Stereochemistry of female-specific normonoterpenes, sex pheromone candidates from the acarid mite, Tyreophagus sp. (Astigmata: Acaridae)

Article abstract of DOI:10.1271/bbb.90364

Two normonoterpenes were detected from an unidentified Tyreophagus sp. as new female-specific components. Both planar structures were identified to be 2,6-dimethyl-5-heptenal (1) and 2,6-dimethyl-5-hepten- 1-ol (2) by GC/MS co-chromatography with synthetic 1 and 2. The stereochemistry of 2 was determined to be R by a GC analysis with a chiral column, while that of 1 was presumed to be similar to 2 based on the biosynthetic aspects.

Full text of DOI:10.1271/bbb.90364

Biosci. Biotechnol. Biochem., 73 (10), 2332–2334, 2009
Note
Stereochemistry of Female-Specific Normonoterpenes,
Sex Pheromone Candidates from the Acarid Mite,
*
Tyreophagus sp. (Astigmata: Acaridae)
Nobuhiro SHIMIZU,1 Kuniaki MIWA,2 Koji yNOGE,2 Ryota YAKUMARU,1
1;
Naoki MORI,2 and Yasumasa KUWAHARA
1Department of Bioscience and Biotechnology, Faculty of Bioenvironmental Science, Kyoto Gakuen University,
1-1 Nanjo, Sogabe, Kameoka 621-8555, Japan
2Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University,
Sakyo-ku, Kyoto 606-8502, Japan
Received May 21, 2009; Accepted June 16, 2009; Online Publication, October 7, 2009
[doi:10.1271/bbb.90364]
Two normonoterpenes were detected from an un-
identified Tyreophagus sp. as new female-specific com-
ponents. Both planar structures were identified to be
2,6-dimethyl-5-heptenal (1) and 2,6-dimethyl-5-hepten-
1-ol (2) by GC/MS co-chromatography with synthetic 1
and 2. The stereochemistry of 2 was determined to be
R by a GC analysis with a chiral column, while that
of 1 was presumed to be similar to 2 based on the
biosynthetic aspects.
temperature programmed from 135 to 250 ꢀC at
4 ꢀC/min, Kovat’s retention index I < ca. 1650)2) from
the female extract was 4 times larger than that from the
males.
Compound 1 (tR 6.62 min) indicated the Mþ ion at
m=z 140 (intensity 9%) and the base ion peak at m=z 82
(100%), with the following ions: 122 (2%), 107 (2%), 98
(2%), 69 (28%), 67 (53%), 55 (19%) and 41 (47%).
Compound 2 (tR 7.77 min) gave the Mþ ion at m=z 142
(intensity 37%) and the base ion at m=z 41 (100%),
together with the following ions: 124 (6%), 109 (53%),
95 (39%), 82 (73%), 69 (99%), 67 (66%) and 55 (42%).
The Mþ and fragment ions higher than m=z 82 of 1
appeared to be m=z 2 less than those of 2, but all other
fragment ions lower than m=z 82 were the same as those
of 2. This suggests that both compounds were structur-
ally related to each other. Compound 1 was suggested to
be an aldehyde and 2 to be an alcohol, judging from the
intensities of each dehydrated ion (Fig. 2). Observation
of a dehydrated ion from the Mþ ion at m=z 124 of 2 and
the high intensity of m=z 69 (supportive of a prenyl
group) looked similar to that of citroneral (Mþ ion at
m=z 156), suggestive of 2 as a one-methylene-unit (m=z
14) shorter analogue. The fragment ion at m=z 82 of 1
could be explained by a McLafferty rearrangement ion
of aldehyde 1, and that of 2 by the elimination of H2O
and propene via a transition state of the six-membered-
ring from alcohol 2. The structure of 1 was, therefore,
elucidated to be 2,6-dimethyl-5-heptenal, and of 2 to be
2,6-dimethyl-5-hepten-1-ol. The elucidated structure of 1
was a compound commercially available, and of 2 was
prepared by LiAlH4 reduction of 1. Authentic compound
1 gave the same GC tR and mass spectral data as those of
natural 1. Synthetic 2 also gave the same GC tR and mass
spectral data as those of natural alcohol 2. As a result,
compound 1 was identified as 2,6-dimethyl-5-heptenal,
and compound 2 as 2,6-dimethyl-5-hepten-1-ol.
Key words: Tyreophagus
sp.;
hepten-1-ol; (R)-2,6-dimethyl-5-heptenal;
(R)-2,6-dimethyl-5-
normonoterpene
Astigmatid mites contain various compounds as
components of the opisthonotal gland exudate compris-
ing terpenes, sesquiterpenes, aromatics, hydrocarbons,
aliphatic compounds and other miscellaneous ones.
Some of those compounds are used as alarm, aggrega-
tion or sex pheromones, although their functions are
mostly obscure.1) As a part of natural product chemistry,
all mites collected from the field have been examined by
mass spectrometry coupled with gas chromatography
(GC/MS) to confirm the nobility of the species as a new
collection.1) During a conventional GC/MS analysis of
an unidentified Tyreophagus sp., two new peaks (1, tR
6.62 min, and 2, tR 7.77 min) were detected as female-
specific components, and these compounds seemed to
be normonoterpenes. Their chemical structures were
elucidated and confirmed by synthesis.
A female extract of the species indicated 12 GC
peaks: tridecane (f), neral (c), heptacosane (i), com-
pound 1, pentadecane (h), compound 2, geranial (d),
nonacosane (k), tetradecane (g), 2-hydroxy-6-methyl-
benzaldehyde (a), neryl formate (e) and cholesterol (l) in
decreasing order, together with squalene (j) originating
from the diet (Fig. 1). A male extract was composed of
10 peaks: tridecane (f), heptacosane (i), nonacosane (k),
neral (c), neryl formate (e), nerol (b), geranial (d),
cholesterol (l), pentadecane (h) and tetradecane (g). The
GC response of the semiochemical zone (tR < 8 min
with a CP-Sil 19CB capillary column, operated at a
In order to determine the stereochemistry, (R)-2,6-
dimethyl-5-hepten-1-ol (2-R) was synthesized by way of
(R)-2,6-dimethyl-5-heptenal (1-R) which had been pre-
pared according to the synthetic route reported,3) starting
from commercially available (R)-3,7-dimethyl-1,6-octa-
*
y
Chemical Ecology of Astigmatid Mites LXXXVI
To whom correspondence should be addressed. Fax: +81-771-29-3429; E-mail: kuwahara@kyotogakuen.ac.jp

Stereochemistry of Female-Specific Normonoterpenes from the Astigmatid Mite
2333
diene (4) with subsequent epoxidation and ozonolysis
(Fig. 3). Racemic acetate 3 displayed two peaks (tR at
45.33 min and 45.85 min) by a GC analysis with a
Chrompack chiral capillary column. Chiral 3-R corre-
sponded to the latter peak (tR at 45.85 min), and natural
3 derived from the mite hexane extract also indicated the
same retention time. Identity was also confirmed by GC
co-chromatography. As a result, the absolute configu-
ration of alcohol 2 was concluded to be R. On the other
hand, the chiral column could not resolve racemic
aldehyde 1, like the case of alcohol 2. A trial GC
analysis of natural 3 via reduction and subsequent
acetylation of natural 1 was not successful, possibly due
to the small amount. From the biosynthetic point of
view, natural aldehyde 1 should also be an R-isomer.
The presence of 1 has been reported in the opistho-
branch mollusk, Melibe leonina, as a putative allo-
mone,4) in ants,5) and in the essential oil of many plants.
However, its absolute structure seems not to have been
clarified. The presence of 2 was first reported in the ant,
Pheidole fallax, by GC and MS,6) and its distribution
has been recognized not only among ants,7–9) but also
in Andrena bees.10) Its distribution has subsequently
been widely demonstrated in plant essential oil as a
principle of its sweet flavor such as in Rutaceae (Citrus
medica L.11) and C. hystrix DC (Swangi)12)), mandarin
orange peel oil,13) Lauraceae (several Lauraceae plants
in China),14) Litsea cubeba Pers.,15) Zingiberaceae
(Zingiber mioga16) and Z. officunalis Roscoe17)) and
Passifloraceae (Passiflora quadrangularis).18) The edible
ascidian, Halocynthia reretzi, also contains 2 as a minor
component.19) As far as we know, there is no report
available on its stereochemistry. This might be the first
study to conclude that the stereochemistry of compound
2 was R.
Compounds 1 and 2 were the fourth and sixth major
components in the female extract, together with tride-
cane and neral, and were not detectable in the male
extract. This fact implies that compounds 1 and 2
function as the female sex pheromone, although this
remains to be confirmed. The female sex pheromone is
distributed between both sexes in the genus Sancassania
(¼ Caloglyphus); however, the amount is biased toward
females, being 1.4–8.4 times more than that in the male
secretion.20) Although the pheromone has not been
studied in the genus Tyreophagus, the pheromone may
also be localized in Tyreophagus females. The presence
of 2 is one of the characteristics of the present species.
The other two related Tyreophagus sp. maintained in
our laboratory do not contain 2 (data not shown). As
acyclic normonoterpenoids other than 1 and 2, sodium
(or potassium) 2,6-dimethylheptyl sulfate has been
reported from the ascidian, Polycitor adriaticus, col-
lected in the northern Adriatic Sea.21) ꢀ-D-Glucosides of
cyclopentano-normonoterpenoids has been found in the
leaves of Cerbera manghas.22) There presently seem to
be four normonoterpenoids known in nature, including
the finding of the present study.
82
1
j
f
6
4
2
67
41
c
Female
Male
69
M+
55
140
107
122
98
1
i
i
d
f
e
2
h
h
a
g
g
k
j
69
l
l
41
100
50
0
2
2
82
e
67
c
k
d
b
109
M+
0
55
95
142
5
10
15
20
25
Retention time (min)
124
Fig. 1. Typical Gas-Liquid Chromatogram of the Hexane Extract
from Unidentified Tyreophagus sp. (Ishigaki).
1, 2,6-dimethyl-5-heptenal; 2, 2,6-dimethyl-5-hepten-1-ol;
a, 2-hydroxy-6-methylbenzaldehyde; b, nerol; c, neral; d, geranial;
e, neryl formate; f, tridecane; g, tetradecane; h, pentadecane;
i, heptacosane; j, squalene; k, nonacosane; l, cholesterol
50
100
150
m/z
Fig. 2. Mass Spectra of 2,6-Dimethyl-5-heptenal (1) and 2,6-
Dimethyl-5-hepten-1-ol (2).
a
b
O
4
5
c
d
OH
O
CHO
O
2-R
1-R
3-R
Fig. 3. Preparative Procedure for Normonoterpenes 1–3.
a, 3-chloroperbenzoic acid, CHCl3; b, O3, Zn, AcOH, NaOAc, NaI; c, LiALH4, Et2O; d, Ac2O, pyridine, CH2Cl2

2334
N. SHIMIZU et al.
(1H, t-like, J ¼ 7:2 Hz, CH=); 13C-NMR (100 MHz) ꢁ: 16.8, 17.7,
Experimental
25.3, 25.7, 32.1, 33.4, 69.4, 124.3, 131.7, 171.3.
(R)-2,6-Dimethyl-5-hepten-1-ol (2-R). Aldehyde 1-R (prepared as
reported3)) was reduced as already described, using LiAlH4 to give
The unidentified Tyreophagus sp. was raised in a culture dish
(90 mm in diameter ꢁ 20 mm in height) by feeding dry yeast in a
humid condition at around 20–25 ꢀC. The species was collected from
organic soil in Ishigaki Island, and its ITS portion was composed of a
324 base pair (measured as reported23)). The culture dish was kept in a
zip-lock plastic bag (240 ꢁ 170 ꢁ 0:04 mm) to maintain moisture and
to prevent any escape or contamination with other mites. Fifty females
and males were collected as a group by a needle into a small conical-
bottomed tube (8 mm in diameter ꢁ 50 mm in height). To each tube,
hexane (4 ml) was added, and then all of the extract was taken up in a
micro-syringe (10 ml, Hamilton Co., USA) after 3 min (exactly), and
subjected to the GC/MS analysis. The GC/MS analysis was performed
at 70 eV by an HP 5989B mass spectrometer coupled with an HP 5890
series II plus gas chromatograph, using an HP-5MS capillary column
(0:25 mm ꢁ 30 m, 0.25 mm in film thickness) in the splitless mode. The
carrier gas was helium at 1.23 ml/min, and the oven temperature was
programmed from 60 ꢀC to 290 ꢀC at 10 ꢀC/min with an initial 2-min
hold. GC was performed with a Hewlett Packard 5890 series II plus gas
chromatograph equipped with a flame ionization detector, using the
same column and conditions as those just stated. The chromatogram
was processed by an HP 3396 series II Integrator. All chemicals used
were of reagent grade. Optical rotation data were taken by a SEPA-300
high-sensitivity polarimeter (Horiba), and column chromatography was
performed on silica gel (Wakosil C-200). 1H- and 13C-NMR spectra
were obtained with a Bruker Biospin AC400M NMR spectrometer,
using TMS as an internal standard.
24
alcohol 2-R. ½ꢂꢂD þ9:1 (c 1.0 CHCl3).
(R)-2,6-Dimethyl-5-heptenyl acetate (3-R). Alcohol 2-R was acety-
lated as already described, using pyridine and acetic anhydride to give
24
acetate 3-R. ½ꢂꢂD ꢃ2:4 (c 1.0 CHCl3).
Determination of the stereochemistry. The absolute structure of
natural 2 in the mite hexane extract was determined after acetylation
by GC co-chromatography, using a Chrompack chiral capillary column
[CP-cyclodextrin-ꢀ-2,3,6-M-19, 0:25 mm i.d. ꢁ 25 m, 0.25 mm in film
thickness, in the split mode (100:1)]. Racemic 3 and chiral acetate 3-R
prepared as described were used as standards. Racemic 3 gave two
peaks at tR 45.33 min and 45.85 min. Chiral 3-R corresponded to the
peak at tR 45.85 min.
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