The synthesis and antibacterial activity of totarol derivatives. Part 2: Modifications at C-12 and O-13

Article abstract of DOI:10.1016/S0968-0896(00)00095-X

Alterations of the C-12 and C-13 aromatic ring substituents of totarol (1) afforded the series of derivatives 2-14, and introduction of substituents at C-12 gave exclusively 2a-14a. The majority of these analogues were tested in vitro against the following organisms: β-lactamase-positive and high level gentamycin-resistant Enterococcus faecalis, penicillin-resistant Streptococcus pneumoniae, methicillin-resistant Staphylococcus aureus (MRSA), and multiresistant Klebsiella pneumoniae. The results were evaluated in terms of structure-activity relationship which reveals that: (a) the phenolic moiety at C-13, in general, is essential for antibacterial activity at <32 μg/mL against Gram-positive species, and (b) derivatization at C-12 has an undesirable effect on the antibacterial activity of this class of compounds, while (c) all compounds tested are ineffective against the Gram-negative Klebsiella pneumoniae. Copyright (C) 2000 Elsevier Science Ltd.

Full text of DOI:10.1016/S0968-0896(00)00095-X

Bioorganic & Medicinal Chemistry 8 (2000) 1653±1662
The Synthesis and Antibacterial Activity of Totarol Derivatives.
Part 2: Modi®cations at C-12 and O-13y
Gary B. Evans* and Richard H. Furneaux
Industrial Research Limited, PO Box 31-310, Lower Hutt, New Zealand
Received 9 December 1999; accepted 9 March 2000
AbstractÐAlterations of the C-12 and C-13 aromatic ring substituents of totarol (1) a€orded the series of derivatives 2 14, and
introduction of substituents at C-12 gave exclusively 2a 14a. The majority of these analogues were tested in vitro against the fol-
lowing organisms: b-lactamase-positive and high level gentamycin-resistant Enterococcus faecalis, penicillin-resistant Streptococcus
pneumoniae, methicillin-resistant Staphylococcus aureus (MRSA), and multiresistant Klebsiella pneumoniae. The results were eval-
uated in terms of structure activity relationship which reveals that: (a) the phenolic moiety at C-13, in general, is essential for
antibacterial activity at <32 mg/mL against Gram-positive species, and (b) derivatization at C-12 has an undesirable e€ect on the
antibacterial activity of this class of compounds, while (c) all compounds tested are ine€ective against the Gram-negative Klebsiella
pneumoniae. # 2000 Elsevier Science Ltd. All rights reserved.
Introduction
translocation in whole cells of a Gram-negative Pseu-
domonas aeruginosa and also NADH oxidation in
membrane preparations from this bacterium.18
The incidence of infection by antibiotic-resistant bacteria
has increased to a threatening extent since penicillin-
resistant Staphylococci were ®rst encountered in the
early 1950s,1,2 and the resistances to multiple antibiotics
of strains of Gram-positive bacteria, e.g., methicillin-
resistant Staphylococcus aureus (MRSA),3 Streptococcus
pneumoniae4 and Enterococcus faecalis3 are now sig-
ni®cant clinical problems. Compounds active against
Gram-positive species are thus of signi®cant interest,
and totarol (1),5 a phenolic diterpene which is the major
constituent of hexane extracts of the heartwood of the
New Zealand native tree Podocarpus totara G. Benn,6,7
but which is also available from other sources,5 is such a
compound. It is active against a variety of Gram-posi-
tive bacteria,8 14 notably including MRSA, with which
it shows an MIC of 2.7 mM in vitro.31
We have been engaged in studies of totarol derivatives
with the objectives of identifying the structural features
on which the biological activity depends and of enhan-
cing understanding of their mode of action. In Part I of
this series19 we attempted (a) to elucidate the minimum
structural requirements for the antibacterial activity of
totarol (1) by investigating a variety of analogues and
homologues carrying modi®cations at O-13 and C-14,
and (b) to enhance the bioavailability of totarol deriva-
tives in vivo by glycosylation at O-13. We now report a
comparative investigation of the antibacterial properties
of totarol (1) and of analogues 2±10 and 2a±14a which
bear new substituents at C-12 and also, in the former
set, O-13.
From mode of action studies on two antibiotic diterpe-
noids structurally related to totarol, pisiferic acid (15)15
and carnosic acid (16),16 it was concluded that their activ-
ity is due to their ability to inhibit bacterial peptidoglycan
synthesis. Another possible mode of action of lipophilic
phenols may involve their ability to uncouple oxidative
phosphorylation in mitochondria.17 Totarol itself inhibits
oxygen consumption and respiratory-driven proton
Results and Discussion
Synthesis of compounds 2±14 and 2a±14a
Since the introduction of electron-withdrawing groups
into the aromatic rings of phenols enhances their anti-
septic properties,20 and also since nitration might aug-
ment any ability to uncouple bacterial oxidative
phosphorylation,17 the known nitro-compound 2a21 was
made by the action of nitric acid in acetic acid on totarol
(1); O-methylation of the product gave the known 2.22
*Corresponding author. Tel.: +64-45690040; fax: +64-4569005;
e-mail: g.evans@irl.cri.nz
yFor Part I see reference 19.
0968-0896/00/$ - see front matter # 2000 Elsevier Science Ltd. All rights reserved.
PII: S0968-0896(00)00095-X

1654
G. B. Evans, R. H. Furneaux / Bioorg. Med. Chem. 8 (2000) 1653±1662
The analogous known 12-bromo-compounds 3a23 and
322 were made directly by electrophilic bromination
reactions. Reduction of 2 and 2a led to the known
amino-compounds 422 and 4a,21 the latter product
a€ording the N,O-diacetyl derivative 5 on treatment with
acetic anhydride in pyridine, and the speci®cally N-sub-
stituted acetanilide 5a with acetic anhydride in methanol.
13C NMR spectra characteristic of a carboxylic acid
group (dC 169.0 (s)) on a similar ring (dH 7.85 (1H, s, H-
11); dC 119.7 (d, C-11)). Lithium aluminum hydride
reduction of the formyl derivative 7 a€orded the hydro-
xymethyl analogue 9 which gave resonances typical of a
hydroxymethyl group (dH 4.70 (2H, s); dC 62.1 (t)) on a
pentasubstituted aromatic ring (dH 7.14 (1H, s, H-11); dC
123.6 (d, C-11)). Demethylation of 7 and 8 was carried
out using boron tribromide in dichloromethane27 to
a€ord 7a and 8a in 68% and 75% yield, respectively, and
lithium aluminum hydride reduction of hydroxyaldehyde
7a gave the diol 9a (85%) (Scheme 1).
Compounds 4a and 5a o€ered the opportunity to
investigate the e€ect of intramolecular hydrogen bond-
ing of the C-13 phenolic group on antibacterial activity,
and likewise for this reason catechol 6a22 was made from
the hydroxyamine 4a by treatment with sodium periodate
in acetic acid. Diazotization of the methoxyamine 4 led
to the methoxyphenol 6 (dispermol24). It is noteworthy
that carnosic acid (16), found in rosemary (Rosmarinus
ocinalis, Linn.) leaves25 and which is structurally
similar to catechol 6a, shows antibiotic activity.26
Several C-12 alkyl compounds were made to probe the
possible relationship between antibacterial activity and
lipophilicity, ortho-directed metalation28 being adopted
to permit the introduction of a methyl and an ethyl
group at this position. Thus 13-methoxytotara-8,11,13-
triene (17)29 was treated sequentially with seco-BuLi
precomplexed with tetra-N-methyl(ethylenediamine)
and methyl or ethyl iodide at 78 ꢀC in hexane to give
the 12-methyl and 12-ethyl derivatives 10 and 11 in 46
and 36% yield, respectively. De-O-methylation by use
of boron tribromide in dichloromethane a€orded phe-
nols 10a (88%) and 11a (86%), respectively (Scheme 2).
The e€ects on the antibiotic activity of totarol (1) of
introducing single carbon groups, in each of the di€er-
ent oxidized states at C-12, were also investigated. In
the event that the formyl, carboxy and hydroxymethyl
groups by themselves confer activity upon the basic
totarol skeleton, the properties of the phenols 7a±9a and
their methyl ethers 7±9 were compared. Lithiation of
compound 3 with n-BuLi followed by quenching sepa-
rately with DMF and solid CO2 led to the O-protected
formyl derivative 7 (55%) and carboxy derivative 8
Analogous synthesis of the isopropyl derivatives 13 and
14 was more problematic because a suitable electrophile
could not be found for direct introduction of the C-
alkyl groups. Therefore, an indirect route was adopted
via the 12-ketone 1230 which was demethylated to give
12a,30 and this with methyllithium gave 2-hydroxyprop-
2-yl compound 13a (Scheme 3). Otherwise, 13a was
made from its C-13 methyl ether 13, which was derived
1
(62%). Compound 7 exhibited resonances in the H and
13C NMR spectra characteristic of an aldehydic group (dH
10.18 (1H, s); dC 190.6 (d)) on a penta-substituted aro-
matic ring (dH 7.58 (1H, s, H-11); dC 124.3 (d, C-11)); the
1
carboxy derivative 8 exhibited resonances in the H and

G. B. Evans, R. H. Furneaux / Bioorg. Med. Chem. 8 (2000) 1653±1662
1655
Scheme 1.
Scheme 2.
Scheme 3.
1
from 3 by lithiation followed by reaction with acetone.
The structure of 13a was con®rmed by its NMR reso-
nances, which were characteristic of a 2-hydroxyprop-2-
yl group (dH 1.69 (6H, s); dC 76.3 (s), 30.3, 30.2 (2q))
bonded to a penta-substituted aromatic ring (dH 6.92
(1H, s, H-11); dC 118.9 (d, C-11)). Quantitative dehy-
dration of 13 was achieved by heating in acetic acid, and
the product 13b showed resonances in the H and 13C
NMR spectra characteristic of a propenyl group (dH
5.10 (2H, brs), 2.15 (3H, s); dC 145.5 (s), 114.4 (t), 22.8
(q)) attached to a penta-substituted aromatic ring (dH
6.98 (1H, s, H-11); dC 124.2 (d, C-11)). Hydrogenation
over palladium on carbon gave 12-isopropyl-13-meth-
oxytotara-8,11,13-triene (14, 91%). Demethylation of

1656
G. B. Evans, R. H. Furneaux / Bioorg. Med. Chem. 8 (2000) 1653±1662
14 using boron tribromide in dichloromethane provided
the desired phenol 14a (82%) (Scheme 3).
4. O-methylation of the phenolic hydroxyl group
usually reduced activity appreciably (compounds 2±
4, 6±10 compared with 2a±4a, 6a±10a, respectively);
5. activity against Streptococcus pneumoniae is less
vulnerable to substitution than is the activity
against the other two Gram-positive strains.
Evaluation of in vitro antibacterial activities and
relationships to structural factors
Listed in Table 1 are the minimum inhibitory concentra-
tions of compounds 2±10 and 2a±14a together with those
of the reference compounds totarol (1), 2,4-dinitrophenol
and carnosic acid (16), against the Gram-positive b-lacta-
mase positive and high level gentamycin-resistant Enter-
ococcus faecalis, penicillin-resistant Streptococcus
pneumoniae, methicillin-resistant Staphylococcus aureus
(MRSA), and the Gram-negative multiresistant Kleb-
siella pneumoniae.
In conclusion, it appears that derivatization of the aro-
matic ring of totarol (1) at C-12 has an undesirable
e€ect on its antibacterial activity in vitro. The carboxy
derivative 8a, which retains appreciable activity, may do
so because it has greater water solubility and hence
better in vivo bioavailability.
As with totarol (1) itself, none of the derivatives dis-
played activity against the Gram-negative organism
Klebsiella pneumoniae, up to the highest concentration
tested (32 mg/mL). Furthermore, none of the analogues
retained totarol's potency against all three Gram-posi-
tive bacteria. The key conclusions are:
Experimental
General methods and syntheses
1H and 13C NMR spectra were recorded on a Bruker
AC300 spectrometer and assignments were made with
the assistance of DEPT and COSY experiments. Infra-
red spectra were obtained (KBr) on a Perkin±Elmer
1600 series FTIR spectrometer. Elemental analyses were
conducted with a Carlo Erba EA 1108 elemental analy-
zer. Low and high resolution (HRMS) mass spectra
were obtained on a VG-70-250S double focusing mag-
netic sector mass spectrometer (VG Analytical) equip-
ped with a standard VG-70S EI/CI ion source. Melting
points were determined on a Reichert hot stage micro-
scope and are uncorrected. All reactions were mon-
itored by thin layer chromatography which was carried
out on Merck 60 PF254 silica gel-coated aluminum
sheets. Puri®cation by ¯ash column chromatography
was conducted using Merck Kieselgel S silica gel. Sol-
vents were dried and puri®ed before use according to
standard procedures.31 ``Petrol'' refers to the fraction of
petroleum ether boiling between 60 and 80 ꢀC.
1. the best tolerated C-12 substituents are the hydroxy
group (compound 6a) and formyl, carboxy and
hydroxymethyl groups (compounds 7a±9a);
2. an electron-withdrawing nitro-substituent at C-12
(2a) strongly diminishes activity, which is con-
sistent with other evidence that the antibacterial
properties are not a result of decoupling of oxida-
tive phosphorylation;17
3. lipophilic substituents at C-12 (Me, Et, i-Pr in 10a,
11a and 14a, respectively) also diminish activity,
but they do not eliminate it;
Table 1. In vitro antibacterial activity MICa, mg/mL (values in
brackets are mM)
Compound
Enterococcus Streptococcus Staphylococcus Klebsiella
faecalis
12-Nitrototara-8,11,13-trien-13-ol (2a).21 A solution of
nitric acid (1.5 mL, 23 mmol) in acetic acid (20 mL) was
added to a stirred suspension of totarol (1, 7.0 g, 24
mmol) in acetic acid (70 mL) at 0 ꢀC. The resulting solu-
tion was stirred for 10 min, the reaction was quenched by
the addition of ice water (900 mL), and the mixture ®l-
tered to yield a yellow solid precipitate. Chromatography
on silica gel using petrol:ethyl acꢀetate (19:1) as theꢀeluent
NMR (CDCl3) d 10.98 (1H, s, OH), 7.90 (1H, s, H-11),
3.33 (1H, m, H-15), 2.79 (2H, m, H-7), 1.38, 1.35
(2Â3H, 2d, J=6.9 Hz, H-16, H-17), 1.17 (3H, s, H-20),
0.96 (3H, s, H-18), 0.93 (3H, s, H-19); 13C NMR
(CDCl3) d 151.9, 144.7, 143.2, 135.0, 132.1, 118.4 (Ar),
49.0 (C-5), 41.3 (C-3), 39.5 (C-1), 37.8 (C-10), 33.2 (C-
4), 33.0 (C-18), 29.2 (C-7), 27.9 (C-15), 25.0 (C-20), 21.6
(C-19), 19.5, 19.5 (C-16, C-17), 19.2 (C-6), 18.8 (C-2).
pneumoniae
aureus
pneumoniae
1
2
3
4
5
6
7
8
9
10
2a
3a
4a
5a
6a
7a
8a
9a
10a
11a
12a
13a
2 (7)
>32
2 (7)
32 (93)
>32
2 (7)
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
32 (102)
2 (5)
>32
>32
8 (25)
32 (93)
>32
>32
32 (97)
2 (5)
>32
32 (93)
>32
>32
>32
>32
32 (93)
>32
>32
>32
>32
1
gave 2a (5.2 g, 64%); mp 78±79 C, lit.21 73±74.5 C; H
>32
>32
8 (27)
>32
>32
>32
32 (106)
32 102)
8 (24)
8 (25)
>32
>32
>32
8 (23)
32 (98)
8 (24)
>32
8 (26)
2 (6)
2 (6)
2 (6)
2 (7)
32 (102)
8 (24)
2 (6)
2 (6)
8 (24)
>32
8 (26)
8 (26)
8 (24)
>32
32 (107)
32 (102)
>32
32 (23)
32 (98)
32 (96)
>32
13-Methoxy-12-nitrototara-8,11,13-triene (2).22 Methyl
iodide (0.94 mL 15 mmol) was added to a stirred sus-
pension of sodium hydride (60% in oil, 0.34 g, 8.5
mmol) and 2a (2.5 g, 7.6 mmol) in DMF (20 mL) at 0 ꢀC
under argon. The suspension was allowed to warm to
14a
Carnosic acid (16)
2,4-Dinitrophenol
aMinimum inhibitory concentrations.

G. B. Evans, R. H. Furneaux / Bioorg. Med. Chem. 8 (2000) 1653±1662
1657
room temperature, stirred for 14 h, diluted with water
(400 mL) and ®ltered to yield a solid. Chromatography
on silica gel with petrol:ethyl acetate (19:1) as the eluent
gave 2 (2.4 g, 92%); mp 104±105 ꢀC, lit.22 mp 104±
106 ꢀC; 1H NMR (CDCl3) d 7.61 (1H, s, H-11), 3.76 (3H,
s, OCH3), 3.36 (1H, m, H-15), 2.98 (1H, dd, J=17.9, 5.8
Hz, H-7b), 2.79 (1H, m, H-7a), 2.24 (1H, br d, J=12.3
Hz, H-1b), 1.35, 1.32 (2Â3H, 2d, J=6.9 Hz, H-16, H-
17), 1.19 (3H, s, H-20), 0.96 (3H, s, H-18), 0.93 (3H, s,
H-19); 13C NMR (CDCl3) d 150.4, 146.4, 141.7, 140.9,
140.9, 119.9 (Ar), 61.8 (OCH3), 49.1 (C-5), 41.4 (C-3),
39.3 (C-1), 38.2 (C-10), 33.4 (C-4), 33.1 (C-18), 29.0 (C-
7), 27.9 (C-15), 24.9 (C-20), 21.5 (C-19), 21.1, 21.1 (C-
16, C-17), 19.2 (C-6), 18.8 (C-2).
138.0, 137.3, 110.6 (Ar), 59.5 (OCH3), 49.5 (C-5), 41.5
(C-3), 39.4 (C-1), 37.8 (C-10), 33.2 (C-4), 33.1 (C-18),
27.9 (C-7), 27.3 (C-15), 24.8 (C-20), 21.7, 21.7 (C-16, C-
17), 21.5 (C-19), 19.4 (C-6), 19.4 (C-2).
12-Acetamido-13-acetoxytotara-8,11,13-triene (5). Acetic
anhydride (2.0 mL) was added to a stirred solution of 4a
(150 mg, 0.5 mmol) in pyridine (2.0 mL) at 0 ꢀC. The
resulting solution was allowed to warm to room tem-
perature, stirred for 14 h, diluted with water (100 mL),
extracted with ethyl acetate (3Â50 mL) and the com-
bined organic extracts were washed with brine (3Â25
mL), dried (MgSO4) and concentrated in vacuo. Silica
gel chromatography using petrol:ethyl acetate (1.5:1) as
the eluent gave 5 (136 mg, 71%); mp 214±216 ꢀC; H
1
12-Bromo-13-methoxytotara-8,11,13-triene (3).22 A solu-
tion of bromine (1.0 g, 6.3 mmol) in acetic acid (10 mL)
was added dropwise to a stirred solution of 13-meth-
oxytotara-8,11,13-triene 1719 (1.0 g, 3.5 mmol) in diethyl
ether (10 mL). After stirring for 3 h the volatiles were
removed in vacuo and the residue was recrystallized
from meꢀthanol to yield 3 as a solid (1.12 g, 84%); mp
130±131 C, lit.22 127.5±129.5 ꢀC; 1H NMR spectral
chemical shifts were identical to published data (dÆ0.1
ppm).22
NMR (CDCl3) d 6.84 (1H, s, H-11), 3.28 (1H, sept,
J=7.0 Hz, H-15), 2.94±2.66 (2H, m, H-7), 2.34 (3H, s,
NCOCH3), 2.09 (3H, s, COCH3), 1.22, 1.22 (2Â3H, 2d,
J=7.0 Hz, H-16, H-17), 1.20 (3H, s, H-20), 0.94 (3H, s,
H-18), 0.92 (3H, s, H-19); 13C NMR (CDCl3) d 169.6,
168.4 (COCH3), 148.6, 136.9, 132.0, 127.2, 123.3, 121.2
(Ar), 49.1 (C-5), 41.3 (C-3), 39.2 (C-1), 38.1 (C-10), 33.2
(C-4), 33.0 (C-18), 28.3 (C-7), 27.2 (C-15), 24.8 (C-20),
23.8 (NCOCH3), 21.4 (C-19), 21.1 (OCOCH3), 20.8,
20.7 (C-16, C-17), 19.2 (C-6), 19.0 (C-2); m/z 385 (M+,
5%), 325 (82), 310 (100), 301 (58), 286 (12), 242 (24),
228 (45), 214 (59); HRMS m/z calcd for C24H35NO3
(M+) 385.2616, found 385.2615.
12-Bromototara-8,11,13-trien-13-ol (3a).23 Bromine (0.11
mL, 2.0 mmol) was added dropwise to a stirred, ice
cooled solution of totarol (1, 0.6 g, 2.1 mmol) in CH2Cl2
(10 mL). After stirring for 1 h the volatiles were
removed in vacuo. Chromatography on silica gel using
petrol:ethyl acetate (100:1) as the eluent gave 3a as an
12-Acetamidototara-8,11,13-trien-13-ol (5a). Acetic anhy-
dride (1.0 mL) was added dropwise to a stirred solution
of 4a (114 mg, 0.38 mmol) in methanol (10 mL) at room
temperature under argon. The mixture was stirred for
14 h, the volatiles were removed under reduced pressure
and the resulting residue was redissolved in ethyl ace-
tate, washed with brine (3Â25 mL), dried (MgSO4) and
concentrated in vacuo. Chromatography on silica gel
1
oil (635 mg, 83%). H NMR spectral chemical shifts
were identical to published data (dÆ0.1 ppm).23
12-Aminototara-8,11,13-trien-13-ol (4a).21 Sodium boro-
hydride (50 mg) was added portionwise to a stirred sus-
pension of 2a (90 mg, 0.27 mmol) and palladium on
charcoal (10%, 20 mg) in methanol at room temperature
under argon. The mixture was stirred for 30 min, ®ltered
through Celite, and concentrated in vacuo. Chromato-
graphy on silica gel using CH2Cl2 as the eluent gave 4a
using petrol:ethyl acetate (19:1) as the eluent gave 5a as
1
an oil; nmax 3501 (OH), 1626 (CO), 1545 (NCO) cm
;
1H NMR (CDCl3) d 8.17 (1H, s, OH), 7.57 (1H, s, NH),
6.69 (1H, m, H-11), 3.27 (1H, m, H-15), 2.92, (1H, dd,
J=17.1, 6.1 Hz, H-7b), 2.73 (1H, ddd, J=17.1, 11.3 7.8
Hz, H-7), 2.20 (3H, s, NCOCH3), 1.37, 1.36 (2Â3H, 2d,
J=7.0 Hz, H-16, H-17), 1.11 (3H, s, H-20), 0.94 (3H, s, H-
18), 0.91 (3H, s, H-19); 13C NMR (CDCl3) d 170.4 (NCO),
146.1, 142.9, 135.7, 132.1, 124.3, 116.4 (Ar), 49.5 (C-5),
41.5 (C-3), 39.7 (C-1), 37.6 (C-10), 33.3 (C-4), 33.2 (C-18),
28.5 (C-7), 28.0 (C-15), 25.2 (C-20), 23.5 (NCOCH3), 21.6
(C-19), 20.2, 20.2 (C-16, C-17), 19.4 (C-6), 19.3 (C-2); m/z
343 (M+, 84%), 325 (56), 310 (68), 301 (100), 286 (31),
240 (53), 228 (48), 214 (75), 190 (26), 69 (22), 55 (21), 41
(32); HRMS m/z calcd for C22H33NO2 (M+) 343.2511,
found 343.2514.
(76 mg, 93%); mp 165±166 ꢀC, lit.21 mp 165±166 ꢀC; H
1
NMR (CDCl3) d 6.68 (1H, m, H-11), 3.26 (1H, sept,
J=7.1 Hz, H-15), 2.98 (1H, dd, J=17.9, 5.8 Hz, H-7b),
2.79 (1H, m, H-7a), 1.35, 1.33 (2Â3H, 2d, J=7.1 Hz, H-
16, H-17), 1.16 (3H, s, H-20), 0.94 (3H, s, H-18), 0.90
(3H, s, H-19); 13C NMR (CDCl3) d 144.3, 142.8, 131.4,
130.5, 126.0, 114.3 (Ar), 49.8 (C-5), 41.7 (C-3), 39.7 (C-
1), 37.7 (C-10), 33.3 (C-18), 33.3 (C-4), 28.3 (C-7), 27.3
(C-15), 25.2 (C-20), 21.6, 21.6 (C-16, C-17), 20.6 (C-19),
19.6 (C-6), 19.5 (C-2).
12-Amino-13-methoxytotara-8,11,13-triene (4).22 Sodium
borohydride (1.50 g) was added portionwise to a stirred
suspension of 2 (1.55 g, 4.5 mmol) and palladium on
carbon (10%, 500 mg) in methanol (50 mL) at room
temperature under argon. The mixture was stirred for 1
h, ®ltered through Celite, and concentrated in vacuo.
Chromatography on silica gel with CH2Cl2 as eluent
gave 4 (1.36 g, 96%); mp 137±138 ꢀC, lit.22 136±138 ꢀC;
1H NMR spectral chemical shifts were identical to pub-
lished data (dÆ0.1 ppm)22; 13C NMR (CDCl3) d 146.5,
13-Methoxytotara-8,11,13-trien-12-ol (6).24 To a stirred
solution of 4 (100 mg, 0.3 mmol) in methanol excess
isoamyl nitrite (0.3 mL) and HCl (0.1 mL, concd) were
added. The reaction mixture was stirred at room tem-
perature overnight, diluted with ethyl acetate (100 mL)
and washed with brine (3Â20 mL), dried (MgSO4) and
concentrated in vacuo. Chromatography on silica gel
using petrol:ethyl acetate (9:1) as the eluent gave 6 (79 mg,
79%); mp 163±165 ꢀC, lit.24 166.5±167.5 ꢀC. 1H NMR

1658
G. B. Evans, R. H. Furneaux / Bioorg. Med. Chem. 8 (2000) 1653±1662
spectral chemical shifts were identical to published data
(dÆ0.1 ppm).24
ddd, J=18.3, 10.9, 8.1 Hz, H-7), 1.38, 1.36 (2 3H, 2d,
J=6.9 Hz, H-16, H-17), 1.20 (3H, s, H-20), 0.98 (3H, s,
H-18), 0.95 (3H, s, H-19); 13C NMR (CDCl3) d 196.7
(CHO), 158.3, 143.8, 142.8, 133.0, 128.0, 119.2 (Ar), 49.3
(C-5), 41.6 (C-3), 39.8 (C-1), 37.7 (C-10), 33.4 (C-4), 33.2
(C-18), 29.5 (C-7), 27.5 (C-15), 25.3 (C-20), 21.7 (C-19),
19.8, 19.8 (C-16, C-17), 19.4 (C-6), 19.1 (C-2); m/z 314
(M+, 51%), 299 (100), 256 (45), 229 (50), 217 (35), 203
(65), 169 (36), 128 (33), 64 (87), 41 (24); anal. calcd for
C21H30O2: C, 80.2; H, 9.6; found: C, 80.3; H, 9.8.
Totara-8,11,13-triene-12,13-diol (6a).32 A solution of 4a
(320 mg, 1.06 mmol) in acetic acid (90 mL) was added
over a period of 3 min to a stirred solution of sodium
periodate (3.0 g) in dilute HCl (0.1 M, 210 mL) at room
temperature. The reaction mixture was stirred for 5 min,
extracted with chloroform (3Â50 mL) and the combined
organic extracts were washed with brine (3Â25 mL) and
shaken with potassium iodide (0.9 g) in acetic acid (2.0
mL) for 2 min. The resulting solution was washed with aq
NaHSO4 (2Â50 mL), brine (2Â50 mL), dried (MgSO4)
and concentrated in vacuo. Chromatography on silica gel
using petrol:ethyl acetate (9:1) as the eluent gave 6a as an
oil (260 mg, 81%); 1H NMR spectral chemical shifts were
identical to published data (dÆ0.1 ppm);32 13C NMR
(CDCl3) d 147.2, 146.8, 144.2, 138.2, 125.7, 110.0 (Ar),
49.6 (C-5), 41.6 (C-3), 39.5 (C-1), 38.1 (C-10), 33.4 (C-4),
33.2 (C-18), 28.0 (C-7), 27.5 (C-15), 24.8 (C-20), 21.7 (C-
19), 21.6, 21.6 (C-16, C-17), 19.5 (C-6), 19.5 (C-2).
12-Carboxy-13-methoxytotara-8,11,13-triene (8). A stir-
red solution of compound 3 (216 mg, 0.78 mmol) in
THF (10 mL) was cooled to 78 ꢀC and treated drop-
wise with a solution of n-BuLi (0.68 mL, 1.3 M in hex-
anes, 0.88 mmol) and the resulting mixture stirred for
an additional 10 min at 78 ꢀC. The reaction was then
quenched with solid CO2 (1 g) and allowed to warm to
room temperature and stirred for an additional 30 min
at this temperature. The mixture was diluted with aq
ammonium chloride (10% w/w, 100 mL), extracted with
ethyl acetate (3Â50 mL) and the combined organic lay-
ers were washed with brine (3Â50 mL), dried (MgSO4)
and concentrated in vacuo to give an oily residue.
Chromatography on silica gel using petrol:ethyl acetate
(1:1) as the eluent gave 8 (165 mg, 62%); mp 179 ꢀC;
12-Formyl-13-methoxytotara-8,11,13-triene (7). A stir-
red solution of bromide 3 (216 mg, 0.78 mmol) in THF
(10 mL) was cooled to 78 ꢀC and treated dropwise
with a solution of n-BuLi (0.68 mL, 1.3 M in hexanes,
0.88 mmol) and the resulting mixture stirred for an
additional 10 min at 78 ꢀC. The reaction was then
quenched with DMF (1 mL) and allowed to warm to
room temperature and stirred for an additional 30 min
at this temperature. The mixture was diluted with aqu-
eous ammonium chloride (10% w/w, 100 mL) and
extracted with ethyl acetate (3Â50 mL) and the com-
bined organic layers were washed with brine (3Â50 mL),
dried (MgSO4) and concentrated in vacuo to give an
oily residue. Chromatography on silica gel using petrol:
1
nmax 3198 (OH), 1728 (CO) cm 1; H NMR (CDCl3) d
7.85 (1H, s, H-11), 3.84 (3H, s, OCH3), 3.43 (1H, sept,
J=7.1 Hz, H-15), 3.01 (1H, dd, J=17.8, 5.3 Hz, H-7b),
2.84 (1H, ddd, J=17.8, 10.5, 7.9 Hz, H-7), 1.36, 1.35
(2Â3H, 2d, J=7.1 Hz, H-16, H-17), 1.19 (3H, s, H-20),
0.96 (3H, s, H-18), 0.94 (3H, s, H-19); 13C NMR
(CDCl3) d 156.0, 147.0, 141.6, 138.9, 127.1, 119.7 (Ar),
63.4 (OCH3), 49.2 (C-5), 41.3 (C-3), 39.2 (C-1), 38.0 (C-
10), 33.2 (C-4), 33.0 (C-18), 28.8 (C-7), 27.0 (C-15), 24.6
(C-20), 21.4 (C-19), 21.3, 21.2 (C-16, C-17), 19.1 (C-6),
18.9 (C-2); m/z 344 (M+, 72%), 329 (100), 311 (26), 269
(25), 259 (64), 247 (47), 233 (82), 69 (27), 41 (29); anal.
calcd for C22H32O3: C, 76.7; H, 9.4; found: C, 76.7; H,
9.4.
ethyl acetate (19:1) as the eluent gave 7 as an oil (142
1
mg, 55%); nmax 1697 (CO), 1592 cm
;
1H NMR
(CDCl3) d 10.18 (1H, s, CHO), 7.58 (1H, m, H-11), 3.80
(3H, s, OCH3), 3.30 (1H, br s, H-15), 2.93 (1H, dd,
J=17.9, 5.7 Hz, H-7b), 2.76 (1H, ddd, J=17.9, 10.5, 8.1
Hz, H-7), 1.32, 1.31 (2Â3H, 2d, J=7.0 Hz, H-16, H-17),
1.11 (3H, s, H-20), 0.88 (3H, s, H-18), 0.86 (3H, s, H-
19); 13C NMR (CDCl3) d 190.6 (CHO), 147.1, 139.3,
127.2, 124.3 (Ar), 65.5 (OCH3), 49.4 (C-5), 41.6 (C-3),
39.6 (C-1), 38.3 (C-10), 33.5 (C-4), 33.3 (C-18), 29.4 (C-
7), 27.4 (C-15), 25.0 (C-20), 21.7 (C-19), 21.6, 21.6 (C-
16, C-17), 19.4 (C-6), 19.2 (C-2); HRMS m/z calcd for
C22H32O2 (M+) 328.2402, found 328.2397.
12-Carboxytotara-8,11,13-trien-13-ol (8a). Boron tri-
bromide (1.0 M in CH2Cl2, 1 mL) was added dropwise
to a stirred solution of 8 (60 mg, 0.17 mmol) in CH2Cl2
(5 mL) at room temperature under an inert atmosphere.
The mixture was stirred for 30 min, diluted with CH2Cl2
(50 mL), washed with aq Na2S2O3 (10%, 3Â25 mL), aq
NaHCO3 (3Â25 mL) and brine (3Â25 mL), dried
(MgSO4), and concentrated in vacuo. Chromatography
on silica gel using petrol:ethyl acetate (2.5:1) as the elu-
ent gave 8a (42 mg, 75%) as an oil; nmax 3108 (OH),
12-Formyltotara-8,11,13-trien-13-ol (7a). Boron tribromide
(1.0 M, 5 mL)27 was added dropwise to a stirred solu-
tion of 7 (500 mg, 1.5 mmol) in CH2Cl2 (20 mL) at
room temperature under argon. The mixture was stirred
for 30 min, diluted with CH2Cl2 (100 mL), washed with
aq Na2S2O3 (10% w/w, 3Â25 mL), aq NaHCO3 (3Â25
mL), brine (3Â25 mL), dried (MgSO4), and concentrated
in vacuo. Chromatography on silica gel using petrol:
ethyl acetate (4:1) as the eluent gave 7a (322 mg, 68%);
mp 119 ꢀC; nmax 3684 (OH), 1688 (CO), 1592 (CˆC)
cm 1; 1H NMR (CDCl3) d 11.15 (1H, s, OH), 9.78 (1H,
s, CHO), 7.30 (1H, s, H-11), 3.29 (1H, sept, J=6.9 Hz,
H-15), 3.01 (1H, dd, J=18.3, 5.9 Hz, H-7b), 2.81 (1H,
1
1647 (CO), 1600 (CˆC), 1446 cm 1; H NMR (CDCl3)
d 10.86 (1H, br s, COOH), 7.68 (1H, s, H-11), 3.24 (1H,
m, H-15), 2.85 (2H, m, H-7ab), 1.30 (2Â3H, t, J=7.2
Hz, H-16, H-17), 1.13 (3H, s, H-20), 0.93 (3H, s, H-18),
0.90 (3H, s, H-19); 13C NMR (CDCl3) d 171.7 (COOH),
158.9, 142.1, 141.6, 132.9, 124.5 (Ar), 49.4 (C-5), 41.6
(C-3), 39.6 (C-1), 37.6 (C-10), 33.3 (C-18), 33.2 (C-4),
29.2 (C-7), 27.0 (C-15), 25.1 (C-20), 21.6, 21.1 (C-16, C-
17), 19.8 (C-19), 19.4 (C-6), 19.2 (C-2); m/z 330 (M+,
42%), 315 (100), 312 (40), 297 (22), 245 (21), 219 (35),
201 (31), 69 (64), 41 (88); HRMS m/z calcd for
C21H30O3 (M+) 330.2195, found 330.2193.

G. B. Evans, R. H. Furneaux / Bioorg. Med. Chem. 8 (2000) 1653±1662
1659
12-Hydroxymethyl-13-methoxytotara-8,11,13-triene (9).
Lithium aluminum hydride (100 mg) was added por-
tionwise to a stirred solution of 7 (200 mg, 0.6 mmol) in
THF (10 mL) under argon at room temperature. The
reaction mixture was stirred for 2 h, quenched with
water (0.1 mL), aq NaOH (15%, 0.1 mL), and water
(0.3 mL), ®ltered and concentrated in vacuo. Chroma-
tography on silica gel using petrol:ethyl acetate (4:1) as
the eluent gave 9 as a solid (93 mg, 40%); mp 101 ꢀC;
(50:1) as the eluent gave 10 (127 mg, 46%); mp 105 C;
1
umax 1637 (CˆC) cm 1; H NMR (CDCl3) d 6.94 (1H,
m, H-11), 3.71 (3H, s, OCH3), 3.35 (1H, m, H-15), 2.97-
2.69 (2H, m, H-7), 2.23 (3H, s, CH3), 1.32, 1.31 (2Â3H,
2d, J=7.1 Hz, H-16, H-17), 1.18 (3H, s, H-20), 0.94
(3H, s, H-18), 0.92 (3H, s, H-19); 13C NMR (CDCl3) d
145.8, 137.7, 131.8, 127.9, 125.4 (Ar), 60.6 (OCH3), 49.6
(C-5), 41.5 (C-3), 39.4 (C-1), 37.8 (C-10), 33.4 (C-4),
33.2 (C-18), 28.4 (C-7), 27.3 (C-15), 24.9 (C-20), 21.7,
21.7 (C-16, C-17), 21.6 (C-19), 19.4 (C-6), 19.4 (C-2),
16.9 (CH3); m/z 314 (M+, 100%), 300 (99), 229 (35),
203 (81), 187 (14), 91 (10), 69 (24). Anal. calcd for
C22H34O: C, 84.0; H, 10.9; found: C, 83.8; H, 11.0.
n
max 3408 (OH), 1600 (CˆC) cm 1; 1H NMR (CDCl3) d
7.14 (1H, m, H-11), 4.70 (2H, s, CH2OH), 3.77 (3H, s,
OCH3), 3.40 (1H, br s, H-15), 2.96 (1H, dd, J=17.2 Hz,
5.4 Hz, H-7b), 2.78, (1H, ddd, J=17.2, 10.6, 7.5 Hz, H-
7), 1.33, 1.32 (2Â3H, 2d, J=7.2 Hz, H-16, H-17), 1.19
(3H, s, H-20), 0.94 (3H, s, H-18), 0.92 (3H, s, H-19); 13C
NMR (CDCl3) d 154.9, 146.6, 138.3, 134.6, 131.2, 123.6
(Ar), 62.1 (OCH3), 62.1 (CH2OH), 49.6 (C-5), 41.6 (C-
3), 39.6 (C-1), 38.1 (C-10), 33.4 (C-4), 33.2 (C-18), 28.6
(C-7), 27.2 (C-15), 25.0 (C-20), 21.7 (C-19), 21.6, 21.6
(C-16, C-17), 19.5 (C-6), 19.4 (C-2); m/z 330 (M+,
100%), 315 (84), 279 (15), 219 (71), 167 (24), 149 (70),
84 (20), 69 (31); HRMS m/z calcd for C22H34O2 (M+)
330.2559, found 330.2553.
12-Methyltotara-8,11,13-trien-13-ol (10a). Boron tri-
bromide (1.0 M in CH2Cl2, 2 mL) was added dropwise
to a stirred solution of 10 (51 mg, 0.16 mmol) in CH2Cl2 (10
mL) at room temperature under argon. The mixture was
stirred for 30 min, diluted with CH2Cl2 (50 mL), washed
with aq Na2S2O3 (10% w/w, 3Â25 mL), aq NaHCO3
(3Â25 mL) and brine (3Â25 mL), dried (MgSO4), and
concentrated in vacuo. Chromatography on silica gel
using petrol:ethyl acetate (19.1) as the eluent gave 10a
(42 mg, 88%); mp 115 C; nmax 3500 (OH), 1648 (CˆC)
1
12-(Hydroxymethyl)totara-8,11,13-trien-13-ol (9a).
Lithium aluminum hydride (100 mg) was added
potionwise to a stirred solution of 7a (200 mg, 0.64
mmol) in THF (10 mL) under argon at room tempera-
ture. The mixture was stirred for 2 h and the reaction
was quenched with water (0.1 mL), aq NaOH (15%, 0.1
mL), and water (0.3 mL). After ®ltration, the ®ltrate
was concentrated in vacuo. Chromatography on silica
gel using petrol:ethyl acetate (9:1) as the eluent gave 9a
(170 mg, 85%); mp 101 ꢀC; nmax 3383 (OH), 1640
(CˆC), 1592 cm 1; 1H NMR (CDCl3) d 7.19 (1H, s, H-
11), 4.78 (2H, d, J=4.6 Hz, CH2OH), 3.30 (1H, sept,
J=7.0 Hz, H-15), 2.95 (1H, dd, J=17.1, 6.4 Hz, H-7b),
2.76, (1H, ddd, J=17.1, 11.2, 7.8 Hz, H-7), 1.39, 1.37 (2
3H, 2d, J=7.0 Hz, H-16, H-17), 1.19 (3H, s, H-20), 0.97
(3H, s, H-18), 0.94 (3H, s, H-19); 13C NMR (CDCl3) d
152.8, 141.8, 133.9, 132.1, 122.4, 121.6 (Ar), 65.4
(CH2OH), 49.5 (C-5), 41.4 (C-3), 39.5 (C-1), 37.4 (C-
10), 33.1 (C-4), 33.1 (C-18), 28.5 (C-7), 27.3 (C-15), 25.0
(C-20), 21.4 (C-19), 20.1, 20.1 (C-16, C-17), 19.3 (C-6),
19.2 (C-2); m/z 316 (M+, 80%), 298 (100), 283 (51), 255
(65), 229 (40), 215 (44), 202 (51), 187 (38), 69 (32), anal.
calcd for C21H32O2: C, 79.6; H, 10.2; found: C, 79.2; H,
10.2.
cm 1; H NMR (CDCl3) d 6.90 (1H, m, H-11), 4.45
(1H, s, OH), 3.35 (1H, m, H-15), 2.90, (1H, dd, J=16.8,
6.5 Hz, H-7b), 2.72, (1H, ddd, J=16.8, 11.2, 7.8 Hz, H-
7), 2.22 (3H, s, CH3), 1.35, 1.33 (2Â3H, 2d, J=7.1 Hz,
H-16, H-17), 1.18 (3H, s, H-20), 0.94 (3H, s, H-18), 0.91
(3H, s, H-19); 13C NMR (CDCl3) d 150.5, 142.6, 131.5,
130.5, 124.4, 120.7 (Ar), 49.6 (C-5), 41.5 (C-3), 39.6 (C-
1), 37.5 (C-10), 33.2 (C-4), 33.2 (C-18), 28.4 (C-7), 27.0
(C-15), 25.1 (C-20), 21.5 (C-19), 20.4, 20.4 (C-16, C-17),
19.4 (C-6), 19.4 (C-2), 15.8 (CH3); m/z 300 (M+, 63%),
285 (100), 215 (31), 189 (51), 69 (18); HRMS m/z calcd
for C21H32O (M+) 300.2453, found 300.2451.
12-Ethyl-13-methoxytotara-8,11,13-triene (11). s-Butyl-
lithium (6.5 mL, 1.3 M in hexanes, 8.5 mmol) was added
dropwise to a stirred solution of Me2NCH2CH2NMe2
(1.3 mL, 8.6 mmol) in hexane (5 mL) at room tempera-
ture under argon. The resulting solution was stirred for
30 min and added via a cannula to a solution of 17 (265
mg, 0.86 mmol) in hexane (5 mL) and this was stirred
for 14 h. The reaction was quenched by the addition of
ethyl iodide (1.0 mL) and the solution diluted with
aqueous ammonium chloride (10% w/w, 50 mL),
extracted with ethyl acetate (3Â50 mL) and the com-
bined organic extracts washed with brine (3Â25 mL),
dried (MgSO4), and concentrated in vacuo. Chromato-
graphy on silica gel using petrol:ethyl acetate (50:1) as
the eluent gave 11 (105 mg, 36%); mp 91 C; nmax 1638
13-Methoxy-12-methyltotara-8,11,13-triene (10). sec-
Butyllithium (6.5 mL, 1.3 M in hexanes, 8.5 mmol) was
added dropwise to a stirred solution of Me2NCH2CH2
NMe2 (1.3 mL, 8.6 mmol) in hexane (5 mL) at room
temperature under argon. The resulting solution was
stirred for 30 min and added via a cannula to a solution
of 1719 (265 mg, 0.86 mmol) in hexane (5 mL) and this
was stirred for 14 h. The reaction was quenched by the
addition of methyl iodide (1.0 mL), and the solution
was diluted with aqueous ammonium chloride (10% w/
w, 50 mL), extracted with ethyl acetate (3Â50 mL) and
the combined organic extracts were washed with brine
(3Â25 mL), dried (MgSO4), and concentrated in vacuo.
Chromatography on silica gel using petrol:ethyl acetate
1
(CˆC) cm 1; H NMR (CDCl3) d 7.00 (1H, m, H-11),
3.72 (3H, s, OCH3), 3.39 (1H, br s, H-15), 2.94 (1H, dd,
J=16.7, 5.9 Hz, H-7b), 2.86 (1H, m, H-7a), 2.63 (2H, q,
J=7.6 Hz, CH2CH3), 1.33, 1.31 (2Â3H, 2d, J=7.2 Hz,
H-16, H-17), 1.23 (3H, t, J=7.6 Hz, CH2CH3) 1.19 (3H,
s, H-20), 0.94 (3H, s, H-18), 0.92 (3H, s, H-19); 13C
NMR (CDCl3) d 146.0, 137.7, 133.9, 131.0, 123.5 (Ar),
61.5 (OCH3), 49.6 (C-5), 41.5 (C-3), 39.5 (C-1), 38.0 (C-
10), 33.4 (C-4), 33.2 (C-18), 28.4 (C-7), 27.2 (C-15), 24.9
(C-20), 22.9 (CH2CH3), 21.7, 21.7 (C-16, C-17), 21.6 (C-
19), 19.4 (C-6), 19.4 (C-2), 14.8 (CH2CH3); m/z 328

1660
G. B. Evans, R. H. Furneaux / Bioorg. Med. Chem. 8 (2000) 1653±1662
(M+, 100%), 313 (95), 271 (14), 243 (30), 217 (62), 69
(17); HRMS m/z calcd for C23H36O (M+) 328.2766,
found 328.2765.
reaction mixture was then quenched with acetone (1.0
mL) and allowed to warm to room temperature and
stirred for an additional 30 min. The mixture was dilu-
ted with aqueous ammonium chloride (10% w/w, 100
mL), extracted with ethyl acetate (3Â50 mL) and the
combined organic layers were washed with brine (3Â50
mL), dried (MgSO4) and concentrated in vacuo to give
an oily residue. Chromatography on silica gel using
petrol:ethyl acetate (19:1) as the eluent gave 13 (190 mg,
12-Ethyltotara-8,11,13-trien-13-ol (11a). Boron tri-
bromide (1.0 M in CH2Cl2, 1 mL) was added dropwise
to a stirred solution of 11 (28 mg, 0.09 mmol) in CH2Cl2
(5 mL) at room temperature under argon. The mixture was
stirred for 30 min, diluted with CH2Cl2 (50 mL), washed
with aq Na2S2O3 (10% w/w, 3Â25 mL), aq NaHCO3
(3Â25 mL), and brine (3Â25 mL), dried (MgSO4), and
concentrated in vacuo. Chromatography on silica gel
41%); mp 149±150 ꢀC; nmax 3439 (OH), 1215 cm 1; H
1
NMR (CDCl3) d 7.10 (1H, s, H-11), 4.96 (1H, s, OH),
3.84 (3H, s, OCH3), 3.61 (1H, sept, J=7.2 Hz, H-15),
3.04 (1H, dd, J=17.0, 6.4 Hz, H-7b), 2.86, (1H, ddd,
17.0, 10.4, 7.6 Hz, H-7), 1.41 (6H, s, (CH3)2), 1.40, 1.38
(2 3H, 2d, J=7.0 Hz, H-16, H-17), 1.19 (3H, s, H-20),
0.96 (3H, s, H-18), 0.95 (3H, s, H-19); 13C NMR
(CDCl3) d 153.9, 146.6, 138.3, 137.3, 135.1, 120.9 (Ar),
73.5 (COH), 63.4 (OCH3), 49.7 (C-5), 41.4 (C-3), 39.4
(C-1), 38.4 (C-10), 33.4 (C-4), 33.2 (C-18), 32.2, 31.9
((CH3)2C), 28.0 (C-7), 26.3 (C-15), 24.8 (C-20), 21.6,
21.5 (C-16,C-17), 21.3 (C-19), 19.4 (C-6), 19.3 (C-2); m/z
358 (M+, 1%), 340 (63), 325 (100), 255 (26), 229 (56), 83
(88), 63 (30). Anal. calcd for C24H38O2: C, 80.4; H, 10.7;
found: C, 80.7; H, 10.9.
using petrol:ethyl acetate (19:1) as the eluent gave 11a as
1
an oil (23 mg, 86%); nmax 3439 (OH), 1639 (CˆC) cm
;
1H NMR (CDCl3) d 6.93 (1H, m, H-11), 4.54 (1H, s,
OH), 3.32 (1H, br s, H-15), 2.91 (1H, dd, J=16.8, 6.6
Hz, H-7b), 2.72 (1H, m, H-7a), 2.55 (2H, q, J=7.6 Hz,
CH2CH3), 1.35, 1.35 (2Â3H, 2d, J=7.0 Hz, H-16, H-
17), 1.24 (3H, t, J=7.6 Hz, CH2CH3), 1.19 (3H, s, H-
20), 0.94 (3H, s, H-18), 0.91 (3H, s, H-19); 13C NMR
(CDCl3) d 150.3, 142.9, 131.0, 130.7, 127.1, 122.8 (Ar),
49.9 (C-5), 41.8 (C-3), 39.8 (C-1), 37.9 (C-10), 33.4 (C-
4), 33.4 (C-18), 28.8 (C-7), 27.3 (C-15), 25.3 (C-20), 23.2
(CH2CH3), 21.7 (C-19), 20.7, 20.7 (C-16, C-17), 19.7 (C-
6), 19.6 (C-2), 14.2 (CH2CH3); m/z 314 (M+, 26%), 299
(28), 286 (42), 217 (41), 191 (100), 177 (40), 163 (33), 109
(44), 81 (51), 69 (60). Anal. calcd for C22H34O: C, 84.0;
H, 10.9; found: C, 83.9; H 11.0.
12-(2-Hydroxyprop-2-yl)totara-8,11,13-trien-13-ol (13a).
Methyllithium (1.2 M in diethyl ether, 1.5 mL) was
added dropwise to a stirred solution of ketone 12a (120
mg, 0.37 mmol) in THF (5 mL) at room temperature.
After 1 h the reaction mixture was diluted with aqueous
ammonium chloride (50 mL, 10%), extracted with ethyl
acetate (3Â20 mL) and the combined organic layers
were washed with brine (3Â20 mL), dried (MgSO4) and
concentrated in vacuo to yield an oily residue. Chro-
matography on silica gel using petrol:ethyl acetate
(19:1) as the eluent gave 13a as an oil (80 mg, 63%);
12-Acetyl-13-methoxytotara-8,11,13-triene (12).30 Tita-
nium tetrachloride (7.5 mL, 1.0 M in CH2Cl2) was added
dropwise to an ice-cooled, stirred solution of compound
17 (1.0 g, 3.4 mmol) and acetyl chloride (0.54 mL) in
CH2Cl2 (20 mL) and the resulting mixture was stirred
for 30 min and the reaction quenched by the addition of
aqueous ammonium chloride (10% w/w, 100 mL). The
solution was further diluted with CH2Cl2 (50 mL),
washed with aq NaHCO3 (3Â20 mL), brine (3Â20 mL),
dried (MgSO4) and concentrated in vacuo. The resulting
residue was puri®ed on silica gel to give 12 (780 mg,
67%); mp 154±155 ꢀC, lit.30 mp 159 ꢀC. 1H and 13C
NMR spectral chemical shifts were identical to pub-
lished data (dÆ0.1 ppm).30
12-Acetyltotara-8,11,13-trien-13-ol (12a).30 Boron tri-
bromide (1.0 M in CH2Cl2, 1 mL) was added dropwise
to a stirred solution of 12 (50 mg, 0.15 mmol) in CH2Cl2
(5 mL) at room temperature under argon. The solution
was stirred for 30 min, diluted with CH2Cl2 (50 mL),
washed with aq Na2S2O3 (10%, 3Â25 mL), aq NaHCO3
(3Â25 mL), and brine (3Â25 mL), dried (MgSO4), and
concentrated in vacuo. Chromatography on silica gel
using petrol:ethyl acetate (100:1) as the eluent gave 12a
(41 mg, 83%), mp 143±145 ꢀC, lit.30 144±146 ꢀC. 1H and
13C NMR spectral chemical shifts were identical to
published data (dÆ0.1 ppm).30
n
max 3334 (OH), 1633 (CˆC) cm 1; 1H NMR (CDCl3) d
8.75 (1H, s, OH), 6.92 (1H, s, H-11), 3.29 (1H, m, H-
15), 2.95, (1H, dd, J=17.0, 5.9 Hz, H-7b), 2.76 (1H,
ddd, J=17.0, 11.3, 7.8 Hz, H-7), 1.69 (6H, s, (CH3)2),
1.39, 1.38 (2Â3H, 2d, J=7.1 Hz, H-16, H-17), 1.21 (3H,
s, H-20), 0.97 (3H, s, H-18), 0.94 (3H, s, H-19); 13C
NMR (CDCl3) d 152.0, 141.1, 133.0, 132.5, 128.5, 118.9
(Ar), 76.3 (COH), 49.7 (C-5), 41.6 (C-3), 39.7 (C-1), 37.7
(C-10), 33.3 (C-4), 33.2 (C-18), 30.3, 30.2 ((CH3)2C),
28.6 (C-7), 27.7 (C-15), 25.3 (C-20), 21.6 (C-19), 20.2,
20.2 (C-16, C-17), 19.5 (C-6), 19.4 (C-2); m/z 344 (M+,
2%), 326 (57), 241 (35), 215 (34), 84 (17), 69 (21), 55
(13), 49 (22); HRMS m/z calcd for C23H36O2 (M+)
344.2715, found 344.2718.
13-Methoxy-12-(prop-2-enyl)totara-8,11,13-triene (13b).
A solution of 13 (150 mg, 0.4 mmol) in acetic acid (10
mL) was heated under re¯ux for 5 min and then con-
centrated under reduced pressure to give 13b (136 mg,
1
100%); mp 148 ꢀC; nmax 1633 (CˆC), 1233, 1018 cm
;
1H NMR (CDCl3) d 6.98 (1H, s, H-11), 5.10 (2H, br s,
CˆCH2), 3.70 (3H, s, OCH3), 3.33 (1H, br s, H-15),
2.95, 2.78 (1H, dd, J=16.9, 6.2 Hz, H-7b), 2.78 (1H,
ddd, J=16.9, 11.0, 7.9 Hz, H-7), 2.15 (3H, s, CH3CˆC),
1.35, 1.34 (2Â3H, 2d, J=7.1 Hz, H-16, H-17), 1.23 (3H,
s, H-20), 0.97 (3H, s, H-18), 0.94 (3H, s, H-19); 13C
NMR (CDCl3) d 145.7 (Ar), 145.5 (CˆCH2), 137.9,
12-(2-Hydroxyprop-2-yl)-13-methoxytotara-8,11,13-triene
(13).30 A stirred solution of bromo-compound 3 (500
mg, 1.3 mmol) in THF (20 mL) was cooled to 78 ꢀC
and treated dropwise with a solution of n-butyllithium
(1.3 mL, 1.3 M in hexanes, 1.6 mmol) and the resulting
mixture stirred for an additional 10 min at 78 ꢀC. The

G. B. Evans, R. H. Furneaux / Bioorg. Med. Chem. 8 (2000) 1653±1662
1661
1
137.3, 134.3, 124.2 (Ar), 114.4 (CH2=C), 60.3 (OCH3),
49.5 (C-5), 41.6 (C-3), 39.5 (C-1), 37.9 (C-10), 33.3 (C-
4), 33.2 (C-18), 28.7 (C-7), 27.6 (C-15), 25.0 (C-20), 22.8
(CH3CˆC), 21.6, 21.6 (C-16, C-17), 21.6 (C-19), 19.5
(C-6), 19.4 (C-2); m/z 340 (M+, 77%), 325 (100), 283
(18), 255 (33), 229 (73), 69 (15). Anal. calcd for
C24H36O: C, 84.7; H, 10.7; found: C, 84.7; H, 10.9.
broth and an inoculum of 104 CFU mL per spot was
applied to agar plates containing graded concentrations
of each compound. After incubation at 37 ꢀC for 18±20
h, the MIC was de®ned as the minimum drug con-
centration which inhibited growth of bacteria.
Acknowledgements
12-Isopropyl-13-methoxytotara-8,11,13-triene (14). A
mixture of 13b (100 mg, 0.29 mmol), Pd/C (10%, 50
mg), isopropanol (5 mL) and hexane (5 mL) was stirred
at room temperature under an atmosphere of hydrogen
for 3 h, after which the solids were removed by ®ltration
through Celite and the volatiles were removed in vacuo.
The residue was chromatographed on silica gel using
petrol:ethyl acetate (100:1) and the eluent gave 14 (92
We wish to thank the Foundation for Research, Science
and Technology for a New Zealand Research, Science and
Technology postdoctoral fellowship awarded to GBE.
Dr M. B. Gravestock, Zeneca Pharmaceuticals PCL,
kindly provided the microbiological test data, and Dr
Herbert Wong measured the NMR spectra. Prof. Robin
Ferrier assisted with the preparation of the manuscript.
mg, 91%); mp 145 ꢀC; H NMR (CDCl3) d 7.08 (1H, s,
1
H-11), 3.74 (3H, s, OCH3), 3.46 (1H, br s, H-15), 3.30
(1H, sept, J=6.9 Hz, (CH3)2CH), 2.96 (1H, dd, J=16.1,
5.7 Hz, H-7b), 2.80 (1H, ddd, J=16.1, 10.9, 7.8 Hz, H-
7), 1.37, 1.36 (2Â3H, 2d, J=6.9 Hz, H-16, H-17), 1.25,
1.23 (2 3H, 2d, J=6.9 Hz, CH(CH3)2), 1.22 (3H, s, H-
20), 0.97 (3H, s, H-18), 0.95 (3H, s, H-19); 13C NMR
(CDCl3) d 154.0, 138.9, 137.6, 137.6, 120.8 (Ar), 62.4
(OCH3), 49.7 (C-5), 41.6 (C-3), 39.5 (C-1), 38.3 (C-10),
33.4 (C-4), 33.2 (C-18), 28.4 (C-7), 27.3 (C-15), 26.4
(CH(CH3)2), 25.0 (C-20), 21.8 (C-19), 21.6, 21.6 (C-16,
C-17), 19.5 (C-6), 19.5 (C-2); m/z 342 (M+, 64%), 327
(100), 257 (39), 231 (82), 69 (18); anal. calcd for
C24H38O: C, 84.2; H, 11.2; found: C, 84.0; H, 11.4.
References and Notes
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12-Isopropyltotara-8,11,13-trien-13-ol (14a). Boron tri-
bromide (1.0 M in CH2Cl2, 1 mL) was added dropwise
to a stirred solution of 14 (60 mg, 0.2 mmol) in CH2Cl2
(5 mL) at room temperature under argon. The mixture
was stirred for 30 min, diluted with CH2Cl2 (100 mL),
washed with aq Na2S2O3 (10% w/w, 3Â25 mL), aq
NaHCO3 (3Â25 mL), and brine (3Â25 mL), dried
(MgSO4), and concentrated in vacuo to give a solid.
Recrystallization from ethyl acetate gave 14a (47 mg,
82%), mp 114 ꢀC; nmax 3431 (OH), 1630 (CˆC), 1215
cm 1; 1H NMR (CDCl3) d 7.02 (1H, s, H-11), 4.65 (1H,
s, OH), 3.36 (1H, sept, J=7.2 Hz, H-15), 3.08 (1H, sept,
J=6.9 Hz, (CH3)2CH), 2.91, (1H, dd, J=16.8, 6.2 Hz,
H-7b), 2.75 (1H, ddd, J=16.8, 11.3, 7.7 Hz, H-7), 1.39,
1.38 (2Â3H, 2d, J=7.2 Hz, H-16, H-17), 1.28 (2Â3H,
2d, J=6.9 Hz, CH(CH3)2), 1.23 (3H, s, H-20), 0.97 (3H,
s, H-18), 0.94 (3H, s, H-19); 13C NMR (CDCl3) d 149.5,
142.6, 131.5, 131.1, 130.4, 119.6 (Ar), 49.7 (C-5), 41.6
(C-3), 39.6 (C-1), 37.9 (C-10), 33.2 (C-4), 33.2 (C-18),
28.6 (C-7), 27.0, 27.0 (C-15,CH(CH3)2), 25.2 (C-20),
23.0, 22.7 (CH(CH3)2), 21.5 (C-19), 20.6, 20.6 (C-16, C-
17), 19.5 (C-6), 19.4 (C-2); m/z 328 (M+, 45%), 313
(100), 256 (39), 243 (82), 217 (18), 192 (32), 160 (60), 128
(52), 96 (20), 64 (85), 43 (26); HRMS m/z calcd for
C23H36O (M+) 328.2766, found 328.2776.
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Microbiological testing
Minimum inhibitory concentrations (MIC) were deter-
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26. Dobrynin, V. N.; Kolosov, M. N.; Chernov, B. K.; Der-
bentseva, N. A Khim. Prir. Soedin 1976, 686.
1
diluted to approximately 108 CFU mL
with fresh

1662
G. B. Evans, R. H. Furneaux / Bioorg. Med. Chem. 8 (2000) 1653±1662
27. Demuynck, M.; De Clercq, P.; Vandewalle, M. J. Org.
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