Supplementary Material
Trail Following Response of Larval Cactoblastis cactorum to 2-Acyl-1,3Cyclohexanediones
Terrence D. Fitzgerald1, Michael Kelly1, Tyler Potter2, James E. Carpenter3, Frank Rossi2
1
Department of Biological Sciences, State University of New York at Cortland,
Cortland, NY, 13045, USA
2
Department of Chemistry, State University of New York at Cortland,
Cortland, NY, 13045, USA
3
U.S. Department of Agriculture Research Service, Crop Protection and Research
Laboratory, Tifton. Ga.
Supporting Information
Table of Contents
I.
Synthesis of Compounds
II.
NMR spectra of compounds synthesized
a. Compound 6
b. Compound 7
c. Compound 8
d. Compound 1
e. Compound 2
I. Synthesis of Compounds
General. Reagents were purchased from Sigma-Aldrich, or Fisher Scientific and were
used without additional purification. Reactions for the preparations of compound 1, 2,
and 7, were carried out under a nitrogen atmosphere. All other reactions were carried out
under an atmosphere of air in capped reaction containers. Column chromatography was
conducted with silica gel 60 (Merck, 230-400 mesh). Melting points were taken on a
Thomas-Hoover melting point apparatus and are uncorrected. NMR was conducted on a
Bruker Avance 600 MHz NMR spectrometer or a Brucker Avance 300 MHz NMR
spectrometer with CDCl3 as the solvent unless otherwise noted. The chemical shift was
calibrated to the residual solvent peak.
HPLC was performed on a Dionex P580
instrument equipped with a PDA-100 diode array detector and a Phenomenex Jupiter
Proteo column (10m, 250 x 10 mm) and eluted with water (solvent A) and acetonitrile
(solvent B) using the following solvent gradient: ramp from 25% A, 75% B to 100% B,
for 5 min; 100 % B, for 20 min; 100 % B to 25 % A 75%B, for 5 min; flow rate 2
mL/min. All other mass spectra were obtained by the University of Wisconsin Mass
Spectrometry Laboratory, Madison Wisconsin.
Preparation of 4-acetoxy-3-methoxy-2-cyclohexenone (6). This procedure is a
modification of the methods reported by Kammath (2013) and Demir (2008). Potassium
permanganate (1.422 g, 9.00 mmol) was added to a mixture of benzene and acetic acid
(10:1, 150 mL). The mixture was stirred under reflux with a Dean-Stark trap until the
purple color of the permanganate turned brown (∼30 min). A solution of 3-methoxy-2cyclohexenone (5) (van der Klei, 2002) (0.378 g, 3.00 mmol) dissolved in benzene was
added to the reaction mixture, and the reflux was continued for 20 h. The reaction
mixture was cooled to room temperature, and neutralized with saturated NaHCO3 (~50
mL) to give a brown emulsion. Sodium bisulfite (10 mL, 3 M) was added to the
emulsion and two layers formed. The organic layer was washed with water (2 x 20 mL)
and the combined aqueous layers were extracted with dichloromethane (2 x 20 mL). The
combined organic layers were washed with brine (30 mL), dried with MgSO4, and
concentrated by rotary evaporation to give crude 6 (0.430 g, 78%) as a viscous yellow oil.
The crude material was purified by flash chromatography (2:3 hexane/ethyl acetate) to
give pure 6 (0.147 g, 35%) as a yellow solid. mp 69 – 71 °C. 1H NMR (600 MHz,
CDCl3) δ 5.40 (d, J = 1.6 Hz, 1H), 5.29 (dd, J = 12.7, 5.2 Hz, 1H), 3.70 (s, 3H), 2.65
(dddd, J = 17.5, 12.1, 5.2, 1.6 Hz, 1H), 2.51 (ddd, J = 17.7, 5.2, 2.9 Hz, 1H), 2.22 (dtd, J
= 13.1, 5.2, 2.9 Hz, 1H), 2.13 (s, 3H), 2.09 (qd, J = 12.4, 5.2 Hz, 1H).
13
C NMR (151
MHz, CDCl3) δ 193.2, 177.5, 170.4, 101.1, 72.6, 56.3, 27.9, 26.9, 21.1. ESI-HRMS calc
for C9H13O4 [M + H]+ 185.0808, found 185.0804.
Preparation of 4-acetoxy-1,3-cyclohexandione (7). Hydrochloric acid (2M, 2.34 mL)
was added to a stirring solution of compound 6 (334 mg, 1.81 mmol, 1 equiv) dissolved
in acetone (8.4 mL). The reaction was monitored by TLC and showed no starting
material after 1 h. The reaction mixture was concentrated by rotary evaporation, taken up
in ethyl acetate (10 mL) and the aqueous layer was extracted. The aqueous layer was
extracted with another portion of ethyl acetate (5 mL) and the combined organic layers
were concentrated by rotary evaporation to yield a crude 7 (250 mg, 81%) as a yellow
solid. The crude product was recrystallized (1:1 chloroform/ether) to yield 7 as a white
solid (156 mg, 51%). mp = 139-140 °C. Proton NMR in CDCl3 shows the presence of
three tautomers in a 15:8:1 ratio. Diketone tautomer: 1H NMR (600 MHz, CDCl3) δ 1.94
(qd, J = 12.9, 5.3 Hz, 1H), 2.11 – 2.18 (m, 0H), 2.18 (s, 1H), 2.20 (s, 3H), 2.70 – 2.83 (m,
2H), 3.50 (dd, J = 16.7, 2.3 Hz, 1H), 3.60 (d, J = 16.8 Hz, 1H), 5.45 (dd, J = 12.4, 5.9 Hz,
1H).
13
C NMR (151 MHz, CDCl3) δ 201.2, 196.4, 170.0, 75.4, 57.0, 38.4, 23.8, 20.7.
Enol tautomer (major isomer): 1H NMR (600 MHz, CDCl3) δ 2.11 – 2.18 (m, 0H), 2.18
(s, 1H), 2.27 (dd, J = 9.6, 3.4 Hz, 1H), 2.31 (ddt, J = 11.9, 5.8, 2.8 Hz, 1H), 2.50 – 2.64
(m, 0H), 2.56 (q, J = 5.5 Hz, 1H), 5.37 (dd, J = 9.0, 4.9 Hz, 0H).
13
C NMR (151 MHz,
CDCl3) δ 186.4, 171.3, 105.0, 71.0, 29.5, 27.1, 21.0,
ESI-HRMS calc for C8H11O4 [M + H]+ 171.0652, found 171.0646.
Synthesis 4-acetoxy-2-oleyl-1,3-cyclohexanedione (8). Oleic acid (141 mg, 0.5 mmol, 1
equiv) was added to a stirring solution of compound 7 (94 mg, 0.55 mmol, 1.1 equiv)
dissolved in dichloromethane (2.5 mL). N,N’-Dicyclohexylcarbodiimide (113 mg, 0.55
mmol, 1.1 equiv), triethylamine (77 uL, 0.55 mmol, 1.1 equiv), and 4-
(dimethylamino)pyridine (6 mg, 0.05 mmol, 0.1 equiv) were successively added to the
reaction mixture. The reaction was stirred and monitored by TLC. After ~30 minutes,
starting material was completely converted to the O-acylated compound (lower spot on
TLC, 5:1 hexane/ethyl acetate w/ 2% acetic acid) and after 15 hours the TLC showed no
O-acylated compound. The reaction mixture was diluted with dichloromethane (25 mL)
and washed with HCl (1M, 10 mL). The aqueous layer was extracted with diethyl ether
(3x5 mL). The combined organic layers were washed with brine (15 mL), dried with
sodium sulfate, and concentrated by rotary evaporation. To remove the DCU, the residue
was dissolved in pentane and filtered through a Hirsch funnel.
The filtrate was
concentrated by rotary evaporation and purified by flash chromatography (7:1
hexane/ethyl acetate with 2% acetic acid) to give 8 (115 mg, 53%) as an oily solid. For
bioassays, 7 was further purified by HPLC.
1
H NMR (600 MHz, CDCl3) δ 0.98 (t,
J=6.96 Hz, 3 H) 1.24 - 1.38 (m, 47 H) 1.56 - 1.67 (m, 5 H) 1.81 (br. s., 2 H) 2.01 (q,
J=6.53 Hz, 9 H) 2.08 - 2.17 (m, 3 H) 2.20 - 2.25 (m, 9 H) 2.78 - 2.90 (m, 2 H) 2.92 - 2.98
(m, 1 H) 3.03 - 3.09 (m, 1 H) 5.30 - 5.38 (m, 7 H) 18.25 (s, 1 H).
13
C NMR (151 MHz,
CDCl3) δ 14.1, 20.8, 22.1, 24.1, 24.5, 27.2, 27.2, 29.1, 29.2, 29.3, 29.3, 29.5, 29.7, 29.8,
31.2, 31.9, 40.4, 72.7, 111.4, 129.8, 129.9, 170.2, 189.5, 96.6, 206.5. ESI-HRMS calc for
C26H43O5 [M + H]+ 435.3105, found 435.3110.
Synthesis of 4-hydroxy-2-oleyl-1,3-cyclohexanedione (1). Potassium carbonate (38
mg, 0.276 mmol, 2 equiv) was added to a stirred solution of 7 (60 mg, 0.138 mmol, 1
equiv) dissolved in methanol (2.7 mL).
The reaction was monitored by TLC until
starting material was consumed (~3 h). The reaction mixture was diluted with hexane (15
mL) and extracted with HCl (1M, 2x5 mL). The aqueous layer was extracted with hexane
(5 mL) and the combined organic extracts were dried with sodium sulfate and
concentrated by rotary evaporation to yield 1 (48 mg, 89%) as an oily solid. Melting
point = ~ room temperature. For bioassays, 1 was further purified by HPLC as described
in the general section. 1H NMR (600 MHz, CDCl3) δ ppm 0.89 (t, J=6.87 Hz, 3 H) 1.18 1.44 (m, 17 H) 1.57 - 1.70 (m, 2 H) 1.83 (qd, J=12.55, 6.59 Hz, 1 H) 1.98 - 2.06 (m, 4 H)
2.35 - 2.42 (m, 1 H) 2.75 - 2.88 (m, 2 H) 2.98 (ddd, J=15.81, 9.03, 6.21 Hz, 1 H) 3.08
(ddd, J=15.72, 9.13, 6.02 Hz, 1 H) 4.09 (dd, J=13.18, 5.46 Hz, 1 H) 5.31 (s, 1 H) 5.32 -
5.42 (m, 2 H) 18.26 (s, 1 H). 13C NMR (151 MHz, CDCl3) δ ppm 14.1, 22.7, 24.5, 24.9,
27.1, 27.2, 27.2, 29.1, 29.1, 29.3, 29.3, 29.3, 29.5, 29.7, 29.8, 31.3, 31.9, 34.1, 40.2,
71.6, 110.3, 129.7, 130.0, 195.5, 197.9, 206.0. ESI-HRMS calc for C24H41O4 [M + H]+
393.3000, found 393.3006.
Synthesis of compound 2. Oleic acid (141 mg, 0.5 mmol, 1 equiv) was added to a
stirring solution of compound 1,3-cyclohexanedione (112 mg, 1.00 mmol, 2.0 equiv)
dissolved in dichloromethane (2.5 mL). N,N’Dicyclohexylcarbodiimide (113 mg, 0.55
mmol, 1.1 equiv), triethylamine (139 uL, 1.00 mmol, 1.0 equiv), and 4(dimethylamino)pyridine (6 mg, 0.05 mmol, 0.1 equiv) were successively added to the
reaction mixture. The mixture reaction was stirred was stirred for 20 h, diluted with
dichloromethane (25 mL) and washed with HCl (1M, 2 x 10 mL) and brine (10 mL),
dried with sodium sulfate, and concentrated by rotary evaporation. The residue was
purified by flash chromatography (hexane/ethyl acetate 10:1with 2% acetic acid) to give
2 (51 mg, 27%). 1H NMR (300 MHz, CDCl3) δ 0.88 (t, J = 6.4 Hz, 4H), 1.29 (d, J = 14.2
Hz, 23H), 1.60 (q, J = 7.2 Hz, 3H), 1.98 (dq, J = 13.0, 7.4, 6.4 Hz, 6H), 2.40 – 2.53 (m,
2H), 2.66 (t, J = 6.4 Hz, 2H), 2.95 – 3.07 (m, 2H), 5.34 (t, J = 5.0 Hz, 2H).
13
C NMR (75
MHz, CDCl3) δ 14.3, 19.2, 22.8, 24.8, 27.4, 27.4, 29.3, 29.47, 29.50, 29.52, 29.7, 29.88,
29.93, 32.1, 33.5, 39.0, 40.7, 77.4, 113.2, 130.0, 130.1, 195.4, 198.8, 206.5. ESI-HRMS
calc for C24H41O3 [M + H]+ 377.3050, found 377.3050.
II. NMR spectra of compounds synthesized. Compound 6
Compound 7
The major isomer is the diketone
Diketone
Literature values3: 1H NMR (CDCl3): 1.93 (1H, ddd, J = 6 Hz, 13 Hz and 13 Hz, 5-H);
2.10-2.35 (1H, m, 5-H); 2.12 (1.6H, s, Ac of diketo form); 2.77 (1.3H, m, 6-H2 of diketo
form); 3.55 (1.3H, dd + d, J = 2 Hz and 17 Hz + 17 Hz, 1⁄2 W ∼
ereotopic
2-H2 of diketo form); 5.45 (0.6H, dd, J = 6 Hz and 12 Hz, 4-H of diketo form).
Found, Diketone: 1H NMR (600 MHz, CDCl3) δ 1.94 (qd, J = 12.9, 5.3 Hz, 1H), 2.11 –
2.18 (m, 0H), 2.18 (s, 1H), 2.20 (s, 3H), 2.70 – 2.83 (m, 2H), 3.50 (dd, J = 16.7, 2.3 Hz,
1H), 3.60 (d, J = 16.8 Hz, 1H), 5.45 (dd, J = 12.4, 5.9 Hz, 1H).
Enol
Literature values3: 1H NMR (CDCl3): 2.09 (1.1H, s, Ac of enolic form); 2.57 (0.8H, dd, J
= 5 Hz and 7 Hz, 4-H2 of enolic form); 5.37 (0.4H, dd, J = 4.8 Hz and 9.6 Hz, 6-H of
enolic form); 5.53 (0.4H, br s, 2-H of enolic form).
Found: 1H NMR (600 MHz, CDCl3) δ 2.11 – 2.18 (m, 0H), 2.18 (s, 1H), 2.27 (dd, J =
9.6, 3.4 Hz, 1H), 2.31 (ddt, J = 11.9, 5.8, 2.8 Hz, 1H), 2.50 – 2.64 (m, 0H), 2.56 (q, J =
5.5 Hz, 1H), 5.37 (dd, J = 9.0, 4.9 Hz, 0H)
13
C NMR of two major tautomers
Compound 8
Compound 1
Compound 2
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