Structure confirmation of a bioactive lactone isolated from Otoba parvifolia through the synthesis of a model compound

Article abstract of DOI:10.1021/np049771x

Synthesis of a model compound 4 structurally related to a bioactive lactone 3 isolated from Otoba parvifolia has been accomplished. The good match between the NMR data of both compounds suggests they have identical bicyclic [3.3.1] carbon skeletons.

Full text of DOI:10.1021/np049771x

J. Nat. Prod. 2004, 67, 1939-1941
1939
Structure Confirmation of a Bioactive Lactone Isolated from Otoba parvifolia
through the Synthesis of a Model Compound
Francisco A. Marques,*,† Cesar A. Lenz,‡ Fabio Simonelli,† Beatriz Helena L. Noronha Sales Maia,†
Adriana P. Vellasco,§ and Marcos N. Eberlin§
Chemistry Department, Federal University of Parana´, P.O. Box 19081, Curitiba, PR, 81.531-990 Brazil,
Parana´ Institute of Technology, TECPAR, Curitiba, PR, 81.350-010 Brazil, and Thomson Mass Spectrometry Laboratory,
State University of Campinas, UNICAMP, Campinas, SP, 13083-970 Brazil
Received July 9, 2004
Synthesis of a model compound 4 structurally related to a bioactive lactone 3 isolated from Otoba parvifolia
has been accomplished. The good match between the NMR data of both compounds suggests they have
identical bicyclic [3.3.1] carbon skeletons.
Gottlieb and co-workers, when studying the fruit kernels
of Otoba parvifolia (Mkgf), A. Gentry, isolated and identi-
fied a number of novel compounds.1 One of these com-
pounds was assigned the novel bicyclic lactone structure
1. This lactone has promise for new antibiotic development
since it shows activity in vitro as pronounced as tetracycline
against important microorganisms such as Staphylococus
aureus, Staphylococus epidermis, and Peptococus sp.2
Attracted by the novel structure 1 and its promising
biological activity, we synthesized the model compound 2
for structural confirmation.3 However, a clear mismatching
of NMR data was observed between the synthetic model
and natural compounds, which led Gottlieb and co-workers
to revise the structure of the natural lactone4 by proposing
the alternative bicyclic lactone structure 3.
To verify the correctness of the revised structure 3,
comparison with another model compound has now been
performed. Herein we present the synthesis and structural
characterization of the model bicyclic lactone 4, which
contains the same bicyclic ring system as the revised
structure 3.
The strategy chosen for synthesis of compound 4 em-
ployed an iodolactonization as the key step (Scheme 1). The
double alkylation of cyclohexenone was carried out under
kinetic conditions and in the presence of HMPA,5 affording
compound 6 in 82% yield for the two steps. When ketone 6
was treated with ethylene glycol in the presence of PTSA
in benzene,6 a mixture of ketals 7 and 8 was obtained in a
9.5:1 ratio, as determined by GC. The regioisomers were
separated by flash chromatography and the migration of
Scheme 1. Synthesis of Model Lactone 4a
1
the double bond was confirmed by the H NMR spectrum
of the major compound, which showed a multiplet for the
vinylic hydrogens. Ester 7 was then hydrolyzed in the
presence of KOH and the acid derivative submitted to
iodolactonization conditions,7,8 furnishing the iodo lactone
9 in 60% yield.
Dehydrohalogenation of the iodolactone 9 with DBU in
benzene9 afforded ketal lactone 10 in 75% yield. Removal
of the ketal protecting group of 10 by transketalization with
PTSA in acetone6 generated the model compound 4 in 65%
yield.
The NMR data of compound 4 showed good agreement
with that of the natural lactone. A good match between
a Reagents: (a) LDA, THF, HMPA, -78 °C, MeI. (b) LDA, THF, HMPA,
-78 °C, BrCH2COOEt. (c) PTSA, HOCH2CH2OH, benzene, reflux 20 h. (d)
KOH, THF/H2O (3:1); HCl. (e) I2, KI, NaHCO3, THF, Et2O. (f) DBU, benzene.
(g) acetone, PTSA.
* To whom correspondence should be addressed. E-mail: tic@ufpr.br.
Tel: 41-361-3174. Fax: 41-361-3186.
† Federal University of Parana´.
the 13C NMR data of both compounds (Table 1) was
observed, except for the peaks attributed to C-2, C-1′, and
‡ Parana´ Institute of Technology.
§ Thomson Mass Spectrometry Laboratory, State University of Campinas.
10.1021/np049771x CCC: $27.50
© 2004 American Chemical Society and American Society of Pharmacognosy
Published on Web 11/03/2004

1940 Journal of Natural Products, 2004, Vol. 67, No. 11
Notes
Table 1. 13C NMR (δ) of Natural Lactone 3 and Model
Compound 4
chromatography (TLC: E. Merck, Type 5554 plates) and gas
chromatography (GC) with a Varian 3800 GC (flame ionization
detector, FID) equipped with a Varian CP-SIL 8 capillary
column (30 m × 0.25 mm × 0.25 µm). Flash column chroma-
tography was carried out with 230-400 mesh silica gel (E.
Merck). IR spectra were recorded on a Bomer model MB 102
spectrometer with internal calibration. Low-resolution mass
spectra were obtained on a GC/MS Varian Saturn 2000
spectrometer. High-resolution and high-accuracy MS and 20
eV MS/MS collision-induced dissociation (CID) spectra were
acquired using electrospray ionization (ESI) on positive ion
mode on a Q-Tof (Micromass, UK) mass spectrometer of hybrid
quadrupole orthogonal time-of-flight configuration. Typical ESI
conditions were as follows: capillary voltage of 1 kV, cone
voltage of 40 V, and dessolvation gas temperature of 100 °C.
1H NMR and 13C NMR spectra were recorded in CDCl3
solutions using a Bruker Advance-400 spectrometer. Deuter-
ated solvents were used as lock and TMS as reference signal.
Chemical shifts (δ) are given in ppm and coupling constants
(J) in Hz. The 13C chemical shifts (δ) are reported in ppm
relative to the center peak of CDCl3 (δ 77.00).
6-Methyl-2-cyclohexenone (5). To a solution of diisopro-
pylamine (1.58 g, 15.6 mmol) in THF (8 mL) at 0 °C was added
n-BuLi in hexanes (5.5 mL, 14.3 mmol). After stirring for 20
min the solution was cooled to -78 °C and 2-cyclohexenone
(1.00 g, 10.4 mmol) in THF (10 mL) was added dropwise. The
solution was stirred for 30 min followed by dropwise addition
of iodomethane (2.96 g, 20.8 mmol). After stirring for 30 min
at -78 °C, HMPA (6 mL) was added and the mixture was kept
for another 2 h at -78 °C. Ether (20 mL) was added to the
reaction mixture at 0 °C, and the organic layer was washed
with saturated NH4Cl (5 × 5 mL) and saturated NaCl (3 × 5
mL) and dried over Na2SO4. After filtration the solvent was
removed by rotatory evaporation and the crude product was
purified by flash chromatography with hexane/ethyl acetate
(10:1 v/v) to give compound 5 (1.07 g, 93%): IR (KBr) νmax 3033,
2964, 2872, 1680 cm-1; 1H NMR (CDCl3, 400 MHz) δ 1.15 (3H,
d, J ) 6.8 Hz), 1.65-1.85 (1H, m), 1.95-2.10 (1H, m), 2.30-
2.45 (3H, m), 5.98 (1H, dt, J ) 10.0, 2.0 Hz), 6.94 (1H, dt, J )
10.0, 4.96 Hz); 13C NMR (CDCl3, 100 MHz) δ 15.0, 25.5, 30.8,
41.6, 129.4, 149.6, 202.3; EIMS m/z 110 [M+] (42), 81 (3), 68
(100), 53 (4), 39 (27); CIMS (CH3CN) m/z 111 [M + 1]+ (100).
Ethyl (1-Methyl-2-oxo-3-cyclohexenyl)acetate (6). To a
solution of diisopropylamine (0.30 g, 2.95 mmol) in THF (3 mL)
at 0 °C was added n-BuLi in hexanes (1.20 mL, 2.85 mmol).
After stirring for 20 min the solution was cooled to -78 °C,
followed by dropwise addition of 3-methyl-2-cyclohexenone
(0.21 g, 1.9 mmol) in THF (3 mL). After 2 h at -78 °C ethyl
bromoacetate (0.65 g, 3.82 mmol) was added dropwise. The
resulting solution was stirred for 30 min at -78 °C, and HMPA
(3 mL) was added. After stirring for 3 h at -78 °C ether (20
mL) was added followed by the addition of saturated NH4Cl
(5 mL). The organic solution was washed with saturated NH4-
Cl (5 × 5 mL) and saturated NaCl (3 × 5 mL), dried over Na2-
SO4, and concentrated under reduced pressure. The crude
product was purified by flash chromatography using a mixture
of hexane/ethyl acetate (5:1, v/v), affording compound 6 (0.33
g, 89%): IR (KBr) νmax 3033, 2977, 2934, 2872, 1735, 1677,
1215 cm-1; 1H NMR (CDCl3, 200 MHz) δ 1.15 (3H, s), 1.20 (3H,
t, J ) 7 Hz,), 1.70-1.90 (2H, m), 2.18-2.42 (2H, m), 2.70 (1H,
s), 2.78 (1H, s), 4.07 (2H, q, J ) 7.0 Hz), 5.94 (1H, dt, J )
10.0, 1.7 Hz), 6.88 (1H, dt, J ) 10.0, 3.9 Hz,); 13C NMR (CDCl3,
50 MHz) δ 14.0, 21.8, 22.9, 32.6, 41.6, 43.3, 60.1, 128.1, 148.7,
171.2, 201.9; EIMS m/z 197 [M + 1]+ (100), 151 (86), 122 (11),
107 (11), 68 (42), 39 (22); CIMS (CH3CN) m/z 197 [M + 1]+
(100).
C
natural lactone 3
model compound 4
1
169.0
39.7
168.5
41.6
2
1′
2′
3′
4′
5′
6′
46.0
42.0
200.3
130.7
144.5
69.8
200.5
130.3
144.4
69.7
33.6
36.9
Table 2. 1H NMR Data of the Natural Lactone 3 and Model
Compound 4
natural lactone
3 (300 MHz)
model compound
4 (400 MHz)
H
2R
2â
3′
2.57 dd
2.72 dd
2.58 d
2.67 d
6.13 d
6.15 d
4′
7.13 ddd
5.00-5.10 m
2.26 ddd
2.20 ddd
7.15 ddd
5.00-5.10 m
2.31 dt
5′
6′R
6′â
2.25 ddd
Figure 1. Long-range coupling (W-coupling) between H-4′ and H-6′â
(J ) 1.57 Hz) and H-6′R and H-2R (J ) 2.02 Hz).
C-6′, which for 3 are under the influence of the farnesyl
substituent and of the methyl substituent for the model
compound 4. The chemical shifts and multiplicity observed
for the vinylic and allylic hydrogens of the model compound
4 are very closely related to those of the natural lactone 3
(Table 2). The double double doublet observed for the signal
at δ 7.15 results from two vicinal couplings of H-4′ with
H-3′ (J ) 9.79 Hz) and with H-5′ (J ) 5.96 Hz), along with
a W-coupling with H-6′â (J ) 1.57 Hz). Another W-coupling
between H-2R and H-6′R (J ) 2.02 Hz) is observed in the
1H NMR spectrum of 4, as shown in Figure 1. The small
difference in the chemical shifts of H-2 and H-6′ observed
between the model and natural substance can be attributed
to the different substituents present on C-1′.
The mass spectrum showed that the protonated bicyclic
lactone 4 dissociated upon collision activation (20 eV CID)
by sequential loss of H2O and CO molecules, and this
dissociation chemistry is typical for protonated lactones.10
Thus, synthesis of model compound 4, structurally
related to bioactive lactone 3 isolated from Otoba parvifolia,
has been performed. Comparison of spectral data suggests
that 3 and 4 have identical bicyclic carbon skeletons. An
enantioselective total synthesis of the natural lactone is
being carried out in our laboratory, which it is hoped will
permit the determination of the absolute configuration of
the natural compound 3.
Experimental Section
General Experimental Procedures. Reagents and sol-
vent were purified and dried using standard methods. Reac-
tions involving organometallic reagents were carried out under
argon in oven-dried glassware. Reagents employed: HMPA
(hexamethylphosphoramide), DBU (1,8-diazabicyclo[5.4.0]-
undec-7-ene, LDA (lithium diisopropylamide), PTSA (p-tolu-
enesulfonic acid). Reactions were monitorated by thin-layer
Ethyl (6-Methyl-1,4-dioxaspiro[4.5]dec-8-en-6-yl)ace-
tate (7). A solution of ketone 6 (0.631 g, 3.21 mmol), ethylene
glycol (1.20 g, 19.3 mmol), and p-toluenesulfonic acid mono-
hydrate (0.077 g, 0.4 mmol) in benzene (15 mL) was refluxed
in a flask connected to a Dean Stark trap for 20 h. The mixture
was diluted with ether (50 mL), washed with saturated
NaHCO3 (2 × 20 mL) and water (1 × 20 mL), and dried over

Notes
Journal of Natural Products, 2004, Vol. 67, No. 11 1941
MgSO4. The filtrate was concentrated in vacuo and purified
by flash chromatography using a mixture of hexane/ether (1:1
v/v) to give ketal 7 (0.487 g, 63%): IR (KBr) νmax 2979, 2360,
2340, 1727, 1027 cm-1; 1H NMR (CDCl3, 400 MHz) δ 1.11 (3H,
s), 1.26 (3H, t, J ) 7.14 Hz), 2.14 (1H, ddd, J ) 17.8, 3.94,
1.91 Hz), 2.21-2.23 (2H, m), 2.34-2.46 (3H, m), 3.94-4.00
(4H, m), 4.11 (2H, q, J ) 7.14 Hz,), 5.55-5.59 (1H, m), 5.64-
5.67 (1H, m); 13C NMR (CDCl3, 100 MHz) δ 14.3, 19.9, 33.3,
36.6, 39.7, 40.2, 60.0, 65.1, 65.2, 111.0, 123.7, 125.9, 172.6;
EIMS m/z 241 [M +1]+ (21), 240 [M+] (25), 195 (31), 186 (33),
152 (14), 125 (100), 113 (58); HRESIMS m/z 241.1411 [M +
H]+ (calcd for C13H21O4, 241.1439).
1719, 1692, 1448, 1397, 1376, 1204, 1107, 1006, 954, 924, 799
cm-1; 1H NMR (CDCl3, 400 MHz) δ 1.09 (3H, s), 1.88 (1H, ddd,
J ) 14.1, 2.88, 1.64 Hz), 2.27 (1H, dt, J ) 14.1, 1.43 Hz), 2.41
(1H, d, J ) 19.8 Hz), 3.10 (1H, dd, J ) 19.8, 1.43 Hz), 3.84-
4.12 (4H, m), 4.76-4.79 (1H, m), 5.79 (1H, d, J ) 9.79 Hz),
6.07 (1H, ddd, J ) 9.79, 5.40, 1.64 Hz); 13C NMR (CDCl3, 100
MHz) δ 22.4, 35.1, 37.5, 39.3, 65.5, 65.8, 70.1, 107.9, 127.4,
131.9, 171.3; EIMS m/z 210 [M+] (3), 151 (2), 139 (5), 128 (100),
112 (8), 99 (10), 83 (12), 55 (21); HRESIMS m/z 211.0937 [M
+ H]+ (calcd for C11H15O4, 211.0970).
5-Methyl-2-oxabicylo[3.3.1]non-7-ene-3,6-dione (4). To
a solution of compound 10 (51 mg, 0.24 mmol) in acetone (15
mL) was added PTSA (5 mg, 0.030 mmol). After stirring for 5
h at room temperature, the solvent was removed under
reduced pressure and ether (20 mL) was added to the residue.
The organic phase was washed with saturated NaHCO3 (2 ×
10 mL) and dried over MgSO4. The filtrate was concentrated
in vacuo and purified by flash chromatography (hexane/ethyl
acetate, 1:1 v/v), giving model compound 4 (26 mg, 65%) as a
white noncrystalline solid: IR (KBr) νmax 2933, 1736, 1690,
4′-Iodo-1′-methyl-7′H-spiro[1,3-dioxolane-2,2′-[6]oxa-
bicyclo[3.3.1]nonan]-7′-one (9). To a solution of 7 (0.143 g,
0.6 mmol) in ethanol (15 mL) was added 20% (w/v) aqueous
KOH solution (0.25 mL, 0.9 mmol). The resulting mixture was
refluxed for 6 h, and water was added (5 mL) followed by
elimination of two-thirds of solvent under reduced pressure.
The solution was acidified by the dropwise addition of 0.1
mol‚L-1 HCl until pH 2 and extracted with ether (3 × 15 mL).
After eliminating the solvent under reduced pressure, the
crude product was dissolved in ether (3.5 mL) and a solution
of saturated NaHCO3 (3.5 mL) was added. After stirring for
30 min, a solution of I2 (0.26 g, 1 mmol) and KI (0.52 g, 3.1
mmol) in water (5 mL) was added. The resulting mixture was
vigorously stirred for 24 h and diluted with water (20 mL),
the phases were separated, and the aqueous phase was
extracted with ether (3 × 20 mL). The organic extracts were
combined and washed with 10% (w/w) Na2S2O3 (2 × 10 mL)
and saturated brine (2 × 10 mL) and dried over MgSO4. After
filtration the solvent was removed under reduced pressure and
the product was purified by flash chromatography using
hexane/ethyl acetate (3:1 v/v) to give iodolactone 9 (0.121 g,
60%): IR (KBr) νmax 2959, 1734, 1368, 1238, 1202, 1106, 1017,
1
1230, 1013 cm-1; H NMR (CDCl3, 400 MHz) δ 1.27 (3H, s),
2.25 (1H, ddd, J ) 14.1, 3.24, 1.57 Hz), 2.31 (1H, dt, J ) 14.1,
2.02 Hz), 2.58 (1H, d, J ) 19.1 Hz), 2.72 (1H, dd, J ) 19.1,
2.02 Hz), 5.00-5.10 (1H, m), 6.15 (1H, d, J ) 9.79 Hz), 7.15
(1H, ddd, J ) 9.79, 5.96, 1.57 Hz); 13C NMR (CDCl3, 100 MHz)
δ 23.3, 36.9, 41.6, 42.0, 69.7, 130.3, 144.4, 168.5, 200.5; EIMS
m/z 167 [M + 1]+ (3), 149 (4), 138 (35), 120 (33), 121 (10), 109
(23), 108 (14), 95 (47), 93 (31), 83 (54), 81 (50), 79 (35), 77 (34),
67 (10), 55 (100), 39 (43); HRESIMS m/z 167.0781 [M + H]+
(calcd for C9H11O3, 167.0708).
Acknowledgment. F.A.M. thanks the Brazilian Council
for the Development of Science and Technology (CNPq) for
financial support. M.N.E. thanks CNPq and the Sa˜o Paulo
State Research Foundation, FAPESP.
945, 840, 807, 688, 589 cm-1 1H NMR (CDCl3, 400 MHz) δ
;
1.01 (3H, s), 1.79 (1H, ddd, J ) 14.6, 4.07, 2.30 Hz), 2.03 (1H,
dd, J ) 16.7, 5.33 Hz), 2.09 (1H, dt, J ) 16.7, 1.48 Hz), 2.31
(1H, d, J ) 19.3 Hz), 2.75 (1H, dt, J ) 14.6, 2.00 Hz), 2.92
(1H, dd, J ) 19.3, 2.30 Hz), 3.98-4.12 (4H, m), 4.54-4.56 (1H,
m), 4.78-4.82 (1H, m); 13C NMR (CDCl3, 100 MHz) δ 20.4,
21.8, 30.7, 34.0, 37.7, 40.2, 65.0, 65.2, 78.9, 109.0, 170.0; EIMS
m/z 339 [M + 1]+ (6), 321 (7), 277 (3), 211 (100), 167 (14), 157
(5), 149 (8), 113 (10), 99 (9), 69 (9); HRESIMS m/z 339.0240
[M + H]+ (calcd for C11H16IO4, 339.0094).
References and Notes
(1) Ferreira, A. G.; Motidome, M.; Gottlieb, O. R.; Fernandes, J. B.; Vieira,
P. C.; Cojocaru, M.; Gottlieb, H. E. Phytochemistry 1989, 28, 579-
583.
(2) Ferreira, A. G. Personal communication. Biological tests were
performed at The Centro de Pesquisas da Rhodia, Paul´ınia, SP,
Brazil.
(3) Ferreira, J. B.; Boscaini, R. C.; Marques, F. A. Nat. Prod. Lett. 1993,
2, 313-316.
1′-Methyl-7′H-spiro[1,3-dioxolane-2,2′-[6]oxabicyclo-
[3.3.1]non[3]en]-7′-one (10). To a solution of iodo lactone 9
(115 mg, 0.34 mmol) in benzene (10 mL) was added DBU (0.10
g, 0.65 mmol). After stirring for 3 h at room temperature, the
mixture was filtered and the solid rinsed with ether (3 × 10
mL). The combined organic extracts were washed with satu-
rated NH4Cl (2 × 10 mL) and saturated NaCl (2 × 10 mL)
and dried over MgSO4. After removing the solvent under
reduced pressure, the crude product was purified by flash
chromatography (hexane/ethyl acetate, 3:1 v/v) to give com-
pound 10 (54 mg, 75%): IR (KBr) νmax 2981, 2962, 2926, 2895,
(4) Ferreira, A. G.; Fernandes, J. B.; Vieira, P. C.; Gottlieb, O. R.; Gottlieb,
H. E. Phytochemistry 1995, 40, 1723-1728.
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(6) Marques, F. A.; Ferreira, J. T. B.; Piers, E. J. Braz. Chem. Soc. 2000,
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NP049771X