Characterization of (1R,4S,4aR,7S,7aR)-dihydronepetalactol as a semiochemical for lacewings, including Chrysopa spp. and Peyerimhoffina gracilis

Article abstract of DOI:10.1023/A:1015201129331

The enantiomerically pure diastereoisomers (1R,4S,4aR,7S,7aR)(1) and (1R,4R,4aR,7S,7aR)-dihydronepetalactol (2) were synthesized diastereoselectively from a renewable resource, (4aS,7S,7aR)-nepetalactone (3), isolated as the main constituent of the essent

Full text of DOI:10.1023/A:1015201129331

Journal of Chemical Ecology, Vol. 28, No. 4, April 2002 (�C 2002)
CHARACTERIZATION OF
(
1R,4S,4aR,7S,7aR)-DIHYDRONEPETALACTOL AS A
SEMIOCHEMICAL FOR LACEWINGS, INCLUDING
Chrysopa spp. AND Peyerimhoffina gracilis
1
2
1
3
A. M. HOOPER, B. DONATO, C. M. WOODCOCK, J. H. PARK,
4
3
2
1,�
R. L. PAUL, K. S. BOO, J. HARDIE, and J. A. PICKETT
1Biological Chemistry Division, IACR-Rothamsted
Harpenden, Herts., AL5 2JQ, United Kingdom
2Aphid Biology Group, Department of Biological Sciences
Imperial College at Silwood Park
Ascot, Berks., SL5 7PY, United Kingdom
3School of Agricultural Biotechnology
Seoul National University
Suwon 441-744, Korea
4School of Chemistry
Cantock’s Close, University of Bristol
Bristol, BS8 1TS, United Kingdom
(Received July 2, 2001; accepted December 9, 2001)
Abstract—The enantiomerically pure diastereoisomers (1R,4S,4aR,7S,7aR)-
1) and (1R,4R,4aR,7S,7aR)-dihydronepetalactol (2) were synthesized diastere-
(
oselectively from a renewable resource, (4aS,7S,7aR)-nepetalactone (3), isolated
as the main constituent of the essential oil of the catmint plant Nepeta cataria.
The stereochemistry of the compounds was determined by NMR spectroscopy
and X-ray crystallography, and the compounds were identified, respectively, as
neomatatabiol and isoneomatatabiol, natural products from Actinidia polygama,
for which the lactol stereochemistry was previously incompletely defined. Com-
pound 1 was found to catch significant numbers of three species of lacewing
in the field: in Korea, Chrysopa cognata, and in the United Kingdom, Nineta
vittata and most notably Peyerimhoffina gracilis. All species caught in signifi-
cant numbers were found more frequently in traps releasing 1 than 2, while
more C. cognata, C. formosa, and C. phyllochroma were found in traps releasing
(1R,4aS,7S,7aR)-nepetalactol (4). The catch of P. gracilis with 1 is of particular
interest as this lacewing has only recently been recorded in the United Kingdom.
�
To whom correspondence should be addressed. E-mail: john.pickett@bbsrc.ac.uk
8
49
0
098-0331/02/0400-0849/0 �C 2002 Plenum Publishing Corporation

8
50
HOOPER ET AL.
Where sexed, the lacewings of all species trapped were found to be male, im-
plying a possible pheromonal role for these or structurally related compounds.
Key Words—Nepeta cataria, Chrysopa cognata, Chrysopa formosa, Chrysopa
phyllochroma, Nineta vittata, Peyerimhoffina gracilis, lacewing, pheromone,
dihydronepetalactol, neomatatabiol, isoneomatatabiol.
INTRODUCTION
The cyclopentanoids (4aS,7S,7aR)-nepetalactone (3) and (1R,4aS,7S,7aR)-
nepetalactol (4) (Figure 1) comprise the sex pheromones of a number of pest
aphids (Aphididae), either as single compounds or as species-specific blends
(Dawson et al., 1987, 1989). Aphid species including Myzus persicae, Aphis
fabae, Acyrthosiphon pisum, Sitobion fragariae, Tuberocephalus momonis, and
Rhopalosiphum padi have been trapped in the field using synthetic samples of these
pheromone components (Dawson et al., 1990; Hardie et al., 1997; Boo et al., 2000).
In addition, predatory species of lacewing such as Chrysopa cognata (Chrysopi-
dae) are caught in traps releasing the same compounds, leading to the suggestion
that, just as many aphid species use similar sex pheromones, lacewing predators
may be using these compounds as kairomones in locating prey (Boo et al., 1998).
Earlier literature reports revealed that the lacewing Chrysopa septempunctata was
attracted to the vine Actinidia polygama (Actinidiaceae), a plant that generates
two dihydronepetalactols previously named neomatatabiol and isoneomatatabiol
(Hyeon et al., 1968), structurally related to 3 and 4. The present work aimed to
synthesize neomatatabiol and isoneomatatabiol and to determine unambiguously
their stereochemistry. Studies were then undertaken to compare the potency of the
aphid sex pheromone components 3 and 4 with the dihydronepetalactols related
stereochemically to 3, and to correlate lacewing catches in the field with the lactol
stereochemistry not reported previously.
FIG. 1. Structures of aphid sex pheromone components (4aS,7S,7aR)-nepetalactone (3) and
(1R,4aS,7S,7aR)-nepetalactol (4).

LACEWING SEMIOCHEMICAL
851
METHODS AND MATERIALS
Experimental Chemistry
NMR spectra were obtained using the Bruker 500 Avance NMR spectrometer.
GC-MS analysis employed the VG-Autospec (Fisons Instruments) with a siloxane
�
�
�
column (50 m � 0.32 mm ID; 0.52 �m) operated from 30 C to 250 C at 5 C/min;
�
EI at 70 EV, 250 C. GC analysis was on an HP-5 siloxane column (30 m �
� � � �
0
.32 mm ID; 0.25 �m) operated from 40 C to 150 C at 5 C/min and from 150 C
�
�
to 250 C at 10 C/min. Compounds eluted at 16.42 (2), 16.93 (1), 31.08 (11), and
1.44 min (10).
1R,4S,4aR,7S,7aR)-Dihydronepetalactol (1). The essential oil from
3
(
N. cataria (10.0 g, 60.2 mmol) was dissolved in ethanol (250 ml) under a hydro-
gen balloon with 10% palladium on charcoal (600 mg) as a catalyst. The mixture
was stirred overnight, filtered, and the solvent removed in vacuo. A portion of
�
the crude product (2.0 g, 11.9 mmol) was dissolved in dry THF (30 ml) at –78 C
under nitrogen. DIBAL (13.1 ml, 1 M in hexanes) was added slowly and the mix-
ture warmed to room temperature over 1 hr. The reaction was quenched without
acidification, extracted with diethyl ether, and the combined organic extracts were
dried (MgSO4), filtered, and concentrated in vacuo. Purification by flash column
chromatography (30% diethyl ether in petroleum ether) and recombination of the
2
0
�
purest fractions gave 1 as a clear colorless oil (1.02 g, 50%). [�] – 1.1 (c = 9.0,
D
1
CHCl3); H NMR (500 MHz, CDCl3) � 4.41 (1H, d, J = 9.1 Hz, H-1), 3.66
(
3
1
1H, dd, J = 5.2, 11.8 Hz, pro-R H-3), 3.28 (1H, t, J = 11.8 Hz, pro-S H-3),
.02 (1H, br s, OH), 2.25 (1H, m, H-4a), 2.14 (1H, m, H-4), 2.06 (1H, m, H-7),
.98 (1H, m, pro-R H-6), 1.59 (1H, m, pro-R H-5), 1.46 (1H, m, H-7a), 1.44 (1H, m,
pro-S H-5), 1.17 (1H, m, pro-S H-6), 1.00 (3H, d, J = 6.9 Hz, 7-CH3), 0.80 (3H,
13
d, J = 7.1 Hz, 4-CH3); C NMR (125 MHz, CDCl3) � 97.1 (C1), 67.3 (C3), 53.2
(
C7a), 41.7 (C4a), 34.5 (C7), 32.3 (C6), 30.1 (C4), 23.9 (C5), 22.4 (7Me), 14.7
+
(
9
4Me); m/z = 170 (0.16%, M ), 81 (100%), 67 (96), 41 (77), 137 (65), 39 (58),
+
5 (55), 109 (50), 79 (44), 152 (42, M -18), 82 (40), 27 (39), 110 (37), 107 (36),
5 (36), 53 (34), 43 (26), 77 (26), 29 (25).
5
(
1R,4R,4aR,7S,7aR)-Dihydronepetalactol (2). The essential oil from Nepeta
�
cataria (10.0 g, 60.2 mmol) was dissolved in methanol (100 ml) at 0 C. A solution
ofsodiumborohydride(2.51g, 66.2mmol)inicecoldmethanol(100ml)wasadded
to the mixture by cannula, and the reaction mixture was stirred until complete. The
pH was adjusted to 7 after the addition of water (250 ml), and the mixture was
extracted with diethyl ether. The combined organic extracts were washed with
water, dried (MgSO4), and concentrated in vacuo. A portion of the crude mixture
(3.0 g, 17.9 mmol) was dissolved in ethanol (75 ml) under a balloon of hydrogen
with 10% palladium on charcoal (200 mg) as a catalyst. The mixture was stirred
overnight, filtered, and the solvent evaporated to yield a colorless oil containing

8
52
HOOPER ET AL.
a 91:9 mixture of diastereoisomers (2.91 g, 96%). Purification of 2.91 g of the
mixture by flash column chromatography (30% ether in petroleum ether) and
recombination of the purest fractions gave 2 as a colorless oil (1.85 g, 64%).
2
0
�
1
[
�] + 9.4 (c = 6.2, CHCl3); H NMR (500 MHz, CDCl3) � 5.12 (1H, br s,
D
H-1), 4.17 (1H, br s, OH), 3.60 (1H, t, J = 11.0 Hz, pro-R H-3), 3.42 (1H, dd,
J = 4.7, 11.3 Hz, pro-S H-3), 2.00 (1H, m, H-7), 1.92 (1H, m, pro-R H-6), 1.88
(
1H, m, H-4a), 1.68 (1H, m, pro-R H-5), 1.52 (1H, m, pro-S H-5), 1.46 (1H, m,
H-4), 1.44 (1H, m, H-7a), 1.19 (1H, m, pro-S H-6), 1.01 (3H, d, J = 6.3 Hz,
1
3
7
(
-CH3), 0.79 (3H, d, J = 6.6 Hz, 4-CH3); C NMR (125 MHz, CDCl3) � 92.8
C1), 64.1 (C3), 51.1 (C7a), 41.5 (C4a), 33.6 (C7), 31.5 (C6), 30.9 (C4), 27.9 (C5),
+
+
2
4
5
0.0 (7Me), 15.5 (4Me); m/z = 170 (0.4%, M ), 67 (100%), 81 (91), 137 (78),
1 (76), 95 (62), 39 (55), 109 (45), 152 (42, M -18), 79 (42), 27 (37), 110 (34),
3 (33), 55 (33), 82 (32), 107 (31), 77 (26).
(1R,4S,4aR,7S,7aR)-Dihydronepetalactol Dimer (10). Isomer 1 (420 mg,
2.47 mmol) was dissolved in diethyl ether (15 ml) and 6 N HCl (9 drops) and
stirred overnight. The solvent was removed by rotary evaporation, and the residue
was purified by flash column chromatography (10% diethyl ether in petroleum
2
0
�
ether) to yield 10 as a clear colorless oil (300 mg, 75%). [�] – 25.5 (c = 10.2,
D
1
CHCl3); H NMR (500 MHz, CDCl3) � 4.51 (1H, d, J = 7.4 Hz, H-1), 3.65
(
2
1H, dd, J = 4.9, 11.3 Hz, pro-R H-3), 3.27 (1H, t, J = 11.3 Hz, pro-S H-3),
.25 (1H, m, H-4a), 2.22 (1H, m, H-4), 1.96 (1H, m, H-7), 1.94 (1H, m, pro-
R H-6), 1.57 (2H, m, pro-R H-5, H-7a), 1.43 (1H, m, pro-S H-5), 1.12 (1H, m,
pro-S H-6), 0.99 (3H, d, J = 6.6 Hz, 7-CH3), 0.80 (3H, d, J = 6.6 Hz, 4-CH3);
13
C NMR (125 MHz, CDCl3) � 98.1 (C1), 66.9 (C3), 50.6 (C7a), 40.9 (C4a),
3
5.0 (C7), 32.9 (C6), 29.6 (C4), 24.3 (C5), 21.8 (7Me), 14.8 (4Me); m/z = 322
+
(0.012%, M ), 81 (100%), 67 (83), 137 (80), 41 (76), 95 (71), 109 (58), 39 (55),
1
7
52 (51), 70 (49), 153 (47), 107 (47), 110 (46), 27 (39), 55 (37), 53 (36), 93 (31),
7 (28), 43 (27).
(1R,4S,4aR,7S,7aR)-Dihydronepetalactol Dimer (11). Isomer 2 (2.41 g,
14.2 mmol) was dissolved in a mixture of diethyl ether (100 ml) and 6 N HCl
(4 ml) and stirred overnight. The solvent was removed by rotary evaporation and
the crude residue purified by flash column chromatography (10% diethyl ether
�
in petroleum ether) to yield 11 as a white solid (1.76 g, 77%). Mp 110–113 C;
20
�
1
[
�] – 39.1 (c = 5.8, CHCl3); H NMR (500 MHz, CDCl3) � 5.02 (1H, br s,
D
H-1), 3.36 (1H, dd, J = 4.9, 11.0 Hz, pro-S H-3), 3.29 (1H, t, J = 11.0 Hz, pro-R
H-3), 2.01 (1H, m, H-7), 1.92 (1H, m, pro-R H-6), 1.79 (1H, m, H-4a), 1.68 (1H, m,
pro-R H-5), 1.50 (1H, m, pro-S H-5), 1.44 (1H, m, H-4), 1.37 (1H, m, H-7a), 1.15
(
4
1H, m, pro-S H-6), 1.00 (3H, d, J = 6.4 Hz, 7-CH3), 0.75 (3H, d, J = 6.4 Hz,
-CH3); 13C NMR (125 MHz, CDCl3) � 92.7 (C1), 64.4 (C3), 50.8 (C7a), 41.8
(
C4a), 33.6 (C7), 31.6 (C6), 31.1 (C4), 27.9 (C5), 19.9 (7Me), 15.4 (4Me);
+
m/z = 322 (0.001%, M ), 67 (100%), 137 (97), 81 (96), 41 (77), 95 (73), 153

LACEWING SEMIOCHEMICAL
853
(
(
59), 39 (59), 109 (51), 152 (50), 79 (49), 110 (43), 107 (39), 27 (37), 53 (35), 55
34), 77 (29), 93 (28), 91 (26).
X-Ray Crystallography
Crystals of 11 suitable for single crystal X-ray structure determination were
grown by recrystallization from hexane. A single crystal of 11 was mounted on
a glass fiber using high-vacuum grease. X-ray measurements were made with a
Bruker SMART CCD area-detector diffractometer with Mo-K� radiation (� =
˚
0
.71073 A); structure solution SHELXTL program system version 5.1, 1998.
Crystal Data for 11. C20H34O3; M = 322.47, orthorhombic, space group
˚
˚
˚
P2(1)2(1)2(1), a = 8.7397(13) A, b = 9.4463(14) A, c = 22.898(3) A, U =
˚
3
3
� 1
1
7
2
890.4(5) A , Z = 4, Dc = 1.133 Mg/m , �(Mo-K�) 0.074 mm , F(000) =
�
12, T = 173(2) K, 3336 independent reflections with 2� ꢀ 50 . Refinement of
12 parameters converged at final R1 = 0.0434, wR2 = 0.0806.
Field Trapping in Korea
Field trapping experiments were carried out at the Suwon Agricultural Cam-
pus of Seoul National University and in the campus arboretum during July and
August 2000. Wing trap dispensers (IMP Technology Inc.) and rubber septa
(
Sigma) were used. The traps were mounted 1.5 m above the ground and 2 m
�
apart. The traps were checked and rotated 90 at 06:00 hr every morning. Anal-
ysis of trap catches was carried by using the GLIM statistics program (Crawley,
1993). The analysis was carried out declaring Poisson errors, with the catch for
each replicate treated as independent.
Field Trapping in the United Kingdom
Lacewings were caught in clear water traps made of 14-cm-diam. � 2-cm-
deep Petri dishes filled with a 1% clear odorless detergent solution. The traps were
placed 2 m apart on 1.1-m-high poles. The volatiles were located in the center of
the Petri dish above the detergent level, in dispensers made of amber glass vials
[Chromacol 08-CPV(A)] with 1-mm holes drilled in the plastic lids. The vials
were suspended below plastic shields to prevent rain entering and affecting release
rates. Vials contained either the test compound (10 mg) in diethyl ether (50 �l)
or ether alone. Traps were located at four woodland edge sites at Silwood Park,
Ascot, Berkshire, with 16 traps at each site. In each trial, there were two traps
for each treatment at each site. Traps were distributed so that identical treatments
did not occur in adjoining traps. During the trial, which was run between May
and August 2000, traps were rotated every four to five days. Traps were visited
every two to three days, when all trapped lacewings were removed, identified, and

8
54
HOOPER ET AL.
counted, and the solution in the traps was replaced. Analysis of water trap catches
was carried with the GLIM statistics program (Crawley, 1993), with the trap totals
for each treatment from each site being averaged, and each site treated as a block.
The analysis was carried out declaring Poisson errors, with each site treated as an
independent replicate.
Electrophysiology
Live adult male Peyerimhoffina gracilis were collected at Silwood Park, from
traps made from clear plastic drinking bottles, during February and March 2001.
Electroantennograms(EAGs)wereobtainedfromexcisedantennae(N = 7), using
Ag–AgCl glass electrodes filled with a saline solution. The signals generated by
the antenna were passed through a high impedance amplifier (Syntech UN-06,
Hilversum, The Netherlands), and data storage and processing were carried out
with a PC-based interface and customised software package (Syntech). The system
for stimulus delivery employed a filter paper strip in a disposable Pasteur pipet
cartridge. Samples of test solutions (1 mg/ml in hexane, 10 �l applied) were
applied to filter paper strips, and the solvent was allowed to evaporate for 30 sec
before the paper strip was placed in the cartridge. A 2-sec stimulus (1 liter/min)
was delivered into a purified airstream flowing continuously over the preparation.
Results were expressed as a percentage of the response to a standard application of
(
E)-2-hexenal at the same concentration, and analysed for significant differences
by Student’s t test.
RESULTS AND DISCUSSION
SynthesisandStructureElucidation. (4aS,7S,7aR)-Nepetalactone(3)isfound
in the essential oil of many species of the Nepeta genus (Regnier et al., 1967;
Eisenbraun et al., 1980; De Pascual Teresa et al., 1987) but, for high yield and
purity, is obtained from N. cataria commercially by our collaborators (Botanix
Limited) at approximately 90% of 3. The essential oil was used here without fur-
ther purification for the synthetic work. Compounds 1 and 2 were synthesized
diastereoselectively from 3, either by hydrogenation followed by lactone reduc-
tion in the case of 1, or by lactone reduction followed by hydrogenation for 2.
Reduction of 3 with 10% palladium on carbon gave a 95:5 ratio of two products
by GC, determined to be dihydronepetalactones by GC-MS (Scheme 1). The ma-
jor component did not have the same spectroscopic properties as that reported
for (4R,4aR,7S,7aR)-dihydronepetalactone (5) (De Pascual Teresa et al., 1987),
but full characterization and stereochemical assignment by NMR spectroscopy
was consistent with (4S,4aR,7S,7aR)-dihydronepetalactone 6, formed by reduc-
tion from the less sterically hindered face. Interestingly, this is the opposite face
selectivity to that reported when palladium on strontium carbonate is used as a

LACEWING SEMIOCHEMICAL
855
SCHEME 1. Divergent diastereoselective synthesis of 1 and 2 from (4aS,7S,7R)-
nepetalactone (3).
hydrogenation catalyst (Regnier et al., 1967). In our hands, however, reduction
with 2% Pd/SrCO3 gave a different result from Regnier et al. (1967) and yielded
the same major isomer 6 in a 6:5 ratio of 96:4 by GC. The crude mixture ob-
tained from reduction with 10% palladium on carbon was then further reduced
with DIBAL, and the crude product was analyzed by GC to reveal a 94:6 ratio of
isomers 1:2. By altering the order of chemical transformation, it was possible to
synthesize 2 diastereoselectively. Borohydride reduction of the essential oil gave
diastereomeric nepetalactols in the ratio 92:8, the stereochemistry of which was
previously elucidated by X-ray crystallography of a derivative (Dawson et al.,
1989). Hydrogenation of this crude mixture then gave a quantitative 91:9 mixture
of 2:1 by GC. The diastereoselectivity of hydrogenation of nepetalactone can be
explained on steric grounds, but, after borohydride reduction, the face selectiv-
ity of hydrogenation is reversed with comparable selectivity. This phenomenon
has been observed previously when hydrogenation of O-ethoxyethyl protected
(1R,4aS,7S,7aR)-nepetalactol (4) was reported to occur with the same facial se-
lectivity (Kigawa et al., 1992).
The relative stereochemistry of compounds 1 and 2 was determined by 1D
gradient NOE spectroscopy (GOESY) (Figure 2) (Stonehouse et al., 1994). In the
case of isomer 1, the H-1 proton gives a NOE to the triplet at H-3 that, in turn, shows
a NOE with a proton at H-5. Due to the known cis ring junction of 3, this is only
possible if H-1, H-3 (triplet), and the H-5 proton are all on the same upper face.
The other H-3 proton, a doublet of doublets, shows a NOE to proton H-4a that is

8
56
HOOPER ET AL.
FIG. 2. Assignment of the stereochemistry of (1R,4S,4aR,7S,7aR)-dihydronepetalactol
1) and (1R,4R,4aR,7S,7aR)-dihydronepetalactol (2), showing the structurally revealing
(
GOESY results.
known to be on the back face, verifying the structure. The methyl stereochemistry
at C-4 is inferred by the large trans diaxial coupling constant of 11.8 Hz between
the H-3 triplet and H-4. H-4a also shows a NOE to H-6 (1.17 ppm) and H-5 (1.59
ppm) and finally, a NOE from H-7 to H-5 (1.44 ppm) completes the assignment
of the C-5 and C-6 protons. Several derivatives were made of 1 in an attempt to
produce crystals for X-ray crystallography (Figure 3). None of compounds 7–9
was suitable for X-ray diffraction, but all showed the same NOEs between protons
H-1, H-3 (triplet), and H-5, suggesting the same stereochemistry as presented in
Figure 3. To ensure that the basic conditions used in generating 7–9 (treatment
FIG. 3. Derivatives of 1 synthesized, which all revealed the same stereochemistry by
GOESY spectroscopy.

LACEWING SEMIOCHEMICAL
857
SCHEME 2. Acid catalyzed dimerization of 1 and 2.
of 1 with n-BuLi in anhydrous solvent followed by the acyl or alkyl chloride)
was not epimerizing the lactol center, 7 was saponified in basic conditions and 1
reisolated with no lactol isomerization. An attempt to epimerize the lactol center
of 1 using acid conditions was also made but instead of epimerization, 10 was
isolated in good yield (Scheme 2). Infrared analysis of compound 10 showed it did
not contain a hydroxyl group, and with one set of NMR peaks, it was characterized
as a symmetric dimer. A very weak parent ion at m/z 322 was detected by electron
impact mass spectroscopy. The ion at m/z 153 could be rationalized by loss of the
C10H17O2 group and the ion at m/z 152 by loss of dihydronepetalactol, analogous to
lossofwaterfromthemonomericcompounds1and2. Again, NOEstudiesrevealed
the same relative stereochemistry as 1, demonstrating the 1R stereochemistry to
be the thermodynamically stable configuration as the reaction mechanism must
involve equilibrium of 1 with an oxonium ion.
Despite purified 2 giving a single peak by three different GC columns (HP-1,
HP-5, and �-cyclodextrin), its NMR spectrum in CDCl3 showed it to be a mix-
ture of two isomers, in a ratio of 90:10. GOESY studies on the major isomer 2
(Figure 2) revealed NOEs between the lactol OH and the H-3 triplet, which, in
turn, gave a NOE to H-4a. As the ring junction stereochemistry is known, H-4a,
H-3 (triplet), and the lactol OH must all be on the same back face. The stereochem-
istry at C-4 was assigned using the trans diaxial coupling of 11.0 Hz between H-3
(triplet) and H-4. Full assignment of the C-5 and C-6 protons was completed as
the H-6 proton at 1.19 ppm shows a NOE to H-7a and the H-5 proton at 1.68 ppm,
which in turn also relaxes H-4a and H-7a. GOESY studies on the minor isomer
of the CDCl3 solution of 2 revealed a NOE between H-1 and the H-3 triplet,
demonstrating it to be the C-1 epimer. The treatment of 2 with acid generated a
crystalline dimer, 11. Mass spectrometry data for 11 were similar to those for 10

8
58
HOOPER ET AL.
FIG. 4. Ortep diagram of 11. Ellipsoids are drawn at the 30% probability level.
and crystals were obtained, suitable for X-ray crystallographic study (Scheme 2).
The X-ray crystallographic results revealed the relative stereochemistry of 2 and
based on the known chiral centers, stereochemistry at C-4 was verified as 4R and
the thermodynamically stable lactol as 1R (Figure 4). By comparing the NMR
data and synthetic methodology with the paper of Hyeon et al. (1968), structure 1,
(1R,4S,4aR,7S,7aR)-dihydronepetalactol, can be termed neomatatabiol and struc-
ture 2, (1R,4R,4aR,7S,7aR)-dihydronepetalactol, can be termed isoneomatatabiol.
Field Results and Electrophysiology. Field trapping experiments in Suwon,
Korea, and at Ascot, United Kingdom, caught several species showing varying
preferences for the compounds described above. The initial study in Korea was
performed between July 17 and 22, 2000. The traps releasing compound 1 caught
significantly more Chrysopa cognata than did those releasing 2. However, com-
pound 4 was more effective than either, and also caught smaller but significant
numbers of Chrysopa formosa and Chrysopa phyllochroma (Table 1). This study
TABLE 1. Chrysopa spp. CAPTURED IN SUWON, JULY 17–22, 2000a
Dispenser treatment
C. cognata
C. formosa
C. phyllochroma
1
2
171.3a
63.7b
20c
1.3ab
0.3b
0b
0.7a
0.3a
0a
N. cataria extract containing 3
4
363.3d
0e
17a
0b
1.7a
0a
Control
a
The insects were not sexed. Values are means. Means within a column followed by a
different letter are significantly different at P ꢀ 0.05.

LACEWING SEMIOCHEMICAL
Dispenser treatment
859
TABLE 2. Chrysopa spp. CAPTURED IN SUWON, AUGUST 2–6,
2
000a
C. cognata
C. formosa and C. phyllochroma
1
2
4
113a
48b
181.3c
0d
1.3a
0.7a
3.7a
0a
control
a
All lacewings captured were male. Values are means. Means within a column
followed by a different letter are significantly different at P ꢀ 0.05.
did not differentiate between male and female insects. The next study, which ran
from August 2 to 6, 2000, caught fewer insects but they were all found to be
male (Table 2). Although the catch means of C. phyllochroma (Table 1) and of
C. formosa and C. phyllochroma (Table 2) for the different treatments did not dif-
fer significantly, analyses of variance showed the response to 4 to be significant at
P ꢀ 0.01. The third trial studied the response of C. cognata to increasing release
rates of 1 and 2 (Table 3).
In the United Kingdom, compounds 1 and 2 were initially tested for lacewing
attraction in the field from May to July 2000 (Table 4). In this study, 11 specimens
of Nineta vittata and 19 of P. gracilis were trapped, all of which were male. Further
trials were performed during July and August 2000 (Table 5), with the addition
of traps containing both 1 and 2 at equal concentrations and both at the same
concentration as compounds in the other traps. Results showed compound 1 was
TABLE 3. Chrysopa cognata CAUGHT IN
SUWON, AUGUST 6–10, 2000
Dispenser treatment
C. cognata/trapa
Compound 1
0
1
1
5
.1 mg
.0 mg
0 mg
0 mg
8.7ab
23.0a
46.7c
183.0d
Compound 2
2
2
2
2
0.1 mg
1.0 mg
10 mg
50 mg
5.3b
5.3b
22.3a
60.3c
110.7e
1.3b
Compound 4, 10 mg
Control
a
Values are means. means within a column fol-
lowed by a different letter are significantly dif-
ferent at P ꢀ 0.05.

8
60
HOOPER ET AL.
TABLE 4. MALE Nineta vittata AND Peyerimhoffina gracilis
a
CAUGHT AT SILWOOD PARK, MAY–JULY 2000
Dispenser treatment
N. vittata
P. gracilis
1
2
3
4
9
1
1
0
0
18
0
0
1
0
Control
a
Catches with compound 1 are significant at P ꢀ 0.001.
highly effective at trapping lacewings compared to the other compounds tested
and that isomer 2 did not influence the response to 1. Of the 198 specimens of
P. gracilis trapped in this study, only one was female.
Previous work (Hyeon et al., 1968) reported the compounds now defined as
1
and 2 to be attractants of adult male C. septempunctata and male Chrysopa
japonica. We have demonstrated that the field response of adult male P. gracilis
and N. vittata can be attributed to the 1R,4S stereochemistry of 1. The preference
of adult male C. cognata is for 1R,4S over 1R,4R, but the overall preference of
C. cognata, C. formosa, and C. phyllochroma is for compound 4.
Results of EAG analysis using male P. gracilis collected from the same
trapping area showed compounds 1 and 2, and the two aphid sex pheromone com-
ponents, (1R,4aS,7S,7aR)-nepetalactol (4) and (4aS,7S,7aR)-nepetalactone (3), to
have significant activity (difference from control, P ꢀ 0.001), the first eliciting a
significantly higher response than the latter three (P ꢀ 0.05) (Table 6). The re-
sults demonstrate that the antennae of P. gracilis show a strong response to all the
compounds tested in the traps, but the largest EAG response to 1 correlates with
the finding that this compound is the most active in the field.
TABLE 5. MALE Peyerimhoffina gracilis CAUGHT
a
AT SILWOOD PARK, JULY–AUGUST 2000
Dispenser treatment
P. gracilis
1
2
3
4
82
1
5
10
95
b
1
+ 2
Control
4
a
Catches with compound 1 and 1 + 2 are significant at
P ꢀ 0.001 and differ from the control at P ꢀ 0.05.
One female was also caught in this treatment.
b

LACEWING SEMIOCHEMICAL
TABLE 6. EAG RESPONSES OF MALE Peyerimhoffina
861
gracilis TO TEST COMPOUNDS (1 mg/ml, 10 �l
APPLIED), EXPRESSED AS PERCENTAGE OF RESPONSE
TO (E)-2-HEXENAL AT THE SAME CONCENTRATION
Stimulus
Responsea
1
2
3
4
266 � 30.52a
160 � 16.38b
175 � 26.09b
185 � 13.42b
47 � 7.29
Hexane control
a
Numbers followed by a different letter are significantly dif-
ferent at P ꢀ 0.05; all treatments are significantly different
from the hexane control at P ꢀ 0.001 (N = 7).
The trapping of large numbers of P. gracilis was of interest for several reasons.
This species was not known to be native to the U.K. and recent surveys at Silwood
Park did not reveal its presence (Hollier and Belshaw, 1992, 1993; see Donato et al.,
2001). In addition, during the period of trapping, no specimens of P. gracilis were
caught in malaise traps or a suction trap present in the same area. Only in the traps
releasing cyclopentanoids were large numbers caught. This species is believed
to be relatively sedentary (Duelli, 1984) and to have a particular preference for
conifer trees, which were not present in the immediate vicinity of the field sites at
Silwood Park. Each of the four sites used in this trial was at least 20 m from a tree
of a species known to support P. gracilis and three sites were approximately 100 m
away. This suggests that either the species is more mobile and less constrained
by habitat than is currently thought or that these compounds are effective over a
considerable distance.
The presence of lacewings in traps releasing aphid sex pheromones and
closely related cyclopentanoids may suggest a kairomonal role for these com-
pounds in prey location, as has previously been proposed (Boo et al., 1998).
Lacewings are polyphagous and feed on a number of insect species, including
aphids. The fact that many aphid species use the same sex pheromone components
would allow their use as indicators of aphid prey. Lacewing larvae are insectivo-
rous predators but only a minority of species is predatory as adults; the majority
feed on honeydew, nectar, and pollen. Of the species recorded in this study, adults
of the genera Nineta and Chrysopa are insectivorous; Peyerimhoffina adults are
not. The compounds tested here, therefore, elicit a response from both predatory
and nonpredatory species, so it is unlikely that food location by adults is the cause
of the observed activity.
It is possible that lacewings are sensitive to these compounds as an aid to
the location of suitable oviposition sites. However, aphids only produce their sex

8
62
HOOPER ET AL.
pheromones during the autumn, as part of the sexual stage in the life cycle that cul-
minates in the production of overwintering eggs. In the United Kingdom, lacewings
usually overwinter as pupae. Of the British species reported here, N. vittata is
typically recorded as an adult from May to August (Plant, 1994), and so would
not normally be exposed to the aphid sex pheromones. Although P. gracilis is
present as an adult in the autumn, studies suggest that its oviposition is stimulated
by increasing day length (Grimal, 1988). Therefore, it might be expected to uti-
lize prey-produced kairomonal cues to locate suitable oviposition sites during the
spring, rather than during the autumn months. If these compounds were function-
ing as prey-location kairomones, it would be expected that the numbers trapped
would either show little sex bias or would be biased towards females looking to
oviposit near a food source, as is the case with Praon parasitoids of aphids (Hardie
et al., 1991). However, only male insects were detected in these trials, suggesting
that these compounds, or analogous structures, may play a pheromonal role in in-
traspecies sexual communication and that their similarity to aphid sex pheromones
may be simply incidental.
The possible role played by pheromones in intraspecific communication
within the Chrysopidae, and the Neuroptera as a whole, is poorly understood. The
only compound identified to date as a lacewing secretion is (Z)-4-tridecene, which
hasrecentlybeenidentifiedfromChrysoperlacarneaandappearstohavedefensive
functions (Zhu et al., 2000). Although close-range communication in the Chrysop-
idae involves species-specific substrate-borne songs (Henry, 1979), it is thought
that sex pheromones are involved in the initial mate location process (Walker et al.,
1994). However, no sex pheromones or glandular sources for pheromones have
been found. In the related Myrmeleontidae, some pheromones with a role in sexual
communication are known, but these species exhibit a lek mating system and the
pheromones eliciting a behavioral response are male-produced. Instead of func-
tioning as sex pheromones, they appear to act as aggregation pheromones, with
females responding to groups of males rather than to individuals (Yasseri et al.,
1998). Although members of the Chrysopidae appear to be solitary and do not
lek, a possible role for these compounds as aggregation pheromones cannot be
discounted. In the Myrmeleontidae, different species utilize different blends of
related compounds, providing species-specific signals (Baeckstrom et al., 1989;
Bergstrom et al., 1992). From the results obtained here, in particular the differ-
ences in response shown to the compounds by the different Chrysopa species, it
would seem possible that a similar pattern might be found in chrysopid lacewings.
The demonstration that P. gracilis antennae respond strongly to all the cyclopen-
tanoids tested, and the behavioral response of males of some species to more than
one cyclopentanoid structure, is consistent with the possibility that these, or similar
structures, may be natural pheromones.
The reasons for the appearance of P. gracilis in the United Kingdom may
be speculated as being due to accidental release, its presence previously being

LACEWING SEMIOCHEMICAL
863
overlooked, wind-borne arrival, or range expansion as a consequence of global
warming (Donato et al., 2001). The identification of cyclopentanoid monoterpenes
as semiochemicals for a number of lacewings will facilitate monitoring the chang-
ing status of the new species and may allow manipulation of natural populations of
these beneficial insects in integrated pest control strategies. Further research into
the ecological and possibly pheromonal roles of these and related compounds is
ongoing.
Supplementary Data—The CIF file for the crystal structure has been deposited at the Cambridge
Crystallographic Centre, 12 Union Road, Cambridge CB2 1EZ, UK (www.ccdc.cam.ac.uk) from which
it is available on request. The deposition number is CCDC 165880.
Acknowledgments—We wish to thank Botanix Limited for supplying essential oils from Nepeta
cataria and Dr. M. D. Ward and Dr. J. C. Jeffery from Bristol University for their assistance. Fund-
ing for this work was provided in part by the Ministry of Agriculture, Fisheries and Food. IACR-
Rothamsted receives grant-aided support from the Biotechnology and Biological Sciences Research
Council (BBSRC). J. Hardie and B. Donato are supported by the BBSRC, K. S. Boo and J. H. Park by
the Brain Korea 21 project, and R. Paul is supported by the Leverhulme Foundation.
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