(5R,7R)-5-Methylheptadecan-7-ol: A novel sex pheromone component produced by a female lichen moth, Miltochrista calamina, in the family Arctiidae

Article abstract of DOI:10.1016/j.tetlet.2011.08.136

A methyl-branched heptadecanol was found in the pheromone gland extract of a female lichen moth, Miltochrista calamina (Arctiidae, Lithosiinae). GC-MS analyses of the alcohol and a hydrocarbon derived from it by subsequent treatments with methanesulfonyl chloride and LiAlD4 in microscale reactions indicated 5-methylheptadecan-7-ol (1) as one possible structure. The four stereoisomers of 1 in a ratio of 4:4:1:1 were prepared from (S)-b-citronellol with 60% ee, and were separated by a combination of achiral and chiral HPLC columns. The absolute configuration of each isomer was determined by the comparison with the chromatographic behaviors of other samples synthesized by a different scheme, which applied the Jacobsen hydrolytic kinetic resolution of racemic 1,2-epoxydodecane to fix the configuration of the 7-hydroxy group. Only the (5R,7R)-isomer attracted male moths; thus, we concluded that M. calamina females secrete (5R,7R)-1 as a sex pheromone, indicating a new chemical class of lepidopteran female sex pheromones.

Full text of DOI:10.1016/j.tetlet.2011.08.136

Tetrahedron Letters 52 (2011) 5808–5811
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(5R,7R)-5-Methylheptadecan-7-ol: a novel sex pheromone component
produced by a female lichen moth, Miltochrista calamina, in the family Arctiidae
⇑
Rei Yamakawa, Ryutaro Kiyota, Tomonori Taguri, Tetsu Ando
Graduate School of Bio-Applications and Systems Engineering, Tokyo University of Agriculture and Technology, Koganei, Tokyo 184-8588, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
A methyl-branched heptadecanol was found in the pheromone gland extract of a female lichen moth,
Miltochrista calamina (Arctiidae, Lithosiinae). GC–MS analyses of the alcohol and a hydrocarbon derived
from it by subsequent treatments with methanesulfonyl chloride and LiAlD4 in microscale reactions indi-
cated 5-methylheptadecan-7-ol (1) as one possible structure. The four stereoisomers of 1 in a ratio of
4:4:1:1 were prepared from (S)-b-citronellol with 60% ee, and were separated by a combination of achiral
and chiral HPLC columns. The absolute configuration of each isomer was determined by the comparison
with the chromatographic behaviors of other samples synthesized by a different scheme, which applied
the Jacobsen hydrolytic kinetic resolution of racemic 1,2-epoxydodecane to fix the configuration of the
7-hydroxy group. Only the (5R,7R)-isomer attracted male moths; thus, we concluded that M. calamina
females secrete (5R,7R)-1 as a sex pheromone, indicating a new chemical class of lepidopteran female
sex pheromones.
Received 18 July 2011
Revised 19 August 2011
Accepted 23 August 2011
Available online 31 August 2011
Keywords:
Lepidoptera
Female sex pheromone
Methyl-branched alcohol
Chiral HPLC
Asymmetric synthesis
Chemical ecology
Ó 2011 Elsevier Ltd. All rights reserved.
Female moths secrete species-specific pheromones to attract
males. Sex pheromones have been identified from adult females
of more than 600 lepidopteran species, and are mainly composed
of fatty alcohols and their derivatives (Type I pheromones) or
polyenyl hydrocarbons and their epoxy derivatives (Type II phero-
mones).1 In addition to these well-known components, many mis-
cellaneous chemicals have been identified, such as secondary
alcohols and methyl-branched hydrocarbons.1,2 Recently, we found
novel methyl-branched ketones from Lyclene dharma dharma in the
subfamily Lithosiinae in the family Arctiidae.3 Larvae of many Lith-
osiinae species feed on lichens and the adults are called lichen
moths. The family Arctiidae is one of the most evolved groups in
the Lepidoptera, and is additionally divided into two other subfam-
ilies, Syntominae and Arctiinae. It is known that the females in
these two subfamilies commonly utilize Type II pheromones.1,2
However, information on sex pheromones of the lichen moths is
still limited, and only the second identification from a lichen moth
has been reported for Eilema japonica japonica. Unexpectedly, E. j.
japonica females secreted unbranched polyenyl hydrocarbons, a
typical Type II pheromone, to attract males.4 We were interested
to know whether or not only L. d. dharma had established an anom-
alous chemical communication system. Lithosiinae includes many
species, with about 80 species recorded from Japan. To more thor-
oughly understand their mating communication, we examined a
pheromone gland extract obtained from another lichen moth, Mil-
tochrista calamina, when a single female was captured on one of
our university campuses in a suburb of Tokyo (June, 2010). In ana-
lytical experiments, the sex pheromone was successfully deter-
mined to be
a novel methyl-branched secondary alcohol.
Furthermore, a field evaluation of all stereoisomers, prepared by
stereoselective syntheses, determined the absolute configuration
of the natural pheromone.
In a GC–MS analysis of an aliquot of the pheromone gland ex-
tract (0.2 female equivalent, FE) on a highly polar capillary column
(DB-23),5 only one abundant component was detected at
16.78 min. The mass spectrum, with characteristic ions at m/z
269 (M-1), 252 (M-18), 171, and 129 (Fig. 1A), suggested a C18 sec-
ondary alcohol structure possessing two alkyl groups (C7H15 and
C10H21), but the GC–MS data did not coincide with those of
octadecan-8-ol (Fig. 1B), which was synthesized by Grignard cou-
pling between octanal and decylmagnesium bromide. Because
the tR of the natural component was shorter than that of authentic
octadecan-8-ol (17.84 min), a methyl-branched skeleton was as-
signed as a possible pheromone candidate. However, the position
of the methyl group could not be determined from the mass spec-
trum. To determine the position, microscale reactions were carried
out with an aliquot of the pheromone extract (0.4 FE). The second-
ary alcohol in the extract was reacted with methanesulfonyl chlo-
ride, and the mesylate produced was reduced with LiAlD4.6 The
GC–MS analysis showed the production of a mono-deuterated
hydrocarbon (C18H37D) with M+ at m/z 255, and the mass spectrum
showed characteristic fragment ions at m/z 198, 170, 85, and 57
(Fig. 1C), indicating a methyl branch at the 5-position. This result
⇑
Corresponding author. Tel./fax: +81 42 388 7278.
E-mail address: antetsu@cc.tuat.ac.jp (T. Ando).
0040-4039/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2011.08.136

R. Yamakawa et al. / Tetrahedron Letters 52 (2011) 5808–5811
5809
171
A
B
C
(%)
100
69
OH
111
43
129
97
83
55
50
0
129
M-1+
269
171
210
252
250
50
100
150
150
150
200
m/z
171
(%)
69
100
50
0
OH
129
55
97
83
111
43
M-1+
269
129
171
85
252
250
50
100
200
m/z
57
(%)
D
43
85
100
50
0
170
198
198
57
M+
255
72
170
50
100
200
250
m/z
Figure 1. Mass spectra: (A) a pheromone candidate (1) extracted from a M. calamina female, (B) synthetic octadecan-8-ol, and (C) hydrocarbon derived from 1 by treatments
with methanesulfonyl chloride and LiAlD4. The spectra were measured by electron impact ionization (EI) using GC–MS with a DB-23 column (0.25 mm ID ꢁ 30 m). The
column temperature program was 50 °C for 2 min, 10 °C/min to 160 °C, and 4 °C/min to 220 °C.
indicates two possible structures, 5-methylheptadecan-7-ol and
13-methylheptadecan-8-ol, for the parent alcohol. Although the
use of LiAlD4 did not reveal the position of the hydroxyl group,
the deuteration contributed to the identification of the hydrocar-
bon derived from the alcohol, among many components in the
crude product after two chemical reactions. To determine the exact
structure of the alcohol, we had to approach it by synthesis be-
cause an insufficient gland extract was available for further analy-
sis. The pheromone candidates included two chiral centers, and so
we examined two synthetic routes to authentic compounds, fixing
the configuration of one of the two centers in each synthesis.
First, (5S,7S)- and (5S,7R)-isomers of 5-methylheptadecan-7-ol
(1) were synthesized from (S)-(ꢀ)-b-citronellol (2, 60% ee) as
shown in Figure 2A. After protection of the hydroxyl group as a
methoxymethyl (MOM) ether, the double bond was oxidized with
Oxone to yield a diol (3),7 which was cleaved to an aldehyde (4) by
the treatment with NaIO4. Wittig reaction between 4 and methyl-
enetriphenylphosphorane produced the terminal olefin (5). By the
catalytic hydrogenation and MOM-deprotection of 5, (S)-3-methyl-
heptan-1-ol (6) was obtained. IBX oxidation and subsequent Grig-
nard coupling with decylmagnesium bromide yielded 1 as a
mixture mainly composing of its (5S,7S)- and (5S,7R)-isomers.
The diastereomers were separable by normal-phase HPLC with
an achiral column (NO2),8 and two fractions (Fr. I, tR 17.8 min;
and Fr. II tR 20.0 min) were obtained by preparative HPLC. GC–
MS analysis of both diastereomers showed the same mass spec-
trum as the natural pheromone candidate, confirming its planar
structure. The diastereomer in Fr. I (GC tR 16.78 min) also had
the same tR as the natural component and was eluted later than
the diastereomer in Fr. II (GC tR 16.66 min) from the DB-23 col-
umn.5,9 Furthermore, enantiomeric resolution by chiral HPLC was
examined with the two fractions (Fig. 3). The analysis revealed that
each fraction included two enantiomers in a ratio of about 4:1, as
expected from the ee of 2 used as the starting material. The four
stereoisomers of 1 had different chromatographic behaviors on a
chiral HPLC column (Chiralpack AY-H); thus, the optical purity of
each synthetic isomer was evaluated by chiral HPLC analysis.
While the configurations of the two enantiomers in each fraction
could not be determined, the analysis showed that each (5S)-iso-
mer eluted faster than its antipode regardless of the configuration
at the 7-position.
The elution order of the four stereoisomers from the chiral col-
umn was proven by comparing with analytical data of the samples
prepared by another synthetic route, which fixed the configuration
of the hydroxyl group at the 7-position, as shown in Figure 2B.
Thus, 1-dodecene (7) was epoxidized with a peracid to yield a race-
mic mixture of 1,2-epoxydodecane (8), which was then subjected
to a hydrolytic kinetic resolution with Jacobsen’s catalyst.10 Hydro-
lysis of 8 was monitored by GC–MS, and the optical purity of the
unhydrolyzed epoxide was analyzed by chiral HPLC.8 Grignard
coupling between (S)-8 (>98% ee) and 2-hexylmagnesium bromide
produced a 1:1 mixture of (5S,7S)-1 and (5R,7S)-1. The two diaste-
reomers were separable by HPLC with the NO2 column,8 and two
fractions (Fr. III and Fr. IV) were obtained. Similarly, another 1:1
mixture of (5R,7R)-1 and (5S,7R)-1 was synthesized from (R)-8
(>98% ee), and two fractions (Fr. V and Fr. VI) were obtained. Chiral
HPLC analyses of all four fractions revealed stereoisomers included
in each fraction as follows: (5S,7S)-1 in Fr. III (tR 16.5 min),

5810
R. Yamakawa et al. / Tetrahedron Letters 52 (2011) 5808–5811
OH
i, ii
A
iv
iii
H
OMOM
OMOM
OH
OH
O
2
3
4
OH
v, vi
vii, viii
ix
Fr. I = (5S,7S)-1
Fr. II = (5S,7R)-1
OMOM
C H
OH
C H
4
C H
10 21
9
4
9
5
(5S,7RS)-1
6
major component
OH
ix
ix
O
xiii
xiii
Fr. III = (5S,7S)-1
Fr. IV = (5R,7S)-1
B
C
H
10 21
C H
C
H
4
9
10 21
xi
x
(S)-8
(5RS,7S)-1
O
C H
10 21
C H
10 21
7
(RS)-8
OH
xii
Fr. V = (5R,7R)-1
Fr. VI = (5S,7R)-1
O
C
H
10 21
C H
C
10 21
H
4
9
(5RS,7R)-1
(R)-8
Figure 2. Synthetic routes to four stereoisomers of 5-methylheptadecan-7-ol (1): (A) synthesis starting from (S)-b-citronellol (2, 60% ee) and (B) synthesis of two
diastereomeric mixtures utilizing the hydrolytic kinetic resolution of an epoxy intermediate (8). Reagents (yield): (i) DMM, p-TsOH, LiBr (91%); (ii) trifluoroacetone, Oxone,
NaHCO3, EDTA, CH3CN, H2O (87%); (iii) NaIO4, Et2O, H2O (81%); (iv) CH2@PPh3, THF (65%); (v) H2, Pd–C, MeOH (quant.); (vi) dry HCl, MeOH (90%); (vii) IBX, DMSO (62%); (viii)
C
10H21MgBr, THF (45%); (ix) HPLC (achiral NO2 column); (x) MCPBA, CH2Cl2 (75%); (xi) (S,S)-(salen)Co(III)OAc, H2O, dimethoxymethane (93%); (xii) (R,R)-(salen)Co(III)OAc,
H2O, dimethoxymethane (91%); (xiii) 2-hexylmagnesium bromide, THF, CuI (65%).
components in Fr. I; thus, the 5S,7S or 5R,7R configuration was as-
signed for the natural component.
(5S,7S)-1
A
(5R,7R)-1
The activity of the synthetic pheromone candidate was evalu-
ated at coppices in the university campus from May, 2011. Each
isomer (0.2 mg) was impregnated into a small rubber septum,
which was placed in a sticky trap as a lure. Traps baited with
(5R,7R)-1 specifically attracted M. calamina males (about 30
males/trap/month), and no males were captured by traps baited
with other isomers. Based on this result, we concluded that mating
communication of M. calamina was mediated by (5R,7R)-1, a new
natural product, although the possibility of 13-methylheptadec-
an-8-ol has not been eliminated. Further experiments with binary
lures indicated that the other isomers neither synergized nor
inhibited responses to (5R,7R)-1. The gland extract and synthetic
diastereomers were also analyzed by GC equipped with an electro-
antennogram detector (GC–EAD)12 using antennae from male li-
chen moths captured by the traps. The antennae clearly
responded to the natural and synthetic (5R,7R)-1 (Fig. 4). Details
of the field and GC–EAD tests will be reported elsewhere.
Fr. I
Fr. III
Fr. V
20
15
t
R (min)
(5S,7R)-1
B
(5R,7S)-1
Fr. II
Methyl-branched hydrocarbon pheromones have been identi-
fied from several species in some families of Lepidoptera, such as
the Lyonetiidae and Geometridae. In the pheromone gland of an
Arctiinae species, 5-methylheptadecane has been interestingly
found together with Type II pheromone components, while the role
in the mating communication has not been reported.13 And, sec-
ondary alcohols with a short straight chain have been identified
from species in the Ereiocraniidae.1,2 However, this is the first iden-
tification of a methyl-branched secondary alcohol as a lepidopter-
an sex pheromone. Further studies will clarify the novelty or
commonness of branched secondary alcohols among the lepidop-
teran sex pheromones. Identifications of the pheromones of many
other lichen moths are expected. In addition to chain length, differ-
ent positions and configurations of methyl and hydroxyl groups
could provide a diversity of species-specific pheromones as do
those of double bonds in Type I pheromones. Some ponerine
ant species secrete methyl-branched secondary alcohols with a
Fr. IV
Fr. VI
20
15
t
R (min)
Figure 3. Chiral HPLC analyses of stereoisomers of synthetic 5-methylheptadecan-
7-ol (1) to determine their absolute configurations. The HPLC was conducted with a
Chiralpack AY-H column (4.6 mm ID ꢁ 25 cm) and an RI detector using 0.3% 2-
propanol in hexane as an eluent (flow rate, 0.5 ml/min). Frs. I and II mainly include
(5S,7SR)-isomers synthesized from (S)-b-citronellol (2, 60% ee), Frs. III and IV
include (5SR,7S)-isomers derived from (S)-8, and Frs. V and VI include (5SR,7R)-
isomers derived from (R)-8.
(5R,7S)-1 in Fr. IV (tR 14.5 min), (5R,7R)-1 in Fr. V (tR 18.0 min), and
(5S,7R)-1 in Fr. VI (tR 15.9 min) (Fig. 3).11 This result clarified the

R. Yamakawa et al. / Tetrahedron Letters 52 (2011) 5808–5811
5811
3. (a) Yamamoto, M.; Kamata, T.; Do, N. D.; Adachi, Y.; Kinjo, M.; Ando, T. Biosci.,
Biotechnol., Biochem. 2007, 71, 2860–2863; (b) Adachi, Y.; Do, N. D.; Kinjo, M.;
Makisako, S.; Yamakawa, R.; Mori, K.; Ando, T. J. Chem. Ecol. 2010, 36, 814–823.
4. Fujii, T.; Nakano, R.; Takubo, Y.; Qian, S.; Yamakawa, R.; Ando, T.; Ishikawa, Y. J.
Insect Physiol. 2010, 56, 1986–1991.
A
B
0.1mV
EAD
FID
5. Mass spectra were measured in EI mode (70 eV) with a GC–MS system equipped
with a split/splitless injector and a DB-23 column (0.25 mm ID ꢁ 30 m, 0.25
lm
film, J&W Scientific). The column temperature program was 50 °C for 2 min,
10 °C/min to 160 °C, and 4 °C/min to 220 °C. The carrier gas was He.
6. A crude gland extract (0.4 FE) dissolved in CH2Cl2 (100 ll) was mixed with
triethylamine (7.5 mg), DMAP (1.0 mg), and methanesulfonyl chloride (33 mg).
After stirring at 0 °C for 2 h and at room temperature for 1 day, a saturated
14
15
13
16
NaHCO3 solution (200
hexane, dried over Na2SO4, and concentrated. The crude produced mesylate
was dissolved in dry THF (100 l) and treated with LiAlD4 (5.5 mg) under an
ll) was added to the mixture, which was extracted with
t
R (min)
l
argon atmosphere. The reaction mixture was acidified with 1 N HCl after 3 h
stirring and then extracted with hexane.
7. Rani, S.; Vankar, Y. D. Tetrahedron Lett. 2003, 44, 907–909.
EAD
FID
8. All HPLC analyses were conducted with an RI detector. For the separation of
diasteromers, a functionalized silica column (Senshu-pak NO2-3151-N, 8 mm
ID ꢁ 15 cm) was used. Eluent, 3% EtOAc in hexane; and flow rate, 1.0 ml/min.
For the enantiomeric resolution of the alcohol (1), a Chiralpak AY-H column
(4.6 mm ID ꢁ 25 cm, Daicel Chemical Industry, Osaka, Japan) was used. Eluent,
0.3% 2-propanol in hexane; and flow rate, 0.5 ml/min. For the enantiomeric
resolution of the epoxide (8), a Chiralpak AS-H column (4.6 mm ID ꢁ 25 cm,
Daicel Chemical Industry) was used. Eluent, 0.1% 2-propanol in hexane; and
flow rate, 0.5 ml/min. tR of (S)-8, 14.3 min; and tR of (R)-8, 11.2 min.
9. NMR data of 1: Isomers in Fr. I [(5S,7S)-1 and (5R,7R)-1], 1H NMR d: 0.88 (6H, t,
J = 6.4 Hz), 0.90 (3H, d, J = 6.4 Hz), ꢂ1.16 (1H, m), ꢂ1.26 (22H, m), ꢂ1.42 (4H,
m), 3.69 (1H, m). 13C NMR d: 14.14, 14.17, 19.30, 22.71, 23.00, 25.72, 29.19,
29.23, 29.36, 29.65 (ꢁ3), 29.74, 31.94, 37.63, 38.41, 45.04, 69.68. Isomers in Fr.
II [(5S,7R)-1 and (5R,7S)-1], 1H NMR d: 0.88 (3H, t, J = 6.5 Hz), 0.89 (3H, t,
J = 6.6 Hz), 0.90 (3H, d, J = 6.4 Hz), ꢂ1.16 (1H, m), ꢂ1.26 (22H, m), ꢂ1.42 (4H,
m), 3.69 (1H, m). 13C NMR d: 14.14, 14.18, 20.43, 22.71, 23.05, 25.58, 29.11,
29.36, 29.58, 29.64 (ꢁ2), 29.67, 29.75, 31.94, 36.31, 37.81, 45.30, 70.04.
10. (a) Schaus, S. E.; Brandes, B. D.; Larrow, J. F.; Tokunaga, M.; Hansen, K. B.; Gould,
A. E.; Furrow, M. E.; Jacobsen, E. N. J. Am. Chem. Soc. 2002, 124, 1307–1315; (b)
Nielsen, L. P. C.; Stevenson, C. P.; Blackmond, D. G.; Jacobsen, E. N. J. Am. Chem.
Soc. 2004, 126, 1360–1362; (c) Stevenson, C. P.; Nielsen, L. P. C.; Jacobsen, E. N.
Org. Synth. 2006, 83, 162–169.
0.1mV
(5R,7S)-1
(5R,7R)-1
14
15
13
16
t
R (min)
Figure 4. GC–EAD analyses: (A) a pheromone gland extract (0.05 FE) and (B) a
mixture of synthetic (5R,7S)-1 and (5R,7R)-1 (1 ng each). The analysis was
conducted with the same GC column and temperature program as for the GC–MS
measurement.
C7–C15 chain skeleton or their acetates as trail pheromones.14 Pher-
omone biosynthesis in these ants has not been reported. The lichen
moth and ants may have similar enzymes that produce these
chemically similar structures. The biosynthetic pathways for these
branched secondary alcohols will be an interesting subject for the
follow-up research.
11. ½ ꢃ : (5S,7R)-Isomer, ꢀ5.12 (c 1.45, CHCl3); (5R,7S)-isomer, +5.59 (c 1.12,
a 2D4
CHCl3); (5S,7S)-isomer, +11.57 (c 0.69, CHCl3); (5R,7R)-isomer, ꢀ10.92 (c 0.65,
CHCl3).
12. (a) Struble, D. L.; Arn, H. In Techniques in Pheromone Research; Hummel, H. E.,
Miller, T. A., Eds.; Springer: New York, 1984; pp 161–178; (b) Inomata, S.;
Watanabe, A.; Nomura, M.; Ando, T. J. Chem. Ecol. 2005, 31, 1429–1442.
13. Schneider, D.; Schulz, S.; Kittmann, R.; Kanagaratnam, P. J. Appl. Entomol. 1992,
113, 280–294.
References and notes
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2. (a) Ando, T. Internet database, 2011. http://www.tuat.ac.jp/~antetsu/
LepiPheroList.htm.; (b) El-Sayed, A. M., Internet database, 2011. http://
www.pherobase.com/.
14. (a) Attygalle, A. B.; Vostrowsky, O.; Bestmann, H. J.; Steghaus-Kovac, S.;
}
Maschwitz, U. Naturwissenshaften 1988, 75, 315–317; (b) Janssen, E.; Ubler, E.;
Bauriegel, L.; Kern, F.; Bestmann, H. J.; Attygalle, A. B.; Steghaus-Kovacˇ, S.;
Maschwitz, U. Naturwissenshaften 1997, 84, 122–125.