Journal of Chemical Ecology, Vol. 19, No. 10, 1993

SECONDARY AMINES ISOLATED FROM VENOM GLAND OF DOLICHODERINE ANT,

Technomyrmex albipes

JOSEPH J. BROPHY, P E T E R S. C L E Z Y , * CHRISTOPHER W.F. L E U N G and PHYLLIS L. ROBERTSON School of Chemistry, University of New South Wales P.O. Box 1, Kensington, NSW 2033 Australia. Received February 17, 1993; accepted May 3, 1993 Abstract--A series of unsaturatedsecondary amines have been isolated from the dolichoderineant Technomyrmex albipes (F. Smith). The major components of the mixturehave been shownby spectroscopicproceduresto be dinon8-enylamine, and N-hept-6-enylnon-8-enamine,and these structural assignments have been confirmed by synthesis. Mass spectrometry indicates the presence of trace amounts of the bis CH amine and the C9-C11 amine. The four amines, present in total at approximately2.8 /~g/ant, are located in the gaster of the insectin a glandthat is consideredto be the venomgland although it is atypical from a morphologicalstandpoint. Key Words--Hymenoptera, Formicidae, ant, Technomyrmex albipes, unsaturated secondary amines, venom gland, GC-MS.

INTRODUCTION The ant subfamily, Dolichoderinae, has not provided a rich source of nitrogencontaining extractives, furnishing so far only the iridoid, actinidine (1), from Iridomyrmex species, e.g., L nitidiceps (Cavill et al., 1982), and a range of pyrazines isolated from the heads of I. humilis (Cavill and Houghton, 1974), the mandibular glands of I. p u r p u r e u s (Cavill et al., 1984), and the heads of other dolichoderine species (Brophy, 1989). Hence, it was of interest to observe that the black house ant, Technomyrmex albipes, which can be something of a household pest in the Sydney area, furnished, by dichloromethane extraction, *To whom correspondence should be addressed. 2183 0098-0331/93/1000-2183507.00/0 9 1993 Plenum Publishing Corporation

2184

BROPHYET AL.

two major products (present in the current extraction in approximately equal amounts), which, from their odd molecular weight and the even mass of their mass spectrometric ions, were likely to contain nitrogen. Two other nitrogenous products, present in only trace amounts, were also detected. The two major extractives were recorded in T. albipes collected from several different sites in the eastern suburbs of Sydney and from the New England Tableland of New South Wales, although the proportions of the two compounds varied.

(1)

This paper describes the isolation, structural determination, and synthesis of these novel compounds and discusses their likely glandular origin.

METHODS AND MATERIALS

Chemical Analyses. IH and J3C NMR data were obtained on a Bruker AC 300F spectrometer with chemical shifts quoted on the 6 scale relative to CHC13 as internal standard. Gas chromatography was obtained on either an OV-1 column (SCOT, 30 m x 0.5 mm) or a DB5 column (FCOT, 30 m x 0.32 mm). Both were programmed from 60~ to 250~ at 5~ Combined gas chromatography-mass spectrometry (GC-MS), both EI and CI, were performed either on: (1) an AEI MS12 mass spectrometer, (2) a Finnigan 4000 mass spectrometer, or (3) a VG Quattro mass spectrometer. In all cases the GLC column conditions were as detailed above. Accurate mass measurements were obtained on either an AEI MS 902 mass spectrometer under CI conditions, using isobutane as reagent gas, by a peak timing method (Brophy et al., 1979) or on a VG Autospec mass spectrometer. Extraction of Ants. The ants (Technomyrmex albipes) were collected on this occasion over several weeks from a location in the eastern suburbs of Sydney and were stored in twice-distilled dichloromethane. The filtration of this suspension of ants in dichloromethane yielded approximately 4500 of the ants (1.58 g) which were ground with anhydrous sodium sulfate (5 g); the solid was transferred to a Soxhlet thimble in which the mixture was extracted with dichloromethane for 24 hr. This dichloromethane extract was combined with the dichloromethane solution obtained from the original ant collection and the total extract (100 ml) was washed with saturated aqueous sodium hydrogen carbonate (2 x 50 ml) and brine (2 x 75 ml). The organic phase was dried (Na2SO4) and

ANTAMINES

2185

evaporated under reduced pressure to give a brown oil (136 mg). A pentane solution (10 ml) of this oil was extracted with aqueous sulfuric acid (0.5 M; 4 x 10 ml); the combined aqueous extracts were made alkaline with sodium hydrogen carbonate and extracted with dichloromethane (4 x 10 ml). Evaporation of the dried (Na2SO4) solution left a gum (12.5 mg) from which the following spectroscopic data were obtained: NMR 6H: 1.25-1.45 (m, CH2), 1.50-1.60 (CHaCHzCH=CH2), 2.00-2.10 (m, CHzCH=CH2) , 2.20 (broad s, NH), 2.65 (t, J = 7 Hz, CH2NH), 4.90-5.05 (m, C H = C H 2 ) , 5.75-5.90 (m, C H_~CH2). ~3c:26.81,27.29, 28.78, 28.88, 29.07, 29.36, 29.73, 33.70, 33.79, 49.61 (CH2), 114.21, 114.41 (=CH2), 138.91, 139.16 ( - - C H = ) . GC/MS (EI) indicated the presence of two major amines and two other amines in trace amounts (Figure 1). The main components showed the following principal ions: (3) m/z(%) 126(100%), 154(87), 196(18), 237(8); (2) re~z(%), 154(100%), 224(15), 264(3), 265(2); (4) m/z(%), 154(100%), 182(55), 293(0.5); (5) m/z(%) 182(100%), 252(5), 321(0). Accurate mass measurements, under CI (isobutane) conditions gave the following results for the protonated parent ions of the two principal compounds: (3) found 238.2518 (C16I-I32N required 238.2534); (2) found 266.2856 (CIsH36N required 266.2847). Non-8-enoic Acid (6). Magnesium turnings (514 nag) were suspended in sodium dried ether (10 ml) and 8-bromoct-l-ene (3.8 g) in dried ether (10 ml) was added portionwise. With the first addition, the mixture was stirred vigorously, with warming, to initiate the reaction. After the addition was complete, the mixture was refluxed for 1 hr, diluted with ether (20 ml) and cooled to - 10~ before excess Dry Ice was added slowly over about 10 min. The mixture was stirred for 15 min, after which aqueous sulfuric acid (50%; 10 ml) was added slowly with cooling, followed by water (50 ml). The product was isolated by extraction with ether (3 • 100 ml), the ethereal phase concentrated (100 ml), and the acid purified by extraction into aqueous sodium hydroxide solution (10%; 2 • 50 ml). The aqueous phase was acidified to pH 1 with hydrochloric acid (10 M) and the acid returned to ether (3 • 100 ml). The organic phase was dried (Na2SO4) and evaporated under reduced pressure to give the acid (1.98 g; 64%) as a pale pink liquid, bp 133-134~ at 3 torr (Ansell and Whitfield, 1971) (bp 110-114~ at 2.5 torr); NMR 6H: 1.25-1.40 (6H, m, 3 • CH2), 1.59 (2H, m, CH2CH2CH~-CH2), 2.00 (2H, m, CH2CH=CH2), 2.29 (2H, t, J = 7.5 Hz, CH2COOH), 4.85--4.98 (2H, m, CH=CH2), 5.74 (1H, qt, Jcis = 10.2 Hz, J~rans = 17.0 Hz, JCH2 = 6.7 Hz, CHzCH~--CH2). 6c: 24.42, 28.54, 28.56, 33.55, 28.73, 33.89, (CH2), 114.11 (=CH2), 138.57 ( = C H - - ) , 180.44 (CO). Hept-6-enenitrile (7). A solution of potassium cyanide (3.7 g) in water (7 ml) was added to 6-bromohex-l-ene (6.3 g) dissolved in ethanol (95 %; 28 ml). The mixture was refluxed for 20 hr, cooled, and diluted with ether (100 ml). The mixture was washed successively with water (2 • 50 ml), hydrochloric

2186

BROPHY

100-

ET AL.

126

806O4020-

,

i..,i , ~,

40

,;..~..L.

60

~-~;~ ~.,~-.... ~

80

100

120

140

100-

,

~,....,....~6 .....~i -~ . ,1 160

180

200 m/z

220

2...,...,...,...,...,..

" 37 240

260

280

300

320

340

154

8O-

(2)

60.40,. 2O-

L

o-

. _ .nil_

.z,a .........

h

' ' 1 ' ' ' 1 ' ' ' 1 ' ' '

40

60

80

I ' ' ' 1 '

100

120

,

224

..........

L.......................

265

' 1 ' ' ' 1 ' ' ' 1 ' ' ' 1 ' ' ' 1 ' ' ' 1 ' ' ' 1 ' ' ' 1 ' ' ' 1 ' ' ' 1 ' '

140

160

180

200

220

240

260

280

300

320

340

m/z I00.-

72

154

8O-

(4)

6O.-

i

4O-

,182 126

I

20.-

.,,L ,,, '''

I'

40

i,,. ,LJ,I , ,Jl .., ,,, ,,, h,~ ,,,,J .......

" 1 ' ' ' 1 ' ' ' 1 ' ' ' 1 ' '

60

80

100

120

140

,

,,.

i, .,., ...........

,

I ' ' ' 1 ' ' ' 1 ' ' ' 1 ' ' ' 1 ' ' ' 1 ' ' ' 1 ' ' ' 1 ' ' ' 1 ' ' ' 1 ' '

160

180

200

220

240

260

280

300

320

340

m/z 100-

182

72

80-

(5) 6O-

40_

II

LL,

20.-

i

0...

~,]l

' ' ' 1 '

40

.

60

'1''

80

~ 1 ' ' ' 1 ' ' ' 1 ' ' ' 1

100

120

140

'~'

160

. . . . . . a..J L.L] . . . . . . . . . . . . . . . . J. . . . . . . . . . . . . . . ' ' ' 1 ' ' ' I ' ' ' 1 ' ' ' I ' ' ' 1

180

20()

220

240

260

0 ........ ' ' ' 1 '''

280

300

I ' ' ' 1

320

'~

340

m/z

F i t . 1~ Mass spectra of the four amines detected in the venom gland of

albipes.

Technomyrmex

ANTAMINES

2187

acid (2 M; 2 • 50 ml), saturated aqueous sodium hydrogen carbonate solution (2 • 50 ml), and finally water (2 • 50 ml). The organic phase was dried (Na2SO4) and the solvent removed under reduced pressure to leave the nitrile (3.5 g; 83%) as a pale yellow liquid, bp 109-112~ at 73 ton" (Vasil'eva and Freidlina, 1966) (bp 82~ at 20 ton"); NMR 6H: 1.40-1.60 (4H, m, 2 • CH2), 1.98-2.08 (2H, m, CH2CH=CH2), 2.28 (2H, t, J = 7 Hz, CH2CN), 4.885.00 (2H, m, C H = C H 2 ) , 5.71 (1H, qt, Jcis = 10.2 Hz, J, . . . . 17.0 Hz, Jcm_ = 6.7 Hz, CH2CH=CH2). 6c: 16.60, 24.39, 27.36, 32.44 (CH2), 114.93 (=CH2), 119.42 (CN), 137.30 ( - - C H = ) . Non-8-enenitrile (8). The nitrile (2.9 g; 85%) was obtained as a yellow liquid, bp 133-135~ at 48 ton", by refluxing a solution of 8-bromoct-l-ene (4.77 g) in aqueous ethanol containing potassium cyanide (2.32 g). The procedure followed that outlined for the lower homolog. NMR 6H: 1.22-1.48 (6H, m, 3 • CH2), 1.55-1.66 (2H, m, CH__2CH2CH=CH2), 1.95-2.05 (2H, m, CH__2CH=CH2), 2.29 (2H, t, J = 7.1 Hz, CH2CN ), 4.88-5.00 (2H, m, CH=CH2), 5.75 (1H, qt, Jcis = 10.3 Hz, Jt ..... = 17.0 Hz, JcH2 = 6.7 Hz, CH2CH=CH2). 6c: 16.88, 25.13, 27.98, 28.28, 28.32, 33.36 (CH2), 114.30 (=CH2), 119.59 (CN), 138.48 ( C H = ) . Hept-6-enamine (9). A well-stirred mixture of sodium (3.25 g) and toluene (40 ml) was heated to reflux whereupon the sodium liquefied. A solution of hept-6-enenitrile (3.5 g) in a mixture of absolute ethanol (15 ml) and toluene (15 ml) was then added slowly over 30 min to the refluxing mixture of sodium in toluene. Further absolute ethanol was added to destroy excess sodium, after which the mixture was refluxed for 1 hr, cooled, and water (50 ml) added, followed by hydrochloric acid (10 M; 25 ml). The mixture was extracted with ether (3 • 80 ml) and the aqueous phase evaporated to dryness under reduced pressure to free the amine salt of residual ethanol. The residue was then treated with aqueous sodium hydroxide (10%; 30 ml) and the free base returned to ether by extraction (3 • 80 ml). The ethereal layer was dried ( N a 2 S O 4 ) and the solvent removed to give the amine (1.53 g; 42%) as a pale yellow liquid, bp 74-78~ at 48 torr. MS 113.1194 (CTHIsN requires 113.1205) NMR 6H: 1.101.30 (8H, 3 • CH2, NH2), 1.88 (2H, m, CH2CH=CH2), 2.50 (2H, t, J = 7 Hz, CH2NH2), 4.73-4.83 (2H, m, C H = C H 2 ) , 5.62 (1H, qt, Jci, = 10.3 Hz, Jtrans = 17.0 Hz, Jcm = 6.7 Hz, CH2CH=CH2). 6c: 26.00, 28.40, 33.27, 33.36, 41.76 (CH2), 113.91 (=CH2), 138.48 ( - - C H = ) . Non-8-enamine (10). The amine (0.69 g; 32%), bp 130-136~ at 76 ton", was obtained by reduction of non-8-enenitrile (2.08 g) in absolute ethanoltoluene (1 : 1, 24 ml) with sodium (2.0 g) suspended in toluene (25 ml) according to the procedure detailed for the lower homolog. MS 141.1515 (C9H19N requires 141.1518) NMR 6H: 1.00-1.50 (12H, 5 • CH2, NH2), 1.92 (2H, dt, CH2CH=CH2), 2.55 (2H, t, J = 7 Hz, CH2NH2), 4.78-4.90 (2H, m, CH=CH2), 5.68 (1H, qt, Jcis = 10.3 Hz, "It. . . . 17.3 Hz, Jcm = 6.7 Hz, =

=

2188

BROPHY ET AL.

CH2CH=CH2): 6c: 26.65, 28.67, 28.89, 29.14, 33.58, 42.00, (CH2), 113.95 (=CH2), 138.88 ( - - C H = ) . N-Hept-6-enylnon-8-enamide (11). Thionyl chloride (2.5 ml; freshly distilled) was added over 10 min to non-8-enoic acid (1.10 g) heated on a water bath maintained at 50-60~ After gas evolution was complete, the temperature of the water bath was raised to 80-90~ and kept at this temperature for 30 min. Excess thionyl chloride was removed under reduced pressure and the residual acid chloride (1.03 g) was dissolved in anhydrous ether (10 ml). A solution of hept-6-enamine (1.06 g) in anhydrous ether (5 ml) was added portionwise to the acid chloride, after which the reaction mixture was washed successively with water, aqueous sodium hydroxide solution, water, dilute hydrochloric acid, and water. The ethereal solution was dried (Na2SO4) and the solvent removed to give a yellow oil (1.31 g) which, after chromatography on silica (60H, Merck Art 7736; eluting with dichloromethane), yielded the amide (1.02 g; 69%) as a white solid mp < 20~ MS (EI) 251.2251 (C16H29NO requires 251.2249), 252(MH +, 3%), 251(1), 210(27), 196(8), 182(6), 168(58), 140(85), 126(30), 114(50), 112(65), 100(48), 69(68), 55(100). NMR fill: 1.25-1.55 (12H, m, 6 X CH2) , 1.60-1.70 (2H, m, CHzCH2CH=CH2), 2.00-2.10 (4H, m, 2 x CH2CH=CH2), 2.16 (2H, t, J = 7.5 H z , C O C H 2 ) , 3.20-3.28 (2H, m, CH__2NH), 4.90-5.03 (4H, m, 2 x CH=CH2), 5.50 (1H, br s, NH), 5.78 (1H, qt), 5.79 (1H, qt) (Jcis = 10.3 Hz, Jt .... = 17.0 Hz, JcH2 ---- 6.7 Hz, 2 • CH2CH=CH2). 8c: 25.71, 26,28, 28.43, 28.64, 28.72, 29.05, 29.41, 30.50, 33.58, 36.67, 39.32 (CH2), 114.15, 114.35 (=CH2), 138.59, 138.83 (2 x - - C H = ) , 173.07 (CO). N-Non-8-enylnon-8-enamide (12). Non-8-enoic acid (517 mg) was converted into its acid chloride by reaction with freshly distilled thionyl chloride (2 ml) as described previously, and a solution of this derivative (455 mg) in anhydrous ether (10 ml) was treated portionwise with non-8-enamine (561 mg) in anhydrous ether (5 ml). The mixture was worked up as described for the lower homolog to give the amide (424 mg, 46%), mp < 25~ MS (EI) 279.2565 (C18H33NO requires 279.2562), 280(MH +, 10%), 279(9), 250(5), 238(13), 210(8), 196(45), 168(20), 156(8), 142(20), 121(18), 100(40), 86(30), 69(55), 55(100). NMR 6H: 1.20-1.40 (14H, m, 7 X CH2) , 1.40-1.50 (2H, m), 1.501.70 (2H, m) (2 • CHzCHzCH=CH2) , 2.02 (4H, m, 2 x CH2CH=CH2) , 2.14 (2H, t, J = 7.5 Hz, C O C H 2 ) , 3.22 (2H, dt, CH2NH), 4.80-5.00 (4H, m, 2 • CH=CH2), 5.60 (1H, br s, NH), 5.78 (1H, qt), 5.79 (1H, qt) (Jcis = 10.3 Hz, Jt .... = 17.0 Hz, JCH2 = 6.7 Hz, 2 x CH2CHz=CH2). 6r 25.76, 26.83, 28.68, 28.74, 28.95, 29.09, 29.59, 33.65, 33.69, 36.77, 39.47 (CH2) , 114.18, 114.22 (=CH2), 138.94, 139.01 (2 • - - C H = ) , 173.19 (CO). N-Hept-6-enylnon-8-enamine (3). A solution of N-hept-6-enylnon-8-enamide (266 mg) in anhydrous ether (5 ml) was added to a suspension of lithium aluminium hydride (65 mg) in anhydrous ether (10 ml), maintained over nitro-

ANTAMINES

2189

gen. The mixture was refluxed for 5 hr, cooled, and excess lithium aluminium hydride destroyed by the careful addition of water. Hydrochloric acid (5 M) was then introduced to acidify the reaction mixture, which was extracted with ether (3 • 30 ml). The combined ethereal extracts were dried (Na2SO4) and evaporated to yield the amine hydrochloride (272 mg) as a white solid. The solid was made alkaline with aqueous sodium hydroxide (10%; 20 ml) and the aqueous mixture extracted with ether (3 • 40 ml); evaporation of the solution left the amine (231 mg; 73%) as a pale yellow liquid, bp 164-167~ at 3 torr, which was identical by gas chromatography and mass spectrometry with the natural material. MS (El) 237(M +, 10%), 208(7), 196(16), 180(11), 154(85), 140(10), 126(100), 55(15). NMR 6H: 1.25-1.50 (16H, m, 8 • e l l 2 ) , 1.70 (1H, hr s, NH), 1.98-2.08 (4H, m, 2 x C H 2 C H = C H 2 ) , 2.56 (2H, t, J = 7 Hz), 2.57 (2H, t, J = 7 Hz) (CH2NHCH2) , 4.88-5.02 (4H, m, 2 • C H = C H 2 ) , 5.78 (2H, qt, Jcis = 10.3 Hz, Jtrans = 17.0 Hz, Jcm = 6.7 Hz, 2 • C H z C H = C H 2 ) . (3c: 26.85, 27.32, 28.81, 28.83, 29.02, 29.37, 29.92, 30.05, 33.67, 33.73, 49.98, 50.03 (CH2) , 114.10, 114.25 (=CH2), 138.90, 139.08 (--CH=). Dinon-8-enylamine (2). A solution of N-non-8-enylnon-8-enamide (340 mg) in anhydrous ether (15 ml) was reduced under nitrogen with lithium aluminium hydride (100 mg) as described for the lower homolog. The reaction mixture was worked up in a similar manner to yield the amine (274 mg, 38%) as a pale yellow liquid, bp 189-195~ at 9 torr, identical by gas chromatography and mass spectrometry with the natural product. MS (EI) 265(M +, 3), 264(4), 224(15), 210(4), 180(3), 168(9), t54(100), 140(6), 126(7). NMR 6n: 1.201.50 (21H, m, 10 • CH2, NH), 2.00 (4H, m, 2 • CH2CH=CH2), 2.56 (4H, t, J = 7 Hz, CH2NHCH2) , 4.70-5.00 (4H, m, 2 x C H = C H 2 ) , 5.78 (2H, qt, Jcis = 10.3 Hz, J, .... = 17.0 Hz, Jcn2 = 6.7 Hz, 2 X C H 2 C H = C H 2 ) . ~c: 27.35, 28.86, 29.05, 29.40, 30.15, 33.76, 50.12 (ell2) , 114.12 (~-CH2), 139.11 (--CH=).

RESULTS AND DISCUSSION A GC-MS study of the dichloromethane extract of whole ants revealed two prominent components, present in approximately equal amounts. The less volatile component fumished a small molecular ion at 265 D with its base peak at m/z 154; the more volatile component, which was marginally the greater compound present, displayed a weak molecular ion at 237 D and provided prominent fragment ions at m/z 154 and 126. Accurate mass determinations indicated molecular formulae of C18H35N and C16H31N, respectively, for these compounds. When a pentane solution of the crude extract was washed with dilute aqueous

2190

BROPHY ET AL,

sulfuric acid, the aqueous phase then made alkaline, and the bases returned to dichloromethane, the latter extract was essentially free of any material apart from the two major nitrogenous compounds referred to above. This fraction was pure enough to give good quality NMR spectra. ~3C NMR spectroscopy revealed the absence of methyl groups and quaternary carbons. Methine carbons at 139 ppm and methylene carbons at 114 ppm suggested the presence of terminal double bonds, while a methylene carbon at 49 ppm indicated a carbon adjacent to nitrogen. A cluster of other methylene carbons were present with chemical shifts ranging between 26 and 33 ppm. Olefinic protons were also clearly evident from the 1H NMR spectrum. The summation of these data suggested that the two major natural products were the unsaturated secondary amines (2) and (3) and their principal fragmentation is outlined in Scheme 1. To confirm these formulations, the amines were synthesized. 8-Bromoct1-ene was converted into the corresponding nitrile (8) by treatment with potas-

CH2=CH(CH2)6-CH2-NH-(CH2)7CH=CH 2

~.~

(2)

+ CH2=CH(CH2)7-NH=CH 2 m/z 154

+ CH2=CH(CH2)5-NH=CH 2

CH2=CH(CH2)6-CH2-NH-CH2(CH2)4CH=CH 2

(3) m/z 126

CH2=CH(CH2)8-CH2-NH-(CH2)9CH=CH 2

~.~

(5)

§ CH2=CH(CH2)9-NH=CH 2 m/z 182

+ CH2=CH(CH2)8-CH2-NH-CH2(CH2)6CH=CH2

CH2=CH(CH2) 7-NH=CH 2

(4) m/z 154 SCHEME. 1.

ANTAMINES

2191

sium cyanide, and this derivative was reduced with sodium in ethanol to produce non-8-enamine (10). Non-8-enoic acid (6) was obtained from the above bromide by carbonation of its Grignard derivative. Reaction of the acid chloride derivative of non-8-enoic acid with non-8-enamine furnished the amide (12), which was reduced with lithium aluminium hydride in ether to give the C18 secondary amine (2). The Cj6 secondary amine (3) was produced in a similar manner utilizing hept-6-enamine (9) to produce the intermediate amide (11) by reaction with the chloride derivative of non-8-enoic acid. Hept-6-enamine (9) was obtained by reduction of the cyanide (7), derived from the commercially available 6-bromohex-l-ene. These reactions are summarized in Scheme 2.

CN"

CH2=CH(CH2) 6Br

l

~-~ ~.~

Na CH2=CH(CH2)6CN

ethanol

,.~ CH2=CH-(CH2)6CH2NH 2

(8)

l. Mg

(10)

2. CO2 3. H+/H20

SOCI2

CH2=CH-(CH2) 6COOH

~.. CH2=CH-(CH2)6COCI

(s) CH2=CH-(CH2)6CH2NHCO(CH 2)6CH=CH 2 CH2=CH(CH2) 4-Br

(12) 1. LiAIH 4 2. H2OPH+

CH2=CH(CH2)4CN

(7)

CH2=CH(CH2)7NH(CH2)7-CH=CH 2

(2) Na/ethanol

CH2=CH(CH2)4CH2NH 2

(9)

CH2=CH(CH2)4CH2NHCO(CH2)6CH=CH2 (11)

1. LiA1H4 CH2=CH(CH2)5NH(CH2)7CH=CH 2 2. H20/H+

(a) SCHEME. 2.

2192

BROPHY ET AL.

Separate examination of disarticulated heads, thoraces, and gasters indicated that the amines were localized in the gaster of Technomyrmex. There are a number of glandular-type structures within the gaster, any one of which might be a focus for amine production. Among them is an unusual gland in Technomyrmex, which must be accepted as a venom gland on morphological grounds, although it does not conform to the characteristic size reduction of this structure within the Dolichoderinae. Dichloromethane extraction of 23 of these glands and 51 glands subsequently isolated by microdissection and analyzed by GC-MS confirmed the presence within them of amines (2) and (3), now recognized as venom constituents. In addition, this study of the gland extracts revealed the presence of two other amines at levels of approximately 1/100 to 1/1000 of the major compounds. The mass spectra of these additional components (the spectra of these two compounds shown in Figure 1 are background-subtracted spectra) indicated that they were probably the C1~-C9 (C2oH39 N ) and the bis Ctl (C22H43 N ) amine analogs (4) and (5) of the major components, although there was insufficient material to carry this characterization further. No trace of the cyclopentanoid monoterpene alkaloid, actinidine (1), characteristic of dolichoderine ant anal (i.e., pygidial) glands, was found in the venom gland, although its presence was confirmed in the extract of the whole gaster of the ant. Acknowledgments--Financial support from the Australian Research Council is gratefully acknowledged. We are grateful to Mr. D. Nelson, of the Australian Jockey Club Laboratories, and Mr. R. Lidgard, of the Biomedical Mass Spectrometry Unit of this University, for some of the mass spectrometric data and to Dr. C.J.R. Fookes, CSIRO Division of Coal and Energy Technology, for his helpful discussions. REFERENCES ANSELL, M.F., and WmTF1ELD, G.F. 1971. Friedel-Crafls reaction of n-alkenoic acids and wchloroalkanoic acids. J. Chem. Soc. (C) 1098-1109. BROPHY, J.J. 1989. Pyrazines obtained from insects: their source, identification, synthesis and function, pp. 221-273, in Atta-Ur-Rahman (ed.). Studies in Natural Products Chemistry 5: Structure Elucidation--Part B. Elsevier, Amsterdam. B~OPHY, J.J., GOLDSACK,R.J., LIDGARD, R.O., MELLY, D.P., and NELSON, D. 1979. Elemental compositions from low resolution magnetic mass spectrometers. Lab. Practice 00:615-619. CAVILL, G.W.K., and HOUGHTON,E. 1974. Volatile constituents of the Argentine ant, lridomyrmex humilis. J. Insect Physiol, 20:2049-2059. CArreL, G.W.K., ROBERTSON, P.L., BROPHY, J.J., CLARK, D.V., DUKE, R., ORTON, C.J., and PLANT, W.D. 1982. Defensive and other secretions of the Australian cocktail ant, Iridomyrmex nitidiceps. Tetrahedron 38:1931-1938. CArreL, G.W.K., ROBERTSON, P.L., BROPHY, J.J., DUKE, R., MCDONALD, J., and PLANT, W.D. 1984. Chemical ecology of the meat ant, Iridomyrmexpurpureus sensfi strict6. Insect Biochem. 14:505-513. VASm'EVA, E.J., and FREIDLINA, R. KH. 1966. Chemical transformations of nitfiles of ~0-chlorocarboxylic acids, lzv. Akad. Naauk SSSR Ser. Khim. 263-267. Chem. Abst. 64:17417h.