Synthesis of stemofoline analogues as acetylcholinesterase inhibitors

Article abstract of DOI:10.1016/j.tet.2012.06.047

Thirty-two new stemofoline analogues were prepared from didehydrostemofoline for studies as AChE inhibitors. C-3 Side-chain modified amino, carbamate, triazole and oxazole stemofoline derivatives were prepared. In general the amine derivatives were found to be stronger inhibitors of AChE than their alcohol analogues that we previously reported. Compounds 5 and 26, with small C-3 side-chain substituents, were two of the most active inhibitors. Preliminary molecular docking studies suggested that these compounds may inhibit AChE by binding horizontally along the passage of the active-site gorge and block access to acetylcholine.

Full text of DOI:10.1016/j.tet.2012.06.047

Tetrahedron 68 (2012) 7103e7115
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Tetrahedron
journal homepage: www.elsevier.com/locate/tet
Synthesis of stemofoline analogues as acetylcholinesterase inhibitors
Kwankamol Sastraruji a, Thanapat Sastraruji a, Alison T. Ung b, Renate Griffith c, Araya Jatisatienr d,
,
Stephen G. Pyne a
*
a School of Chemistry, University of Wollongong, Wollongong, New South Wales 2522, Australia
b School of Chemistry and Forensic Science, University of Technology Sydney, Sydney 2007, Australia
c School of Medical Sciences, University of New South Wales, Sydney 2052, Australia
d Department of Biology, Chiang Mai University, Chiang Mai 50202, Thailand
a r t i c l e i n f o
a b s t r a c t
Article history:
Thirty-two new stemofoline analogues were prepared from didehydrostemofoline for studies as AChE
inhibitors. C-3 Side-chain modified amino, carbamate, triazole and oxazole stemofoline derivatives were
prepared. In general the amine derivatives were found to be stronger inhibitors of AChE than their
alcohol analogues that we previously reported. Compounds 5 and 26, with small C-3 side-chain sub-
stituents, were two of the most active inhibitors. Preliminary molecular docking studies suggested that
these compounds may inhibit AChE by binding horizontally along the passage of the active-site gorge
and block access to acetylcholine.
Received 29 March 2012
Received in revised form 25 May 2012
Accepted 12 June 2012
Available online 21 June 2012
Ó 2012 Elsevier Ltd. All rights reserved.
1. Introduction
the insecticidal activity of these alkaloids and the crude extracts of
Stemona plants. Our earlier studies focused on C-3 hydroxyalkyl
One of the primary roles of acetylcholinesterase (AChE) is the
hydrolysis of the neurotransmitter acetylcholine (ACh) to inactive
choline and acetate in cholinergic synapses. Thus AChE is essential
for the rapid modulation of synaptic acivity.1 Hydrolysis of ACh
occurs through a protease-like action of an active-site serine resi-
due. This site resides at the end of a relatively long and narrow
gorge. Inhibitors of AChE can bind irreversibly (e.g., some pesti-
cides) or reversibly to the active site or at other sites in the gorge
(e.g., the peripheral anionic site (PAS)) and block the access of ACh
to the active site.1 Reversible AChE inhibitors, for example, the al-
kaloid galanthamine (reminyl), have been used in the treatment of
patients with Alzheimer’s disease (AD) to alleviate the symptoms of
reduced ACh concentration in the brain.2 More recent AD drug
development strategies involve targeting microtubule-associated
derivatives, including the synthesis of rare Stemona alkaloids,
where it was found that the AChE inhibitory activity was dependent
upon the length of the C-3 alkyl chain, the position of the hydroxyl
group and in some cases the configuration of the carbinol carbon.10
This study also revealed that the butyrolactone ring of stemofoline
was essential for AChE inhibitory activity.10 We report here the
synthesis of several novel C-3 side-chain amino, carbamate, triazole
and oxazole stemofoline derivatives and their activities as AChE
inhibitors.
2. Results and discussion
2.1. Synthesis of compounds
s
-protein, metal ion dyshomeostasis and the various
(A
) pathological mechanisms of this disease.2,3 AChE colocalizes
with A , which then promotes and accelerates A
aggregation.4e6
b-amyloid
In order to prepare the amine derivatives 3e19, the known al-
dehyde 2 was prepared from didehydrostemofoline 1 (Fig. 1), fol-
lowing our previously described procedures9 and used as a key
b
b
b
This has renewed an intense interest in AChE inhibitors, including
dual binding AChE inhibitors7 and those that can be activated by
AChE8 and have A
b-antiaggregating action. We have recently
reported that the Stemona alkaloids stemofoline and didehy-
drostemofoline and some of their C-3 side-chain derivatives are
inhibitors of AChE.9e11 Such activity is most likely associated with
* Corresponding author. Tel.: þ61 2 42213511; fax: þ61 2 42214287; e-mail ad-
dress: spyne@uow.edu.au (S.G. Pyne).
Fig. 1. Didehydrostemofoline 1 and its aldehyde derivative 2.
0040-4020/$ e see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.tet.2012.06.047

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K. Sastraruji et al. / Tetrahedron 68 (2012) 7103e7115
scaffold for reductive amination reactions to prepare the 17 amine
derivatives as shown in Scheme 1.
Another keyscaffold, thealkyne 26, was prepared from a one-step
alkynylation reaction14 of the aldehyde 2 using the BestmanneOhira
reagent15,16 (dimethyl-1-diazo-2-oxopropylphosphonate), which
was prepared as described by Ghosh et al.17 (Scheme 5).
Scheme 5. Synthesis of the alkyne 26. Reagents and conditions: (i) BestmanneOhira
reagent (MeCOC(N2)P(OMe)2), K2CO3, MeOH/MeCN (1:3), rt, 24 h (yield 76%).
The 1,3-dipolar cyclization (‘click’) reactions of the alkyne 26
(Scheme 6) were examined under three different reaction condi-
Scheme 1. Synthesis of amine derivatives 3e19. Reagents and conditions: (i) amine,
NaBH(OAc)3, CH2Cl2, 0.1% HOAc, rt, 24 h (yields 25e93%).
Using the reductive amination procedure described by Abdel-
Magid et al.,12 the amines 3e19 were obtained in yields ranging
from 25 to 93%. A further methylation reaction of the amine 6
(Scheme 2) gave the tertiary amine 20 in 94% yield. Carbamylation
reactions of the amines 15e17 (Scheme 3) gave the carbamates
21e23 in yields ranging from 56 to 80%. The HCl salt of the gua-
nidine derivative 25 was prepared in two steps via a key guanidi-
nation reaction13 of the amine 3 (Scheme 4).
Scheme 6. Click reactions of the alkyne 26. Reagents and conditions: (i) RBr, NaN3, Cu
powder, CuSO4, tBuOH/H2O (1:1), MW 125 ꢀC, 10 min (yields 18e23%); (ii) RN3,
Cu(OAc)2, sodium ascorbate, MeOH/H2O (1:1), rt, 4 h (yields 33e46%); (iii) RC(Cl)]
NOH, Cu(OAc)2, sodium ascorbate, EtOH/H2O (1:1), NaOH, rt, 4 h (yield 59%).
tions to provide the four triazoles 27e30 and the two isoxazoles
31e32 in yields ranging from 18 to 59%. The alkyne 26 was treated
under Sonogashira coupling conditions18 to yield the phenyl
substituted alkyne 33 in 62% yield and the self-condensation
product 34 in 4% yield (Scheme 7 (i)). The dimer 34 was more ef-
fectively obtained in 89% yield under Eglington coupling condi-
tions19 (Scheme
7 (ii)). The Sonogashira product 33 was
Scheme 2. Synthesis of amine derivative 20. Reagents and conditions: (i) formalde-
hyde, NaBH(OAc)3, dichloroethane, 0.1% HOAc, rt, 24 h (yield 94%).
Scheme 3. Synthesis of amine derivatives 21e23. Reagents and conditions: (i) ethyl
chloroformate, THF/NaHCO3 (2:1), 0 ꢀC, 3 h (yields 56e80%).
Scheme 4. Synthesis of guanidine derivative 25. Reagents and conditions: (i) guani-
dine triflate, CH2Cl2, rt, 24 h; (ii) TFA/CH2Cl2 (1:1), rt, 3 h (yield 54%); (iii) precipitated
from MeOH by HCl in ether (yield 52%).
Scheme 7. Synthesis of compounds 33e35. Reagents and conditions: (i) PhI,
PdCl2(PPh3)2, CuI, Et3N, THF, N2, rt, 24 h (yields 33: 62%, 34: 4%); (ii) Cu(OAc)2, MeCN,
argon gas, 40 ꢀC, 4 h (yield 89%); (iii) Pd/C, H2, EtOAc, rt, 24 h (yield 74%).

K. Sastraruji et al. / Tetrahedron 68 (2012) 7103e7115
Table 1 (continued )
7105
hydrogenated over Pd/C under a H2 atmosphere for 24 h to give the
Minimum inhibitory requirementsb
reduced product 35 in 74% yield (Scheme 7 (iii)).
Entry Compound
Side chain
ng
nmol
2.2. Biological assays
15
20
10
0.024
We initially used the TLC bioautographic method of Hos-
tettmann20 to determine the minimum inhibitory requirements
(MIRs) of these compounds against electric eel AChE (eeAChE)
using galanthamine as a positive control. The results are reported in
Tables 1e3. In our assay, galanthamine had a MIR of 1 ng. This assay
revealed that the carbamate 22 and the tertiary amino compound 4
(Table 1, entries 1 and 2) were essentially equipotent to galanth-
amine and more potent than their parent compound
16
17
6
8
10
10
0.025
0.025
18
19
15
10
10
50
0.025
0.100
Table 1
AChE inhibitory activities of didehydrostemofoline 1 and the amine derivatives
3e23 and 25 against eeAChEa
Entry Compound
Side chain
Minimum inhibitory requirementsb
20
12
50
0.115
ng
nmol
1
22
1
0.002
21
22
14
11
50
0.120
0.225
100
2
3
4
1
5
0.003
0.011
19
23
7
100
0.232
a
Galanthamine was used as a positive control having a MIR of 1 ng or 0.003 nmol.
Entries listed in order of decreasing activity.
4
5
5
5
5
0.012
0.012
b
25
didehydrostemofoline 1. The secondary amino compounds 19, 5
and 3 and the guanidine derivative 25 were equipotent to dide-
hydrostemofoline 1 (Table 1, entries 3e7). The carbamates 23 and
21 and the secondary amines, 13, 18, 16, 17 and 20 and the qua-
ternary amines 9 and 20, all had MIR values of 10 ng (Table 1, en-
tries 8e18). The dibasic-amino compounds 11 and 17 were the least
active amine derivatives (Table 1, entries 22 and 23). None of the
click products 27e32 (Table 2) were as potent as didehy-
drostemofoline 1 or the amino derivatives in Table 1, entries 1e7.
The N-benzyl-triazole derivative 27 was the most potent of this
series of compounds while the others with longer and more flexible
linkers (28 and 30) or no linker (29, 31 and 32) between the het-
erocyclic triazole or isoxazole moiety and the aryl substituent were
significantly less active.
The terminal alkyne 26 and the phenylethyl derivative 35 were
essentially equipotent to galanthamine and didehydrostemofoline
1, respectively (Table 3, entries 1 and 2). Substitution of the ter-
minal alkyne CH of 26 with a phenyl group, as in compound 33, had
an adverse effect on inhibitory activity. The dimer of 26 had very
poor activity (Table 3). Compared to our earlier results,10 this study
showed that in general the amine derivatives were more active
than our previously reported alcohol derivatives.10
6
7
1
3
5
5
0.013
0.013
8
23
10
0.019
9
21
13
18
10
10
10
0.021
0.022
0.022
10
11
12
13
14
9
10
10
10
0.023
0.023
0.023
We attempted to determine the IC50 values of many of the more
active compounds, and some of the alcohol analogues we reported
earlier,10 against eeAChE and human AChE (hAChE) using Ellman’s
assay.21 We found however, that the majority of these compounds
were not soluble in the assay medium (DMSO, pH 7.0 phosphate
buffer). The IC50 results determined on those that were soluble in
the assay medium are shown in Table 4. In our assay, galanthamine
16
17

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K. Sastraruji et al. / Tetrahedron 68 (2012) 7103e7115
Table 2
AChE inhibitory activities of the click products 27e32 against eeAChEa
Entry
Compound
Side chain
Minimum inhibitory requirementsb
ng
nmol
0.020
1
27
10
50
2
30
0.085
3
4
5
28
32
50
50
0.097
0.102
31
29
50
0.105
0.211
6
100
a
Galanthamine was used as a positive control having a MIR of 1 ng or 0.003 nmol.
Entries listed in order of decreasing activity.
b
Table 3
AChE inhibitory activities of miscellaneous derivatives against eeAChEa
Entry
Compound
Side chain
Minimum inhibitory requirementsb
ng nmol
1
2
26
35
1
5
0.003
0.011
3
33
34
50
0.116
0.141
4
100
a
Galanthamine was used as a positive control having a MIR of 1 ng or 0.003 nmol.
Entries listed in order of decreasing activity.
b
had IC50 values of 0.912
hAChE, respectively (Table 4, entry 1). Didehydrostemofoline 1 and
compound 5, which both had MIRs of 5 ng against eeAChE, showed
m
M and 0.597
m
M against eeAChE and
MIR values. However, their IC50 values against hAChE were rela-
tively similar (ca. 37e52 M).
m
To inhibit AChE, inhibitors can bind to one or more sites of the
enzyme. AChE has been reported to have at least two binding sites,
the active site and the peripheral anionic site (PAS).22 The active site
similar IC50 values (ca. 12e25 mM) against eeAChE and hAChE,
which indicated they were significantly less potent than galanth-
amine (Table 4, entries 2 and 3). Compounds 36,10 8, 9 and 37,10 all
with MIRs of 10 ng against eeAChE, were less active than com-
pounds 1 and 5 (Table 4, entries 4e7). Their IC50 values against
eeAChE however, did not correlate as well to their corresponding
is buried at the bottom of a 20 A deep narrow gorge23 and includes
ꢀ
four catalytic subsites, the esteractic site (which contains Ser200,
His440 and Glu327), the oxyanion hole (which contains Gly118,
Gly119 and Ala201), the acyl pocket (which contains Phe288 and

K. Sastraruji et al. / Tetrahedron 68 (2012) 7103e7115
7107
Table 4
Acetylcholinesterase inhibitory activity of stemofoline derivatives
Entry
Compound
Minimum inhibitory requirements
IC50 values
eeAChE
m
M (R2)
ng
1
nmol
0.003
hAChE
1
2
3
galanthamine
0.902ꢁ0.04 (0.9953)
19.20ꢁ0.26 (0.8749)
12.94ꢁ0.08 (0.9883)
0.597ꢁ0.07 (0.9877)
24.98ꢁ0.13 (0.9714)
19.93ꢁ0.17 (0.9455)
1
5
5
0.013
0.012
5
OH
OMe
H
N
(R)
4
10
0.025
302.3ꢁ0.29 (0.9245)
41.17ꢁ0.22 (0.9114)
O
O
O
O
5
6
8
9
10
10
0.025
0.025
52.45ꢁ0.14 (0.9668)
77.19ꢁ0.22 (0.9274)
37.49ꢁ0.16 (0.9540)
28.72ꢁ0.19 (0.9210)
OMe
H
N
H
O
7
10
0.028
108.1ꢁ0.15 (0.9659)
52.42ꢁ0.21 (0.9215)
O
O
O
O
Phe290) and the anionic subsite (which contains Trp84, Phe330
and Glu199).24 In contrast, the PAS, which consists of Tyr70, Asp72,
Tyr121, Tyr334 and Trp279, is located at the entrance of the active-
site gorge. The binding of ligands to the PAS may block the passage
of ACh or change the conformation of the active site allosterically
and inhibit its function.24 Preliminary molecular docking studies
using GOLD suite versions 4.1 and 5.0 (CCDC, Cambridge, UK)25
indicated that the more potent derivatives 5 and 26, having small
side chains, fitted ‘horizontally’ in the active site while those with
longer side chains (e.g., 33) fitted only ‘vertically’ into the active-
site gorge (Fig. 2). Further studies to understand the mode of ac-
tion of these compounds are in progress.
amine 4, carbamate 22 and the terminal alkyne 26) were found to
be as active as galanthamine using the TLC bioautographic method
while Ellman’s assay, against eeAChE and human AChE (hAChE),
identified didehydrostemofoline 1 and compound 5 as the most
active of the compounds tested with IC50 values in the range of
12e25 mM against eeAChE and hAChE. These activities were sig-
nificantly less than that of galanthamine.
3. Experimental
3.1. General experimental procedures
In conclusion, 32 new stemofoline analogues were prepared to
study as AChE inhibitors. In general the amine derivatives were
found to be stronger inhibitors of AChE than their alcohol ana-
logues that we previously reported.9,10 Three compounds (tertiary
All reactions, unless otherwise stated, were performed in oven
dried, single-necked round bottom flasks under an atmosphere of
dry nitrogen. Reagents and analytical grade solvents were pur-
chased from commercial sources. Progress of reactions was
Fig. 2. Overlay of the alkyne 26 (yellow carbons, ‘horizontal’ binding near the active site at the base of the active-site gorge) and the phenyl alkyne derivative 33 (green carbons,
‘vertical’ binding, along the walls of the gorge) docked into the active-site gorge of Torpedo californica AChE (TcAChE). The amino acid residues at the binding sites of TcAChE are
coloured pink for the active site and blue for the PAS. This view is expanded in the lower figure.

7108
K. Sastraruji et al. / Tetrahedron 68 (2012) 7103e7115
monitored by TLC using aluminium backed Merck F254 sorbent
silica gel with UV detection at 254 nm and/or Dragendorff’s re-
agent. Compounds were purified by column chromatography using
59.0 (OeCH3), 49.7 (C-7), 47.9 (C-5, C-9), 47.3 (NeCH3), 34.7 (C-10),
33.3 (C-1), 27.1 (C-6), 18.5 (C-17), 9.3 (C-16). ESIMS m/z 389.0 (100%)
[MþH]þ, 390.2 (20%). HRESIMS m/z 389.2059 [MþH]þ, calcd for
Merck flash silica gel (40e63
mm). Purity of compounds was de-
C21H29N2O5 389.2076.
termined by 1H NMR spectroscopy and HPLC (see Supplementary
data for details), and was always ꢂ95%. 1H and 13C NMR spectra
were recorded on a Varian Inova-500 spectrometer (500 MHz 1H,
125 MHz 13C) in deuterochloroform (CDCl3), unless otherwise
specified. NMR assignments were based on gCOSY, gHSQC, gHMBC
and DEPT or APT experiments. 1H and 13C NMR assignments are
based on the numbering system used for stemofoline and not on
the systematic name and numbering used in the naming of
many stemofoline derivatives and analogues in the experimental
section. Low-resolution mass spectra were obtained on a Waters
LCZ single quadrupole (ESI). High-resolution mass spectra were
obtained on a Waters QTOF (ESI). The microwave reactions were
performed on a CEM Discovery Microwave Synthesis System
(NC, USA).
3.1.3. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-(N-iso-
propylaminomethyl)-9-methyl-4H-2,2,6-(epoxy[1]propanyl[3]yli-
dene)furo[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-3-methyl-
2(5H)-furanone (5). Prepared using the general method described
above, using the aldehyde 2 (24.3 mg, 0.068 mmol), dichloroethane
(2.0 mL), acetic acid (0.2 mL), isopropylamine (11.6 mL, 0.135 mmol)
and NaBH(OAc)3 (43.0 mg, 0.203 mmol). The purified product was
obtained as a colourless gum (21.1 mg, 0.052 mmol, 78% yield).
Rf¼0.23 in MeOH/CH2Cl2 (1:9). ½a D25
ꢄ
þ246.9 (c 1.26, CHCl3). IR nmax
3383, 2961, 2885, 2356, 2337, 1742, 1621 cmꢅ1. 1H NMR
d
4.43 (br s,
1H, H-2), 4.13 (s, 3H, OeCH3), 3.45 (br s, 1H, H-9a), 3.12e3.06 (m,
2H, H-5, H-10), 3.02e2.97 (m, 1H, H-5), 2.77e2.74 (m, 1H, H-100),
2.74 (d, J 10.0 Hz, 1H, H-10), 2.69 (d, J 10.0 Hz, 1H, H-10), 2.67 (d, J
5.0 Hz, 1H, H-7), 2.06 (s, 3H, H-16), 1.94 (d, J 11.5 Hz, 1H, H-1a),
1.91e1.87 (m, 1H, H-9), 1.85e1.81 (m, 2H, H-6), 1.71 (d, J 12.0 Hz, 1H,
H-1b),1.50 (br s, 1H, NH), 1.37 (d, J 7.0 Hz, 3H, H-17),1.05 (d, J 6.5 Hz,
3.1.1. General reaction procedure and preparation of (5Z)-5-
[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-(N-methyl-
aminomethyl)-9-methyl-4H-2,2,6-(epoxy[1]propanyl[3]ylidene)furo
[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-3-methyl-2(5H)-fur-
anone (3). To a solution of aldehyde 2 (26.4 mg, 0.074 mmol) in dry
dichloroethane (2.0 mL) and acetic acid (0.2 mL) at rt was added
3H, H-40), 1.02 (d, J 6.0 Hz, 3H, H-200). 13C NMR
d 169.9 (C-15), 163.0
(C-13), 148.6 (C-12), 128.0 (C-11),112.6 (C-8), 98.7 (C-14), 82.9 (C-3),
79.1 (C-2), 61.3 (C-9a), 59.0 (OeCH3), 49.9 (C-7), 49.4 (C-100), 48.0
(C-10), 47.8 (C-5), 34.7 (C-10), 33.7 (C-1, C-9), 27.2 (C-6), 23.4 (C-200þ),
22.8 (C-40), 18.5 (C-17), 9.3 (C-16). ESIMS m/z 403.0 (100%) [MþH] ,
404.1 (20%). HRESIMS m/z 403.2234 [MþH]þ, calcd for C22H31N2O5
403.2233.
methylamine (8.03 M in MeOH, 16.3 mL, 0.147 mmol) and then
NaBH(OAc)3 (46.7 mg, 0.220 mmol). The reaction mixture was left
to stir for 24 h. The mixture was quenched with saturated aqueous
NaHCO3 solution (10 mL) and was directly extracted with CH2Cl2
(3ꢃ20 mL). The combined organic extracts were washed with brine
and dried (MgSO4) before being concentrated in vacuo. The con-
centrated residue was purified by column chromatography using
gradient elution from CH2Cl2 to CH2Cl2/MeOH (9:1) to give the
amine 3 (13.8 mg, 0.037 mmol, 60% yield) as a yellow gum. Rf¼0.10
3.1.4. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-(N-allyla-
minomethyl)-9-methyl-4H-2,2,6-(epoxy[1]propanyl[3]ylidene)furo
[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-3-methyl-2(5H)-fur-
anone (6). Prepared using the general method described above,
using the aldehyde
(2.0 mL), acetic acid (0.2 mL), allylamine (10.0 m
2
(24.0 mg, 0.067 mmol), dichloroethane
L, 0.134 mmol) and
in MeOH/CH2Cl2 (1:9). ½a D25
ꢄ
þ311.1 (c 0.92, CHCl3). IR nmax 3350,
4.42 (s, 1H, H-2),
2941, 2921, 2880, 2794, 1756, 1623 cmꢅ1. 1H NMR
d
NaBH(OAc)3 (42.5 mg, 0.200 mmol). The purified product was
obtained as a yellow gum (18.7 mg, 0.047 mmol, 70% yield). Rf¼0.11
4.12 (s, 3H, OeCH3), 3.44 (br s, 1H, H-9a), 3.12e3.07 (m, 2H, H-5a,
H-10), 3.02e2.98 (m, 1H, H-5b), 2.70 (s, 2H, H-10), 2.67 (d, J 6.5 Hz,
1H, H-7), 2.45 (s, 3H, H-30), 2.06 (s, 3H, H-16), 1.94 (d, J 12.0 Hz, 1H,
H-1b), 1.92e1.87 (m, 1H, H-6a), 1.85e1.80 (m, 2H, H-6b, H-9), 1.70
(d, J 12.0 Hz,1H, H-1a),1.58 (br s,1H, NH),1.36 (d, J 7.0 Hz, 3H, H-17).
in MeOH/EtOAc (2:8). ½a D25
þ437.6 (c 1.25, CHCl3). IR nmax 3402,
ꢄ
2952, 2925, 2880, 2847, 1742, 1619 cmꢅ1 1H NMR
. d 5.89e5.81
(m, 1H, H-200), 5.15 (dd, J 16.0 Hz, 1.5 Hz, 1H, H-300(E)), 5.08 (dd, J
10.0 Hz, 1.0 Hz, 1H, H-300(Z)), 4.44 (br s, 1H, H-2), 4.12 (s, 3H,
OeCH3), 3.44 (br s, 1H, H-9a), 3.29e3.22 (m, 2H, H-100), 3.11e3.06
(m, 2H, H-5b, H-10), 3.02e2.96 (m, 1H, H-5a), 2.71 (ABq, J 10.0 Hz,
2H, H-10), 2.66 (d, J 6.0 Hz, 1H, H-7), 2.05 (s, 3H, H-16), 1.94 (d, J
12.0 Hz, 1H, H-1a), 1.90e1.86 (m, 1H, H-6b), 1.84e1.79 (m, 2H, H-6a,
H-9), 1.71 (dt, J 12.0 Hz, 3.0 Hz, 1H, H-1b), 1.63 (br s, 1H, NH), 1.36 (d,
13C NMR
d 169.8 (C-15), 163.0 (C-13), 148.5 (C-12), 128.0 (C-11),
112.6 (C-8), 98.7 (C-14), 82.8 (C-3), 79.0 (C-2), 61.3 (C-9a), 59.0
(OeCH3), 52.6 (C-10), 49.8 (C-7), 48.0 (C-9), 47.8 (C-5), 37.2 (NeCH3),
34.7 (C-10), 33.7 (C-1), 27.2 (C-6), 18.4 (C-17), 9.3 (C-16). ESIMS m/z
375.0 (100%) [MþH]þ, 376.1 (20%). HRESIMS m/z 375.1938 [MþH]þ,
calcd for C20H27N2O5 375.1920.
J 6.5 Hz, 3H, H-17). 13C NMR
d 169.8 (C-15), 163.0 (C-13), 148.5
(C-12), 136.9 (C-200), 128.0 (C-11), 116.1 (C-300), 112.6 (C-8), 98.7
(C-14), 82.9 (C-3), 79.0 (C-2), 61.3 (C-9a), 59.0 (OeCH3), 52.8 (C-100),
49.8 (C-7), 49.5 (C-10), 48.0 (C-9), 47.8 (C-5), 34.6 (C-10), 33.7 (C-1),
27.2 (C-6), 18.4 (C-17), 9.3 (C-16). ESIMS m/z 401.0 (100%) [MþH]þ,
402.1 (20%). HRESIMS m/z 401.2073 [MþH]þ, calcd for C22H29N2O5
401.2076.
3.1.2. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-(N,N-di-
methylaminomethyl)-9-methyl-4H-2,2,6-(epoxy[1]propanyl[3]yli-
dene)furo[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-3-methyl-
2(5H)-furanone (4). Prepared using the general method described
above, using the aldehyde 2 (16.6 mg, 0.046 mmol), dichloroethane
(2.0 mL), acetic acid (0.2 mL), dimethylamine (2.0 M in THF, 47 mL,
0.092 mmol) and NaBH(OAc)3 (29.4 mg, 0.139 mmol). The purified
3.1.5. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-[N-(2-(di-
methylamino)ethyl)aminomethyl]-9-methyl-4H-2,2,6 (epoxy[1]prop-
anyl[3]ylidene)furo[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-3-
methyl-2(5H)-furanone (7). Prepared using the general method
described above, using the aldehyde 2 (23.9 mg, 0.067 mmol), di-
chloroethane (2.0 mL), acetic acid (0.2 mL), 2-(dimethylamino)
product was obtained as a yellow gum (7.0 mg, 0.018 mmol, 39%
yield). Rf¼0.26 in MeOH/CH2Cl2 (1:9). ½a D25
ꢄ
þ281.3 (c 0.47, CHCl3). IR
nmax 2936, 2768, 2356, 2337, 1743, 1620 cmꢅ1. 1H NMR
d
4.38 (s, 1H,
H-2), 4.13 (s, 3H, OeCH3), 3.45 (br s, 1H, H-9a), 3.17e3.11 (m, 1H,
H-5a), 3.11e3.06 (m,1H, H-10), 3.03e2.97 (m, 1H, H-5b), 2.78 (d, J
6.0 Hz, 1H, H-7), 2.48 (d, J 13.5 Hz, 1H, H-10a), 2.35 (d, J 13.5 Hz, 1H,
H-10b), 2.30 (s, 6H, NeCH3), 2.06 (s, 3H, H-16), 1.95 (d, J 11.5 Hz, 1H,
H-1a), 1.91e1.86 (m, 1H, H-6a), 1.84e1.81 (m, 1H, H-6b), 1.79e1.76
ethylamine (15.3 mL, 0.133 mmol) and NaBH(OAc)3 (42.3 mg,
0.200 mmol). The purified product was obtained as a colourless
gum (8.2 mg, 0.019 mmol, 28% yield). Rf¼0.17 in MeOH/CH2Cl2
(m, 1H, H-9), 1.76e1.74 (m, 1H, H-1b), 1.36 (d, J 7.0 Hz, 3H, H-17). 13
C
(4:6). ½a 2D5
þ178.5 (c 0.55, CHCl3). IR nmax 3384, 2962, 2932, 2360,
ꢄ
NMR
d
169.8 (C-15), 163.0 (C-13), 148.6 (C-12), 128.0 (C-11), 112.7
1740, 1618 cmꢅ1 1H NMR (300 MHz)
. d 4.43 (br s, 1H, H-2), 4.13
(C-8), 98.8 (C-14), 83.2 (C-3), 79.0 (C-2), 60.8 (C-9a), 59.9 (C-10),
(s, 3H, OeCH3), 3.46 (br s, 1H, H-9a), 3.16e3.06 (m, 2H, H-5, H-10),

K. Sastraruji et al. / Tetrahedron 68 (2012) 7103e7115
7109
3.04e2.95 (m, 1H, H-5), 2.75 (s, 2H, H-10), 2.74 (t, J 6.0 Hz, 2H, H-100),
dichloroethane
(2.0 mL),
acetic
acid
(0.2 mL),
ethyl
2.69 (d, J 5.4 Hz, 1H, H-7), 2.44 (t, J 6.0 Hz, 2H, H-200), 2.25 (s, 6H,
NeCH3), 2.20 (br s, 1H, NH), 2.06 (s, 3H, H-16), 1.94 (d, J 12.0 Hz, 1H,
H-1a), 1.90e1.84 (m, 2H, H-6), 1.79 (dd, J 10.2 Hz, 3.6 Hz, 1H, H-9),
1-piperazinecarboxylate (22.0 mL, 0.148 mmol) and NaBH(OAc)3
(47.1 mg, 0.222 mmol). The purified product was obtained as
a colourless gum (28.9 mg, 0.058 mmol, 78% yield). Rf¼0.33 in
1.73 (dt, J 12.3 Hz, 3.3 Hz, 1H, H-1b), 1.36 (d, J 6.6 Hz, 3H, H-17). 13
C
MeOH/CH2Cl2 (1:9). ½a D25
þ236.7 (c 1.93, CHCl3). IR nmax 2964, 2932,
ꢄ
NMR (75 MHz)
d
169.9 (C-15), 163.0 (C-13), 148.5 (C-12), 128.0
2356, 2337, 1743, 1694, 1620 cmꢅ1 1H NMR
. d 4.34 (br s, 1H, H-2),
(C-11), 112.6 (C-8), 98.7 (C-14), 82.9 (C-3), 78.9 (C-2), 61.2 (C-9a),
59.0 (OeCH3), 58.9 (C-200), 50.3 (C-10), 49.8 (C-7), 48.3 (C-9), 47.9
(C-5, C-100), 45.6 (NeCH3), 34.6 (C-10), 33.6 (C-1), 27.1 (C-6), 18.4
(C-17), 9.3 (C-16). ESIMS m/z 432.0 (100%) [MþH]þ, 433.2 (20%).
HRESIMS m/z 432.2487 [MþH]þ, calcd for C23H34N3O5 432.2498.
4.12 (s, 3H, OeCH3), 4.11e4.08 (m, 2H, NCO2CH2CH3), 3.42 (br s, 5H,
H-9a, H-300), 3.14e3.04 (m, 2H, H-5a, H-10), 3.00e2.95 (m, 1H,
H-5b), 2.76 (br s, 2H, H-200), 2.72 (d, J 6.0 Hz, 1H, H-7), 2.46 (s, 2H,
H-10), 2.34e2.32 (m, 2H, H-200), 2.05 (s, 3H, H-16), 1.93 (d, J 12.0 Hz,
1H, H-1a), 1.89e1.84 (m, 1H, H-6a), 1.82e1.78 (m, 1H, H-6b), 1.75
(dd, J 10.0 Hz, 2.5 Hz, 1H, H-9), 1.69 (d, J 12.0 Hz, 1H, H-1b), 1.35 (d, J
7.0 Hz, 3H, H-17), 1.23 (t, J 7.5 Hz, 3H, NCO2CH2CH3). 13C NMR
3.1.6. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-[N-(2-
hydroxyethyl)aminomethyl]-9-methyl-4H-2,2,6-(epoxy[1]propanyl
[3]ylidene)furo[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-3-methyl-
2(5H)-furanone (8). Prepared using the general method described
above, using the aldehyde 2 (18.6 mg, 0.052 mmol), dichloroethane
d
169.8 (C-15), 162.9 (C-13), 155.6 (NCO2CH2CH3), 148.4 (C-12),
128.0 (C-11), 112.7 (C-8), 98.6 (C-14), 83.2 (C-3), 79.0 (C-2), 61.4
(NCO2CH2CH3), 60.8 (C-9a), 59.0 (OeCH3), 58.5 (C-10), 54.4 (C-200),
49.7 (C-7), 47.8 (C-5, C-9), 44.0 (C-300), 34.6 (C-10), 33.3 (C-1), 27.2
(C-6), 18.4 (C-17), 14.8 (NCO2CH2CH3), 9.3 (C-16). ESIMS m/z 502.0
(100%) [MþH]þ, 503.1 (20%). HRESIMS m/z 502.2546 [MþH]þ, calcd
for C26H36N3O7 502.2553.
(2.0 mL), acetic acid (0.2 mL), ethanolamine (8.5 mL, 0.104 mmol)
and NaBH(OAc)3 (32.9 mg, 0.155 mmol). The purified product was
obtained as a colourless gum (10.0 mg, 0.025 mmol, 48% yield).
Rf¼0.20 in MeOH/CH2Cl2 (1:9). ½a D25
þ267.7 (c 0.67, CHCl3). IR nmax
ꢄ
3392, 2961, 2936, 1740, 1618, 1143 cmꢅ1. ESIMS m/z 405.0 (100%)
[MþH]þ, 406.1 (15%). HRESIMS m/z 405.2015 [MþH]þ, calcd for
3.1.9. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-[(4-methyl
piperazine)methyl]-9-methyl-4H-2,2,6-(epoxy[1]propanyl[3]ylidene)
furo[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-3-methyl-2(5H)-fur-
anone (11). Prepared using the general method described above,
C
21H29N2O6 405.2026. 1H NMR
d 4.42 (br s, 1H, H-2), 4.13 (s, 3H,
OeCH3), 3.66e3.58 (m, 2H, H-200), 3.48 (br s, 1H, H-9a), 3.16e3.12
(m, 1H, H-5a), 3.11e3.06 (m, 1H, H-10), 3.04e2.99 (m, 1H, H-5b),
2.87e2.83 (m, 1H, H-10), 2.84 (d, J 12.0 Hz, 1H, H-100), 2.81e2.76 (m,
1H, H-10), 2.76 (d, J 12.0 Hz,1H, H-100), 2.67 (d, J 6.0 Hz,1H, H-7), 2.11
(br s, 1H, NH), 2.06 (s, 3H, H-16), 1.96 (d, J 12.0 Hz, 1H, H-1a),
1.94e1.88 (m, 1H, H-6a), 1.88e1.85 (m, 1H, H-6b), 1.84e1.81 (m, 1H,
H-9), 1.74 (d, J 12.0 Hz, 1H, H-1b), 1.37 (d, J 6.0 Hz, 3H, H-17). 13C
using the aldehyde
(2.0 mL), acetic acid (0.2 mL), 1-methylpiperazine (16.2
2
(26.0 mg, 0.072 mmol), dichloroethane
L,
m
0.145 mmol) and NaBH(OAc)3 (46.0 mg, 0.217 mmol). The purified
product was obtained as a colourless gum (20.7 mg, 0.047 mmol,
64% yield). Rf¼0.09 in MeOH/CH2Cl2 (1:9). ½a D25
ꢄ
þ279.4 (c 1.38,
CHCl3). IR nmax 2961, 2939, 2356, 2325, 1742, 1618 cmꢅ1
.
1H NMR
NMR
d
169.9 (C-15), 163.0 (C-13), 148.4 (C-12), 128.1 (C-11), 112.5
d 4.42 (br s, 1H, H-2), 4.12 (s, 3H, OeCH3), 3.41 (br s, 1H, H-9a),
(C-8), 98.8 (C-14), 83.2 (C-3), 78.9 (C-2), 61.2 (C-9a, C-200), 59.0
(OeCH3), 51.8 (C-100), 49.7 (C-7), 49.6 (C-10), 48.0 (C-9), 47.8 (C-5),
34.6 (C-10), 33.6 (C-1), 27.1 (C-6), 18.4 (C-17), 9.3 (C-16). ESIMS m/z
405.0 (100%) [MþH]þ, 406.1 (15%). HRESIMS m/z 405.2015 [MþH]þ,
calcd for C21H29N2O6 405.2026.
3.16e3.10 (m,1H, H-5a), 3.08e3.04 (m,1H, H-10), 3.00e2.94 (m,1H,
H-5b), 2.80 (br s, 4H, H-200), 2.75 (d, J 6.0 Hz, 1H, H-7), 2.46 (s, 2H,
H-10), 2.42 (br s, 4H, H-300), 2.26 (s, 3H, NeCH3), 2.05 (s, 3H, H-16),
1.92 (d, J 12.0 Hz, 1H, H-1a), 1.89e1.85 (m, 1H, H-6a), 1.81e1.77
(m, 1H, H-6b), 1.75 (dd, J 9.0 Hz, 2.5 Hz, 1H, H-9), 1.70 (d, J 12.0 Hz,
1H, H-1b), 1.35 (d, J 6.5 Hz, 3H, H-17). 13C NMR
d 169.8 (C-15), 162.9
3.1.7. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-
morpholinomethyl-9-methyl-4H-2,2,6-(epoxy[1]propanyl[3]ylidene)
furo[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-3-methyl-2(5H)-fur-
anone (9). Prepared using the general method described above,
(C-13), 148.5 (C-12), 128.0 (C-11),112.8 (C-8), 98.6 (C-14), 83.2 (C-3),
79.0 (C-2), 60.8 (C-9a), 59.0 (OeCH3), 58.4 (C-10), 55.4 (C-300), 54.5
(C-200), 49.5 (C-7), 47.9 (C-5), 47.8 (C-9), 46.0 (NeCH3), 34.6 (C-10),
33.2 (C-1), 27.2 (C-6), 18.4 (C-17), 9.3 (C-16). ESIMS m/z 444.0
(100%) [MþH]þ, 445.2 (10%). HRESIMS m/z 444.2501 [MþH]þ, calcd
for C24H34N3O5 444.2498.
using the aldehyde
2
(24.8 mg, 0.069 mmol), dichloroethane
L, 0.138 mmol)
(2.0 mL), acetic acid (0.2 mL), morpholine (12.2
m
and NaBH(OAc)3 (43.9 mg, 0.207 mmol). The purified product was
obtained as a colourless gum (18.7 mg, 0.043 mmol, 63% yield).
3.1.10. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-
anilinomethyl-9-methyl-4H-2,2,6-(epoxy[1]propanyl[3]ylidene)furo
[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-3-methyl-2(5H)-fur-
anone (12). Prepared using the general method described above,
Rf¼0.28 in MeOH/CH2Cl2 (1:9). ½a D25
ꢄ
þ317.6 (c 0.99, CHCl3). IR nmax
2951, 2932, 2359, 2337, 1740, 1620 cmꢅ1. 1H NMR
d
4.35 (br s, 1H,
H-2), 4.13 (s, 3H, OeCH3), 3.69e3.63 (m, 4H, H-200), 3.42 (br s, 1H,
H-9a), 3.16e3.11 (m, 1H, H-5b), 3.09e3.05 (m, 1H, H-10), 3.01e2.96
(m, 1H, H-5a), 2.79 (br s, 2H, H-300), 2.75 (d, J 5.5 Hz, 1H, H-7),
2.49e2.43 (m, 2H, H-10), 2.40e2.37 (m, 2H, H-300), 2.06 (s, 3H, H-16),
1.94 (d, J 12.5 Hz, 1H, H-1a), 1.92e1.86 (m, 1H, H-6b), 1.83e1.80 (m,
1H, H-6a), 1.79e1.75 (m, 1H, H-9), 1.71 (br s, 1H, H-1b), 1.36 (d, J
using the aldehyde
(2.0 mL), acetic acid (0.2 mL), aniline (13.2
2
(15.9 mg, 0.044 mmol), dichloroethane
L, 0.089 mmol) and
m
NaBH(OAc)3 (28.2 mg, 0.133 mmol). The purified product was
obtained as a colourless gum (3.0 mg, 0.007 mmol, 25% yield).
Rf¼0.74 in MeOH/CH2Cl2 (1:9). ½a D25
ꢄ
þ272.1 (c 0.51, CHCl3). IR nmax
6.5 Hz, 3H, H-17). 13C NMR
d
169.8 (C-15), 162.9 (C-13), 148.4 (C-12),
3380, 2964, 2923, 2363, 2331, 1742, 1621 cmꢅ1. 1H NMR
d 7.18 (t, J
128.0 (C-11), 112.7 (C-8), 98.7 (C-14), 83.2 (C-3), 79.0 (C-2), 67.2
(C-200), 60.8 (C-9a), 59.0 (OeCH3), 58.9 (C-10), 55.2 (C-300), 49.6 (C-7),
47.8 (C-5, C-9), 34.6 (C-10), 33.3 (C-1), 27.2 (C-6), 18.4 (C-17), 9.3
(C-16). ESIMS m/z 431.2 (100%) [MþH]þ, 432.3 (5%). HRESIMS m/z
431.2163 [MþH]þ, calcd for C23H31N2O6 431.2182.
7.5 Hz, 2H, ArH), 6.73 (t, J 7.5 Hz,1H, ArH), 6.66 (d, J 7.5 Hz, 2H, ArH),
4.42 (s, 1H, H-2), 4.14 (s, 3H, OeCH3), 3.62 (br s, 1H, H-9a), 3.45 (d, J
10.5 Hz, 1H, H-10), 3.19e3.16 (m, 2H, H-5a, H-10), 3.14e3.10 (m, 2H,
H-5b, H-10), 2.84 (d, J 6.0 Hz, 1H, H-7), 2.07 (s, 3H, H-16), 2.02 (d, J
12.5 Hz, 1H, H-1a), 2.05e1.98 (m, 1H, H-6a), 1.94e1.90 (m, 1H,
H-6b), 1.89 (d, J 3.5 Hz, 1H, H-9), 1.86e1.84 (m, 1H, H-1b), 1.40 (d, J
3.1.8. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-[(4-
ethoxycarboxyl piperazine)methyl]-9-methyl-4H-2,2,6-(epoxy[1]pro
panyl[3]ylidene)furo[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-
3-methyl-2(5H)-furanone (10). Prepared using the general method
described above, using the aldehyde 2 (26.6 mg, 0.074 mmol),
7.0 Hz, 3H, H-17). 13C NMR
d 169.8 (C-15), 162.8 (C-13), 148.2 (C-12,
ArCeN),129.4 (ArCH),128.2 (C-11),118.2 (ArCH),113.4 (ArCH),112.4
(C-8), 98.9 (C-14), 83.1 (C-3), 79.0 (C-2), 61.7 (C-9a), 59.0 (OeCH3),
49.9 (C-7), 48.0 (C-9), 47.8 (C-5), 44.2 (C-10), 34.6 (C-10), 33.7 (C-1),
26.9 (C-6), 18.5 (C-17), 9.3 (C-16). ESIMS m/z 437.0 (100%) [MþH]þ,

7110
K. Sastraruji et al. / Tetrahedron 68 (2012) 7103e7115
438.1 (20%). HRESIMS m/z 437.2077 [MþH]þ, calcd for C25H29N2O5
1H, H-6b),1.86 (d, J 12.0 Hz,1H, H-1a), 1.80e1.74 (m, 2H, H-6b, H-9),
1.63 (d, J 12.0 Hz, 1H, H-1b), 1.30 (d, J 6.5 Hz, 3H, H-17), 0.36 (d, J
6.5 Hz, 2H, H-200), 0.29e0.26 (m, 1H, H-200), 0.23e0.21 (m, 1H, H-200).
437.2076.
3.1.11. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-(N-benzy-
laminomethyl)-9-methyl-4H-2,2,6-(epoxy[1]propanyl[3]ylidene)furo
[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-3-methyl-2(5H)-fur-
anone (13). Prepared using the general method described above,
13C NMR
d 169.9 (C-15), 163.0 (C-13), 148.5 (C-12), 128.0 (C-11),
112.5 (C-8), 98.6 (C-14), 82.9 (C-3), 78.9 (C-2), 61.2 (C-9a), 59.0
(OeCH3), 49.7 (C-10), 49.6 (C-7), 47.9 (C-9), 47.8 (C-5), 34.6 (C-10),
33.5 (C-1), 31.0 (C-100), 27.0 (C-6), 18.4 (C-17), 9.2 (C-16), 6.4 (C-200).
ESIMS m/z 400.7 (100%) [MþH]þ, 401.9 (25%). HRESIMS m/z
401.2083 [MþH]þ, calcd for C22H29N2O5 401.2076.
using the aldehyde
(2.0 mL), acetic acid (0.2 mL), N-benzylamine (16.0
2
(26.0 mg, 0.072 mmol), dichloroethane
L, 0.145 mmol)
m
and NaBH(OAc)3 (46.0 mg, 0.217 mmol). The purified product was
obtained as a yellow gum (27.9 mg, 0.062 mmol, 86% yield). Rf¼0.46
3.1.14. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-(N-cyclo-
pentylaminomethyl)-9-methyl-4H-2,2,6-(epoxy[1]propanyl[3]yli-
dene)furo[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-3-methyl-
2(5H)-furanone (16). Prepared using the general method described
above, using the aldehyde 2 (22.6 mg, 0.063 mmol), dichloroethane
in MeOH/CH2Cl2 (1:9). ½a D25
þ196.9 (c 1.86, CHCl3). IR nmax 3388,
ꢄ
2958, 2945, 2356, 2337, 1736, 1625 cmꢅ1 1H NMR (300 MHz)
.
d
7.34e7.29 (m, 4H, ArH), 7.25e7.23 (m, 1H, ArH), 4.43 (br s, 1H,
H-2), 4.13 (s, 3H, OeCH3), 3.82 (s, 2H, H-100), 3.45 (br s, 1H, H-9a),
3.11e3.08 (m, 1H, H-10), 3.06e2.95 (m, 2H, H-5), 2.74 (q, J 12.0 Hz,
1H, H-10), 2.69 (d, J 12.0 Hz,1H, H-10), 2.65 (d, J 6.0 Hz,1H, H-7), 2.06
(s, 3H, H-16), 1.95 (d, J 12.0 Hz, 1H, H-1a), 1.82e1.79 (m, 3H, H-6,
H-9), 1.74 (dt, J 12.0 Hz, 3.0 Hz,1H, H-1b), 1.37 (d, J 6.0 Hz, 3H, H-17).
13C NMR (75 MHz, one ArCH was not observed due to peak overlap)
(2.0 mL), acetic acid (0.2 mL), cyclopentylamine (12.9 mL,
0.126 mmol) and NaBH(OAc)3 (40.0 mg, 0.189 mmol). The purified
product was obtained as a yellow gum (24.4 mg, 0.057 mmol, 90%
yield). Rf¼0.21 in MeOH/CHCl3 (1:9). ½a D25
ꢄ
þ206.6 (c 0.35, CHCl3). IR
nmax 3351, 2951, 2885, 2359, 2337, 1743, 1620 cmꢅ1. 1H NMR
d
4.38
d
169.8 (C-15), 163.0 (C-13), 148.5 (C-12), 140.4 (ArC), 128.5 (ArCH),
(br s, 1H, H-2), 4.07 (s, 3H, OeCH3), 3.39 (br s, 1H, H-9a), 3.06e3.00
(m, 2H, H-5, H-10), 2.98e2.94 (m, 2H, H-5, H-100), 2.67 (s, 2H, H-10),
2.61 (d, J 6.0 Hz, 1H, H-7), 2.18 (br s, 1H, NH), 1.99 (s, 3H, H-16), 1.87
(d, J 12.0 Hz, 1H, H-1a), 1.84e1.81 (m, 1H, H-6a), 1.76e1.74 (m, 3H,
H-6b, H-200), 1.65 (d, J 12.0 Hz, 1H, H-1b), 1.60e1.56 (m, 2H, H-300),
1.48e1.44 (m, 2H, H-300), 1.30 (d, J 6.5 Hz, 3H, H-17), 1.28e1.21
128.0 (C-11), 127.0 (ArCH), 112.6 (C-8), 98.6 (C-14), 82.9 (C-3), 79.1
(C-2), 61.3 (C-9a), 59.0 (OeCH3), 54.1 (C-100), 49.7 (C-7), 49.2 (C-10),
48.0 (C-9), 47.8 (C-5), 34.6 (C-10), 33.7 (C-1), 27.2 (C-6), 18.4 (C-17),
9.2 (C-16). ESIMS m/z 451.0 (100%) [MþH]þ, 452.3 (20%). HRESIMS
m/z 451.2245 [MþH]þ, calcd for C26H31N2O5 451.2233.
(m, 2H, H-200). 13C NMR
d 169.8 (C-15), 162.9 (C-13), 148.5 (C-12),
3.1.12. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-(N-cyclo-
propylmethylaminomethyl)-9-methyl-4H-2,2,6-(epoxy[1]propanyl[3]
ylidene)furo[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-3-methyl-
2(5H)-furanone (14). Prepared using the general method described
above, using the aldehyde 2 (24.8 mg, 0.069 mmol), dichloroethane
127.9 (C-11), 112.5 (C-8), 98.5 (C-14), 82.8 (C-3), 79.0 (C-2), 61.2
(C-9a), 60.2 (C-100), 58.9 (OeCH3), 49.7 (C-7), 48.7 (C-10), 47.9 (C-9),
47.7 (C-5), 34.6 (C-10), 33.6 (C-1), 33.1 (C-200), 33.0 (C-200), 27.1 (C-6),
24.1 (C-300), 24.0 (C-300), 18.4 (C-17), 9.2 (C-16). ESIMS m/z 428.9
(100%) [MþH]þ. HRESIMS m/z 429.2414 [MþH]þ, calcd for
(2.0 mL), acetic acid (0.2 mL), (aminomethyl)cyclopropane (12.4
mL,
C24H33N2O5 429.2389.
0.138 mmol) and NaBH(OAc)3 (43.9 mg, 0.207 mmol). The purified
product was obtained as a colourless gum (17.3 mg, 0.042 mmol,
3.1.15. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-(N-cyclo-
hexylaminomethyl)-9-methyl-4H-2,2,6-(epoxy[1]propanyl[3]ylidene)
furo[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-3-methyl-2(5H)-fur-
anone (17). Prepared using the general method described above,
60% yield). Rf¼0.28 in MeOH/CH2Cl2 (1:9). ½a D25
ꢄ
þ276.5 (c 1.15,
CHCl3). IR nmax 3347, 2999, 2951, 2359, 2337, 1742, 1619 cmꢅ1
.
1H
NMR d 4.45 (br s, 1H, H-2), 4.12 (s, 3H, OeCH3), 3.46 (br s, 1H, H-9a),
3.12e3.06 (m, 2H, H-5b, H-10), 3.02e2.97 (m, 1H, H-5a), 2.77 (s, 2H,
H-10), 2.68 (d, J 6.0 Hz, 1H, H-7), 2.54 (dd, J 12.5 Hz, 6.5 Hz, 1H,
H-100a), 2.44 (dd, J 12.5 Hz, 6.5 Hz, 1H, H-100b), 2.05 (s, 3H, H-16),
1.99 (br s, 1H, NH), 1.94 (d, J 12.5 Hz, 1H, H-1a), 1.91e1.86 (m, 1H,
H-6a), 1.84e1.79 (m, 2H, H-6b, H-9), 1.73 (dt, J 12.5 Hz, 3.5 Hz, 1H,
H-1b), 1.36 (d, J 6.5 Hz, 3H, H-17), 0.96e0.89 (m, 1H, H-200),
using the aldehyde
(2.0 mL), acetic acid (0.2 mL), cyclohexylamine (15.0
0.129 mmol) and NaBH(OAc)3 (41.1 mg, 0.194 mmol). The purified
2
(23.2 mg, 0.065 mmol), dichloroethane
L,
m
product was obtained as a yellow gum (24.6 mg, 0.056 mmol, 86%
yield). Rf¼0.30 in MeOH/CHCl3 (1:9). ½a D25
ꢄ
þ250.9 (c 1.05, CHCl3). IR
nmax 3377, 2929, 2856, 2363, 2337, 1743, 1621 cmꢅ1. 1H NMR
d
4.43
0.46e0.45 (m, 2H, H-300), 0.10e0.08 (m, 2H, H-300). 13C NMR
d
169.8
(br s, 1H, H-2), 4.12 (s, 3H, OeCH3), 3.45 (br s, 1H, H-9a), 3.11e3.06
(m, 2H, H-5b, H-10), 3.02e2.96 (m, 1H, H-5a), 2.79 (d, J 12.0 Hz, 1H,
H-10), 2.72 (d, J 12.0 Hz, 1H, H-10), 2.67 (d, J 6.0 Hz, 1H, H-7),
2.42e2.38 (m, 1H, H-100), 2.06 (s, 3H, H-16), 1.93 (d, J 11.5 Hz, 1H,
H-1a), 1.90e1.87 (m, 1H, H-6b), 1.84e1.80 (m, 4H, H-6a, H-9, H-200),
1.73e1.69 (m, 3H, H-1b, H-300), 1.58 (d, J 12.0 Hz, 1H, H-400), 1.36 (d, J
6.0 Hz, 3H, H-17), 1.28e1.21 (m, 2H, H-300), 1.18e1.11 (m, 1H, H-400),
(C-15), 163.0 (C-13), 148.5 (C-12), 128.0 (C-11), 112.6 (C-8), 98.7
(C-14), 83.9 (C-3), 79.0 (C-2), 61.3 (C-9a), 59.0 (OeCH3), 55.5 (C-100),
49.9 (C-10), 49.8 (C-7), 48.0 (C-9), 47.8 (C-5), 34.6 (C-10), 33.7 (C-1),
27.2 (C-6), 18.4 (C-17), 11.1 (C-200), 9.3 (C-16), 3.6 (C-300), 3.4 (C-300).
ESIMS m/z 415.1 (100%) [MþH]þ, 416.2 (10%). HRESIMS m/z
415.2252 [MþH]þ, calcd for C23H31N2O5 415.2233.
1.09e1.04 (m, 2H, H-200). 13C NMR
d 169.8 (C-15), 163.0 (C-13), 148.5
3.1.13. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-(N-cyclo-
propylaminomethyl)-9-methyl-4H-2,2,6-(epoxy[1]propanyl[3]yli-
dene)furo[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-3-methyl-
2(5H)-furanone (15). Prepared using the general method described
above, using the aldehyde 2 (21.6 mg, 0.060 mmol), dichloroethane
(C-12), 128.0 (C-11), 112.6 (C-8), 98.7 (C-14), 83.0 (C-3), 79.1 (C-2),
61.3 (C-9a), 59.0 (OeCH3), 57.5 (C-100), 49.8 (C-7), 48.1 (C-9), 47.8
(C-5), 47.3 (C-10), 34.6 (C-10), 33.7 (C-200), 33.6 (C-1), 33.3 (C-200),
27.2 (C-6), 26.3 (C-400), 25.1 (C-300), 25.0 (C-300), 18.4 (C-17), 9.3
(C-16). ESIMS m/z 443.0 (100%) [MþH]þ. HRESIMS m/z 443.2555
[MþH]þ, calcd for C25H35N2O5 443.2546.
(2.0 mL), acetic acid (0.2 mL), cyclopropylamine (8.2 mL,
0.120 mmol) and NaBH(OAc)3 (38.3 mg, 0.181 mmol). The purified
product was obtained as a yellow gum (22.4 mg, 0.056 mmol, 93%
3.1.16. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-[N-(2S-
phenylethyl)aminomethyl]-9-methyl-4H-2,2,6-(epoxy[1]propanyl[3]
ylidene)furo[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-3-methyl-
2(5H)-furanone (18). Prepared using the general method described
above, using the aldehyde 2 (29.7 mg, 0.083 mmol), dichloroethane
yield). Rf¼0.36 in MeOH/CHCl3 (1:9). ½a D25
ꢄ
þ281.4 (c 0.25, CHCl3). IR
nmax 3364, 2961, 2936, 2356, 2334, 1742, 1620 cmꢅ1. 1H NMR
d
4.42
(br s, 1H, H-2), 4.07 (s, 3H, OeCH3), 3.42 (br s, 1H, H-9a), 3.10e3.01
(m, 2H, H-5a, H-10), 2.99e2.93 (m, 1H, H-5b), 2.81 (d, J 12.0 Hz, 1H,
H-10), 2.75 (d, J 12.0 Hz, 1H, H-10), 2.63 (d, J 6.0 Hz, 1H, H-7), 2.03
(apparent sept, J 3.0 Hz, 1H, H-100), 2.00 (s, 3H, H-16), 1.89e1.83 (m,
(2.0 mL), acetic acid (0.2 mL), (S)-1-aminoethylbenzene (21.3
mL,
0.165 mmol) and NaBH(OAc)3 (105.2 mg, 0.496 mmol). The purified

K. Sastraruji et al. / Tetrahedron 68 (2012) 7103e7115
7111
product was obtained as a yellow gum (31.1 mg, 0.067 mmol, 81%
(C-7), 47.9 (C-5, C-9), 44.2 (NeCH3), 34.7 (C-10), 33.3 (C-1), 27.1
(C-6), 18.5 (C-17), 9.3 (C-16). ESIMS m/z 415.2 (100%) [MþH]þ,
416.3 (10%). HRESIMS m/z 415.2220 [MþH]þ, calcd for
yield). Rf¼0.38 in MeOH/CHCl3 (1:9). ½a D25
ꢄ
þ166.0 (c 2.07, CHCl3). IR
nmax 3015, 2958, 2359, 2334, 1742, 1621 cmꢅ1. 1H NMR
d
7.34e7.30
(m, 2H, ArH), 7.26 (s, 2H, ArH), 7.25e7.20 (m, 1H, ArH), 4.45 (br s,
1H, H-2), 4.13 (s, 3H, OeCH3), 3.71 (d, J 6.0 Hz, 1H, H-100a), 3.44 (br s,
1H, H-9a), 3.09 (br s, 1H, H-10), 2.93 (br s, 2H, H-5), 2.64 (d, J
12.0 Hz, 1H, H-10), 2.62 (d, J 6.0 Hz, 1H, H-7), 2.53 (d, J 12.0 Hz, 1H,
H-10), 2.06 (s, 3H, H-16), 1.98e1.93 (m, 1H, H-1a), 1.80e1.72 (m, 4H,
H-1b, H-6a,b, H-9), 1.36 (d, J 6.0 Hz, 3H, H-17), 1.33 (d, J 6.0 Hz, 3H,
C23H31N2O5 415.2233.
3.1.19. Carbamylation reaction: preparation of (5Z)-5-[(2S,2aR,6-
S,7aS,7bR,8R,9S)-hexahydro-7b-1-(N-ethoxycarboxyl-N-cyclo-
propylaminomethyl)-9-methyl-4H-2,2,6-(epoxy[1]propanyl[3]yli-
dene)furo[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-3-methyl-
2(5H)-furanone (21). To a solution of the amine 15 (11.2 mg,
0.028 mmol) in THF/saturated aqueous NaHCO3 solution (2:1)
100-CH3). 13C NMR
d 169.8 (C-15), 162.9 (C-13), 148.5 (C-12), 145.9
(ArC), 128.5 (ArCH), 127.9 (C-11), 126.9 (ArCH), 126.4 (ArCH), 112.5
(C-8), 98.6 (C-14), 82.8 (C-3), 79.1 (C-2), 61.2 (C-9a), 59.0 (OeCH3),
58.5 (C-100), 49.6 (C-7), 47.8 (C-5, C-9, C-10), 34.6 (C-10), 33.6 (C-1),
27.1 (C-6), 24.3 (100-CH3), 18.4 (C-17), 9.2 (C-16). ESIMS m/z 464.6
(100%) [MþH]þ, 465.2 (25%). HRESIMS m/z 465.2398 [MþH]þ, calcd
for C27H33N2O5 465.2389.
(3.0 mL) at 0 ꢀC was added ethyl chloroformate (4.5
mL,
0.056 mmol) and the reaction was left to stir for 3 h. The mixture
was quenched with saturated aqueous NaHCO3 solution (10 mL)
and was directly extracted with CH2Cl2 (3ꢃ20 mL). The combined
organic extracts were washed with brine and dried (MgSO4) before
being concentrated in vacuo. The concentrated residue was puri-
fied by column chromatography using gradient elution from CH2Cl2
to CH2Cl2/MeOH (9:1) to give 21 as a colourless gum (10.0 mg,
3.1.17. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-[N-(2R-
phenylethyl)aminomethyl]-9-methyl-4H-2,2,6-(epoxy[1]propanyl[3]
ylidene)furo[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-3-methyl-
2(5H)-furanone (19). Prepared using the general method described
above, using the aldehyde 2 (30.9 mg, 0.086 mmol), dichloroethane
0.021 mmol, 76% yield). Rf¼0.45 in MeOH/CHCl3 (1:9). ½a D25
þ164.0
ꢄ
(c 0.91, CHCl3). IR nmax 2961, 2932, 2359, 1744, 1690, 1621 cmꢅ1. 1H
NMR
d 4.63 (br s, 1H, H-2), 4.16 (q, J 7.5 Hz, 2H, NCO2CH2CH3), 4.13
(2.0 mL), acetic acid (0.2 mL), (R)-1-aminoethylbenzene (21.0
0.172 mmol) and NaBH(OAc)3 (109.5 mg, 0.517 mmol). The purified
m
L,
(s, 3H, OeCH3), 3.62 (d, J 13.0 Hz, 1H, H-10), 3.44 (br s, 1H, H-9a),
3.29 (d, J 13.0 Hz, 1H, H-10), 3.28e3.22 (m, 1H, H-5a), 3.10e2.99
(m, 2H, H-5b, H-10), 2.77 (d, J 6.0 Hz, 1H, H-7), 2.64 (br s, 1H, H-100),
2.10e2.02 (m, 1H, H-6a), 2.05 (s, 3H, H-16), 1.94 (d, J 12.0 Hz, 1H,
H-1a), 1.84e1.80 (m, 2H, H-6b, H-9), 1.72 (d, J 12.0 Hz, 1H, H-1b),
1.36 (d, J 6.5 Hz, 3H, H-17), 1.26 (t, J 7.5 Hz, 3H, NCO2CH2CH3),
0.88e0.82 (m, 1H, H-200), 0.78e0.74 (m, 1H, H-200), 0.74e0.67 (m,
1H, H-200), 0.65e0.56 (m, 1H, H-200). 13C NMR (the carbamate C]O
product was obtained as a yellow gum (24.7 mg, 0.053 mmol, 62%
yield). Rf¼0.41 in MeOH/CHCl3 (1:9). ½a D25
ꢄ
þ100.9 (c 1.65, CHCl3). IR
nmax 3030, 2961, 2936, 2356, 2340, 1742, 1620 cmꢅ1
.
1H NMR
d
7.34e7.28 (m, 2H, H-60), 7.30 (d, J 2.5 Hz, 2H, ArH), 7.24e7.20 (m,
1H, ArH), 4.38 (br s, 1H, H-2), 4.12 (s, 3H, OeCH3), 3.74 (q, J 6.5 Hz,
1H, H-100b), 3.46 (br s, 1H, H-9a), 3.10e3.04 (m, 2H, H-5, H-10),
3.00e2.95 (m, 1H, H-5), 2.62 (d, J 12.0 Hz, 1H, H-10), 2.55 (d, J 4.5 Hz,
1H, H-7), 2.46 (d, J 12.0 Hz, 1H, H-10), 2.04 (s, 3H, H-16), 1.95 (d, J
13.0 Hz, 1H, H-1a), 1.79e1.77 (m, 4H, H-1b, H-6, H-9), 1.36 (d, J
signal was not observed)
d 169.7 (C-15), 162.9 (C-13), 148.3 (C-12),
128.1 (C-11), 112.5 (C-8), 98.8 (C-14), 83.5 (C-3), 78.2 (C-2), 60.7
(C-9a), 61.7 (NCO2CH2CH3), 59.0 (OeCH3), 49.1 (C-7), 48.2 (C-5),
47.7 (C-9), 47.0 (C-10), 34.6 (C-10), 33.0 (C-1), 29.3 (C-100), 27.1 (C-6),
18.4 (C-17), 14.7 (NCO2CH2CH3), 10.0 (C-200), 9.3 (C-16), 8.8 (C-200).
ESIMS m/z 472.6 (100%) [MþH]þ, 473.7 (10%). HRESIMS m/z
473.2300 [MþH]þ, calcd for C25H33N2O7 473.2288.
6.5 Hz, 3H, H-17), 1.31 (d, J 6.5 Hz, 3H, 100-CH3). 13C NMR
d 170.0
(C-15), 163.1 (C-13), 148.7 (C-12), 145.9 (ArC), 128.8 (ArCH), 128.0
(C-11), 127.2 (ArCH), 126.8 (ArCH), 112.7 (C-8), 98.7 (C-14), 83.1
(C-3), 79.1 (C-2), 61.3 (C-9a), 59.1 (OeCH3), 59.0 (C-100), 49.8 (C-7),
48.1 (C-9, C-10), 47.9 (C-5), 34.8 (C-10), 33.8 (C-1), 27.2 (C-6), 25þ.2
(100-CH3), 18.5 (C-17), 9.4 (C-16). ESIMS m/z 464.8 (100%) [MþH] ,
465.9 (25%). HRESIMS m/z 465.2390 [MþH]þ, calcd for C27H33N2O5
465.2389.
3.1.20. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-(N-
ethoxycarboxyl-N-cyclopentylaminomethyl)-9-methyl-4H-2,2,6-(ep-
oxy[1]propanyl[3]ylidene)furo[2,3,4-gh]pyrrolizin-10-ylidene]-4-
methoxy-3-methyl-2(5H)-furanone (22). Prepared using the general
method described above, using the amine 16 (11.2 mg, 0.026 mmol)
2:1 THF/saturated aqueous NaHCO3 solution (3.0 mL) and ethyl
3.1.18. Methylation reaction: preparation of (5Z)-5-[(2S,2aR,6-
S,7aS,7bR,8R,9S)-hexahydro-7b-1-(N-methyl-N-allylaminomethyl)-
9-methyl-4H-2,2,6-(epoxy[1]propanyl[3]ylidene)furo[2,3,4-gh]pyrro-
lizin-10-ylidene]-4-methoxy-3-methyl-2(5H)-furanone
(20). Prepared using the general method described above, using
chloroformate (5.0 mL, 0.052 mmol). The purified product was
obtained as a yellow gum (7.3 mg, 0.015 mmol, 56% yield). Rf¼0.45
in MeOH/CHCl3 (1:9). ½a D25
ꢄ
þ205.6 (c 0.61, CHCl3). IR nmax 2958,
formaldehyde solution (ca. 400 g/L, 3.75
m
L, 0.245 mmol) as the
2888, 2362, 2337, 1737, 1675, 1616 cmꢅ1. 1H NMR
d 4.50 (br s, 1H,
aldehyde component, dichloroethane (2.0 mL), acetic acid
(0.2 mL), the amine 6 (9.8 mg, 0.024 mmol) as the amine com-
ponent and NaBH(OAc)3 (31.2 mg, 0.147 mmol). The reaction was
left for 16 h. The purified product was obtained as a yellow gum
(9.5 mg, 0.023 mmol, 94% yield). Rf¼0.37 in MeOH/CH2Cl2 (1:9).
H-2), 4.18e4.10 (m, 2H, NCO2CH2CH3), 4.13 (s, 3H, OeCH3),
3.70e3.63 (m, 2H, H-10, H-100), 3.46 (br s, 1H, H-9a), 3.24 (d, J
15.5 Hz, 1H, H-10), 3.22e3.16 (m, 1H, H-5), 3.09e3.00 (m, 2H, H-5,
H-10), 2.82 (d, J 5.5 Hz, 1H, H-7), 2.10e2.02 (m, 1H, H-6a), 2.06 (s,
3H, H-16), 1.93 (d, J 11.5 Hz,1H, H-1a), 1.84e1.72 (m, 7H, H-1b, H-6b,
H-9, H-200), 1.48 (br s, 4H, H-300), 1.36 (d, J 6.0 Hz, 3H, H-16), 1.26 (t, J
6.0 Hz, 3H, NCO2CH2CH3). 13C NMR (the carbamate C]O signal was
½
a 2D5
1621 cmꢅ1
ꢄ
þ263.6 (c 0.63, CHCl3). IR nmax 2955, 2920, 2847, 2359, 1743,
.
1H NMR (300 MHz) 5.89e5.75 (m, 1H, H-200), 5.15
d
(d, J 17.0 Hz, 1H, H-300(E)), 5.10 (d, J 9.0 Hz, 1H, H-300(Z)), 4.38 (br s,
1H, H-2), 4.13 (s, 3H, OeCH3), 3.44 (br s, 1H, H-9a), 3.23 (dd, J
13.8 Hz, 6.3 Hz, 1H, H-100b), 3.18e3.14 (m, 1H, H-5a), 3.13e3.06
(m, 1H, H-10), 3.03e2.98 (m, 1H, H-5b), 2.92 (dd, J 13.8 Hz, 6.6 Hz,
1H, H-100a), 2.76 (d, J 5.7 Hz, 1H, H-7), 2.53 (d, J 13.8 Hz, 1H, H-10),
2.42 (d, J 13.8 Hz, 1H, H-10), 2.30 (s, 3H, NeCH3), 2.06 (s, 3H,
H-16), 1.94 (d, J 12.0 Hz, 1H, H-1a), 1.91e1.81 (m, 2H, H-6),
1.79e1.72 (m, 2H, H-1b, H-9), 1.36 (d, J 6.6 Hz, 3H, H-17). 13C NMR
not observed) d 169.8 (C-15),162.9 (C-13),148.4 (C-12),128.0 (C-11),
112.6 (C-8), 98.8 (C-14), 83.7 (C-3), 77.6 (C-2), 62.6 (C-100), 61.4
(NCO2CH2CH3), 60.9 (C-9a), 59.0 (OeCH3), 49.4 (C-7, C-10), 48.2
(C-5), 47.8 (C-9), 34.7 (C-10), 33.0 (C-1), 27.1 (C-6), 24.9 (C-200), 24.8
(C-300), 18.4 (C-17), 14.8 (NCO2CH2CH3), 9.3 (C-16). ESIMS m/z 500.7
(100%) [MþH]þ, 501.9 (20%). HRESIMS m/z 501.2622 [MþH]þ, calcd
for C27H37N2O7 501.2601.
(75 MHz)
d
169.9 (C-15), 163.0 (C-13), 148.6 (C-12), 136.0 (C-200),
3.1.21. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-(N-ethox-
ycarboxyl-N-cyclohexylaminomethyl)-9-methyl-4H-2,2,6-(epoxy[1]
propanyl[3]ylidene)furo[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-
128.0 (C-11), 117.6 (C-300), 112.8 (C-8), 98.7 (C-14), 83.4 (C-3), 79.0
(C-2), 62.4 (C-100), 60.8 (C-9a), 59.0 (OeCH3), 56.9 (C-10), 49.7

7112
K. Sastraruji et al. / Tetrahedron 68 (2012) 7103e7115
3-methyl-2(5H)-furanone (23). Prepared using the general method
described above, using the amine 17 (11.1 mg, 0.025 mmol), 2:1
THF/saturated aqueous NaHCO3 solution (3.0 mL) and ethyl chlor-
not observed. d
172.0 (C-15), 164.8 (C-13), 160.0 (C-100), 149.7 (C-12),
129.4 (C-11), 112.5 (C-8), 99.6 (C-14), 76.5 (C-2), 58.9 (OeCH3), 51.0
(C-7), 47.3 (C-10), 38.8 (C-10), 34.3 (NeCH3), 32.2 (C-1), 24.4 (C-6),
16.8 (C-17), 7.9 (C-16). ESIMS m/z 416.7 (100%) [MþH]þ. HRESIMS m/
z 417.2155 [MþH]þ, calcd for C21H29N4O5 417.2138.
oformate (4.0 mL, 0.050 mmol). The purified product was obtained
as a yellow gum (10.2 mg, 0.020 mmol, 80% yield). Rf¼0.45 in
MeOH/CHCl3 (1:9). ½a D25
ꢄ
þ192.6 (c 0.78, CHCl3). IR nmax 2929, 2366,
1744, 1684, 1621 cmꢅ1
.
1H NMR
d
4.50 (br s, 1H, H-2), 4.18e4.10
3.1.24. Alkynylation reaction: preparation of (5Z)-5-[(2S,2aR,6-
S,7aS,7bR,8R,9S)-hexahydro-7b-1-ethynyl-9-methyl-4H-2,2,6-(epoxy
[1]propanyl[3]ylidene)furo[2,3,4-gh]pyrrolizin-10-ylidene]-4-
methoxy-3-methyl-2(5H)-furanone (26). To a mixture of the alde-
hyde 2 (151 mg, 0.420 mmol) and K2CO3 (69.7 mg, 0.504 mmol) in
MeOH/MeCN (1:3) (8.0 mL) at rt was added the BestmanneOhira
reagent (97 mg, 0.504 mmol) and the reaction mixture was left to
stir for 24 h. The mixture was quenched with saturated aqueous
NaHCO3 solution (10 mL) and was extracted with diethyl ether
(3ꢃ20 mL). The combined organic extracts were washed with brine
and dried over MgSO4 before being concentrated in vacuo. The
concentrated residue was purified by column chromatography us-
ing gradient elution from CH2Cl2 to CH2Cl2/MeOH (98:2) to give 26
as a white gum (113.2 mg, 0.319 mmol, 76% yield). A small sample
was crystallized from CH2Cl2. Mp 174e176 ꢀC (decomposed).
(m, 2H, NCO2CH2CH3), 4.13 (s, 3H, OeCH3), 3.64 (br s, 1H, H-10), 3.46
(br s, 1H, H-9a), 3.41 (s, 1H, H-100), 3.20 (d, J 15.0 Hz, 1H, H-5),
3.12e3.01 (m, 2H, H-5, H-10), 2.82 (d, J 6.0 Hz, 1H, H-7), 2.07 (s, 3H,
H-16), 1.94 (d, J 12.5 Hz, 1H, H-1a), 1.87e1.72 (m, 11H, H-1b, H-6, H-
9, H-300, H-400), 1.61 (apparent d, J 9.0 Hz, 1H, H-200), 1.37 (d, J 6.5 Hz,
3H, H-17), 1.27 (t, J 7.0 Hz, 3H, NCO2CH2CH3), 1.22 (apparent d, J
13.0 Hz, 2H, H-300), 1.14e1.08 (m, 1H, H-200). 13C NMR (the carbamate
C]O signal was not observed) d 169.8 (C-15), 162.9 (C-13), 148.4 (C-
12), 128.0 (C-11), 112.7 (C-8), 98.8 (C-14), 83.7 (C-3), 77.6 (C-2), 70.8
(C-100), 61.4 (C-9a), 61.2 (NCO2CH2CH3), 59.0 (OeCH3), 48.5 (C-10),
48.2 (C-5, C-7), 47.8 (C-9), 34.7 (C-10), 33.0 (C-1), 27.2 (C-400), 26.6
(C-6, C-300), 25.6 (C-200), 18.4 (C-17), 14.8 (NCO2CH2CH3), 9.3 (C-16).
ESIMS m/z 514.5 (100%) [MþH]þ, 515.3 (40%). HRESIMS m/z
515.2726 [MþH]þ, calcd for C28H39N2O7 515.2757.
a 25
Rf¼0.30 in MeOH/EtOAc (1:9). ½ ꢄD þ292.8 (c 0.66, CHCl3). IR nmax
3.1.22. Guanidination reaction: preparation of (5Z)-5-[(2S,2aR,6-
S,7aS,7bR,8R,9S)-hexahydro-7b-1-[[[[[(1,1-dimethylethoxy)carbonyl]
amino][[(1,1-dimethylethoxy)carbonyl]imino]methylamino]methyl]-
9-methyl-4H-2,2,6-(epoxy[1]propanyl[3]ylidene)furo[2,3,4-gh]pyrro-
lizin-10-ylidene]-4-methoxy-3-methyl-2(5H)-furanone (24). To a so-
3249, 2958, 2917, 2366, 1744, 1627 cmꢅ1. 1H NMR (CDCl3)
d 4.57 (br
s, 1H, H-2), 4.13 (s, 3H, OeCH3), 3.51 (br s, 1H, H-9a), 3.40e3.34 (m,
1H, H-5a), 3.11e3.05 (m, 2H, H-5b, H-10), 3.04 (d, J 6.0 Hz, 1H, H-7),
2.54 (s, 1H, H-20), 2.17e2.10 (m, 1H, H-6a), 2.06 (s, 3H, H-16), 2.00
(d, J 12.5 Hz, 1H, H-1a), 1.91 (dt, J 12.5 Hz, 3.0 Hz, 1H, H-1b),
1.89e1.84 (m, 1H, H-6b), 1.77 (dd, J 10.0 Hz, 3.0 Hz, 1H, H-9), 1.36
lution of the amine
3 (8.8 mg, 0.024 mmol) in dry CH2Cl2
(2.0 mL) was added 1,3-di-Boc-2-(trifluoromethylsulfonyl)-guani-
dine (9.2 mg, 0.024 mmol) at rt and the reaction mixture was left to
stir for 24 h. The mixture was quenched with saturated aqueous
NaHCO3 solution (10 mL) and then extracted with CH2Cl2
(3ꢃ20 mL). The combined organic extracts were washed with brine
and dried (MgSO4) before being concentrated in vacuo. The con-
centrated residue was purified by column chromatography using
gradient elution from CH2Cl2 to CH2Cl2/MeOH (9:1) to give a white
gum (7.9 mg, 0.013 mmol, 54% yield). Rf¼0.45 in MeOH/CH2Cl2
(d, J 6.5 Hz, 3H, H-17). 13C NMR (CDCl3)
d 169.7 (C-15), 162.8 (C-13),
147.8 (C-12), 128.2 (C-11), 112.0 (C-8), 98.9 (C-14), 81.2 (C-3), 80.9
(C-2), 75.2 (C-10), 74.6 (C-20), 60.7 (C-9a), 59.0 (OeCH3), 54.4 (C-7),
48.6 (C-5), 47.3 (C-9), 34.6 (C-10), 33.5 (C-1), 27.0 (C-6), 18.3 (C-17),
9.3 (C-16). ESIMS m/z 356.1 (100%) [MþH]þ. HRESIMS m/z 356.1459
[MþH]þ, calcd for C20H22NO5 356.1498.
3.1.25. Click reaction method A: preparation of (5Z)-5-[(2S,2aR,6-
S,7aS,7bR,8R,9S)-hexahydro-7b-1-(1-benzyl-1H-1,2,3-triazol-4-yl)-
9-methyl-4H-2,2,6-(epoxy[1]propanyl[3]ylidene)furo[2,3,4-gh]pyrro-
lizin-10-ylidene]-4-methoxy-3-methyl-2(5H)-furanone (27). A mix-
(1:9). 1H NMR
d 4.44 (br s, 1H, H-2), 4.15 (s, 3H, OeCH3), 3.73 (d, J
13.5 Hz, 2H, H-10), 3.54 (br s, 1H, H-9a), 3.28e3.22 (m, 1H, H-5a),
3.12e3.04 (m, 2H, H-5b, H-10), 3.08 (s, 3H, NeCH3), 2.74 (d, J 5.5 Hz,
1H, H-7), 2.08 (s, 3H, H-16), 2.01 (d, J 12.5 Hz, 1H, H-1a), 1.99e1.89
(m, 2H, H-6), 1.83 (dd, J 10.0 Hz, 3.5 Hz, 1H, H-9), 1.75 (d, J 12.0 Hz,
1H, H-1b), 1.67 (br s, 1H, NH), 1.47 (s, 18H, CO2C(CH3)3), 1.39 (d, J
ture of the alkyne 26 (12.7 mg, 0.034 mmol), benzylbromide (4.8 mL,
0.040 mmol), sodium azide (2.6 mg, 0.040 mmol) and copper
powder (2.0 mg) in tBuOH/H2O (1:1) (1.0 mL) was treated with 1 M
aqueous CuSO4 (0.1 mL). The mixture was stirred and heated in
a microwave reactor at 125 ꢀC for 10 min (10 W). The mixture was
quenched with saturated aqueous NaHCO3 solution (10 mL) and
was directly extracted with diethyl ether (3ꢃ20 mL). The combined
organic extracts were washed with brine and dried (MgSO4) before
being concentrated in vacuo. The concentrated residue was purified
by column chromatography using gradient elution from CH2Cl2 to
CH2Cl2/MeOH (9:1) to give a yellow gum (3.8 mg, 0.008 mmol, 23%
13
6.5 Hz, 3H, H-17). C NMR (the signals for C-100 and the Boc car-
bonyls were not observed.) d 169.6 (C-15), 162.6 (C-13), 147.6 (C-12),
128.0 (C-11), 112.0 (C-8), 98.7 (C-14), 83.8 (C-3), 76.5 (C-2), 60.2 (C-
9a), 58.9 (OeCH3), 52.4 (C-10), 50.4 (C-7), 47.8 (C-5), 47.5 (C-9), 40.0
(NeCH3), 34.2 (C-10), 33.4 (C-1), 28.2 (CO2C(CH3)3), 26.4 (C-6), 18.2
(C-17), 9.1 (C-16). ESIMS m/z 616.6 (100%) [MþH]þ, 617.4 (80%).
HRESIMS m/z 617.3188 [MþH]þ, calcd for C31H45N4O9 617.3187.
yield). Rf¼0.11 in MeOH/EtOAc (5:95). ½a D25
þ207.6 (c 0.25, CHCl3).
ꢄ
3.1.23. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-[[(amino-
iminomethyl)methylamino]methyl]-9-methyl-4H-2,2,6-(epoxy[1]
propanyl[3]ylidene)furo[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-
3-methyl-2(5H)-furanoyl hydrochloride (25). To a solution of 24
(7.9 mg, 0.013 mmol) in dry CH2Cl2 (1.0 mL) was added TFA (1.0 mL)
at rt and the reaction mixture was left to stir for 3 h. The solvent was
removed under vacuum. To the residue was added hydrogen chlo-
ride in ether (2.0 mL, 1 M) and concentrated under vacuum. The
desired product was isolated as its hydrochloride salt by dissolution
in MeOH and precipitation by the addition of diethyl ether as awhite
1H NMR 7.34 (s,1H, C-50), 7.32e7.28 (m, 3H, ArH), 7.22e7.21 (m, 2H,
d
ArH), 5.44 (d, J 2.5 Hz, 2H, C-100), 4.41 (br s, 1H, H-2), 4.07 (s, 3H,
OeCH3), 3.51 (br s, 1H, H-9a), 3.30 (d, J 5.5 Hz, 1H, H-7), 3.09e3.04
(m, 1H, H-10), 3.03e2.99 (m, 2H, H-5), 2.00 (s, 3H, H-16), 1.96 (d, J
12.0 Hz,1H, H-1a),1.92e1.86 (m, 3H, H-1b, H-6a, H-9),1.84e1.81 (m,
1H, H-6b), 1.33 (d, J 6.5 Hz, 3H, H-17). 13C NMR
d 169.8 (C-15), 162.9
(C-13), 148.3 (C-12), 147.8 (ArC), 134.5 (C-40), 129.2 (ArCH), 128.9
(ArCH), 128.3 (ArCH), 128.1 (C-11), 120.9 (C-50), 112.7 (C-8), 98.7 (C-
14), 81.0 (C-2), 79.9 (C-3), 61.1 (C-9a), 59.0 (OeCH3), 54.4 (C-100), 52.1
(C-7), 48.6 (C-5), 47.7 (C-9), 34.6 (C-10), 33.3 (C-1), 27.2 (C-6), 18.4
(C-17), 9.2 (C-16). ESIMS m/z 489.1 (100%) [MþH]þ. HRESIMS m/z
489.2121 [MþH]þ, calcd for C27H29N4O5 489.2138.
solid salt (4.3 mg, 0.010 mmol, 52% yield). ½a D25
þ191.2 (c 0.23,
ꢄ
CH3OH). 1H NMR (CD3OD, a very broad spectrum was observed,
therefore only the methyl signals are reported)
3.25 (s, 3H, NeCH3), 2.07 (s, 3H, H-16),1.47 (d, J 6.0 Hz, 3H, H-17). 13
NMR (CD3OD) the signals for carbons at C-3, C-5, C-9 and C-9a were
d 4.23 (s, 3H, OeCH3),
C
3.1.26. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-[1-(3-
phenylpropyl)-1H-1,2,3-triazol-4-yl]-9-methyl-4H-2,2,6-(epoxy[1]

K. Sastraruji et al. / Tetrahedron 68 (2012) 7103e7115
7113
propanyl[3]ylidene)furo[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-
3-methyl-2(5H)-furanone (28). Prepared using method A, from the
1.93e1.89 (m, 1H, H-1b), 1.85e1.80 (m, 4H, H-1a, H-6, H-9), 1.33 (d,
J 6.5 Hz, 3H, H-17). 13C NMR 169.5 (C-15), 168.0 (C-100, C-300), 162.7
d
alkyne 26 (15.0 mg, 0.042 mmol), bromopropylbenzene (9
m
L,
(C-13), 148.1 (C-12), 147.0 (C-40), 134.0 (ArCH), 131.6 (ArC), 127.6 (C-
11), 123.1 (ArCH), 121.6 (C-50), 112.4 (C-8), 98.2 (C-14), 80.7 (C-2),
79.5 (C-3), 60.8 (C-9a), 58.7 (OeCH3), 51.8 (C-7), 48.2 (C-5), 47.6 (C-
1000), 47.3 (C-9), 34.8 (C-3000), 34.4 (C-10), 33.0 (C-1), 29.0 (C-2000),
26.9 (C-6), 18.0 (C-17), 8.9 (C-16). ESIMS m/z 586.4 (100%) [MþH]þ.
HRESIMS m/z 586.2318 [MþH]þ, calcd for C31H32N5O7 586.2302.
0.060 mmol), sodium azide (3.9 mg, 0.060 mmol), copper powder
(2.5 mg), tBuOH/H2O (1:1) (1.0 mL) and 1 M aqueous CuSO4
(0.1 mL). The mixture was heated in a microwave reactor at 125 ꢀC
for 10 min (10 W). The product was obtained as a white gum
(4.0 mg, 0.008 mmol,18% yield). Rf¼0.22 in MeOH/EtOAc (2:8). ½a D25
ꢄ
þ178.4 (c 0.27, CHCl3). 1H NMR
d
7.40 (s, 1H, H-50), 7.21 (d, J 8.0 Hz,
2H, ArH), 7.13 (t, J 7.5 Hz, 2H, ArH), 7.09 (d, J 7.5 Hz, 1H, ArH), 4.42
(br s, 1H, H-2), 4.27 (t, J 7.0 Hz, 2H, H-100), 4.08 (s, 3H, OeCH3), 3.53
(br s, 1H, H-9a), 3.30 (d, J 6.0 Hz, 1H, H-7), 3.10e3.06 (m, 1H, H-10),
3.06e3.02 (m, 2H, H-5), 2.60 (t, J 7.5 Hz, 2H, H-300), 2.18 (quint, J
7.5 Hz, 2H, H-200), 2.00 (s, 3H, H-16), 1.99 (d, J 14.0 Hz, 1H, H-1a),
1.90e1.87 (m, 2H, H-1b, H-6a), 1.86e1.82 (m, 2H, H-6b, H-9), 1.34
3.1.29. Click reaction method C: preparation of (5Z)-5-[(2S,2aR,6-
S,7aS,7bR,8R,9S)-hexahydro-7b-1-(3-phenylisoxazol-5-yl)-9-methyl-
4H-2,2,6-(epoxy[1]propanyl[3]ylidene)furo[2,3,4-gh]pyrrolizin-10-
ylidene]-4-methoxy-3-methyl-2(5H)-furanone (31). To a solution of
the alkyne 26 (15.1 mg, 0.042 mmol), Cu(OAc)2 (3.1 mg,
0.017 mmol) and sodium ascorbate (12.6 mg, 0.064 mmol) in EtOH/
H2O (1:1) (1.0 mL) at rt was added N-hydroxybenzenecarbox-
imidoyl chloride (10 mg, 0.064 mmol) and sodium hydroxide
(2.6 mg, 0.064 mmol). The reaction was left to stir for 3 h. The
mixture was quenched with saturated aqueous NaHCO3 solution
(10 mL) and was directly extracted with CH2Cl2 (3ꢃ20 mL). The
combined organic extracts were washed with brine and dried
(MgSO4) before being concentrated in vacuo. The concentrated
residue was purified by column chromatography using gradient
elution from CH2Cl2 to CH2Cl2/MeOH (9:1) to give a white gum
(11.9 mg, 0.025 mmol, 59% yield). Rf¼0.37 in MeOH/EtOAc (1:9).
(d, J 6.5 Hz, 3H, H-17). 13C NMR
d 169.8 (C-15), 162.9 (C-13), 148.3
(C-12), 147.3 (C-40), 140.2 (ArC), 128.6 (ArCH), 128.5 (ArCH), 128.0
(C-11), 126.4 (ArCH), 121.0 (C-50), 112.6 (C-8), 98.6 (C-14), 81.0 (C-2),
79.8 (C-3), 61.1 (C-9a), 59.0 (OeCH3), 52.1 (C-7), 49.7 (C-100), 47.7
(C-9), 47.5 (C-5), 34.6 (C-10), 33.3 (C-1), 32.6 (C-300), 31.6 (C-200), 27.2
(C-6), 18.3 (C-17), 9.2 (C-16). ESIMS m/z 517.4 (100%) [MþH]þ.
HRESIMS m/z 517.2439 [MþH]þ, calcd for C29H33N4O5 517.2451.
3.1.27. Click reaction method B: preparation of (5Z)-5-[(2S,2aR,6-
S,7aS,7bR,8R,9S)-hexahydro-7b-1-(1-phenyl-1H-1,2,3-triazol-4-yl)-
9-methyl-4H-2,2,6-(epoxy[1]propanyl[3]ylidene)furo[2,3,4-gh]pyrro-
lizin-10-ylidene]-4-methoxy-3-methyl-2(5H)-furanone (29). To a so-
lution of the alkyne 26 (16.2 mg, 0.046 mmol), Cu(OAc)3 (0.8 mg,
0.005 mmol) and sodium ascorbate (13.6 mg, 0.068 mmol) in
MeOH/H2O (1:1) (1.0 mL) at rt was added 0.09 M phenylazide in
MeOH (1.0 mL) and the reaction mixture was left to stir for 3 h. The
mixture was quenched with saturated aqueous NaHCO3 solution
(10 mL) and was directly extracted with CH2Cl2 (3ꢃ20 mL). The
combined organic extracts were washed with brine and dried
(MgSO4) before being concentrated in vacuo. The concentrated
residue was purified by column chromatography using gradient
elution from CH2Cl2 to CH2Cl2/MeOH (9:1) to give a yellow gum
(7.2 mg, 0.015 mmol, 33% yield). Rf¼0.37 in MeOH/CH2Cl2 (1:9).
½
a 2D5
ꢄ
þ203.4 (c 0.79, CHCl3). 1H NMR
d 7.82e7.79 (m, 2H, ArH), 7.45
(br s, 3H, ArH), 6.56 (s, 1H, H-40), 4.62 (s, 1H, H-2), 4.16 (s, 3H,
OeCH3), 3.65 (br s, 1H, H-9a), 3.43 (d, J 5.5 Hz, 1H, H-7), 3.29e3.24
(m, 1H, H-5b), 3.19e3.14 (m, 2H, H-5a, H-10), 2.11e2.08 (m, 1H,
H-1a), 2.09 (s, 3H, H-16), 2.03e1.96 (m, 3H, H-1a, H-6), 1.93 (dd, J
9.5 Hz, 2.5 Hz, 1H, H-9), 1.43 (d, J 6.0 Hz, 3H, H-17). 13C NMR
d 171.2
(C-50), 169.7 (C-15), 162.8 (C-13), 162.6 (C-30), 147.8 (C-12), 130.3
(ArC), 128.9 (ArC), 128.8 (ArC), 128.3 (C-11), 127.5 (ArC), 112.5 (C-8),
100.2 (C-40), 99.0 (C-14), 80.6 (C-3), 80.1 (C-2), 61.3 (C-9a), 59.0
(OeCH3), 52.3 (C-7), 49.0 (C-5), 47.7 (C-9), 34.6 (C-10), 33.3 (C-1),
27.2 (C-6), 18.4 (C-17), 9.3 (C-16). ESIMS m/z 475.1 (100%) [MþH]þ.
HRESIMS m/z 475.1871 [MþH]þ, calcd for C27H27N2O6 475.1869.
½
a 2D5
ꢄ
þ229.9 (c 0.48, CHCl3). 1H NMR
d
7.95 (s, 1H, H-50), 7.74 (d, J
3.1.30. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-[3-(4-
fluorophenyl)isoxazol-5-yl]-9-methyl-4H-2,2,6-(epoxy[1]propanyl[3]
ylidene)furo[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-3-methyl-
2(5H)-furanone (32). Prepared using method C, from the alkyne 26
(10.6 mg, 0.030 mmol), N-hydroxy-4-fluorobenzenecarboximidoyl
chloride (20 mg, 0.153 mmol), sodium ascorbate (8.8 mg,
0.045 mmol), sodium hydroxide (1.8 mg, 0.045 mmol) and
Cu(OAc)2 (2.2 mg, 0.012 mmol). The product was obtained as
a yellow gum (6.3 mg, 0.013 mmol, 43% yield). Rf¼0.62 in MeOH/
8.0 Hz, 2H, ArH), 7.54e7.51 (m, 2H, ArH), 7.44 (t, J 7.5 Hz, 1H, ArH),
4.58 (s, 1H, H-2), 4.16 (s, 3H, OeCH3), 3.64 (br s, 1H, H-9a), 3.50 (d, J
5.5 Hz, 1H, H-7), 3.24e3.12 (m, 3H, H-5, H-10), 2.12e2.07 (m, 1H,
H-1a), 2.09 (s, 3H, H-16), 2.05e2.02 (m, 2H, H-1b, H-6a), 2.00e1.98
(m, 1H, H-6b), 1.97e1.93 (m, 1H, H-9), 1.43 (d, J 7.0 Hz, 3H, H-17).
13C NMR (one ArCH was not observed due to peak overlap)
d 169.8
(C-15), 163.0 (C-13), 148.3 (C-12), 137.1 (ArC), 129.9 (ArCH), 128.9
(C-11), 128.2 (C-40), 120.6 (ArCH), 119.4 (C-50), 112.7 (C-8), 98.9
(C-14), 81.2 (C-2), 80.0 (C-3), 61.3 (C-9a), 59.0 (OeCH3), 52.3 (C-7),
48.7 (C-5), 47.9 (C-9), 34.7 (C-10), 33.4 (C-1), 27.3 (C-6), 18.5 (C-17),
9.3 (C-16). ESIMS m/z 475.1 (100%) [MþH]þ. HRESIMS m/z 475.1972
[MþH]þ, calcd for C26H27N4O5 475.1981.
EtOAc (2:8). ½a 2D5
ꢄ
þ236.1 (c 0.36, CHCl3). 1H NMR
d 7.80e7.77 (m,
2H, ArH), 7.15 (t, J 8.5 Hz, 2H, ArH), 6.52 (s, 1H, H-40), 4.61 (br s, 1H,
H-2), 4.16 (s, 3H, OeCH3), 3.65 (br s, 1H, H-9a), 3.42 (d, J 5.5 Hz, 1H,
H-7), 3.29e3.23 (m, 1H, H-5), 3.19e3.12 (m, 2H, H-5, H-10), 2.11 (s,
1H, H-1a), 2.09 (s, 3H, H-16), 2.04e1.96 (m, 3H, H-1b, H-6), 1.93 (dd,
J 10.0 Hz, 3.0 Hz, 1H, H-9), 1.42 (d, J 6.5 Hz, 3H, H-17). 13C NMR
3.1.28. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-[3-(2-
propylisoindoline-1,3-dionyl)-1H-1,2,3-triazol-4-yl]-9-methyl-4H-
2,2,6-(epoxy[1]propanyl[3]ylidene)furo[2,3,4-gh]pyrrolizin-10-
ylidene]-4-methoxy-3-methyl-2(5H)-furanone (30). Prepared using
method B, from the alkyne 26 (12.1 mg, 0.034 mmol), N-pro-
pylphthalimide azide (0.50 mL, 0.187 M in THF, 0.068 mmol), so-
dium ascorbate (10 mg, 0.051 mmol) and Cu(OAc)2 (0.6 mg,
0.003 mmol). The product was obtained as a colourless gum
(9.1 mg, 0.016 mmol, 46% yield). Rf¼0.15 in MeOH/EtOAc (1:9).
d
171.5 (C-50), 169.7 (C-15), 164.0 (ArCeF, d, JCeF 250 Hz), 162.8
(C-13), 161.7 (C-30), 147.8 (C-12), 128.9 (ArCH d, JCeF 9.8 Hz), 128.8
(ArCH), 128.3 (C-11), 125.1 (ArC, d, JCeF 3.3 Hz), 116.3 (ArCH, d, JCeF
22.0 Hz), 116.1 (ArCH), 112.4 (C-8), 100.1 (C-40), 99.0 (C-14), 80.6
(C-3), 80.1 (C-2), 61.3 (C-9a), 59.0 (OeCH3), 52.3 (C-7), 49.0 (C-5),
47.7 (C-9), 34.6 (C-10), 33.3 (C-1), 27.2 (C-6), 18.4 (C-17), 9.3 (C-16).
ESIMS m/z 492.9 (100%) [MþH]þ. HRESIMS m/z 493.1789 [MþH]þ,
calcd for C27H26N2O6F 493.1775.
½
a 2D5
ꢄ
þ131.4 (c 0.61, CHCl3). 1H NMR
d 7.76e7.74 (m, 2H, ArH),
7.67e7.66 (m, 2H, ArH), 7.65 (s, 1H, H-50), 4.41 (br s, 1H, H-2), 4.34
(t, J 7.0 Hz, 2H, H-1000), 4.08 (s, 3H, OeCH3), 3.66 (t, J 6.5 Hz, 2H, H-
3000), 3.53 (br s, 1H, H-9a), 3.26 (d, J 6.0 Hz, 1H, H-7), 3.10e3.00 (m,
3H, H-5, H-10), 2.27 (quint, J 6.5 Hz, 2H, H-2000), 1.98 (s, 3H, H-16),
3.1.31. Sonogashira coupling: preparation of (5Z)-5-[(2S,2aR,6-
S,7aS,7bR,8R,9S)-hexahydro-7b-1-(2-phenylethyn-1-yl)-9-methyl-
4H-2,2,6-(epoxy[1]propanyl[3]ylidene)furo[2,3,4-gh]pyrrolizin-10-
ylidene]-4-methoxy-3-methyl-2(5H)-furanone
(33). A
solution

7114
K. Sastraruji et al. / Tetrahedron 68 (2012) 7103e7115
mixture of the alkyne26 (13.4 mg, 0.038 mmol), iodobenzene (4.2
mL,
13C NMR
d 169.8 (C-15), 162.9 (C-13), 148.5 (C-12), 141.7 (ArC), 128.6
0.038 mmol), PdCl2(PPh3)2 (0.5 mg, 0.001 mmol), CuI (0.3 mg,
0.002 mmol) and Et3N (19 mg, 0.189 mmol) in THF (1.0 mL) in a glass
tube was bubbled with argon gas and then the tube was sealed. The
reaction was left to stir at rt for 24 h. The mixture was extracted with
EtOAc (3ꢃ20 mL). The combined organic phase was dried (Na2SO4).
The crude residue was purified by PTLC with MeOH/CH2Cl2 (1:99) as
a mobile phase to give a phenyl alkyne product 33 as a yellow gum
(10.0 mg, 0.023 mmol, 62% yield) and the dimeric by-product 34
(0.6 mg, 0.0001 mmol, 4% yield), which was directly prepared via the
(ArCH), 128.3 (ArCH), 128.0 (C-11), 126.1 (ArCH), 112.7 (C-8), 98.6 (C-
14), 82.7 (C-3), 78.5 (C-2), 61.0 (C-9a), 58.9 (OeCH3), 50.4 (C-7), 47.7
(C-5, C-9), 34.6 (C-10), 33.7 (C-20), 33.4 (C-1), 31.4 (C-10), 26.7 (C-6),
18.4 (C-17), 9.2 (C-16). ESIMS m/z 436.1 (100%) [MþH]þ. HRESIMS m/z
436.2118 [MþH]þ, calcd for C26H30NO5 436.2124.
3.1.34. AChE inhibition studies.
3.1.34.1. TLC bioautographic method. The AChE used in this assay
was extracted from electric eels and purchased from Sigma-
eAldrich (EC 3.1.1.7). The enzyme stock solution was prepared from
a solution of AChE (1000 U) in 0.05 M trisehydrochloric acid buffer
(150 mL) at pH 7.8 to which was added bovine serum albumin
(150 mg) to stabilize the enzyme. The stock solution was kept at
4 ꢀC. TLC plates used for the bioautography were washed with ac-
etone and then thoroughly dried. The samples were prepared as
solutions in MeOH at concentrations of 1000, 100, 10, 1 and
0.1 ppm. The samples were applied to the TLC plates in varying
Eglington coupling reaction. Rf¼0.42 in MeOH/EtOAc (1:9). ½a D25
ꢄ
þ235.9 (c 0.44, CHCl3). 1H NMR
d 7.45e7.44 (m, 2H, ArH), 7.31e7.30
(m, 3H, ArH), 4.66 (br s,1H, H-2), 4.15 (s, 3H, OeCH3), 3.56 (br s,1H, H-
9a), 3.48e3.42 (m,1H, H-5a), 3.14 (d, J 6.0 Hz, 1H, H-7), 3.11e3.08 (m,
2H, H-5b, H-10), 2.23e2.17 (m,1H, H-6a), 2.08 (s, 3H, H-16), 2.06 (d, J
12.0 Hz, 1H, H-1a), 1.98 (d, J 12.0 Hz, 1H, H-1b), 1.93e1.88 (m, 1H, H-
6b), 1.83 (d, J 10.0 Hz, 1H, H-9), 1.39 (d, J 6.5 Hz, 3H, H-17). 13C NMR
d
169.8 (C-15), 162.9 (C-13), 148.0 (C-12), 131.9 (ArCH), 128.6 (ArCH),
128.4 (ArCH), 128.2 (C-11), 122.5 (ArC), 112.2 (C-8), 98.9 (C-14), 86.3
(C-20), 86.1 (C-10), 81.2 (C-2), 76.0 (C-3), 60.8 (C-9a), 59.0 (OeCH3),
54.6 (C-7), 48.7 (C-5), 47.4(C-9), 34.6(C-10), 33.6 (C-1), 27.1 (C-6),18.4
(C-17), 9.3 (C-16). ESIMS m/z 432.1 (100%) [MþH]þ. HRESIMS m/z
432.1821 [MþH]þ, calcd for C26H26NO5 432.1811.
quantities using Camag Nanomat 4 TLC spotter with 0.5 mL capil-
laries and sprayed with AChE enzyme stock solution and thor-
oughly dried again. The plates were laid flat on plastic plugs in
a covered water bath (to avoid the plates from contacting the H2O
directly) and then incubated in a humid atmosphere at 37 ꢀC for
20 min. The plates were taken out and sprayed with a freshly
prepared indicator solution which was a mixture of two solutions,
a solution of 1-naphthyl acetate (25 mg) in EtOH (10 mL) and
a solution of Fast Blue B salt (40 mg) in H2O (16 mL). After 1e2 min,
a purple colouration on the TLC plates appeared and white spots
indicated inhibition of AChE by the samples.
3.1.32. Eglington coupling: preparation of bis-(5Z)-5-[(2S,2aR,6-
S,7aS,7bR,8R,9S)-hexahydro-7b-1-ethynyl-9-methyl-4H-2,2,6-(epoxy
[1]propanyl[3]ylidene)furo[2,3,4-gh]pyrrolizin-10-ylidene]-4-
methoxy-3-methyl-2(5H)-furanone (34). A solution of the alkyne 26
(13.5 mg, 0.038 mmol) and Cu(OAc)2 (35 mg, 0.190 mmol) in dry
MeCN (1.0 mL) in a sealed tube was bubbled with argon gas and then
the tube was sealed. The reaction mixture was heated and stirred at
40 ꢀC for 4 h. The mixture was cooled to rt and filtered through a thin
pad of Celite. The filtrate was treated with an aqueous solution of
NaHCO3 and extractedwithCH2Cl2 (3ꢃ20 mL). The combined organic
phase was washed with brine and dried (MgSO4). The evaporated
residue was purified by PTLC with MeOH/CH2Cl2 (2:98) as a mobile
phase to give a dimer 34 as a yellow gum (12.0 mg, 0.017 mmol, 89%
3.1.34.2. Spectroscopic-based method. Acetylthiocholine iodide
(ATChI) was used as a substrate while 5,50-dithiobis[2-nitrobenzoic
acid] (DTNB) was used as a reagent. Two stock solutions were used
in this assay, buffer solution and substrate solution. The pH 7.0
phosphate buffer solution was prepared from a mixture of 37.2 mM
NaH2PO4$H2O and 62.7 mM Na2HPO4$2H2O, in Milli Q H2O. The
substrate solution was prepared as a 4.73 mM ATChI solution in
phosphate buffer pH 7.0. The reagent solution was prepared as
a 3.15 mM DTNB solution in phosphate buffer pH 7.0.
yield). Rf¼0.50 in MeOH/CH2Cl2 (1:9). ½a D25
ꢄ
þ286.3 (c 0.39, CHCl3). 1H
NMR d 4.56 (br s, 2H, H-2), 4.13 (s, 6H, OeCH3), 3.50 (br s, 2H, H-9a),
3.37e3.31 (m, 2H, H-5a), 3.10e3.02 (m, 4H, H-5b, H-10), 3.05 (d, J
6.0 Hz, 2H, H-7), 2.14e2.08 (m, 2H, H-6a), 2.06 (s, 6H, H-16), 2.00 (d, J
12.0 Hz, 2H, H-1a), 1.89e1.86 (m, 2H, H-1b), 1.85e1.83 (m, 2H, H-6b),
1.76(dd, J10.5 Hz, 3.5 Hz, 2H, H-9),1.35(d, J6.5 Hz, 6H, H-17).13C NMR
The assay was performed in 96-well plates. In each well, 120
of phosphate buffer pH 7.0 was mixed with 20 L of reagent and
20 L of substrate, then 20 L of AChE (0.75 U/mL) in phosphate
buffer pH 7.0 was added with 20 L of the sample (which was
prepared in concentrations of 5000, 1000, 200, 40, 8, 1.6 and
0.32 M in DMSO). The final concentrations were 500, 100, 20, 4,
mL
m
m
m
d
169.7 (C-15),162.8 (C-13),147.6 (C-12),128.2 (C-11),111.9 (C-8), 99.0
m
(C-14), 80.9 (C-2), 77.4 (C-3), 75.8 (C-10), 70.4 (C-20), 60.8 (C-9a), 59.0
(OeCH3), 54.8 (C-7), 48.9 (C-5), 47.3 (C-9), 34.6 (C-10), 33.6 (C-1), 27.0
(C-6), 18.3 (C-17), 9.3 (C-16). ESIMS m/z 709.5 (100%) [MþH]þ. HRE-
SIMS m/z 709.2791 [MþH]þ, calcd for C40H41N2O10 709.2761.
m
0.8, 0.16 and 0.032, respectively. Then the well plate was directly
put into the microplate reader, which was thermostated at 25 ꢀC.
The absorbances were read using a SPECTRAmaxÒ PLUS384 micro-
plate thermostated spectrophotometer (California, USA) at 412 nm,
every 15 s for 30 min continuously. Enzyme activity was calculated
as a percentage compared to an assay using a buffer without any
inhibitor. The AChE inhibitory data were analysed with the soft-
ware package GraphPad PrismÒ (Graph Pad Inc., San Diego, USA).
IC50 values are meansꢁSD of three individual determinations each
performed in triplicate.
3.1.33. (5Z)-5-[(2S,2aR,6S,7aS,7bR,8R,9S)-Hexahydro-7b-1-(2-
phenylethyl)-9-methyl-4H-2,2,6-(epoxy[1]propanyl[3]ylidene)furo
[2,3,4-gh]pyrrolizin-10-ylidene]-4-methoxy-3-methyl-2(5H)-fur-
anone (35). To
a mixture of the phenyl alkyne 33 (4.7 mg,
0.011 mmol) and Pd/C (0.5 mg, 0.002 mmol) in EtOAc (1.0 mL) at rt,
a hydrogen gas was bubbled into the solution and the reaction was let
to stir for 24 h. The reaction mixture was then filtered through a thin
pad of Celite. The concentrated residue was purified by PTLC with
MeOH/EtOAc(1:99)asamobilephasetogiveaproductasawhitegum
Acknowledgements
(3.5 mg, 0.008 mmol, 74% yield). Rf¼0.32 in MeOH/CH2Cl2 (1:9). ½a D25
ꢄ
þ248.4 (c 0.23, CHCl3). 1H NMR
d 7.20 (d, J 7.0 Hz, 2H, ArH), 7.13e7.10
We thank the Australian Research Council and the University of
Wollongong for financial support.
(m, 3H, ArH), 4.24 (br s,1H, H-2), 4.06 (s, 3H, OeCH3), 3.43 (br s,1H, H-
9a), 3.12e3.06 (m, 1H, H-5), 3.05e3.00 (m, 1H, H-10), 2.99e2.93 (m,
1H, H-5), 2.72 (td, J 11.5 Hz, 5.0 Hz,1H, H-10), 2.66 (d, J 6.0 Hz,1H, H-7),
2.50(td, J 13.5 Hz, 5.5 Hz,1H, H-10), 2.00 (s, 3H, H-16),1.90 (d, J12.0 Hz,
1H, H-6a),1.90e1.79 (m, 3H, H-1a, H-9, H-20), 1.78e1.74 (m, 2H, H-6b,
H-20), 1.67 (dt, J 12.0 Hz, 3.5 Hz, 1H, H-1b), 1.31 (d, J 6.5 Hz, 3H, H-17).
Supplementary data
Compound purity analysis by HPLC, synthesis of the Best-
manneOhira reagent and the coupling components for the Click

K. Sastraruji et al. / Tetrahedron 68 (2012) 7103e7115
7115
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