Sex Attractant Pheromone of the Luna Moth, Actiasluna (Linnaeus)

Supplementary Information

Sex Attractant Pheromone of the Luna Moth, Actiasluna (Linnaeus)

Jocelyn G. Millar1,*, Kenneth F. Haynes2, Aaron T. Dossey3, J. Steven McElfresh1, and Jeremy D. Allison4

1Departments of Entomology and Chemistry, University of California, Riverside CA 92521, USA

2Department of Entomology, University of Kentucky, Lexington, KY 40546, USA

3All Things Bugs LLC, 120 Mark Twain Circle, APT# L 5, Athens, GA 30605, USA

4Natural Resources Canada - Canadian Forest Service, Great Lakes Forestry Centre, Sault Ste. Marie, ON, P6A 2E5, Canada

General synthesis details

THF was dried by distillation from sodium/benzophenone ketyl under argon. All other solvents were Optima grade (Fisher Scientific, Pittsburgh PA, USA) and were used as received. Mass spectra were taken with a Hewlett-Packard (HP) 6890 GC (Hewlett-Packard, now Agilent, Avondale, PA, USA) interfaced to an HP 5973 mass selective detector, in EI mode (70 eV) with helium carrier gas. The GC was equipped with a DB17-MS column (17% phenylmethylsiloxane, 25 m × 0.20 mm i.d., 0.33 μm film). 1H and 13C NMR spectra were taken with a Varian INOVA-400 (400 and 100.5 MHz, respectively) spectrometer (Palo Alto, CA, USA), as CDCl3 solutions. 1H NMR chemical shifts are expressed in ppm relative to residual CHCl3 (7.27 ppm) and 13C NMR chemical shifts are reported relative to CDCl3 (77.16 ppm). Reactions with air- or water-sensitive reagents were carried out in oven-dried glassware under argon. Solutions of crude products were dried over anhydrous Na2SO4, and concentrated by rotary evaporation under reduced pressure. Crude products were purified via vacuum flash column chromatography on silica gel.

Synthesis of (11Z)-11-octadecenal (7).

A solution of 10-chlorodecanol 1(25 g, 130 mmol), dihydropyran (13.7 ml, 150 mmol), and p-toluenesulphonic acid (PTSA, 0.5 g) in 200 ml dry ether was stirred overnight at room temp. The solution was then extracted twice with 1M aqueous NaHCO3 and once with brine, then dried and concentrated. The residue was purified by Kugelrohr distillation, bp ~95ºC/0.05 mm Hg, yielding 32.2 g of chloride 2 as a colorless oil (89%).

Lithium acetylide-ethylenediamine complex (1.84 g, 20 mmol) and NaI (0.25 g, 1 mmol) were added to 20 ml dry DMSO in a dry flask under Ar, and chloride 2 (4.07 g, 14.7 mmol) was added in portions over 10 min. The resulting slurry was stirred at room temp for 6 h, then poured into cold water and extracted twice with hexane. The combined hexane extracts were washed with brine, dried, and concentrated. The residue was purified by Kugelrohr distillation, bp~100ºC/0.07 mm Hg, yielding the terminal acetylene 3as a colorless oil (3.14 g, 80%).

The terminal acetylene 3(1.07 g, 4 mmol) and ~50 mg triphenylmethane were dissolved in 20 ml THF in a dry flask under Ar, the solution was cooled in an ice-bath, and treated dropwise with BuLi in hexanes (2.5 M, 1.9 ml) until a pink-red color persisted. The mixture was stirred for 15 min, then hexyl iodide (0.67 ml, 4.5 mmol) was added dropwise. The solution was warmed to 50ºC and stirred overnight, then cooled, quenched with water and extracted with hexane. The hexane solution of crude 4was dried and concentrated, then taken up in 20 ml MeOH, PTSA (~50 mg) was added, and the solution was stirred overnight. The solution was then poured into 100 ml 1M aqueous NaHCO3 and extracted with hexane. The hexane solution was washed with brine, dried, and concentrated. The residue was Kugelrohr distilled, taking a forerun (THP-protected MeOH) at <60ºC at 0.1 mm Hg. After changing the collection bulb, alcohol 5was distilled over at ~120ºC at 0.07 mm Hg, yielding 0.89 g of product (84%). This was recrystallized from 12 ml hexane at -20ºC, yielding 0.60 g of the desired alcohol 5(98% pure by GC). 1H NMR (CDCl3): 3.60 (t, 2H, J = 6.7 Hz), 2.10 (t, 4H, J = 7.0 Hz), 1.53 (m, 2H), 1.44 (m, 4H), 1.4-1.2 (m, 18 H), 0.86 (t, 3H, J = 6.96 Hz). 13C NMR (CDCl3): 80.43, 63.23, 32.98, 31.57, 29.74, 29.66, 29.61, 29.33, 29.03, 28.72, 25.92, 22.76, 18.94, 14.23 ppm. MS (m/z: abundance): 210 (1), 192 (1), 163 (2), 149 (4), 138 (10), 135 (10), 124 (65), 109 (36), 96 (52), 95 (83), 82 (67), 81 (100), 79 (44), 67 (95), 55 (66), 43 (31), 41 (59).

Ni(OAc)2· 6H2O (62 mg, 0.25 mmol) was dissolved in 5 ml 95% EtOH, and the solution was degassed under Ar. The flask was fitted with a balloon of H2, and with vigourous stirring, 0.25 ml of a solution of NaBH4 (1M, prepared by dissolving 0.4 g NaBH4 in 9.5 ml EtOH and 0.5 ml 1 M aqueous NaOH and filtering) was added quickly. The resulting black slurry was stirred for 15 min, then 0.05 ml ethylenediamine was added, followed by a solution of alkynol5 (0.3 g, 1.1 mmol) in 2 ml EtOH. The mixture was stirred until all starting material had been consumed (70 min), then filtered through a pad of Celite (top layer) and activated charcoal (bottom layer). The pad was rinsed well with EtOH, and the filtrate was diluted with 300 ml 1M HCl and extracted with hexane. The hexane layer was washed with saturated aqueous NaHCO3 and brine, dried, and concentrated, and the residue was Kugelrohr distilled, bp~120/0.07 mm Hg, yielding alkenol6 (0.18 g, 60%). Attempted recrystallization from hexane at -20ºC to remove traces of over-reduced material failed, so the product was used as is in the next step. 1H NMR (CDCl3): 5.36 (m, 2H), 3.65 (t, 2H, J = 6.6 Hz), 2.02 (m, 4H), 1.59 (m, 2H), 1.4-1.23 (m, 22H), 0.89 (t, 3H, J = 7.0 Hz). 13C NMR (CDCl3): 130.1, 63.3, 33.04, 32.0, 20.0, 29.8, 29.6, 29.5, 29.2, 27.4, 25.9, 22.9, 14.3.

Alkenol6 (50 mg, 0.19 mmol) was added to a slurry of pyridinium dichromate (70 mg, 0.19 mmol) in 1 ml methylene chloride and the mixture was stirred for 2 h at room temp. The mixture thenwas diluted with 5 ml hexane, stirred for 10 min, and filtered through a layered plug of activated charcoal (top) and celite (bottom), rinsing well with hexane. After concentration, the residue was purified by vacuum flash chromatography, eluting with 5% EtOAc in hexane, yielding (Z)-11-octadecenal7 (27 mg, 54%). 1H NMR (CDCl3): 9.76 (t, 1H, J = 1.9 Hz), 5.34 (m, 2h), 2.41 (td, 2H, J = 7.4, 1.9 Hz), 2.01 (m, 4H), 1.62 (m, 2H), 1.37-1.22 (m, 20H), 0.87 (t, 3H, J = 6.9 Hz). MS (m/z: abundance) (Fig. S3): 266 (M+, 4), 248 (9), 237 (1), 223 (2), 166 (3), 149 (6), 135 (12), 121 (20), 111 (23), 109 (19), 98 (40), 97 (37), 96 (32), 95 (41), 83 (47), 82 (36), 81 (48), 69 (63), 67 (54), 55 (100), 41 (79).

Synthesis of (6E)-6-octadecenal

(6E)-6-octadecenoic acid 8(petroselaidic acid, 100 mg, 0.35 mmol; Sigma Chemical Co., St. Louis MO, USA) was added to an ice-cooled slurry of LiAlH4 (80 mg, 2 mmol) in 5 ml dry THF under Ar. The cooling bath was removed, and the mixture was stirred for 4 h at room temp, then quenched by sequential addition of water (84 µl), 20% NaOH (63 µl), and water (294 µl). The resulting slurry was stirred for 30 min, then filtered through a pad of celite. Concentration of the filtrate gave alcohol 9 as a white solid, which was used without further purification in the next step.MS (m/z: abundance): 268 (M+, trace), 250 (12), 222 (3), 194 (3), 180 (2), 166 (3), 152 (4), 138 (7), 137 (9), 123 (15), 109 (27), 96 (65), 95 (68), 82 (98), 81 (65), 67 (100), 55 (76), 43 (57), 41 (71).

Alkenol9 (49 mg, 0.18mmol) was added to a slurry of pyridinium dichromate (300 mg, 0.79mmol) and 300 mg powdered 4Å molecular sieve. The mixture was stirred for 2 h at room temp, then diluted with 20 ml ether, stirred 10 min, and filtered through a pad of celite, rinsing with ether. After concentration, the residue was purified by vacuum flash chromatography on silica gel, eluting with 5% EtOAc in hexane, yielding (E)-6-octadecenal10 (19 mg, 39%). The ~50% unreacted starting material (estimated by TLC of the crude product) was not recovered. 1H NMR (CDCl3):  9.74 (t, 1H, J = 1.9 Hz), 5.37 (m, 2H), 2.40 (td, 2H, J = 7.4, 1.9 Hz), 1.96 (m, 4H), 1.62 (quint, 2H, J = 7.6 Hz), 1.35-1.22 (m, 20H), 0.86 (t, 3H, J = 6.9 Hz).MS (m/z: abundance) (Fig. S4): 266 (M+, 1), 248 (22), 222 (6), 194 (2), 180 (5), 166 (6), 152 (4), 149 (6), 135 (12), 121 (26), 112 (46), 98 (72), 97 (67), 84 (44), 83 (55), 82 (47), 81 (54), 79 (47), 70 (44), 69 (46), 67 (63), 57 (45), 55 (95), 43 (76), 41 (100).

Synthesis of (6E,11Z)-6,11-octadecadienal

The alcohol function of 4-pentyn-1-ol 11was protected as the THP derivative12, the terminal alkyne was alkylated with hexyl iodide to give 13, and the THP protecting group was removed as described above in the syntheses of 2, 4, and 5, respectively, and the resulting 4-undecyn-1-ol 14was Kugelrohr distilled, taking a forerun <50º at 0.4 mm Hg to remove THP-protected MeOH, followed by distillation of the product14 (bp~95ºC/1.3 mm Hg; 76% over 3 steps). 1H NMR (CDCl3)  3.76 (t, 2H, J = 6.0 Hz), 2.29 (tt, 2H, J = 7.0, 2.2 Hz), 2.14 (tt, 2H, J = 7.0, 2.4 Hz), 1.74 (tt, 2H, J~6.8 Hz), 1.67 (br s, OH), 1.44-1.52 (m, 2H), 1.24-1.40(m, 6H), 0.89 (t, 3H, J = 6.8 Hz).13C NMR (CDCl3)  81.39, 79.45, 62.32, 31.81, 31.57, 29.23, 28.76, 22.77, 18.93, 15.67, 14.24 ppm.

Alkynol14 (2.52 g, 15 mmol) was reduced with P2 nickel and hydrogen, as described for 6 above, and the resulting alkenol15 was purified by Kugelrohr distillation (bp~85º/0.6 mm Hg; 2.28 g, 89%). 1H NMR (CDCl3): 5.39 (m, 2H), 3.67 (t, 2h, J = 6.6 Hz), 2.13 (m, 2H), 2.04 (m, 2H), 1.64 (tt, 2H, J ~6.8 Hz), 1.25-1.40 (m, 8H), 0.89 (t, 3H, J = 7.2 Hz). 13C NMR (CDCl3): 131.05, 129.00, 62.93, 32.87, 31.99, 29.90, 29.20, 27.44, 23.83, 22.86, 14.30 ppm.

Mesyl chloride (1.37 g, 12 mmol) was added dropwise to an ice-cooled solution of Et3N (1.8 ml, 13 mmol) and alkenol15 (1.7 g, 10 mmol) in 30 ml CH2Cl2, and the mixture was stirred until all the starting alcohol had been consumed (2 h). The solution was then poured into 100 ml water, shaken, and the CH2Cl2 layer was removed. The aqueous layer was extracted again with CH2Cl2 and the combined organic phases were washed with 1 M HCl and brine, then dried and concentrated to give the crude mesylate as a yellow oil. The oil was taken up in 30 ml dry acetone, LiBr (3 g, 35 mmol) was added, and the mixture was stirred at 55ºC for 2 h. The mixture was cooled, poured into 120 ml water, and extracted with hexane. The hexane layer was washed with saturated aqueous NaHCO3 and brine, dried, concentrated, and Kugelrohr distilled (bp~75-80º/1.2 mm, 2.07 g, 89%) to give bromide 16. 1H NMR (CDCl3):5.45 (dtt, 1H, J = 11.2, 6.8, 2.2 Hz), 5.34 (dtt, 1H, J = 11.2, 7.2, 1.4 Hz), 3.42 (t, 2H, J = 6.6 Hz), 2.20 (br. q, 2H, J ~7 Hz), 2.06 (br. q, 2H, J ~ 6.6 Hz), 1.92 (quin, 2H, J = 7.0 Hz), 1.24-1.40 (m, 8H), 0.90 (t, 3H, J = 6.8 Hz). 13C NMR (CDCl3): 132.02, 127.56, 33.63, 32.89, 31.99, 29.87, 29.20, 27.53, 25.85, 22.86, 14.33 ppm.

An oven-dried flask was charged with bis(cyclopentadienyl)zirconium (IV) chloride hydride (1 g, 3.88 mmol), and evacuated and refilled with Ar 6 times. Dry THF (4 ml) was then added, followed by THP-protected 6-heptyn-1-ol 17(0.392 g, 3.88 mmol), and the mixture was stirred2 h, giving a pale yellow solution of alkenyl intermediate 18, which was diluted with 4 ml dry N-methylpyrrolidine. A second dry flask was charged with anhydrous LiBr (0.35 g, 4 mmol) and Pd(acac)2 (30 mg, 0.08 mmol), and flushed thoroughly with Ar. The solution of the zirconium intermediate 18thenwas added by syringe, followed by (Z)-4-undecenyl bromide 16 (0.47 g, 2 mmol), and the mixture was heated at 55ºC overnight under Ar. The resulting black solution was cooled and poured into 100 ml water, and extracted with hexane. The hexane layer was washed with brine, dried, and treated with decolorizing charcoal. Filtration through a pad of celite yielded a yellow solution, which was concentrated and the residue was purified by vacuum flash chromatography eluting with 5% EtOAc in hexane. The product 19was then stirred overnight in 10 ml MeOH with ~20 mg PTSA, and the solution was poured into dilute aqueous NaHCO3 and extracted with hexane. The hexane solution was washed with brine, dried, and concentrated, then purified by Kugelrohr distillation, taking a forerun to remove the THP-protected MeOHsideproduct (oven temp <60 mm/0.4 mm Hg), followed by dienol20 (~115-120/0.25 mm Hg,0.345 g, 65%) as a viscous oil. Attempts to recrystallize alcohol 20from hexane at -20ºC failed, so the product was used directly in the next step. 1H NMR (CDCl3):5.37 (m, 2H), 5.35-5.28 (m, 2H), 3.61 (t, 2H, J = 6.6 Hz), 1.97 (m, 8H), 1.55 (quint, 2H, J = 6.9 Hz), 1.42-1.20 (m, 14 H), 0.86 (t, 3H, J = 6.9 Hz). 13C NMR (CDCl3): 130.87, 130.50, 130.34, 129.72, 63.18, 32.85, 32.73, 32.33, 31.97, 29.92, 29.87, 29.59, 29.18, 27.43, 26.87, 25.43, 22.85, 14.28 ppm.MS (m/z: abundance): 266 (9), 166 (12), 149 (8), 138 (16), 121 (14), 110 (20), 109 (29), 96 (47), 95 (56), 93 (35), 83 (28), 82 (61), 81 (79), 79 (45), 68 (38), 67 (100), 55 (72), 41 (64).

Alcohol 20 was oxidized with PDC in CH2Cl2 as described above for 7 and 10, giving (6E,11Z)-6,11-octadecadienal (87 mg, 50%, 97% pure by GC) as a colorless oil. This was dilutedimmediately with hexane, a small crystal of BHT was added, and the solution was sealed in an ampoule and stored at -20ºC until used in experiments. 1H NMR (CDCl3):  9.74 (t, 1H, J = 1.8 Hz), 5.44-5.28 (m, 4H), 2.40 (td, 2H, J = 7.4,1.8 Hz), 2.05-1.93 (m, 8H), 1.61 (m, 2H), 1.45-1.20 (m, 12H), 0.87 (t, 3H, J = 6.8 Hz). 13C NMR (CDCl3): 202.99, 130.99, 130.38, 129.88, 129.67, 43.96, 32.44, 32.32, 31.97, 29.92, 29.83, 29.24, 29.19, 27.44, 26.88, 22.85, 21.75, 14.30 ppm. MS (m/z: abundance): 264 (M+, 3), 246 (4), 235 (1), 221 (2), 175 (6), 166 (12), 161 (14), 148 (17), 138 (28), 123 (14), 121 (13), 119 (11), 110 (24), 109 (35), 95 (74), 84 (51), 83 (36), 82 (49), 81 (81), 79 (64), 67 (100), 55 (92), 43 (46), 41 (95).

Figure legend

Figure S1.Synthesis of (11Z)-11-octadecenal, (6E)-6-octadecenal, and (6E,11Z)-6,11-octadecadienal. a) Dihydropyran, PTSA; b) Lithium acetylide-ethylene diamine complex, NaI, DMSO; c) BuLi, hexyl iodide, THF, 50ºC; d) PTSA, MeOH; e) P-2 nickel, ethylenediamine, H2, EtOH; f) PDC, CH2Cl2; g) LiAlH4, THF; h) Mesyl chloride, Et3N, CH2Cl2, then LiBr, acetone; i) Cp2ZrHCl, THF, NMP; j) LiBr, Pd(acac)2, (4Z)-4-undecenyl bromide 16.

Figure S2. EI mass spectrum of (6E,11Z)-6,11-octadecadienal

Figure S3. EI mass spectrum of (11Z)-11-octadecenal

Figure S4. EI mass spectrum of (6E)-6-octadecenal

Fig. S2

Fig. S3

Fig. S4