Isolation and total syntheses of two new alkaloids, dubiusamines-A, and -B, from Pandanus dubius

Article abstract of DOI:10.1016/j.tet.2010.02.073

Chemical investigation of the crude base of Pandanus dubius led to the isolation of two new alkaloids, dubiusamine-A (1) and dubiusamine-B (2). The structures of the two alkaloids including their absolute configuration were unambiguously confirmed by 1D a

Full text of DOI:10.1016/j.tet.2010.02.073

Tetrahedron 66 (2010) 3353–3359
Contents lists available at ScienceDirect
Tetrahedron
journal homepage: www.elsevier.com/locate/tet
Isolation and total syntheses of two new alkaloids, dubiusamines-A, and -B,
from Pandanus dubius
,
Mario A. Tan a, Mariko Kitajima a, Noriyuki Kogure a, Maribel G. Nonato b, Hiromitsu Takayama a
*
a Graduate School of Pharmaceutical Sciences, Chiba University, 1-33 Yayoi-cho, Inage-ku, Chiba 263-8522, Japan
b College of Science, University of Santo Tomas, Espana, Manila 1015, Philippines
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 9 February 2010
Received in revised form 22 February 2010
Accepted 22 February 2010
Available online 25 February 2010
Chemical investigation of the crude base of Pandanus dubius led to the isolation of two new alkaloids,
dubiusamine-A (1) and dubiusamine-B (2). The structures of the two alkaloids including their absolute
configuration were unambiguously confirmed by 1D and 2D NMR analysis and asymmetric total
synthesis.
Ó 2010 Elsevier Ltd. All rights reserved.
Keywords:
Pandanus
Alkaloid
Isolation
Total synthesis
1. Introduction
contain alkaloids. To date, 16 alkaloids were identified from the
genus Pandanus. As part of our research on novel biologically active
The genus Pandanus (family Pandanaceae) comprises approxi-
mately 700 species distributed in tropical and sub-tropical regions.
Several Pandanus species are used as medicinal plants for treating
leprosy, rheumatism, epilepsy, and sore throat.1 The isolation of
alkaloids with a novel C9–N–C9 unit from Pandanus amaryllifolius
had impelled interest in other Pandanus plants that may also
alkaloids from the genus Pandanus,2 we conducted a phytochemical
analysis on Pandanus dubius. In this paper, we report the isolation of
dubiusamine-A (1) and dubiusamine-B (2) from the crude base of
P. dubius (Fig. 1), their structure elucidation by means of spectro-
scopic analysis, and their asymmetric total synthesis to determine
their absolute configuration.
H3C21
O
3
2
4
O
5
O
O
6
2'O
O
2
11
CH3
H31C1'
7
3
3'
9'
9
7
8
7'
5
5'
4
N
H
4'
9
6
6'
8'
8
H
N
O
11
12
14
O
1
18
15
H
17
13
16
CH3
2
20
Figure 1. Structures of new alkaloids from P. dubius.
* Corresponding author: Tel./fax: þ81 43 2902901; e-mail address: htakayam@
p.chiba-u.ac.jp.
0040-4020/$ – see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2010.02.073

3354
M.A. Tan et al. / Tetrahedron 66 (2010) 3353–3359
2. Results and discussion
Examination of the structure of 1 revealed the presence of two
stereocenters d one at C-3 and the other at C-5. Detailed analysis of
both 1H and 13C NMR spectra showed the occurrence of an in-
separable 5:1 diastereomeric mixture. In particular, additional
The crude base that was obtained using the conventional acid-
base extraction procedure from a MeOH extract of P. dubius was
separated by extensive column chromatography to obtain two new
alkaloids, dubiusamine-A (1) and dubiusamine-B (2). In addition,
seven known alkaloids, including pandamarilactonines-A,2b -B,2b
and -C,2c pandamarilactones-1,3 -313 and -32,3 and pandamar-
ilactam-3y,4 were isolated. The known alkaloids were identified by
comparing their spectroscopic data (NMR and MS) with reported
values. The structures of 1 and 2 were unambiguously identified
minor signals at d d
4.34 and 2.49 (1H) and 37.3, 35.9, and 15.1 (13C)
were observed in the NMR spectra. To determine the stereochem-
ical relationship of the substituents at C-3 and C-5, differential NOE
experiment was carried out. Irradiation of the proton at
H-50) led to a clear peak enhancement of the methyl protons at
1.28 (H-11, H-110). Hence, an anti relationship between the C-3
d 4.51 (H-5,
d
and C-5 methine protons is disclosed for the major diastereomer
and a syn relationship is found5 for the minor diastereomer.
Although natural product 1 was obtained as a diastereomeric
mixture, the total synthesis allowed for its isolation and charac-
terization in pure form.
using spectroscopic and synthetic methods.
20
Dubiusamine-A (1) was obtained as an optically active {[a]
D
þ29.5 (c 0.07, CHCl3)} amorphous solid in 0.12% yield (based on the
crude alkaloid fraction). The molecular formula was established as
C18H31NO4 using HR-FABMS with its protonated molecular ion peak
[MþH]þ at m/z 326.2341 (calcd for C18H32NO4, 326.2331). The 13C
NMR spectrum revealed the presence of nine carbons, symptomatic
of a symmetrical structure. Through 13C NMR and DEPT experi-
To unambiguously confirm the structure including its absolute
configuration, the asymmetric total synthesis of 1 was conducted
(Scheme 1). The synthesis started with the monoprotection of
1,6-hexanediol with TBSCl6 and the oxidation of the remaining
alcohol to give aldehyde 3.7 A three-step reaction that included the
ments, a lactone carbonyl (
d
180.1), an oxygenated methine (
d 78.3),
a methylene associated to a nitrogen atom (
d
49.7), and further
proline-mediated a
-aminoxylation8 of aldehyde 3 followed by the
upfield methylenes, methine, and methyl were detected. The IR
absorption band at 1759 cmꢀ1 validated the presence of a carbonyl
of a lactone moiety. The 1H NMR spectrum showed a total count of
15 hydrogen atoms in the molecule. Supported by 1H–1H COSY data
Horner–Emmons reaction and the O–N bond cleavage utilizing
CuSO4 in MeOH9 afforded highly enantioselective ester 4 in 77%
yield (over three steps) and 99% ee as determined by chiral HPLC
analysis.10 Based on the well-established mechanistic consideration
(Fig. 2), a 3,5-disubstituted
g-butyrolactone was identified via
for proline-catalyzed aldehyde
chemistry of the chiral center (C-4) in 4 was concluded to be R.
Corresponding -butyrolactone 5 was furnished by subsequent
hydrogenation using 10% Pd/C in MeOH. Treatment of 5 with LHMDS
and MeI allowed the diastereoselective -methylation of the lactone
a
-aminoxylation,8,9,11 the stereo-
correlations of protons associated to the alpha going to the gamma
carbons. The linkage of the methyl group at C-3 with the n-butyl
group at C-5 was also observed by further examination of the COSY
spectrum. The gross structure of 1 was established by linking two
units of the nine carbon segment to a nitrogen atom. This was
g
a
ring to give 6 in a 7:1 (anti/syn) ratio.12 The removal of the TBS group
in 6 was easily achieved using camphorsulfonic acid in MeOH at 0 ꢁC
to afford alcohol 7 in quantitative yield. The Mitsunobu reaction13
employing a mixture of alcohol 7, 2-nitrobenzenesulfonamide,
PPh3, and DEAD in THF/toluene gave, in 71% yield, corresponding
nosyl-protected amine 8. This compound was again treated with
PPh3 and DEAD in the presence of alcohol 7 using toluene as solvent
to provide nosyl-protected symmetrical amide 9. To complete the
synthesis of 1, deprotection of the nosyl group in amide 9 was ac-
complished with K2CO3 and PhSH. The product was carefully puri-
corroborated by the chemical shift of the terminal carbon (d 49.7,
C-9, C-90) of the butyl moiety. The HMBC experiment further
supported the proposed structure. Long-range couplings were
observed between the doublet methyl protons (H-11) and C-2, C-3,
and C-4, and between the H-4 protons and C-2, C-3, and C-5. HMBC
cross-peaks were also observed between the H-9 protons and C-7
and C-8 carbons.
O
O
2'
2
1H-1H COSY
Key HMBC
11
fied using silica gel chromatography to afford diastereomerically
11'
3
O
5'
O
5
CH3
23
pure 1 {[a]
D
þ61.2 (c 0.05, CHCl3)}. The spectroscopic data (IR, 1H
7
H3C
9
8'
3'
6'
and 13C NMR, HRMS) of synthetic 1 were identical to those of the
natural product, thereby establishing the structure including the
absolute configuration (3R, 5R) of dubiusamine-A.
N
H
9'
7'
4
4'
8
6
Figure 2. COSY and selected HMBC correlations of 1.
OR
a
O
b
OH
TBSO
O
CO2CH3
HO
TBSO
R=NHPh: 4a
3
e
1,6-Hexanediol
R=H:
4
O
O
O
c
h
CH3
R
R
R=OH:
5a
R=OTBS: 6
7
d
f
R=OTBS: 5
R=OH:
g
R=NHNs: 8
O
O
O
O
O
O
O
O
i
CH3
CH3
N
H3C
N
H3C
H
Ns
1
9
Scheme 1. Synthetic route for dubiusamine-A. Reagents and conditions: (a) (i) TBSCl, NaH, THF, rt, 3 h; (ii) SO3-pyridine complex, DMSO, Et3N, CH2Cl2, 0 ꢁC to rt, 3 h, 80% in two
steps; (b) (i) L-Proline, Nitrosobenzene, CH3CN, ꢀ20 ꢁC, 20 h; (ii) Methyl diethyl phosphonoacetate, NaH, THF, 0 ꢁC, 1 h; (iii) CuSO4, MeOH, rt, 15 h, 77% in three steps; (c) 10% Pd/C,
MeOH, H2, rt, 45 h; (d) TBSCl, Imidazole, CH2Cl2, rt, 13 h, 96% in two steps; (e) LHMDS, MeI, THF, ꢀ78 ꢁC, 6 h, 67% (dr 7:1); (f) CSA, MeOH, 0 ꢁC, 1 h, quant.; (g) NsNH2, PPh3,
DEAD,THF/Toluene, rt, 1 h, 71%; (h) Compound 7, PPh3, DEAD, Toluene, 60 ꢁC, 1 h, 80%; (i) K2CO3, PhSH, DMF/CH3CN, rt, 3 h, 96%.

M.A. Tan et al. / Tetrahedron 66 (2010) 3353–3359
3355
Dubiusamine-B (2) was obtained as an optically active, amor-
To confirm the structure and establish the absolute stereo-
chemistry of 2, its total synthesis was attempted. As the absolute
configuration at C-14 and C-15 in natural pandamarilactonine-A
has already been demonstrated to be R and R,2f respectively, we
18
phous solid {[
a
]
þ13.6 (c 0.025, CHCl3)}. In its HR-FABMS spec-
D
trum, the protonated molecular ion peak [MþH]þ at m/z 322.2028
(calcd for C18H28NO4, 322.2018) agreed well with the molecular
formula C18H27NO4. 1H, 13C (Table 1) and DEPT NMR spectra
indicated two lactone carbonyls, five methines (two oxygen-
bearing, one nitrogen-bearing and one sp2), eight methylenes
(two nitrogen-bearing), two methyls, and one olefinic quaternary
carbon. The observed 1H and 13C NMR signals at {dH 7.06 (m, H-16),
4.81 (ddd, J¼5.0, 1.8, 1.6, H-15), 1.94 (d, J¼0.4, H-20); dC 174.4 (C-18),
146.9 (C-16), 131.3 (C-17), 83.7 (C-15), 10.8 (C-20)} and at {dH 2.80
(ddd, J¼8.0, 5.5, 5.0, H-14); dC 54.3 (C-11), 25.9 (C-12), 23.9 (C-13),
have decided to use
D-prolinol as our chiral starting material
(Scheme 2).
D
-Prolinol was converted into known aldehyde 112f in
two steps, and this was followed by vinylation of the aldehyde
group to obtain diastereomeric adducts 12 in a 1:2 (i.e., less polar/
more polar) ratio. After chromatographic separation of the
diastereomers, their absolute configuration was determined by
converting them into their oxazolidinone derivatives,14 16a and 16b
(Scheme 3). Measurement of the coupling constants (JH1,7a) and
NOE crosspeak observation15 between H-7a and H-1 for both 16a
and 16b clearly indicated a (20R, 1S, erythro) configuration for the
less polar adduct and (20R, 1R, threo) configuration for the more
polar adduct. Removal of the Cbz group in the more polar adduct
using TMSI in CH3CN afforded amino alcohol 13. Condensation of 13
65.4 (C-14)} are characteristic of a pyrrolidinyl-
a-methyl-a,b-un-
saturated -lactone moiety. This is a unique structural feature of
g
known alkaloids pandamarilactonines-A and -C having a threo
configuration at C-14 and C-15 positions.2b,c The structure of 2 was
further concluded on the basis of 1H–1H COSY, HMQC, and HMBC
spectroscopic analysis (Fig. 3). Both 1H and 13C NMR signals for
with the g-butyrolactone iodide 10 having a 3R, 5R configuration in
upper portion of 2 having the 3,5-disubstituted
coherent to those of 1. In the HMBC spectrum, correlations in the
pyrrolidinyl- -methyl- -unsaturated -lactone between H-14
and C-9/C-16; H-9 and C-11; H-15 and C-16; H-16 and C-18/C-20;
and H-20 and C-17/C-18 were observed. Thus, the gross structure of
dubiusamine-B was determined as in 2.
g
-butyrolactone are
the presence of Ag2CO3 in CH3CN led to the formation of in-
termediate adduct 14. Esterification of the hydroxyl group in 14
with methacrylic anhydride in the presence of Et3N and DMAP in
CH2Cl2 easily gave ester 15. The synthesis of 2 was completed by
employing Grubbs second-generation catalyst in the presence of
p-TsOH under reflux.16 The spectroscopic data (1H NMR, 13C NMR)
a
a,
b
g
including the high-resolution mass spectrometric and optical
17
rotation {[
a
]
þ22 (c 0.30, CHCl3)} data of the synthetic material,
D
were in excellent agreement with those of the natural product.
Thus, the structure including the absolute configuration (3R, 5R,
14R, 15R) of 2 was established.
Table 1
1H and 13C NMR data for 2 in CDCl3
Position
dH (500 MHz)
dC (125 MHz)
2
3
4
180.0
34.0
3. Experimental section
2.70, m
2.03, ddd (10.0, 7.2, 6.6)
2.12, ddd (10.5, 7.0, 4.0)
4.51, m
1.70–1.80, 2H, overlapped
1.46–1.63, 2H, overlapped
1.46–1.63, 2H, overlapped
2.87, ddd (10.8, 7.9, 7.5)
2.40, ddd (11.0, 7.5, 6.0)
3.12, dd (7.5, 6.5)
35.5a
3.1. General experimental procedures
5
6
7
8
9
78.3
35.4a
23.2
28.7
55.6
Optical rotations were measured on a JASCO P-1020 polarimeter.
IR spectra were recorded on JASCO FTIR-230 spectrophotometer.
Low- and high-resolution FABMS were recorded on JEOL JMS-
HX110 or JEOL JMS-AX500 mass spectrometer. HR-ESIMS were
recorded on a Thermo Fisher Scientific Exactive spectrometer. NMR
spectra were recorded on JEOL JNM A-500 and JEOL JNM ECP400
spectrometers. The chemical shifts are given in
pling constants, in hertz. Kieselgel 60 [Merck, 70–230 mesh
(for open column chromatography)], Silica gel 60 N [Kanto Chem-
11
12
54.3
25.9
2.20, m
1.70–1.80, overlapped
1.46–1.63, overlapped
1.70–1.80, 2H, overlapped
2.80, ddd (8.0, 5.5, 5.0)
4.81, ddd (5.0, 1.8, 1.6)
7.06, m
d (ppm) and cou-
13
14
15
16
17
18
20
21
23.9
65.4
83.7
146.9
131.3
174.4
10.8
ical, 40–50 mm (for flash column chromatography)], Chromatorex
NH [Fuji Silysia Chemical, (100–200 mesh), and Al2O3 90 (Merck,
Basic, Activity II–III) were used for column chromatography.
Medium-pressure liquid chromatography was carried out on a sil-
ica gel prepacked column CPS-HS-221-05 (Kusano Kagakukikai).
1.94, 3H, d (0.4)
1.28, 3H, d (7.5)
15.9
a
Interchangeable.
3.2. Plant material
The fresh leaves of P. dubius were collected at PICC, Manila,
Philippines, and identified by Assistant Professor Rosie Madulid,
Department of Biological Sciences, College of Science, University of
Santo Tomas. A voucher specimen (USTH-4846) was deposited at
the Plant Sciences Herbarium, Research Center for the Natural
Sciences, University of Santo Tomas.
21
H3C
3
O
2
4
O
5
1H-1H COSY
Key HMBC
3.3. Extraction and isolation
H
N
O
18
Air-dried and ground leaves of P. dubius (4.0 kg) were extracted
with 5 L of MeOH (3ꢂ24 h) at room temperature and filtered. The
combined filtrates were concentrated in vacuo to obtain the crude
alcoholic extract (370 g). A portion of the crude extract (155 g) was
partitioned between EtOAc and 1 N HCl. The aqueous layer was
basified with Na2CO3 (pH 9–10) and exhaustively extracted with 5%
O
11
14
15
17
H
CH3
20
Figure 3. COSY and selected HMBC correlations of 2.

3356
M.A. Tan et al. / Tetrahedron 66 (2010) 3353–3359
O
O
3
O
O
a
CH3
5
CH3
HO
I
7
10
OH
OH
H
Cbz
N
H
N
Cbz
H
H
N
1
b
N
c
2
d
CHO
OH
2'
H
H
3
13
12
11
D-Prolinol
H3C
O
O
3
O
18
H3C
O
O
H3C
O
2
5
O
5'
CH3
g
10, e
f
3'
H
H
O
OH
16
2'
N
N
14
H
N
O
14
O
4'
15
15
H
17
CH3
14
15
2
Scheme 2. Synthetic route for dubiusamine-B. Reagents and conditions: (a) I2, PPh3, Imidazole, CH2Cl2, rt, 3.5 h, 96%; (b) CbzCl, K2CO3, CH3CN, ꢀ20 ꢁC, 4 h; then SO3–pyridine
complex, DMSO, Et3N, CH2Cl2, 0 ꢁC, 3.5 h, 92% in two steps; (c)Vinyl magnesium bromide, THF, 0 ꢁC, 3 h, 73% (dr 2:1); (d) TMSI, CH3CN, ꢀ15 ꢁC to 0 ꢁC, 30 min 53%; (e) 10, Ag2CO3,
CH3CN, rt, 45 h, 65%; (f) Methacrylic anhydride, Et3N, DMAP, CH2Cl2, rt, 4 h, 71%; (g) p-TsOH, Grubbs second-generation catalyst, CH2Cl2, reflux, 19 h, 61%.
20
3.3.1. Dubiusamine-A (1). Amorphous solid; [
a
]
D
þ29.5 (c 0.07,
CHCl3); IR (ATR) nmax 2936, 1759, 1160 cmꢀ1 1H NMR (CDCl3,
;
O
O
500 MHz)
d
4.51 (2H, dddd, J¼13.0, 8.0, 8.0, 5.0 Hz, H-5, H-50), 2.69
OH
1
N
N
Cbz
N
2'
K2CO3, 2-Propanol / H2O (1:1)
reflux, 70 oC
O
(2H, m, H-3, H-30), 2.61 (4H, dd, J¼6.8, 6.8 Hz, H2-9, H2-90), 2.12
(2H, ddd, J¼10.4, 6.6, 4.0 Hz, H-4b, H-40b), 2.00 (2H, ddd, J¼10.4, 6.0,
6.0 Hz, H-4a, H-40a), 1.70 (2H, m, H-6, H-60), 1.60–1.46 (10H, m,
overlapped, H-6, H-60, H2-7, H2-70, H2-8, H2-80),1.28 (6H, d, J¼7.0 Hz,
O
7a
2
7a
1
1
H
H
16a
H
16b
H
H
3
12
(from less polar adduct) (from more polar adduct)
JH1,7a = 7.0 Hz
JH1,7a = 4.2 Hz
H3-11, H3-110); 13C NMR (CDCl3, 125 MHz) 180.1 (C, C-2, C-20), 78.3
d
NOE = not observed
NOE = 5.5%
(CH, C-5, C-50), 49.7 (CH2, C-9, C-90), 35.5 (CH2, C-4, C-40), 35.4
(CH2, C-6, C-60), 34.0 (CH, C-3, C-30), 29.7 (CH2, C-8, C-80), 23.3
(CH2, C-7, C-70), 15.9 (CH3, C-11, C-110); HR-FABMS: calcd for
C18H32NO4 [MþH]þ: 326.2331, found: 326.2341.
Scheme 3. Determination of absolute configuration.
MeOH/CHCl3. The organic layer was dried with anhydrous MgSO4
and evaporated to afford the crude alkaloidal fraction (2.1 g).
A portion of the base (2 g) was subjected to silica gel flash column
chromatography using CHCl3/MeOH gradient to yield nine frac-
tions: F1 (48.1 mg), CHCl3; F2 (103.7 mg), 1% MeOH/CHCl3; F3
(119.0 mg), 1–3% MeOH/CHCl3; F4 (121.0 mg), 3–5% MeOH/CHCl3;
F5 (266.9 mg), 5–10% MeOH/CHCl3; F6 (408.0 mg), 10–15% MeOH/
CHCl3; F7 (148.8 mg), 15–20% MeOH/CHCl3; F8 (299.2 mg), 20–50%
MeOH/CHCl3; F9 (92.4 mg), 50% MeOH/CHCl3/MeOH. F2 was sub-
jected to silica gel open column chromatography (twice, 5% MeOH/
CHCl3) and MPLC (2% EtOH/CHCl3) to afford pandamarilactone-1
(1.8 mg). F3 was subjected to gradient CHCl3/MeOH silica gel
open column chromatography to obtain sub-fractions F3A–F3D.
Sub-fraction F3A was purified by repeated silica gel open column
chromatography (CHCl3/MeOH gradient) to afford pandamari
lactonine-A (2.4 mg) and pandamarilactonine-B (1.1 mg). Sub-
fraction F3B was purified by MPLC (50% EtOAc/hexaned2% EtOH/
CHCl3) to afford pandamarilactam-3y (3.1 mg). F4 was subjected to
silica gel open column chromatography (CHCl3/MeOH gradient),
MPLC (twice, 3% MeOH/CHCl3) and amino-silica gel column
chromatography (CHCl3) to afford pandamarilactone-31 (1.2 mg)
and dubiusamine-B (2) (0.5 mg). Repeated silica gel open column
chromatography of F5 (CHCl3/MeOH gradient) and further
purification by amino-silica gel column chromatography gave
pandamarilactonine-A (1.1 mg), pandamarilactonine-C (0.8 mg),
and pandamarilactone-32 (1.3 mg). Silica gel open column chro-
matography of F7 (CHCl3/MeOH gradient) gave mixtures of
pandamarilactonines-A and -B or pandamarilactonines-A and -C.
Fraction F9 was purified by silica gel (15–30% MeOH/
CHCl3dMeOH) and amino-silica gel (thrice, 5% MeOH/CHCl3) open
column chromatography to give dubiusamine-A (1) (2.3 mg).
18
3.3.2. Dubiusamine-B (2). Amorphous solid; [
a]
þ13.6 (c 0.025,
D
CHCl3); UV lmax 209, 243, 273 nm; 1H and 13C NMR data, see
Table 1; FABMS (NBA) m/z 322 [MþH]þ; HR-FABMS: calcd for
C18H28NO4, [MþH]þ: 322.2018, found: 322.2028.
3.4. Preparation and characterization of the new compounds
3.4.1. Ester (4). To a stirred solution of 3 (100.8 mg, 0.440 mmol)
and nitrosobenzene (94.0 mg, 0.880 mmol, 2 equiv) in CH3CN
(1.1 mL) was added L-proline (20.2 mg, 0.180 mmol, 0.4 equiv) at
ꢀ20 ꢁC and the mixture was stirred for 20 h. In another reaction
flask, methyl diethyl phosphonoacetate (0.16 mL, 0.880 mmol,
2 equiv) was added dropwise to a stirred solution of NaH (35.2 mg,
0.880 mmol, 2 equiv) in THF (1 mL) in an ice bath. The mixture was
stirred at room temperature for 45 min before transferring to the
above solution of 3 at 0 ꢁC via a cannula. The resulting mixture was
stirred for 1 h at 0 ꢁC and then quenched with cold saturated NH4Cl,
and the resulting aqueous layer was extracted three times with
EtOAc. The combined organic layers were washed with brine, dried
over MgSO4, and concentrated under reduced pressure. Purification
by silica gel flash column chromatography (20% EtOAc/hexane)
afforded 4a (149.0 mg, 86% yield) as an orange-amorphous solid. To
a stirred solution of 4a (149.0 mg, 0.380 mmol) in MeOH (1.9 mL)
was added CuSO4 (18.1 mg, 0.114 mmol, 0.3 equiv) at room tem-
perature and the mixture was stirred for 15 h. The reaction mixture
was quenched with saturated NH4Cl and the aqueous layer was
extracted three times with CHCl3. The combined CHCl3 layers were
washed with brine, dried over MgSO4, and concentrated under
reduced pressure. Title compound 4 was obtained after silica gel

M.A. Tan et al. / Tetrahedron 66 (2010) 3353–3359
3357
flash column chromatography (20% EtOAc/hexane) in 89% yield
(102.0 mg, 77% yield over three steps) as an orange-amorphous
6.3 Hz, H2-9), 2.69 (1H, m, H-3), 2.12 (1H, ddd, J¼10.8, 7.0, 6.9 Hz,
H-4a), 2.00 (1H, ddd, J¼10.7, 7.0, 7.0 Hz, H-4b), 1.76–1.43 (6H, m,
H2-6, H2-7, H2-8), 1.28 (3H, d, J¼7.0 Hz, H3-10); 13C NMR (CDCl3,
24
solid; [
a
]
ꢀ44.1(c 0.3, CHCl3); UV (MeOH) lmax 206 nm; IR (ATR)
D
nmax 3437, 2929, 2857, 1725, 1659, 1471, 1253 cmꢀ1; 1H NMR (CDCl3,
400 MHz)
125 MHz) d 180.1 (C, C-2), 78.3 (CH, C-5), 62.5 (CH2, C-9), 35.4
d
6.95 (1H, dd, J¼15.6, 4.8 Hz, H-3), 6.05 (1H, dd, J¼16.0,
(CH2, C-4), 35.2 (CH2, C-6), 34.0 (CH, C-3), 32.2 (CH2, C-8), 21.7
(CH2, C-7), 15.9 (CH3, C-10); HRESIMS: calcd for C9H16O3Na
[MþNa]þ: 195.0992, found: 195.0992.
2.0 Hz, H-2), 4.32 (1H, m, H-4), 3.74 (3H, s, OCH3), 3.62 (2H, dd, J¼6.0,
6.0 Hz, H2-8), 1.63–1.47 (6H, m, H2-5, H2-6, H2-7), 0.90 (9H, s,
C(CH3)3), 0.05 (6H, s, Si(CH3)2); 13C NMR (CDCl3, 100 MHz)
d 166.9
(C, C-1), 150.4 (CH, C-3), 119.7 (CH, C-2), 71.0 (CH, C-4), 70.0 (CH2,
C-8), 51.6 (CH3, OCH3), 36.3 (CH2, C-7), 32.3 (CH2, C-5), 25.9 (CH3,
C(CH3)3), 21.6 (CH2, C-6), 18.3 (C, C(CH3)3), ꢀ5.5 (CH3, Si(CH3)2);
HRESIMS: calcd for C15H31O4Si, [MþH]þ: 303.1986, found: 303.1979.
3.4.4. Nosyl-protected amine (8). To a stirred solution of 7 (15.0 mg,
0.087 mmol) in THF (0.4 mL) and toluene (0.6 mL) were added
triphenylphosphine (29.7 mg, 0.113 mmol, 1.3 equiv) and 2-nitro-
benzenesulfonamide (44.0 mg, 0.218 mmol, 2.5 equiv), respectively,
at room temperature. The solution was allowed to cool in an ice bath
3.4.2.
g-Butyrolactone (5). To a stirred solution of 4 (102.0 mg,
and DEAD (40% in toluene, 49 mL, 0.113 mmol, 1.3 equiv) was added
0.337 mmol) in MeOH (6.7 mL) was added 10% Pd/C (50.0 mg).
The reaction mixture was stirred under hydrogen atmosphere
(balloon) for 45 h and filtered using a pad of Celite (EtOAc). The
filtrate was concentrated under reduced pressure. The crude
material was subjected to silica gel flash column chromatography
(EtOAc) to afford 5a (54.1 mg) as colorless oil in quantifiable yield.
To a solution of 5a (54.1 mg, 0.342 mmol) in CH2Cl2 (0.68 mL) was
added imidazole (46.6 mg, 0.684 mmol, 2 equiv) and the resulting
mixture was stirred in an ice bath. TBSCl (62.0 mg, 0.410 mmol,
1.2 equiv) was added and the mixture was stirred for an addi-
tional 1 h in an ice bath. The temperature of the mixture was
allowed to return to room temperature and stirring was
commenced for 13 h. The reaction mixture was quenched with
saturated NH4Cl and the aqueous layer was extracted three times
with CHCl3. The combined organic layers were dried over MgSO4
and concentrated under reduced pressure. Purification by silica
gel flash column chromatography (30% EtOAc/hexane) afforded
dropwise. The reaction mixture was stirred at room temperature for
1 h. The mixture was filtered using a Celite pad and the filtrate was
concentrated under reduced pressure. Title compound 8 was
obtained after purification using MPLC (50% EtOAc/hexane) and
silica gel flash column chromatography (hexane/EtOAc/CHCl3¼1/
23
0.5/0.5) as a light-yellow oil in 71% yield (22.1 mg); [
a
]
D
þ31.8
(c 0.033, CHCl3); UV (MeOH) lmax 204, 221 (sh) nm; IR (ATR) nmax
2935, 2866, 1755, 1537, 1338, 1161 cmꢀ1; 1H NMR (CDCl3, 400 MHz)
d
8.14 (1H, m, Nosyl group), 7.88 (1H, m, Nosyl group), 7.76 (2H, m,
Nosyl group), 5.27 (1H, dd, J¼5.8, 5.8 Hz, NH), 4.46 (1H, m, H-5), 3.10
(2H, q, J¼6.4, 2.0 Hz, H2-9), 2.65 (1H, m, H-3), 2.07 (1H, ddd, J¼12.4,
8.8, 5.6 Hz, H-4a), 1.99 (1H, ddd, J¼12.2, 8.8, 5.6 Hz, H-4b), 1.67–1.47
(6H, m, H2-6, H2-7, H2-8), 1.27 (3H, d, J¼7.2 Hz, H3-10); 13C NMR
(125 MHz, CDCl3)
d 179.8 (C, C-2), 148.1 (C), 133.7 (CH), 133.6 (C),
132.8 (CH), 131.1 (CH), 125.4 (CH) [Nosyl group], 77.9 (CH, C-5), 43.5
(CH2, C-9), 35.4 (CH2, C-6), 34.9 (CH2, C-4), 34.0 (CH, C-3), 29.3 (CH2,
C-8), 22.5 (CH2, C-7), 15.9 (CH3, C-10); HRESIMS: calcd for
C15H20N2O6NaS [MþNa]þ: 379.0934, found: 379.0922.
title compound 5 as a colorless oil in 96% yield over two steps
22
(89.6 mg); [
a
]
ꢀ35.4 (c 0.3, CHCl3); IR (ATR) nmax 2922, 2858,
D
1750, 1213 cmꢀ1
;
1H NMR (CDCl3, 400 MHz)
d
4.50 (1H, m, H-5),
3.4.5. Nosyl-protected symmetrical amide (9). To a stirred mixture
of 8 (17.5 mg, 0.050 mmol), 7 (12.9 mg, 0.075 mmol, 1.5 equiv), and
triphenylphosphine (17.0 mg, 0.065 mmol, 1.3 equiv) in toluene
3.62 (2H, dd, J¼6.2, 6.2 Hz, H2-9), 2.33 (1H, ddd, J¼13.6, 6.8,
6.8 Hz, H-4), 2.53 (2H, m, H2-3), 1.91–1.49 (7H, m, H-4, H2-6, H2-7,
H2-8), 0.90 (9H, s, C(CH3)3), 0.05 (6H, s, Si(CH3)2); 13C NMR (CDCl3,
(0.32 mL) was added DEAD (40% in toluene, 28 mL, 0.065 mmol,
100 MHz)
d
177.2 (C, C-2), 81.0 (CH, C-5), 62.8 (CH2, C-9), 35.3
1.3 equiv) dropwise at 0 ꢁC (ice bath). The mixture was heated at
60 ꢁC for 1 h. After cooling, the reaction mixture was filtered using
a Celite pad and the filtrate was concentrated under reduced pres-
sure. The crude residue was purified by MPLC (90% EtOAc/hexane) to
(CH2, C-6), 32.4 (CH2, C-8), 28.8 (CH2, C-3), 28.0 (CH2, C-4), 25.9
(CH3, C(CH3)3), 21.7 (CH2, C-7), 18.3 (C, C(CH3)3), ꢀ5.3 (CH3,
Si(CH3)2); HRESIMS: calcd for C14H29O3Si [MþH]þ: 273.1886,
found: 273.1895.
obtain title compound 9 as a light-yellow amorphous solid in 80%
22
yield (20.5 mg); [
a]
þ20.9 (c 0.02, CHCl3); UV (MeOH) lmax
1H NMR
8.00 (1H, m, Nosyl group), 7.71 (2H, m, Nosyl
D
3.4.3. Alcohol (7). A solution of 5 (124.0 mg, 0.455 mmol) in THF
(1.0 mL) was added to lithium bis(trimethylsilyl)amide (1.0 M in
THF, 0.5 mL, 0.500 mmol, 1.1 equiv) at ꢀ78 ꢁC via a cannula. The
mixture was stirred for 1 h at ꢀ78 ꢁC and methyl iodide (0.426 mL,
6.82 mmol, 15 equiv) was added dropwise to it. The mixture was
stirred for another 5 h before it was allowed to warm at ꢀ20 ꢁC
and quenched with 1 N HCl. The resulting mixture was extracted
three times with CHCl3 and the combined organic layers were
washed with brine, dried over MgSO4, and evaporated under
reduced pressure. Compound 6 was obtained after silica gel flash
column chromatography (20% EtOAc/hexane) as a colorless oil in
67% yield (87.0 mg, dr 7:1, anti/syn; FABMS (NBA): m/z 287
[MþH]þ). To a stirred solution of 6 (56.0 mg, 0.200 mmol) in
MeOH (2.0 mL) was added camphorsulfonic acid (CSA, 19.1 mg,
0.040 mmol, 0.2 equiv) at 0 ꢁC. After 1 h, the reaction mixture was
quenched with saturated NaHCO3 and the aqueous layer was
extracted three times with CHCl3. The combined organic layers
were dried over MgSO4 and evaporated under reduced pressure.
Purification by silica gel flash column chromatography (20%
206 nm; IR (ATR) nmax 2937, 1759, 1542, 1455, 1159 cmꢀ1
(CDCl3, 400 MHz)
;
d
group), 7.62 (1H, m, Nosyl group), 4.46 (2H, m, H-5, H-50), 3.28 (4H,
m, H2-9, H2-90), 2.67 (2H, m, H-3, H-30), 2.07 (2H, ddd, J¼12.4, 8.8,
5.6 Hz, H-4b, H-4b0), 1.99 (2H, ddd, J¼12.2, 8.8, 5.6 Hz, H-4a, H-4a0),
1.65–1.38 (12H, m, H2-6, H2-60, H2-7, H2-70, H2-8, H2-80), 1.27 (6H, d,
J¼7.2 Hz, H3-10, H3-100); 13C NMR (CDCl3, 125 MHz)
d 180.0 (C, C-2,
C-20), 148.0 (C), 133.5 (CH), 133.4 (C), 132.7 (CH), 130.7 (CH), 124.1
(CH) [Nosyl group], 78.0 (CH, C-5, C-50), 47.1 (CH2, C-9, C-90), 35.4
(CH2, C-6, C-60), 34.8 (CH2, C-4, C-40), 33.9 (CH, C-3, C-30), 27.8 (CH2,
C-8, C-80), 22.5 (CH2, C-7, C-70), 15.9 (CH3, C-10, C-100); HRESIMS:
calcd for C24H34N2O8NaS [MþNa]þ: 533.1928, found: 533.1916.
3.4.6. Synthetic dubiusamine-A (1). To a stirred solution of 9
(20.0 mg, 0.039 mmol) in DMF (50
added K2CO3 (10.8 mg, 0.078 mmol, 2 equiv) and thiophenol
(5.2 L, 0.051 mmol, 1.3 equiv) and the mixture was stirred at room
mL) and CH3CN (80 mL) were
m
temperature for 3 h. The reaction mixture was filtered using a Celite
pad and the filtrate was evaporated under reduced pressure. The
crude residue was purified by silica gel open column chromatog-
MeOH/CHCl3) afforded title compound 7 as a colorless oil in
22
quantifiable yield; [
a
]
þ40 (c 0.01, CHCl3); IR (ATR) nmax 3419,
raphy (5% MeOH*/CHCl3, MeOH*¼10% NH3 in MeOH) to afford
D
23
2934, 2866, 1752, 1455, 1185 cmꢀ1
;
1H NMR (CDCl3, 400 MHz)
synthetic 1 in 96% yield (12.3 mg); [
a
]
þ61.2 (c 0.05, CHCl3);
D
d
4.51 (1H, dddd, J¼13.0, 8.0, 8.0, 5.5 Hz, H-5), 3.67 (2H, dd, J¼6.3,
HRESIMS: calcd for C18H32NO4 [MþH]þ: 326.2326, found:

3358
M.A. Tan et al. / Tetrahedron 66 (2010) 3353–3359
326.2317; IR, 1H, and 13C NMR data of synthetic 1 were identical
with those of natural 1.
2966, 2875, 1631, 1536 cmꢀ1 1H NMR (CDCl3, 400 MHz)
; d 5.90
(1H, ddd, J¼17.0,10.0, 6.8 Hz, H-2), 5.33 (1H, dd, J¼17.0,1.6 Hz, H-3a),
5.17 (1H, dd, J¼10.0, 1.6 Hz, H-3b), 3.73 (1H, dd, J¼9.5, 6.8 Hz, H-1),
3.11 (1H, ddd, J¼8.0, 7.2, 7.2 Hz, H-20), 2.97 (1H, m, H-50a), 2.88
(1H, m, H-50b), 1.85–1.56 (4H, m, H2-30, H2-40); 13C NMR (CDCl3,
3.4.7.
g-Butyrolactone iodide (10). To a stirred solution of 7
(16.0 mg, 0.093 mmol) in CH2Cl2 (4.7 mL) were added triphenyl-
phosphine (44.0 mg, 0.170 mmol, 1.8 equiv) and imidazole (17.0 mg,
0.240 mmol, 2.6 equiv). The mixture was cooled in an ice bath and
then I2 (43 mg, 0.170 mmol, 1.8 equiv) was added. The mixture was
stirred at room temperature for 3.5 h. Saturated aqueous Na2S2O3
(4.7 mL) was added to the mixture and the mixture was further
stirredfor15 min. The mixturewasquenchedwithsaturatedNaHCO3
and extracted three times with CHCl3. The combined organic layers
were washed with brine, dried over MgSO4, and concentrated under
reduced pressure. Title compound 10 was obtained after purification
100 MHz) d 135.4 (CH, C-2),119.7 (CH2, C-3), 72.1 (CH, C-1), 64.6 (CH,
C-20), 45.7 (CH2, C-50), 27.2 (CH2, C-40), 24.1 (CH2, C-30); HRESIMS:
calcd for C7H14NO [MþH]þ: 128.1070, found: 128.1076.
3.4.10. Condensed adduct (14). To a stirred solution of 13 (10.0 mg,
0.079 mmol) and 10 (27.0 mg, 0.095 mmol, 1.2 equiv) in CH3CN
(1.0 mL) was added Ag2CO3 (50 wt % on Celite, 65.3 mg, 0.119 mmol,
1.5 equiv). The reaction mixture was stirred at room temperature for
45 h. The heterogeneous mixture was then filtered using a Celite pad
and the filtrate was concentrated under reduced pressure. Purifica-
tion by silica gel open column chromatography (50% EtOAc/hexane to
using MPLC (30% EtOAc/hexane) as a colorless oil in 96% yield
28
(25.0 mg); [
a
]
þ72 (c 0.02, CHCl3); IR (ATR) nmax 1755 cmꢀ1
;
1H
D
NMR (CDCl3, 400 MHz)
d
4.52 (1H, dddd, J¼12.8, 7.6, 7.6, 4.8 Hz, H-5),
5% MeOH/CHCl3) afforded title compound 14 as a colorless oil in 65%
19
3.20 (2H, dd, J¼7.0, 7.0 Hz, H2-9), 2.70 (1H, m, H-3), 2.13 (1H, ddd,
J¼12.8, 9.0, 5.2 Hz, H-4a), 2.03(1H, ddd, J¼12.8, 7.6, 7.6 Hz,H-4b),1.87
(2H, quin, J¼7.2 Hz, H2-8), 1.75–1.49 (4H, m, H2-6, H2-7), 1.29 (3H, d,
yield (14.2 mg); [
d
a
]
þ56.4 (c 0.17, CHCl3); 1H NMR (CDCl3, 400 MHz)
D
5.83 (1H, ddd, J¼17.0,10.0, 5.5 Hz, H-16), 5.31 (1H, dd, J¼17.1,1.5 Hz,
H-17a), 5.13 (1H, dd, J¼10.0,1.5 Hz, H-17b), 4.50(1H, dddd, J¼13.0, 8.0,
8.0, 5.5 Hz, H-5), 3.73 (1H, dd, J¼5.5, 5.5 Hz, H-15), 3.12 (1H, ddd,
J¼10.0, 6.5, 4.0 Hz, H-11a), 2.75 (1H, m, H-9a), 2.68 (2H, overlapped,
H-3, H-14), 2.43 (1H, m, H-9b), 2.37 (1H, m, H-11b), 2.10 (1H, ddd,
J¼12.5, 8.5, 5.0 Hz, H-4a), 2.00 (1H, m, H-4b),1.89 (1H, m, H-12a),1.76
(2H, m, H2-13),1.72–1.51 (6H, m, H2-6, H-7, H2-8, H-12b),1.38 (1H, m,
J¼7.2 Hz, H3-11); 13C NMR (CDCl3, 125 MHz)
d 179.9 (C, C-2), 78.0
(CH, C-5), 35.4 (CH2, C-6), 34.3 (CH2, C-4), 34.0 (CH, C-3), 32.9 (CH2,
C-8), 26.4 (CH2, C-7),15.9 (CH3, C-11), 6.2 (CH2, C-9); HRESIMS: calcd
for C9H15O2INa [MþNa]þ: 305.0002, found: 305.0009.
3.4.8. Adduct (12). To a stirred solution of 11 (1.2 g, 5.15 mmol) in
THF (10.3 mL) at 0 ꢁC was added vinyl magnesium bromide (7.7 mL,
7.73 mmol, 1.5 equiv) and the reaction mixture was stirred for 3 h.
The mixture was quenched with saturated NH4Cl and the resulting
aqueous mixture was extracted three times with CHCl3. The com-
bined organic layers were washed with brine, dried over MgSO4,
and evaporated under reduced pressure. Title compound 12 was
obtained as a 2:1 diastereomeric mixture after silica gel flash col-
umn chromatography (40% EtOAc/hexane) in 73% yield (990.0 mg).
H-7), 1.28 (3H, d, J¼7.0 Hz, H3-18); 13C NMR (CDCl3, 100 MHz)
d 180.0
(C, C-2), 139.8 (CH, C-16), 115.2 (CH2, C-17), 78.3 (CH, C-5), 74.2 (CH,
C-15), 68.8 (CH, C-14), 57.1 (CH2, C-9), 54.1 (CH2, C-11), 35.4 (CH2, C-4),
35.3 (CH2, C-6), 34.0 (CH, C-3), 28.4 (CH2, C-12), 28.2 (CH2, C-8), 24.3
(CH2, C-13), 23.0 (CH2, C-7), 15.9 (CH3, C-18); HRESIMS: calcd for
C16H28NO3 [MþH]þ: 282.2064, found: 282.2057.
3.4.11. Ester (15). To a stirred solution of 14 (11.4 mg, 0.041 mmol) in
CH2Cl2 (0.41 mL) at 0 ꢁC were added Et3N (17
m
L, 0.123 mmol, 3 equiv)
and DMAP (5.0 mg, 0.041 mmol, 1 equiv). Methacrylic anhydride
(14 L, 0.090 mmol, 2.2 equiv) was then added dropwise and the
Separation of the diastereomers was achieved using MPLC (20%
22
EtOAc/hexane); Less polar diastereomer (erythro): [
a
d
]
þ56.2
m
D
(c 0.5, CHCl3); 1H NMR (DMSO-d6, VT 100, 400 MHz)
7.35–7.28
resulting mixture was stirred at room temperature for 4 h. The reaction
was quenched with saturated NH4Cl and the aqueous layer was
extracted three times with CHCl3. The combined organic layers were
washed with brine, dried over MgSO4, and concentrated under
reduced pressure. Purification by silica gel column chromatography
(5H, m, Ph of Cbz), 5.80 (1H, ddd, J¼16.0, 10.8, 5.2 Hz, H-2), 5.19
(1H, dd, J¼17.2, 2.0 Hz, H-3a), 5.10 (2H, s, CH2 of Cbz), 5.05 (1H, dd,
J¼10.4, 1.8 Hz, H-3b), 4.63 (1H, d, J¼5.2 Hz, OH), 4.38 (1H, br s, H-1),
3.79 (1H, m, H-20), 3.46 (1H, m, H-50a), 3.31 (1H, m, H-50b), 1.81
(4H, m, H2-30, H2-40); 13C NMR (DMSO-d6, VT 100, 150 MHz)
d
153.3
(3% MeOH/CHCl3) afforded title compound 15 as a colorless oil in 71%
19
(C]O of Cbz), 136.2 (C), 127.2 (CH), 126.5 (CH), 126.3 (CH) [Ph of
yield (10.2 mg); [
d
a
]
þ16.4 (c 0.06, CHCl3); 1H NMR (CDCl3, 400 MHz)
D
Cbz], 138.7 (CH, C-2), 113.1 (CH2, C-3), 70.7 (CH, C-1), 64.8 (CH2 of
6.14 (1H, br s, H-40a), 5.95 (1H, ddd, J¼15.6, 10.0, 4.8 Hz, H-16), 5.57
Cbz), 60.7 (CH, C-20), 45.9 (CH2, C-50), 23.9 (CH2, C-30), 22.5 (CH2,
(1H, br d, J¼1.6 Hz, H-40b), 5.35 (1H, dd, J¼5.2, 5.2 Hz, H-15), 5.26
(1H, dd, J¼15.6, 1.6 Hz, H-17a), 5.22 (1H, dd, J¼9.8, 1.6 Hz, H-17b), 4.51
(1H, dddd, J¼13.0, 8.0, 8.0, 5.5 Hz, H-5), 3.09 (1H, ddd, J¼8.8, 6.0, 2.8 Hz,
H-11a), 2.83 (1H, m, H-9a), 2.70 (2H, overlapped, H-3, H-14), 2.39
(1H, m, H-9b), 2.16 (1H, m, H-11b), 2.09 (1H, ddd, J¼13.0, 8.8, 4.4 Hz,
H-4a), 2.00 (1H, m, H-4b), 1.96 (3H, dd, J¼1.4, 1.0 Hz, H3-50), 1.75–1.47
(10H, m, H2-6, H2-7, H2-8, H2-12, H2-13), 1.28 (3H, d, J¼7.2 Hz, H3-18);
23
C-40); More polar diastereomer (threo): [
a
]
þ64.9 (c 0.22, CHCl3);
D
IR (ATR) nmax 3422, 2975,1671,1497,1412 cmꢀ1; 1H NMR (DMSO-d6,
VT 100, 400 MHz) 7.35–7.28 (5H, m, Ph of Cbz), 5.80 (1H, ddd,
d
J¼16.4, 10.8, 5.2 Hz, H-2), 5.17 (1H, dd, J¼17.2, 2.0 Hz, H-3a), 5.10
(2H, s, CH2 of Cbz), 5.07 (1H, dd, J¼10.4, 2.0 Hz, H-3b), 4.67 (1H, d,
J¼4.4 Hz, OH), 4.36 (1H, q, J¼5.0 Hz, H-1), 3.93 (1H, m, H-20), 3.46
(1H, m, H-50a), 3.24 (1H, m, H-50b), 1.79 (4H, m, H2-30, H2-40); 13C
13C NMR (CDCl3, 100 MHz)
d
180.1 (C, C-2), 166.4 (C, C-20), 136.5 (CH,
NMR (DMSO-d6, VT 100, 150 MHz)
d
153.6 (C]O of Cbz), 136.2 (C),
C-16), 133.5 (C, C-30),125.4 (CH2, C-40),116.8 (CH2, C-17), 78.4 (CH, C-5),
76.0 (CH, C-15), 65.7 (CH, C-14), 55.6 (CH2, C-9), 54.3 (CH2, C-11), 35.4
(CH2, C-4), 35.3 (CH2, C-6), 34.0 (CH, C-3), 28.7 (CH2, C-8), 26.0 (CH2,
C-12), 23.9 (CH2,C-13),23.2(CH2,C-7),18.3(CH3,C-50),15.9 (CH3,C-18);
HRESIMS: calcd for C20H32NO4 [MþH]þ: 350.2326, found: 350.2312.
127.2 (CH), 126.5 (CH), 126.3 (CH) [Ph of Cbz], 137.3 (CH, C-2), 114.0
(CH2, C-3), 70.6 (CH, C-1), 64.9 (CH2 of Cbz), 60.5 (CH, C-20), 45.9
(CH2, C-50), 24.6 (CH2, C-30), 22.3 (CH2, C-40); HRESIMS: calcd for
C15H19NO3Na [MþNa]þ: 284.1250, found: 284.1257.
3.4.9. Amino alcohol (13). To a stirred solution of 12 (163.0 mg,
0.700 mmol) in CH3CN (4.7 mL) at ꢀ15 ꢁC was added TMSI (0.5 mL,
3.50 mmol, 5 equiv). The reaction mixture was stirred for 15 min at
ꢀ15 ꢁC and another 15 min at 0 ꢁC. The mixture was quenched with
MeOH and evaporated under reduced pressure. Purification by silica
gel flash column chromatography and Al2O3 column chromatogra-
3.4.12. Synthetic dubiusamine-B (2). A mixture of 15 (14.0 mg,
0.040 mmol) and p-TsOH (7.6 mg, 0.040 mmol, 1 equiv) was
dissolved with CH2Cl2 (4.0 mL). The resulting mixture was refluxed
for 30 min and Grubbs second-generation catalyst (5 mg, 6.0 mmol,
0.15 equiv) in 0.5 mL of CH2Cl2 was then added. The mixture was
stirred under reflux for 19 h. The mixture was cooled to room
temperature and quenched with 1 M NaOH, and the aqueous layer
was extracted three times with 5% MeOH/CHCl3. The combined
phy (10% MeOH/CHCl3) afforded title compound 13 as a colorless oil
23
in 53% yield (47.0 mg); [
a
]
ꢀ5.5 (c 0.29, CHCl3); IR (ATR) nmax 3316,
D

M.A. Tan et al. / Tetrahedron 66 (2010) 3353–3359
3359
d
179.5 (C, C-2, C-20), 78.5 (CH, C-5, C-50), 49.8 (CH2, C-9, C-90), 37.3 (CH2, C-4,
organic layers were washed with 1 M NaOH and then brine, dried
over MgSO4, and evaporated under reduced pressure. The crude
residue was purified with amino-silica gel (CHCl3), followed by
C-40), 35.9 (CH2, C-3, C-30), 35.4 (CH, C-6, C-60), 29.9 (CH2, C-8, C-80), 23.2 (CH2,
C-7, C-70), 15.1 (CH3, C-11, C-110); HR-FABMS: calcd for C18H32NO4 [MþH]þ: 326.
2331, found 326.2341.
MPLC (2% EtOH/CHCl3) to give synthetic 2 in 61% yield (7.8 mg);
17
[
a]
þ22 (c 0.30, CHCl3); HRESIMS calcd for C18H28NO4 [MþH]þ:
D
322.2013, found: 322.2013; 1H and 13C NMR data of synthetic 2
were identical with those of natural 2.
O
O
2
11
CH3
O
O
5
7
3
9
H3C
N
H
4
8
6
Acknowledgements
Figure 4. Structure of (ꢃ)-17, the minor syn diastereomer of 1.
This work was supported by a Grant-in-Aid for Scientific
Research from the Japan Society for the Promotion of Science.
6. McDougal, P. G.; Rico, J. G.; Oh, Y. I.; Condon, B. D. J. Org. Chem. 1986, 51,
3388–3390.
References and notes
7. Kaiser, F.; Schwink, L.; Velder, J.; Schmalz, H. G. J. Org. Chem. 2002, 67,
9248–9256.
8. (a) Hayashi, Y.; Shoji, M.; Ishikawa, H.; Yamaguchi, J.; Tamura, T.; Imai, H.;
Nishigaya, Y.; Takabe, K.; Kakeya, H.; Osada, H. Angew. Chem., Int. Ed. 2008, 47,
6657–6660; (b) Mangion, I. K.; MacMillan, D. W. C. J. Am. Chem. Soc. 2005, 127,
3696–3697.
9. Zhong, G.; Yu, Y. Org. Lett. 2004, 6, 1637–1639.
10. For reference, the chiral HPLC conditions (Daicel Chiralcel AD-H; hexane/EtOH
1. Quisumbing, E. Medicinal Plants of the Philippines; Bureau of Printing: Manila,
1978.
2. (a) Nonato, M. G.; Takayama, H.; Garson, M. J., Chapter 4 In The Alkaloids;
Cordell, G. A., Ed.; Academic: New York, NY, 2008; Vol. 66, pp 215–249; (b)
Takayama, H.; Ichikawa, T.; Kuwajima, T.; Kitajima, M.; Seki, H.; Aimi, N.;
Nonato, M. G. J. Am. Chem. Soc. 2000, 122, 8635–8639; (c) Takayama, H.; Ichi-
kawa, T.; Kitajima, M.; Nonato, M. G.; Aimi, N. Chem. Pharm. Bull. 2002, 50,
1303–1304; (d) Takayama, H.; Ichikawa, T.; Kitajima, M.; Nonato, M. G.; Aimi, N.
J. Nat. Prod. 2001, 64, 1224–1225; (e) Takayama, H.; Ichikawa, T.; Kitajima, M.;
Aimi, N.; Lopez, D.; Nonato, M. G. Tetrahedron Lett. 2001, 42, 2995–2996; (f)
Takayama, H.; Sudo, R.; Kitajima, M. Tetrahedron Lett. 2005, 46, 5795–5797.
3. Nonato, M. G.; Garson, M. J.; Truscott, R. J. W.; Carver, J. A. Phytochemistry 1993,
34, 1159–1163.
4. Sjaifullah, A.; Garson, M. J. ACGC Chemical Research Commun. 1996, 5, 24–27.
5. In order to confirm and characterize spectroscopically the minor syn di-
astereomer of compound 1, we also synthesized it in its racemic form. The NOE
experiment for synthesized (ꢃ)-17 (Fig. 4) clearly supported the proposed
configuration, i.e., a clear NOE enhancement was observed between the two
95/5; flow rate¼0.5 mL/min; ¼254 nm; tmajor¼9.9 min., tminor¼12.6 min) were
l
established using a racemic sample that was prepared in the same way using
DL-proline as catalyst.
11. (a) Zhong, G. Angew. Chem., Int. Ed. 2003, 42, 4247–4250; (b) Zhong, G. Chem.
Commun. 2004, 606–607; (c) Hayashi, Y.; Yamaguchi, J.; Hibino, K.; Shoji, M.
Tetrahedron Lett. 2003, 44, 8293–8296; (d) Brown, S. P.; Brochu, M. P.; Sinz, C. J.;
Macmillan, D. W. C. J. Am. Chem. Soc. 2003, 125, 10808–10809.
12. In the 1H NMR spectrum, two sets of peaks having a 7:1 integration for the H-5
proton were observed (
assigned to be anti because a clear NOE peak enhancement was observed
between the H-5 methine ( 4.51) and the H-11 doublet methyl ( 1.28).
d 4.51, anti and 4.34, syn). The major diastereomer was
d
d
13. Kurosawa, W.; Kan, T.; Fukuyama, T. J. Am. Chem. Soc. 2003, 125, 8112–8113.
14. St-Denis, Y.; Chan, T. H. J. Org. Chem. 1992, 57, 3078–3085.
15. Bonini, B. F.; Comes-Franchini, M.; Fochi, M.; Laboroi, F.; Mazzanti, G.; Ricci, A.;
Varchi, G. J. Org. Chem. 1999, 64, 8008–8013.
16. (a) Jakubec, P.; Cockfield, D. M.; Dixon, D. J. J. Am. Chem. Soc. 2009, 131, 16632–
16633; (b) Honda, T.; Ushiwata, M.; Mizutani, H. Tetrahedron Lett. 2006, 47,
6251–6254; (c) Cheng, X.; Waters, S. P. Org. Lett. 2010, 12, 205–207.
methine protons at
d
4.34 (H-5, H-50) and 2.67 (H-3, H-30). (ꢃ) ꢀ17: amorphous
solid; IR (ATR) nmax 2935, 1761 cmꢀ1 1H NMR (CDCl3, 500 MHz)
; d 4.34 (2H,
dddd, J¼10.0, 7.5, 5.0, 5.0 Hz, H-5, H-50), 2.67 (2H, m, H-3, H-30), 2.61 (4H, dd,
J¼7.0, 7.0 Hz, H2-9, H2-90), 2.49 (2H, ddd, J¼12.4, 8.4, 5.6 Hz, H-4b, H-40b), 1.74
(2H, m, H-6, H-60), 1.65 (2H, m, H-6, H-60), 1.47 (10H, m, H-4, H-40, H2-7, H2-70,
H2-8, H2-80), 1.27 (6H, d, J¼7.0 Hz, H3-11, H3-110); 13C NMR (CDCl3, 125 MHz)