296 (58), 281 (16); HREIMS m/z 367.1779 (calcd
for C22H25NO4, 367.1783).
N-Bu tyln or liteba m in e (15): mp 115-118 °C; H-
NMR (DMSO-d ) δ 8.91 (1H, s, H-5), 7.60 (1H, d, J )
1
6
9.1 Hz, H-10), 7.43 (1H, d, J ) 9.1 Hz, H-9), 7.19 (1H,
P r ep a r a tion of N-Isop r op ylsecola u r olitsin e (9a ).
To a solution of 2a .HCl (200 mg, 0.57 mmol), MeOH (5
mL) and acetone (0.5 mL) was added NaBH4 (200 mg)
portionwise and the resulting suspension was stirred
for 2 h at room temperature. The reaction mixture, after
the removal of organic solvents, was diluted with water
s, H-8), 3.93 (3H, s, 6-OMe), 3.70 (3H, s, 4-OMe), 3.47
(2H, s, H-2a), 3.04 (2H, m, H-11), 2.72 (2H, m, H-12),
2.50 (2H, m, N-CH2C3H7), 1.52 and 1.33 (2H each, m,
N-CH2CH2CH2CH3), 0.89 (3H, t, J ) 7.3 Hz, N-C3H7CH3);
+
EIMS (20 eV) m/z 382 (24), 381 [M] (100), 366 (10),
3
55 (10), 339 (40), 338 (92), 296 (55), 281 (8); HREIMS
m/z 381.1946 (calcd for C23H27NO4, 381.1940).
(30 mL) and was adjusted to pH 8.0 with 25% ammonia
water. The mixture was passed over an Amberlite
XAD-2 column (40 g) washed with distilled water (300
mL) to remove the inorganic salt, then was eluted with
MeOH (250 mL). Evaporation of the solvent afforded
N-Isobu tyln or liteba m in e (16): mp 120-124 °C;
1
H-NMR (ΜeÃΗ-d4) δ 8.92 (1H, s, H-5), 7.61 (1H, d, J
) 9.0 Hz, H-10), 7.44 (1H, d, J ) 9.0 Hz, H-9), 7.19 (1H,
s, H-8), 3.93 (3H, s, 6-OMe), 3.71 (3H, s, 4-OMe), 3.54
(2H, s, H-2a), 3.04 (2H, m, H-11), 2.70 (2H, m, H-12),
2.27 (2H, d, J ) 7.2 Hz, N-CH2iC3H7), 1.90 [1H, m,
N-CH2CH(CH3)2], 0.88 [6H, d, J ) 6.5 Hz, N-CH2CH-
1
pure 9a (182 mg, 89.8% yield): mp 135-140 °C; H-
NMR (MeOH-d4) δ 9.12 (1H, s, H-5), 7.73 (1H, d, J )
9
.1 Hz, H-10), 7.44 (1H, d, J ) 9.1 Hz, H-9), 7.18 (1H,
+
s, H-8), 7.08 (1H, s, H-2), 4.04 (3H, s, 6-OMe), 3.84 (3H,
s, 4-OMe), 3.21 (2H, m, H-11), 2.95 (2H, m, H-12), 2.93
(CH3)2]; EIMS (20 eV) m/z 382 (8), 381 [M] (33), 339
(22), 338 (100), 296 (4); HREIMS m/z 381.1939 (calcd
for C23H27NO4, 381.1940).
[
6
2
1H, septet, J ) 6.3 Hz, N-CH(CH3)2], 1.13 [6H, d, J )
.3 Hz, N-CH(CH3)2]; EIMS (20 eV) m/z 355 [M] (18),
84 (100), 269 (8), 72 (44); HREIMS m/z 355.1749 (calcd
+
N-Allyln or liteba m in e (17): mp 151-154 °C; 1H-
NMR (MeOH-d ) δ 9.12 (1H, s, H-5), 7.67 (1H, d, J )
4
for C21H23NO4, 355.1783).
9.0 Hz, H-10), 7.45 (1H, d, J ) 9.0 Hz, H-9), 7.19 (1H,
s, H-8), 4.04 (3H, s, 6-OMe), 3.78 (3H, s, 4-OMe), 3.77
P r ep a r a tion of N-Eth yl- (13), N-P r op yl- (14),
N-Bu tyl- (15), N-Isobu tyl- (16), N-Allyl- (17), N-
Ben zyl- (18), a n d N-Isop r op yln or liteba m in es (19).
The preparation of 13 is given as an example. The
mixture of N-ethylsecolaurolitsine (3a ) (300.0 mg, 0.88
mmol), MeOH (20 mL), 0.1 M AcOH (8 mL), 1 M NaOAc
(
2H, s, H-2a), 3.22 (2H, ψt, J ) 5.8 Hz, H-11), 2.90 (2H,
ψt, J ) 5.8 Hz, H-12), 3.29 (2H, br d, J ) 6.6 Hz, N-CH2-
CHdCH2), 6.02 (1H, ddt, J ) 17.2, 10.2, 6.6 Hz,
N-CH2CHdCH2), 5.37 (1H, br d, J ) 17.2 Hz, N-CH2-
CHdCHEHZ), 5.30 (1H, br d, J ) 10.2 Hz, N-CH2-
+
CHdCHEHZ); EIMS (20 eV) m/z 366 (22), 365 [M]
(8 mL), and 37% HCHO (1 mL) in a 100-mL round-
(
(
100), 350 (16), 296 (58), 281 (14); HREIMS m/z 365.1650
calcd for C22H23NO4, 365.1627).
N-Ben zyln or liteba m in e (18): mp 130-132 °C; H-
bottom flask was stirred at room temperature for 5 h.
After removal of MeOH, the aqueous residue was
diluted with distilled H2O (20 mL) and partitioned
against CHCl3 (20 mL × 3). The combined CHCl3 layers
were dried (Na2SO4) and evaporated to give crude 13.
The residue was washed with CHCl3 and MeOH to give
1
NMR (DMSO-d6) δ 8.92 (1H, s, H-5), 7.63 (1H, d, J )
9.1 Hz, H-10), 7.46 (1H, d, J ) 9.1 Hz, H-9), 7.39 (1H,
s, H-8), 7.39 (5H, m, N-CH2C6H5), 3.94 (3H, s, 6-OMe),
3.70 (3H, s, 4-OMe), 3.73 (2H, s, N-CH2C6H5), 3.58 (2H,
s, H-2a), 3.10 (2H, m, H-11), 2.80 (2H, m, H-12); EIMS
1
pure 13 (289.0 mg, 93.0% yield): mp 154-156 °C; H-
NMR (DMSO-d6) δ 8.90 (1H, s, H-5), 7.61 (1H, d, J )
+
9
.1 Hz, H-10), 7.43 (1H, d, J ) 9.1 Hz, H-9), 7.18 (1H,
(20 eV) m/z 416 (14), 415 [M] (58), 400 (22), 296 (78),
s, H-8), 3.92 (3H, s, 6-OMe), 3.72 (3H, s, 4-OMe), 3.51
2H, s, H-2a), 3.06 (2H, ψt, J ) 5.8 Hz, H-11), 2.72 (2H,
281 (48), 210 (10), 165 (18), 91 (100); HREIMS m/z
415.1774 (calcd for C26H25NO4, 415.1784).
N-Isop r op yln or liteba m in e (19): mp 225-227 °C;
H-NMR (DMSO-d6) δ 9.03 (1H, s, H-5), 7.68 (1H, d, J
(
ψt, J ) 5.8 Hz, H-12), 2.55 (2H, q, J ) 7.1 Hz, N-CH2-
CH3), 1.30 (3H, t, J ) 7.1 Hz, N-CH2CH3); EIMS (20
1

5
0
J ournal of Natural Products, 1998, Vol. 61, No. 1
Chiou et al.
)
9.0 Hz, H-10), 7.45 (1H, d, J ) 9.0 Hz, H-9), 7.19 (1H,
DU-650 spectrophotometer. All operations were per-
formed in the dark due to the photosensitivity of the
AChE.
s, H-8), 4.04 (3H, s, 6-OMe), 3.79 (3H, s, 4-OMe), 3.87
(2H, s, H-2a), 3.23 (2H, m, H-11), 2.97 (2H, m, H-12),
2
.98 [1H, m, N-CH(CH3)2], 1.25 [1H, d, J ) 6.5 Hz,
Ack n ow led gm en t. This research was supported by
NSC, R.O.C., under grant NSC 85-2331-B002-241 and
NSC 87-2314-B002-005.
+
N-CH(CH3)2]; EIMS (20 eV) m/z 368 (13), 367 [M] (87),
3
m/z 367.1777 (calcd for C22H25NO4, 367.1784).
52 (100), 310 (25), 296 (37), 281 (22), 165 (23); HREIMS
P r ep a r a tion of N-P r op yln or liteba m in e N-Meth -
oiod id e (20). The mixture of N-propylnorlitebamine
Refer en ces a n d Notes
(1) Wu, Y. C.; Liou, J . Y.; Duh, C. Y.; Lee, S. S.; Lu, S. T. Tetrahedron
Lett. 1991, 32, 4169-4170.
(14) (50 mg, 136 µmol), CH3CN (4.0 mL), and MeI (4.0
(
2) Teng, C. M.; Hsueh, C. M.; Chang, Y. L.; Ko, F. N.; Lee, S. S.;
mL) was stirred at 40 °C for 6 h. After evaporation of
Liu, K. C. S. J . Pharm. Pharmacol. 1997, 49, 706-711.
the solvents, the precipitate was washed with MeOH
(3) Kang, J . J ; Lee, S. S. unpublished data.
1
(4) Lee, S. S.; Lin, Y. J .; Chen, M. Z.; Wu, Y. C.; Chen, C. H.
Tetrahedron Lett. 1992, 33, 6309-6310.
(
5 mL) to give a pure product (20) (61.2 mg, 92.5%): H-
NMR (DMSO-d6) δ 8.94 (1H, s, H-5), 7.68 (1H, d, J )
.1 Hz, H-10), 7.59 (1H, d, J ) 9.1 Hz, H-9), 7.26 (1H,
s, H-8), 4.69 (1H, d) and 4.63 (1H, d) (J AB ) 16.2 Hz)
(
5) Lee, S. S.; Chiou, C. M.; Lin, H. Y.; Chen, C. H. Tetrahedron
Lett. 1995, 36, 1531-1532.
9
(
6) Hara, H.; Kaneko, K.; Endoh, M.; Uchida, H.; Hoshino, O.
Tetrahedron 1995, 51, 10189-10192.
(H-2a), 3.96 (3H, s, 6-OMe), 3.80 (2H, m, H-12), 3.75
(3H, s, 4-OMe), 3.45 (4H, m, H-11 and N-CH2C2H5), 3.16
(3H, s, N-Me), 1.91 (2H, m, N-CH2CH2CH3), 1.03 (3H,
(7) Lee, S. S.; Tsai, F. Y.; Chen, I. S.; Liu, K. C. S. J . Chin. Chem.
Soc. 1993, 40, 209-212.
(8) Neumeyer, J . L.; Law, S. J .; Meldrum, B.; Anlezark, G.; Watling,
K. J . J . Med. Chem. 1981, 24, 898-899.
t, J ) 7.4 Hz, N-C2H4CH3); EIMS (70 eV) m/z 381 [M -
(9) Shamma, M. The Isoquinoline Alkaloids; Academic Press: New
York, 1972; p 219.
10) Ibid. pp 259-264.
11) For review of the phenanthrene alkaloids, see Guinaudeau, H;
Lebœuf, M.; Cav e´ , A. J . Nat. Prod. 1975, 38, 275-338; 1979,
+
+
+
HI] (8), 367 [M - MeI] (12), 338 [M - C3H7I] (18),
(
(
+
3
24 (24), 296 (30), 281 (32), 142 [MeI] (100), 127 (58).
Assa y of a n tiACh E Activity. The enzymatic activ-
ity of AChE (EC 3.1.1.7) purified from electric eel (type
4
1
2, 325-360; 1983, 46, 761-835; 1988, 51, 389-474; 1994, 57,
033-1135.
V-S, Sigma Co.) was determined by the colorimetric
(12) Hufford, C. D.; Sharma, A. S.; Oguntimein, B. O. J . Pharm. Sci.
1980, 69, 1180-1183.
13) Ellman, G. L.; Courtney, K. D.; Andres, V., J r.; Featherstone,
1
3
method using acetylcholine iodide as substrate and
(
5
,5′-dithio-(bis-2-nitrobenzoic acid) (DTNB) as a coupler.
R. M. Biochem. Pharmacol. 1961, 7, 88-95.
(14) Molecular modeling was performed on an Indy workstation
Briefly, AChE (0.125 U) was incubated in phosphate
buffer (0.1 M, 1 mL, pH 8.0) containing DTNB (0.3 mM)
and test samples for 2 min at 37 °C. The reaction was
initiated by adding acetylcholine (0.5 mM), and the
activity was measured spectrophotometrically at 412 nm
as a function of time using a Beckman (Fullerton, CA)
(
(
Silicon Graphic Inc.) using SYBYL software (version 6.03)
Tripos Associates Inc.) to obtain a conformation of minimum
energy for litebamine and a zigzag extended conformation of
minimum energy for acetylcholine, of which the distance be-
tween the corresponding O and N atoms was measured.
NP970298F