Asymmetric Synthesis of Oxygenated Monoterpenoids of Importance for Bark Beetle Ecology

Article abstract of DOI:10.1021/acs.jnatprod.0c00669

Herein we report the asymmetric syntheses of a number of oxygenated terpenoids that are of importance in the chemical ecology of bark beetles. These are pinocamphones, isopinocamphones, pinocarvones, and 4-thujanols (= sabinene hydrates). The camphones were synthesized from isopinocampheol, the pinocarvones from β-pinene, and the thujanols from sabinene. The NMR spectroscopic data, specific rotations, and elution orders of their stereoisomers on a chiral GC-phase (β-cyclodextrin) are also reported. This enables facile synthesis of pure compounds for biological activity studies and identification of stereoisomers in mixed natural samples.

Full text of DOI:10.1021/acs.jnatprod.0c00669

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Article
Asymmetric Synthesis of Oxygenated Monoterpenoids of
Importance for Bark Beetle Ecology
Suresh Ganji, Fredric G. Svensson, and C. Rikard Unelius*
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sı Supporting Information
ABSTRACT: Herein we report the asymmetric syntheses of a
number of oxygenated terpenoids that are of importance in the
chemical ecology of bark beetles. These are pinocamphones,
isopinocamphones, pinocarvones, and 4-thujanols (= sabinene
hydrates). The camphones were synthesized from isopinocam-
pheol, the pinocarvones from β-pinene, and the thujanols from
sabinene. The NMR spectroscopic data, specific rotations, and
elution orders of their stereoisomers on a chiral GC-phase (β-
cyclodextrin) are also reported. This enables facile synthesis of
pure compounds for biological activity studies and identification of stereoisomers in mixed natural samples.
he colonization of trees by bark beetles is generally
influenced by an intricate release of chemical signals in
T
strict chronological order. The signals recruit conspecifics to a
suitable host tree, and later in the colonization, other
compounds are produced to convey to conspecifics that this
1
tree is becoming overexploited. Thereby competition for food
and larvae is avoided. These attractant chemical signals can
originate from metabolized monoterpenoids, like for the
noxious larger spruce bark beetle, Ips typographus, where cis-
verbenol is converted to verbenone, but the pheromones can
2
also be synthesized de novo. In recent work with semi-
ochemicals for tree-killing bark beetles we have encountered a
number of oxygenated monoterpenoids that are physiologically
active (antiattractive, i.e., reduce the effect of aggregation
3
,4
pheromone) in these beetles (Figure 1).
We recently published that the production of oxygenated
monoterpenoids is related to tree stress and that it might be a
3
signal for a suitable or nonsuitable host for bark beetles.
Figure 1. Absolute configuration of the synthesized oxygenated
terpenoids.
Investigations by gas chromatographic electroantennographic
3
detection (GC-EAD) of monoterpenones by us and Kalinova
5
et al. revealed that both isopinocamphones and pinocam-
8
inseparable mixture of oxygenated monoterpenoids. These
phones elicit antennal responses in I. typographus. There are
relatively few syntheses of pinocamphone and isopinocam-
phone reported. In one report in Chinese, Wang et al. reacted
α-pinene with borane to obtain diisopinocampheylborane,
which was oxidized to isopinocampheol by sodium perborate
syntheses do not yield pure stereoisomers or are tedious and
have low yields. For short and convenient synthesis without
heating, we developed a simple method using pure
isopinocampheol stereoisomers that are commercially avail-
able.
6
to afford isopinocampheol. The isopinocampheol was finally
oxidized by H O with vanadium phosphorus oxide as catalyst
2
2
́
̌
Received: June 16, 2020
to yield isopinocamphone. Pitinova-Stekrova and co-workers
́
́
utilized different titanosilicate catalysts to convert α-pinene to
7
obtain campholenic aldehyde. Some of these catalysts
produced pinocamphone as side-product. In another report,
thermolysis of α-pinene epoxide in supercritical anhydrous
isopropanol afforded up to ∼25% pinocamphone, but in an
©
XXXX American Chemical Society
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A
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Pinocarvone is reported as a pheromone for the southern
pine beetle (Dendroctonus frontalis) and has been found in the
Scheme 1. Synthesis of the Four Stereoisomers of
Pinocamphone
9
hindguts of male white pine cone beetles, Conophthorus
1
0
coniperda. In the GC-EAD analysis of I. typographus,
pinocarvone gave strong antennal responses, indicating bio-
3
logical activity. Pinocarvone has been isolated from
Eucalyptus oil and has been produced by oxidation of β-
pinene with SeO . In this reaction, myrtenal is formed by a
2
rearrangement, and this byproduct was either overlooked or
11,12
lost during spinning band distillation.
In previous reports, we reported that one of the two trans-4-
thujanol stereoisomers showed strong GC-EAD activity for
3
bark beetles and that this (+)-trans-thujanol is a field-active
13
Specific Rotation of Isopinocamphones and Pinocam-
phones. The sign of the specific rotation changed when going
from isopinocampheols (1 and 2) to isopinocamphones (3 and
semiochemical for the bark beetle, I. typographus. Blazyte-
Cereskiene et al. reported that young spruce trees release more
-thujanol than older trees and that 4-thujanol plays an
important role in both host defense and tree choice by bark
̇
̇
4
4
), but not during epimerization from isopinocamphones to
1
4
pinocamphones (5 and 6). The specific rotations are listed in
beetles. Thus, 4-thujanol is seemingly an indicator of healthy
strong trees which should be avoided and could be of interest
in forest protection. Several publications on the synthesis of 4-
thujanol have been published, including the biotransformation
of α-pinene to cis-4-thujanol using the microorganism
Table 1.
Table 1. Enantioselective GC-FID and Specific Rotations of
Isopinocampheols, Isopinocamphones, and Pinocamphones
1
5
Fusarium saloni, Baeckstro
̈
m’s synthesis of trans-4-thujanol
s.
specific
1
6
from 3-thujol, Galopin’s synthesis of the trans isomers from
no
compound
(1S,2S,3S,5R)-
t
R
rotation
class
17
18,19
methyl vinyl ketone, Cheng’s syntheses of cis-thujanol,
1
2
3
+34 (c 1.0, isopinocampheols
DCM)
20
(
+)-isopinocampheol
(1R,2R,3R,5S)-
−)-isopinocampheol
and Fanta’s synthesis of trans-thujanol. However, all these
synthetic procedures involve many steps and/or expensive
starting materials, and there are no effective synthetic routes
for all possible stereoisomers. In order to develop a short
synthesis of all stereoisomers of 4-thujanol for the investigation
of GC-EAD activity, herein these were synthesized from
commercial sabinene. The absolute configuration of each
stereoisomer was unambiguously assigned by the deduction
from the original sabinene in combination with NMR
spectroscopy and chiral-phase GC-MS analysis.
−36 (c 1.0,
(
DCM)
(1S,2S,5R)-
11.69 −11.4 (c
isopinocamphones
(IPC)
(−)-isopinocamphone
1.0,
EtOH)
4
5
6
(1R,2R,5S)-
11.61 +11.2 (c
1.0,
(
+)-isopinocamphone
EtOH)
(1S,2R,5R)-
(−)-pinocamphone
11.37 −20.2 (c
pinocamphones
(PC)
1.0,
EtOH)
(1R,2S,5S)-
11.53 +22.7 (c
1.0,
These compounds are obviously important in bark beetle
ecology, and some of them act as indicators of tree health; they
are also of interest for managing bark beetle populations. It is
well known that the stereochemistry of pheromones and other
(
+)-pinocamphone
EtOH)
GC Elution Order of Isopinocamphones and Pinocam-
phones. In the analysis by GC equipped with HP-5MS or DB-
21−24
semiochemicals is often extremely important.
Thus, there
is a need to develop analytical procedures to be able to use
enantioselective gas chromatography to differentiate between
the stereoisomers of these semiochemicals in a biological
sample, as well as their facile synthesis. We herein report the
syntheses, specific rotations, and elution orders on a chiral GC
phase (β-cyclodextrin) of pinocamphones, isopinocamphones,
pinocarvones, and 4-thujanols (sabinene hydrate).
5
MS columns, the pinocamphones eluted before the
isopinocamphones. In the GC analysis using a chiral-phase
column (Cyclosil B), (−)-pinocamphone (5) eluted before
(
(
+)-pinocamphone (6) and (+)-isopinocamphone (4) before
−)-isopinocamphone (3) (Table 1 and Figure 2).
Synthesis of Pinocarvone Stereoisomers. There are
only a few syntheses of pinocarvone published, and usually
RESULTS AND DISCUSSION
■
Synthesis of Isopinocamphones and Pinocam-
phones. Scheme 1 summarizes the syntheses of the four
stereoisomers of pinocamphone. The pure enantiomers of
isopinocampheol (1 and 2) were separately oxidized with
pyridinium dichromate (PDC) to obtain both enantiomers of
isopinocamphone in >98% optical purity. The oxidation was
improved by adding silica gel to the reaction mixture, which
prevents the formation of lumps and tar and in turn leads to
higher yields and easier filtration at workup.
To produce pinocamphones, NaOEt was used to epimerize
C-2 of isopinocamphone. The thermodynamic equilibrium
seems to be 4:1 in favor of pinocamphone, and thus, 20% of
isopinocamphone had to be removed by chromatography to
obtain pure pinocamphone (Scheme 1).
Figure 2. Mix of pinocamphones and isopinocamphones separated by
enantioselective GC. The temperature program was isothermal 110
°C on a Cyclosil B column.
B
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pinocarvone has been synthesized by oxidation of α-pinene or
(12). Chirality of the sabina ketone was confirmed by use of
specific rotation and a GC column (β-cyclodextrin phase).
25,26
29
β-pinene (Scheme 2).
One example is the Crich synthesis
The ketone was reacted with MeLi. The methyl group attacked
stereoselectively from the sterically less hindered side of the
carbonyl, resulting in a 10:1 excess of cis-forms (13 plus
enantiomer 16) over the corresponding trans-forms of thujanol
Scheme 2. Synthesis of Pinocarvones
(
14 plus enantiomer 15) (i.e., cis-4-thujanol is the major
diasteromer formed).
As the ratio of stereoisomers in the sabinene (10) was
(
(
−)-93:(+)-7, it was easy to differentiate the (+)-(1R)- and
−)-(1S)-forms of sabina ketone by enantioselective GC. The
reaction with MeLi yielded a mixture of all four 4-thujanol
stereoisomers, which could be defined as (+)-trans-(1R,4S)-4-
thujanol (15), (−)-trans-(1S,4R)-4-thujanol (14), (+)-cis-
(
(
1R,4R)-4-thujanol (16), and (−)-cis-(1S,4S)-4-thujanol
13), in a ratio of 1:9:4:86, by use of the enantioselective
GC column.
of pinocarvone from β-pinene (7) using perfluorooctyl selenic
The NMR spectrum and specific rotation proved that the
isomer purchased from Sigma-Aldrich was the (+)-trans-
isomer, and the major product could be assigned as (−)-cis-4-
thujanol (13), based on the retention of ring configuration in
the synthesis sequence, as well as regioselective considerations
acid, where focus was on the preparation of the catalyst, and
27
the yield was ca. 40%. However, a serious drawback with
pinene as starting material is the formation of myrtenal, which
cannot be removed by silica column chromatography. Here we
report a process where each pinocarvone enantiomer was
separately synthesized from enantiopure β-pinene isomers (7)
by oxidation with SeO2 (Scheme 2). The mixture of
pinocarvone and myrtenal was subsequently oxidized with
H O /NaH PO and NaClO to remove the myrtenal in the
30,31
and reported NMR spectroscopic data.
GC Elution Order of 4-Thujanol Stereoisomers. On the
HP-5MS GC column, the trans diastereomers eluted first. On
the β-cyclodextrin column the first peak of four synthetic
isomers coeluted with the commercial (+)-trans-4-thujanol
stereoisomer (15) purchased from Sigma-Aldrich, and the last
peak coeluted with the isolated (−)-cis-thujanol (13). The
elution order of all isomers was (+)-trans, (−)-trans, (+)-cis,
2
2
2
4
2
form of myrtenic acid by silica gel chromatography (see GC-
chromatogram, Figure 3). On the enantioselective GC-column
(
−)-cis (Figure 5). The elution order is in accordance with
31 32
those reported by Larkov et al. and Marriott et al.
Specific Rotation of Sabina Ketone and 4-Thujanol
Stereoisomers. It should be noted that (−)-sabinene (10)
yields (+)-sabina ketone (12), which is subsequently trans-
formed to thujanols with (−)-cis-thujanol (13) as the major
isomer (Scheme 3). The commercial sabinene (apparently
from a natural source) has a specific rotation of −73 (c 1.0,
EtOH) and −81 (c 1.0, DCM). Moreover, the chemical purity
of the commercial sabinene (10) was only 75%, with 25% β-
pinene as an impurity and with an ee of 86%. Sabina ketone
(
[
12) was obtained in 86% ee and with specific rotations of
2
3
23
α]
+24 (c 1.0, EtOH) and [α]
+33.5 (c 1.0, EtOAc)
D
D
after removal of byproducts (pinene ketones) by chromatog-
raphy. The (−)-cis-thujanol isomer (13) produced in the last
step had, after chromatography, an optical purity of 91% ee
and a specific rotation of −40 (c 0.5, DCM). The commercial
Figure 3. Gas chromatograms before and after oxidation of myrtenal
by NaClO /H O .
2
2
2
(
+)-trans-4-thujanol (15) (Sigma-Aldrich) had a specific
rotation of +29.8 (c 0.5, DCM).
phase (−)-pinocarvone (9) elutes before (+)-pinocarvone (8)
1
(
Figure 4). The H NMR spectroscopic data of pinocarvone
28
13
have been published, but here we also provide C NMR
spectroscopic data.
EXPERIMENTAL SECTION
■
General Experimental Procedures. Optical rotations were
recorded in EtOH, EtOAc, and DCM on a 2019 model Rudolph
automatic polarimeter (APIII) manufactured by Rudolph Research
Analytical (Hackettstown, NJ, USA). NMR spectra were recorded in
Specific Rotation of Pinocarvone Stereoisomers. The sign
of specific rotation changed when going from pinene to
pinocarvone, i.e., (−)-β-pinene yielded (+)-pinocarvone (8)
and (+)-β-pinene yielded (−)-pinocarvone (9). The specific
CDCl on Bruker 400 and Varian 500 MHz spectrometers. The GC-
3
2
3
D
MS instrument was an Agilent 6890 GC and 5973 mass detector and
a Hewlett-Packard with a FID detector (Palo Alto, CA, USA). Helium
was used as carrier gas. Two types of columns, a nonpolar column
rotations were as follows: (+)-pinocarvone (8) [α] +30.8 (c
23
1
.0, EtOAc); (−)-pinocarvone (9) [α]
−29.6 (c 2.0,
D
EtOAc).
(
4
HP-5MS, film thickness = 0.25 μm; Agilent Technologies 19091S-
33) and a chiral-phase capillary column (Cyclosil-B, 30 m × 0.25
Synthesis of 4-Thujanol (Sabinene Hydrate) Stereo-
isomers. (−)-Sabinene (10) (86% ee) was subjected to a mild
permanganate oxidation yielding sabinenediol (11) (Scheme
μm, i.d. 0.25 mm, J&W Scientific, via Scantech Nordic AB, Jonsered,
Sweden), were used. Mass spectra were obtained by electron impact
ionization (70 eV). The general gas chromatography temperature
3
). The diol was cleaved using periodate to yield sabina ketone
C
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Figure 4. GC elution order of pinocarvone stereoisomers. Separated on GC equipped with a Cyclosil B column. In the chiral analysis
(
−)-pinocarvone eluted earlier than (+)-pinocarvone. (A) (−)-Pinocarvone (9), (B) (+)-pinocarvone (8), and (C) mix of pinocarvone
enantiomers. The temperature program: Initial oven temperature was 40 °C (held for 5 min) increased to 150 °C at 3 °C/min and finally increased
to 220 °C at 10 °C/min (held for 5 min at the final temperature).
Scheme 3. Synthesis of 4-Thujanol Stereoisomers
program for both columns was as follows: initial temperature 50 °C
(
hold for 2 min), raised to 200 °C with 10 °C/min (hold for 15 min)
(splitless). When a different temperature program was used for
resolution of enantiomers (on the Cyclosil-B column), the temper-
ature program is described in the figure legends of the GC
chromatograms. The synthesized compounds were purified on silica
gel column chromatography using 230−400 mesh ultra pure silica.
Synthesis of the Four Stereoisomers of Pinocamphone
(
2
Scheme 1). Synthesis of (−)-Isopinocamphone (3). (1S,2S,3S,5R)-
,6,6-Trimethylbicyclo[3.1.1]heptan-3-ol [(1S,2S,3S,5R)-(+)-isopino-
campheol] (1) (Sigma-Aldrich, Schnelldorf, Germany) (12.32 g, 80.0
mmol, chemical purity 98% and optical purity 95% ee) was dissolved
32
in CH Cl (150 mL). Silica gel (18 g) and PDC (60 g, 160 mmol)
2
2
were added, and the mixture was stirred for 3.5 h at RT before leaving
it in a fridge overnight. The slurry was diluted with cyclohexane and
filtered. The solid material was washed twice with 1:1 cyclohexane/
CH Cl (50 mL). The filtrate was concentrated and subjected to
2
2
MPLC, yielding 87% (10.6 g, 69.7 mmol) of (1S,2S,5R)-2,6,6-
trimethylbicyclo[3.1.1]heptan-3-one ((1S,2S,5R)-(−)-isopinocam-
phone) (3). The chemical purity was 99% with 98% ee.
Figure 5. Chromatograms of chiral-phase GC of 4-thujanol isomers in
the mixture and chromatograms of isolated (+)-trans- and (−)-cis-4-
thujanol isomers. The temperature program: Initial oven temp 40 °C
23
1
Specific rotation [α] = −11.4 (c 1.0, EtOH); H NMR (CDCl ,
D
3
5
2
−
(
00 MHz) δ (in ppm) 2.68−2.58 (2H, m), 2.54−2.45 (2H, m),
(
hold for 3 min) and increased to 150 °C at 3 °C/min and finally
.15−2.10 (1H, m), 2.08−2.05 (1H, m), 1.42 (1H, br s), 1.33 (3H, s,
increased to 250 °C at 15 °C/min. It was kept for 10 min at the final
temperature of 250 °C.
13
CH ), 1.21 (3H, d, −CH ), 0.87 (3H, s, −CH ); C NMR
3
3
3
CDCl , 125 MHz) δ (in ppm) 215.3, 51.4, 45.1, 45.0, 39.1, 34.5,
3
2
6
7.2, 27.1, 22.1, 17.0; GC-MS m/z 83 (100%), 69, 55, 95, 41, 81, 97,
7, 152 (M ), 110 (decreasing order of intensity).
+
D
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(
+)-Isopinocamphone (4). In analogy with the procedure used for
d, J = 2.6 Hz), 2.50 (1H, dd, J = 19.3 and 2.7 Hz), 2.18 (1H, dd, J =
5.8 and 3.0 Hz), 1.33 (3H, s, H-8), 1.27 (1H, d, J = 10.5 Hz), 0.78
the (−)-antipode, (1R,2R,3R,5S)-2,6,6-trimethylbicyclo[3.1.1]heptan-
1
3
3
-ol [(1R,2R,3R,5S)-(−)-isopinocampheol] (2) (12.21 g, 79.3 mmol)
(3H, s, H-9); C NMR (CDCl , 125 MHz) δ (in ppm) 200.1, 149.1,
3
was oxidized with PDC (60 g, 160 mmol) to yield 75% (9.03 g, 59.4
117.5, 48.3, 42.6, 40.9, 38.6, 32.5, 26.1, 21.6; GC-MS t 9.06 m/z 81
R
+
mmol) (1R,2R,5S)-2,6,6-trimethylbicyclo-[3.1.1]heptan-3-one
(100%), 108, 53, 107, 135, 79, 41, 77, 150 (M ), 69, 91, 122
[
(1R,2R,5S)-(+)-isopinocamphone] (4) after column chromatogra-
(intensity of decreasing order). NMR spectrometry data are in
23
16,28
phy. [α]
= +11.2 (c 1.0, EtOH), 98% ee. The NMR data were
accordance with reported values.
D
2
3
1
identical to the data for the (−)-isomer. GC-MS m/z. See other
enantiomer.
(−)-Pinocarvone (9). [α]
= −29.6 (c 2.0, EtOAc); H NMR
D
(CDCl , 500 MHz) δ (in ppm) 5.94 (1H, s, H-10 a), 4.99 (1H, s. H-
3
(
−)-Pinocamphone (5). (1S,2R,5R)-2,6,6-Trimethylbicyclo[3.1.1]-
heptan-3-one. To a solution of (−)-isopinocamphone (3) (3.04 g,
0.0 mmol) dissolved in EtOH (10 mL) was added NaOEt in EtOH
21% w/w, 11 mL, 34 mmol), and the mixture stirred for 24 h at RT.
Water (50 mL) and Et O (50 mL) were added after 24 h, when the
10b), 2.74 (1H, t, J = 5.9 Hz), 2.69−2.64 (1H, m), 2.63 (1H, d, J =
2.6 Hz), 2.52 (1H, dd, J = 19.3 and 2.7 Hz), 2.24−2.17 (1H, m), 1.33
1
3
2
(
(3H, s, H-8), 1.27 (1H, d, J = 10.5 Hz), 0.78 (3H, s, H-9); C NMR
(CDCl , 125 MHz) δ (in ppm) 200.2, 149.1, 117.6, 48.3, 42.6, 40.41,
3
2
38.6, 32.5, 26.1, 21.6; GC-MS t 9.06 m/z 108 (100%), 81, 53, 107,
R
+
4
:1 equilibrium ratio between (−)-pinocamphone (5) and (−)-iso-
pinocamphone (3) had been established. The aqueous phase was
extracted with Et O (2 × 50 mL), and the combined ether phases
were washed with 15 mL of water to remove EtOH. After drying over
135, 79, 41, 150 (M ), 69, 91, 122 (intensity of decreasing order).
Synthesis of 4-Thujanol Isomers (Scheme 3). Synthesis of
Sabinenediol (11). To a solution of (−)-sabinene (10) 86% ee (1 g,
2
7.4 mmol) in THF (3 mL) was added KMnO (2.3 g, 14.6 mmol) in
4
MgSO , filtration, and evaporation, 15 mL of toluene was added
water (4 mL) over a period of 2.5 h. The mixture was stirred for
another hour before the precipitate was filtered off. The filtrate was
extracted with EtOAc (2 × 50 mL), and the combined organic layers
4
before subsequent evaporation. The removal of water/EtOH by
azeotropic distillation with toluene was repeated twice. The clear
amber-colored residue was subjected to MPLC, yielding 2.9 g (19.1
mmol) of (1S,2R,5R)-(−)-pinocamphone) (5) as a 95:5 mixture with
were washed with brine and dried over Na
SO . The solution was
2
4
concentrated by rotatory evaporation to yield 875 mg (5.1 mmol) of
2
3
1
(
1S,2S,5R)-(−)-isopinocamphone (3). [α] = −20.2 (c 1.0, EtOH);
crude sabinene diol (11) (70% yield). H NMR (CDCl , 500 MHz) δ
3
D
1
H NMR (CDCl , 500 MHz) δ (in ppm) 2.67−2.59 (2H, m), 2.49
(in ppm) 3.55 (2H, app t, J = 11.6 Hz), 2.45 (1H, br s, −OH), 2.26
(1H, br s, −OH), 1.95−1.89 (1H, m), 1.67−1.61 (1H, m), 1.54 (1H,
dd, 14.1 and 8.6 Hz), 1.45 (1H, heptet, J = 6.9 Hz), 1.25−1.18 (1H,
m), 1.11 (1H, ddd, J = 8.5, 3.6, and 1.4 Hz), 0.98 (3H, d, J = 6.8 Hz),
3
(
(
−
2H, m), 2.14−2.07 (1H, m), 1.92 (1H, td, J = 6.1 and 2.1 Hz), 1.33
3H, s, −CH ), 1.16 (1H, d, J = 10.8 Hz), 1.10 (3H, d, J = 7.3 Hz,
3
1
3
CH ), 0.89 (3H, s, −CH ); C NMR (CDCl , 125 MHz) δ (in
3
3
3
ppm) 215.7, 46.5, 44.4 (2C), 39.4, 38.2, 29.1, 26.4, 19.8, 15.1; GC-
0.89 (3H, d, J = 6.8 Hz), 0.41 (1H, dd, J = 8.3 and 5.3 Hz), 0.24 (1H,
+
13
MS m/z 83 (100%), 55, 69, 41, 95, 81, 97, 67, 152 (M ), 53
dd, J = 5.2 and 3.5 Hz); C NMR (CDCl
3
, 125 MHz) δ (in ppm)
(
decreasing order of intensity).
83.5, 68.1, 34.5, 32.5, 32.3, 30.4, 25.5, 20.2, 20.1, 12.9.
Synthesis of Sabina Ketone (12). To a stirred solution of
(
+)-Pinocamphone (6). In analogy with the (−)-antipode,
(1R,2R,5S)-2,6,6-trimethylbicyclo[3.1.1]heptan-3-one [(1R,2R,5S)-
+)-isopinocamphone, 4) (3.04 g, 20.0 mmol)] was epimerized
sabinenediol diastereomers (11) (800 mg, 4.7 mmol) in THF/H
O
2
(
(1:1, 5 mL) was added NaIO (3.7 g, 17.3 mmol) in five portions
4
with NaOEt in EtOH. The yield of (1R,2S,5S)-2,6,6-trimethylbicyclo-
3.1.1]heptan-3-one ((1R,2S,5S)-(+)-pinocamphone, 6) as a 4:1
mixture with (1R,2R,5S)-(+)-isopinocamphone, 4), after column
during 30 min at RT. After 4 h, the mixture was diluted with water
[
(20 mL) and extracted with EtOAc (2 × 30 mL). The combined
organic layers were dried over Na SO and concentrated in vacuo to
2 4
2
3
chromatography, was 95% (2.89 g, 19.0 mmol). [α] = +22.7 (c 1.0,
obtain a crude mixture, which was purified on silica gel column
chromatography using 10% EtOAc in n-hexane as eluent. The yield
was 650 mg (4.7 mmol) of sabina ketone (12, 99% yield, 86% ee,
D
1
EtOH); H NMR (CDCl , 500 MHz) δ (in ppm) 2.69−2.58 (2H,
3
m), 2.50−2.36 (2H, m), 2.11 (1H, td, J = 5.8 and 2.8 Hz), 1.92 (1H,
2
3
D
1
td, J = 6.1 and 2.2 Hz), 1.33 (3H, s, −CH ), 1.16 (1H, d, J = 10.8
chemical purity 99%). [α]
= +24.4 (c 1.0, DCM); H NMR
3
1
3
Hz), 1.10 (3H, d, J = 7.3 Hz, −CH ), 0.89 (3H, s, −CH ).; C NMR
(CDCl , 400 MHz) δ (in ppm) 2.14−2.06 (2H, m), 1.97−1.94 (2H,
3
3
3
(
CDCl , 125 MHz) δ (in ppm) 215.7, 46.5, 44.4 (2C), 39.4, 38.2,
m), 1.63 (1H, dd, J = 8.8 and 2.8 Hz), 1.56 (1H, app quin, J = 6.8
Hz), 1.17 (1H, dd, J = 9.2 and 4.8 Hz), 1.06 (1H, dd, J = 4.8 and 3.2
3
2
9.1, 26.3, 19.8, 15.1; GC-MS m/z identical to the (−)-isomer 5 of
1
3
pinocamphone (see above).
Hz), 0.98 (3H, d, J = 6.8 Hz), 0.93 (3H, d, J = 6.8 Hz); C NMR
(CDCl , 100 MHz) δ (in ppm) 214.9, 39.5, 33.7, 33.2, 32.2, 23.6,
19.5, 19.3, 19.1; GC-MS t 24.00 (5.9%), 24.42 (94.1%) (Figure S29,
Synthesis of the Pinocarvone Stereoisomers (Scheme 2). To
3
a solution of (−)-β-pinene (7) (0.25 g, 1.8 mmol) in DCM (3 mL)
R
was added SeO (0.20 g, 1.8 mmol), and the mixture was refluxed for
Supporting Information), m/z 81 (100%), 96, 95, 67, 55, 123, 41, 138
2
+
2.5 h until GC-MS showed complete transformation. The solution
(M ), 109, 110 (decreasing order of intensity).
was filtered through silica gel (in a Pasteur pipet), and the product
washed out of the silica gel with additional aliqouts of DCM. The
product was concentrated under reduced pressure at 30 °C to obtain
a mixture of pinocarvone and myrtenal. These two compounds were
close on TLC and difficult to purify by column chromatography. To
the concentrate, MeCN (2 mL), NaH PO (70 mg) in Milli-Q-water
Synthesis of Sabinene Hydrates (4-Thujanols 13−16). To a
solution of sabina ketone 12 (200 mg, 1.4 mmol) in anhydrous Et O
2
(5 mL) was carefully added MeLi (1.8 mL, 2.8 mmol, 2.0 equiv, 1.6
M in Et O) at −78 °C. The mixture was stirred at the same
temperature for 1 h and another 1 h at RT. The reaction was
quenched by the addition of aqueous NH Cl (20 mL). The mixture
was extracted with Et O (2 × 30 mL), before the organic phase was
dried over Na SO , filtered, and concentrated in vacuo to afford a
2
2
4
4
(
1 mL), and 35% H O (0.2 mL) were added. The solution was
2
2
2
stirred for approximately 1 h until the solution became clear. On an
2
4
ice bath, NaClO (0.32 g) in MQ-water (3 mL) was added dropwise,
and the mixture was stirred overnight. One spatula of anhydrous
crude mixture (∼91% conversion) of four sabinene hydrates (4-
thujanols). The diastereomeric ratio was 90% cis-isomers [(−)-cis-
(1S,4S)-4-thujanol (13 plus its enantiomer 16)] and 10% trans-
isomers [(−)-trans-(1S,4R)-4-thujanol (14 plus 15)]. The cis- and
trans-thujanol diastereomers could be separated by column
chromatography on silica gel. Isolated yield of the cis-diastereomers
was 150 mg (0.97 mmol). GC-MS: The retention times of the trans-
isomers were (19.86, 20.02) and (21.08, 21.26) for the cis-isomers
(Figure S29, Supporting Information).
2
Na SO was added, and the mixture was extracted with DCM (3 × 5
2
3
mL). After removal of the solvents, the concentrate was purified on
silica gel. The combined fractions were concentrated by rotatory low-
vacuum evaporation to afford (+)-pinocarvone (8) (yield 60%, 165
mg, 1.1 mmol). The same experimental procedure was followed to
produce (−)-pinocarvone (9) from (+)-β-pinene. The chemical
purity of (+)-pinocarvone was 95%, and the optical purity was 97%
ee.
2
3
(−)-cis-(1S,4R)-4-Thujanol (13). [α] = −40 (c 0.5, DCM) (91%
D
2
3
1
(
+)-Pinocarvone (8). [α]D = +30.8 (c 1.0, EtOAc) and +27.5 (c
ee); H NMR (CDCl , 400 MHz) δ (in ppm) 1.69 (1H, br s, OH),
3
1
1
4
.0, DCM); H NMR (CDCl , 500 MHz) δ (in ppm) 5.93 (1H, s),
1.61−1.49 (3H, m), 1.29 (3H, s), 1.31−1.28 (2H, m), 1.01 (1H, dd, J
= 8.0 and 3.6 Hz), 0.89 (3H, d, J = 6.8 Hz), 0.84 (3H, d, J = 6.8 Hz),
3
.98 (1H, s), 2.74 (1H, t, J = 5.9 Hz), 2.69−2.64 (1H, m), 2.63 (1H,
E
https://dx.doi.org/10.1021/acs.jnatprod.0c00669
J. Nat. Prod. XXXX, XXX, XXX−XXX

Journal of Natural Products
pubs.acs.org/jnp
Article
0
.62 (1H, dd, J = 5.2 and 3.6 Hz), 0.28 (1H, dd, J = 8.0 and 5.2 Hz);
(4) Zhao, T.; Ganji, S.; Schiebe, C.; Bohman, B.; Weinstein, P.;
Krokene, P.; Borg-Karlson, A. K.; Unelius, C. R. ISME J. 2019, 13,
1535−1545.
13
C NMR (CDCl , 100 MHz) δ (in ppm) 79.2, 36.0, 33.5, 33.1, 32.5,
3
2
1
7.9, 25.4, 19.5, 19.4, 11.1; GC-MS m/z 71 (100%), 111, 93, 81, 43,
+
39, 121, 79, 69, 55, 136, 154 (M ) (decreasing order of intensity).
(5) Kalinova, B.; Brizova, R.; Knizek, M.; Turcani, M.; Hoskovec, M.
́ ́
(
+)-trans-(1S,4S)-4-Thujanol (15) (from Sigma-Aldrich, Schnell-
Arthopod-Plant Interact. 2014, 8, 305−316.
(6) Wang, S.-F.; Li, Y.-P.; Zhang, M.-G. Chin. J. Org. Chem. 2007,
1227, 1612−1617.
2
3
1
dorf, Germany). [α] = +29.8 (c 0.5, DCM) (99% ee); H NMR
D
(
CDCl , 400 MHz) δ (in ppm) 1.88−1.80 (1H, m), 1.58 (1H, dd, J =
3
1
2.0 and 8.0 Hz),1.52 (1H, dd, J = 14.0 and 8.4 Hz), 1.48−1.39 (2H,
(7) Pitínova-
́
S
̌
tekrova,
̌
, Z.; Cejka, J. Catal. Sci. Technol. 2018, 8, 4690−4701.
M.; Eliasova, P.; Weissenberger, T.; Shamzhy,
̌
́ ́ ́
m), 1.29 (3H, s), 1.26 (1H, ddd, J = 16.0,5.6 and 0.3 Hz), 1.07−1.03
M.; Musilovaa
́
(
(
(
1H, ddd, J = 8.4, 3.6, and 1.2 Hz), 0.96 (3H, d, J = 6.8 Hz), 0.88
3H, d, J = 6.8 Hz), 0.38 (1H, ddd, J = 8.0, 5.2, and 0.8 Hz), 0.20
(8) Anikeev, V. I.; Il’inab, I. V.; Volchob, K. P.; Yermakovaa, A.; S,
N. F. J. Supercrit. Fluids 2010, 52, 71−75.
(9) Rudinsky, J. A.; Morgan, M. E.; Libbey, L. M.; Putnam, T. B.
Environ. Entomol. 1974, 3, 90−98.
1
3
1H, dd, J = 5.2 and 3.6 Hz); C NMR (CDCl , 100 MHz) δ (in
3
ppm) 80.5, 36.6, 34.6, 34.4, 32.2, 25.9, 24.9, 20.0, 19.9, 13.3; GC-MS
m/z 71 (100%), 43, 93, 111, 81, 121, 139, 69, 55, 79, 136, 154 (M )
+
(10) Birgersson, G.; Debarr, G. L.; de Groot, P.; Dalusky, M. J.;
Pierce, H. D., Jr.; Borden, J. H.; Meyer, H.; Francke, W.; Espelie, K.
E.; Berisford, C. W. J. Chem. Ecol. 1995, 21, 143−167.
(11) Whittaker, D.; Banthorpe, D. V. Chem. Rev. 1972, 72, 305−313.
(decreasing order of intensity).
ASSOCIATED CONTENT
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(
12) Hartshorn, M. P.; Wallis, A. F. A. J. Chem. Soc. 1964, 0, 5254−
260.
13) Jirosova, A.; Kalinova, B.; Modlinger, R.; Jakus, R.; Unelius, C.
R.; M Blazenec, M.; Schlyter, F. Submitted, 2020.
*
sı Supporting Information
5
(
The Supporting Information is available free of charge at
https://pubs.acs.org/doi/10.1021/acs.jnatprod.0c00669.
GC chromatograms, 13C NMR and 1H NMR spectra
̌
́
́
̌
̌
(14) Blazyte-Cereskiene, L.; Apsegaite, V.; Radziute, S.; Mozuraitis,
R.; Buda, V.; Peciulyte, D. Ann. For. Sci. 2016, 73, 247−256.
(
PDF)
GC chromatograms, 13C NMR and H NMR spectra
PDF)
(15) Cecati, F. M.; Magallanes-Noguera, C.; Tonn, C. E.; Ardanaz,
1
C. E.; Kurina-Sanz, M. Process Biochem. 2018, 64, 93−102.
16) Baeckstrom, P.; Koutek, B.; Saman, D.; Vrkoc, J. Bioorg. Med.
Chem. 1996, 4, 419−421.
17) Galopin, C. C. Tetrahedron Lett. 2001, 42, 5589−5591.
(18) Cheng, D.; Kreethadumrongdat, T.; Cohen, T. Org. Lett. 2001,
, 2121−2123.
19) Cheng, D.; Knox, K. R.; Cohen, T. J. Am. Chem. Soc. 2000, 122,
(
(
(
AUTHOR INFORMATION
■
3
(
Corresponding Author
C. Rikard Unelius − Department of Chemistry and
Biomedical Sciences, Linnaeus University, 392 31 Kalmar,
Sweden; orcid.org/0000-0001-7158-6393;
Email: rikard.unelius@lnu.se
412−413.
(20) Fanta, W. I.; Erman, W. F. J. Org. Chem. 1968, 33, 1656−1658.
(21) Witzgall, P.; Bengtsson, M.; Unelius, C. R.; Lofqvist, J. J. Chem.
Ecol. 1993, 19, 1917−1928.
(22) Unelius, C. R.; El-Sayed, A. M.; Twidle, A.; Bunn, B.; Zaviezo,
Authors
T.; Flores, M. F.; Bell, V.; Bergmann, J. J. Chem. Ecol. 2011, 37, 166−
72.
23) Witzgall, P.; Backman, A. C.; Svensson, M.; Bengtsson, M.;
1
(
Suresh Ganji − Department of Chemistry and Biomedical
Sciences, Linnaeus University, 392 31 Kalmar, Sweden
Fredric G. Svensson − Department of Chemistry and
Biomedical Sciences, Linnaeus University, 392 31 Kalmar,
Sweden
Unelius, C. R.; Vrkoc, J.; Kirsch, P. A.; Ioriatti, C.; Lofqvist, J. J. Appl.
Entomol. 1996, 120, 611−614.
(24) Lofqvist, J. In Mechanisms in Insect Olfaction; Payne, T. L.,
̈
Birch, M. C., Kennedy, E. J., Eds.; Oxford Univ. Press, 1986.
(25) Chelucci, G.; Saba, A. Tetrahedron: Asymmetry 1998, 9, 2575−
2578.
Complete contact information is available at:
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(26) Malkov, A. V.; Baxendale, I. R.; Bella, M.; Langer, V.; Fawcett,
J.; Russell, D. R.; Mansfield, D. J.; Valko, M.; Kocovsky, P.
Notes
Organometallics 2001, 20, 673−690.
The authors declare no competing financial interest.
(
(
27) Crich, D.; Zou, Y. Org. Lett. 2004, 6, 775−777.
28) Abraham, R. J.; Cooper, M. A.; Indyk, H.; Siverns, T. M.;
ACKNOWLEDGMENTS
Whittake, D. Org. Magn. Reson. 1973, 5, 373−377.
29) Barberis, M.; Perez-Prieto, J. Tetrahedron Lett. 2003, 44, 6683−
685.
30) Krause, S. T.; Kollner, T. G.; Asbach, J.; Degenhardt, J. Arch.
Biochem. Biophys. 2013, 529, 112−121.
31) Larkov, O.; Dunkelblum, E.; Zada, A.; Lewinsohn, E.; Freiman,
L.; Dudai, N.; Ravid, U. Flavour Fragrance J. 2005, 20, 109−114.
32) Marriott, P. J.; Shellie, R.; Cornwell, C. J. Chromatogr A 2001,
936, 1−22.
■
(
6
(
This work was supported by two grants from The Carl Trygger
Foundation (Grant CTS 13:487 and CTS 14:493 awarded to
C.R.U. and S.G.) and the Linnaeus University, Kalmar,
Sweden. The authors are thankful to Dr. R. Naidu, SU,
Stockholm, for providing 400 MHz NMR data and Dr. J.
Wiklander, Linnaeus University, Kalmar, for providing 500
MHz NMR data.
(
(
REFERENCES
■
(
1) Symonds, M. R. E.; Gitau-Clarke, C. W. In Advances in Insect
Physiology; Tittiger, C., Blomquist, G. J., Eds.; Elsevier, 2016; Vol. 50,
Chapter 5, pp 195−234.
2) Blomquist, G. J.; Figueroa-Teran, R.; Aw, M.; Song, M.;
(
Gorzalski, A.; Abbott, N. L.; Chang, E.; Tittiger, C. Insect Biochem.
Mol. Biol. 2010, 40, 699−712.
(
3) Schiebe, C.; Unelius, C. R.; Ganji, S.; Binyameen, M.;
Birgersson, G.; Schlyter, F. J. Chem. Ecol. 2019, 45, 474−489.
F
https://dx.doi.org/10.1021/acs.jnatprod.0c00669
J. Nat. Prod. XXXX, XXX, XXX−XXX