232 J . Org. Chem., Vol. 67, No. 1, 2002
Ung et al.
brown solid (0.059 g, 86%). 1H NMR (D2O) δ 6.82 (br s, 1H,
H-3), 2.68-2.57 (m, 2H), 2.48-2.39 (m, 1H), 2.18-2.09 (m, 1H);
13C NMR (D2O) δ 175.98 (CO), 169.91 (CO), 146.42 (CH),
4.79 (1H, m), 4.98 (1H, br s), 6.70 (1H, br s), 9.15 (2H, br s);
13C ΝΜR (D2O) δ major isomer: 13.8 (CH3), 15.7 (CH3), 20.6
(CH3), 21.7 (CH3), 23.0 (CH2), 26.1 (CH), 31.2 (CH), 34.2 (CH2),
40.0 (CH2), 41.8 (CH2), 43.4 (CH2), 46.9 (CH), 61.9 (CH2), 62.9
(C), 77.6 (CH), 132.6 (C), 140.5 (CH), 165.0 (C), 170.8 (C) ppm.
MS (CI +ve) m/z 338 (M + 1); HRMS Calcd for C19H32NO4
(MH+) 338.2331, Found 338.2330.
22
137.72 (C), 70.63 (C), 33.55 (CH2), 30.08 (CH2); [R]D +11 (c
0.4 H2O); MS (ES +ve) m/z 171.8 (M + 1, 20%); HRMS (ES
+ve) Calcd for C7H10NO4 (MH+): 172.0610. Found: 172.0618.
Met h yl 1-[Dip h en ylm et h ylid en ea m in o]-3-cyclop en -
ten e 1,3-d ica r boxyla te (21). A solution of ethyl butynoate
20 (0.196 g, 2 mmol), methyl 2-[N-(diphenylmethylidene-
amino)propenoate 4 (0.26 g, 1 mmol), and tributylphosphine
(40 mg, 0.2 mmol) in benzene (5 mL) was stirred at RT for 24
h. The solvent was removed to give a black thick oil which
was purified by column chromatography (25% ethyl acetate/
hexane) to give a bright yellow thick oil (0.372 g, 98%) which
upon triteration in cold petroleum spirit gave a white crystal-
1-(1R,2S,5R)-Men th yl 3-Eth yl (1R)-1-Ben zyloxyca r bon -
yla m in o-3-cyclop en ten ed ica r boxyla te (32) a n d 1-(1R,2S,
5R)-Men th yl 3-Eth yl (1S)-1-Ben zyloxyca r bon yla m in o-3-
cyclop en ten ed ica r boxyla te (33). The above mixture of the
salts 30 and 31 (250 mg, 0.669 mmo1) was dissolved in THF
(10 mL), and the solution was cooled to 0 °C. Saturated sodium
carbonate solution (10 mL), benzyl chloroformate (0.2 mL, 1.40
mmol) was added, and the mixture was stirred at 0 °C for 2 h.
The volatiles were removed in vacuo, and the residue was
diluted with water and extracted with chloroform (2×). The
combined organic extracts were dried (MgSO4), filtered, and
evaporated in vacuo to give a clear syrup. The pure product
was obtained by column chromatography (9:1 petroleum spirit:
ethyl acetate as eluent) which gave the title compounds (220
mg, 0.467 mmol, 69.7%) as a clear syrup. The two isomers
could be partially separated by HPLC (6.5% ethyl acetate in
petroleum spirit as eluent, 20 mL min-1, 10 mg injections),
which afforded 30 mg and 15 mg of 33 and 32, respectively.
1
line solid. H NMR δ 7.58-7.11 (m, 10 H), 6.65 (m, 1H), 4.18
(q, 2H), 3.38-3.32 (m, 1H), 3.29 (s, 3H), 3.28-3.18 (m, 1H),
3.15-3.03 (m, 2H), 1.28 (t, 3H); 13C NMR δ 173.97 (CO), 167.93
(CO), 164.15 (CN), 140.13 (C), 139.56 (CH), 136.67 (C), 133.58
(C), 128.47 (CH), 128.24 (2 CH), 128.29 (2 CH), 127.76 (2 CH),
127.66 (2 CH), 72.38 (C), 59.94 (CH2), 51.36 (CH3), 47.92 (CH2),
46.30 (CH2), 13.99 (CH3); MS (ES +ve) m/z 378.2 (M + 1+).
Anal. Calcd for C23H23NO4: C, 73.19; H, 6.14; N, 3.71. Found:
C, 73.40; H, 6.34; N, 3.48%.
1-Am in o-3-cyclop en ten e 1,3-d ica r boxylic Acid (h yd r o-
ch lor id e sa lt) (r a c-2). Cycloadduct 21 (0.50 g, 1.32 mmol)
was dissolved in 6 N HCl (3 mL). The solution was heated at
under reflux for 6 h. After cooling, the aqueous layer was
extracted with ether (2×), and the water was removed under
reduced pressure to give a white solid (0.235 g, 91%). 1H NMR
(D2O) δ 6.63 (s, 1H), 3.20 (dd, 2H, J 3.0, 19.8 Hz), 2.78 (dd,
2H, J 2.4, 19.4 Hz); 13C NMR (D2O) δ 172.13 (CO), 166.23 (CO),
140.69 (CH), 131.71 (C), 62.96 (C), 43.06 (CH2), 41.19 (CH2);
32: [R]D -10 (c 0.5, CHCl3); 1Η ΝΜR δ 0.74 (3H, d, J 7.2
23
Hz), 0.86 (3H, d, J 7.2 Hz), 0.89 (3H, d, J 7.2 Hz), 1.28 (3H, t,
J 6.9 Hz), 0.70-2.10 (9H, m), 2.85-3.00 (2H, m), 3.10-3.30
(2H, m), 4.20 (2H, q, J 6.9 Hz), 4.71 (1H, td, J 10.5, 4.2 Hz),
5.09 (2H, AB system, J 12.3 Hz), 5.56 (1H, br. s), 6.69 (1H, br.
s), 7.30-7.40 (5H, m); 13C ΝΜR δ 14.2 (CH3), 15.9 (CH3), 20.8
(CH3), 22.0 (CH3), 23.1 (CH2), 26.1 (CH), 31.3 (CH), 34.1 (CH2),
40.2 (CH2), 43.8 (CH2), 44.6 (CH2), 46.8 (CH), 60.5 (CH2), 64.2
(C), 66.7 (CH2), 76.1 (CH), 128.0, 128.0, 128.2, 128.5, 128.5
(Ar CH’s), 133.3 (C), 136.0 (C), 139.9 (CH), 155.0 (C), 164.0
MS (ES +ve) m/z 172.0 (M + 1, 100%). HRMS: Calcd for C7H10
-
NO4 (MH+): 172.0616.
(C), 172.8 (C). 33:[R]D -43 (c 1.0, CHCl3); 1Η ΝΜR δ 0.74
23
1-(1R,2S,5R)-Men th yl 3-Eth yl (1R)-1-Dip h en ylm eth yl-
en eam in o-3-cyclopen ten e dicar boxylate (28) an d 1-(1R,2S,
5R)-Men th yl 2-Eth yl (1S)-1-Dip h en ylm eth ylen ea m in o-3-
cyclop en ten e d ica r boxyla te (29). The acrylate 27 (305 mg,
0.783 mmol) was dissolved in dry benzene (8.0 mL), and then
ethyl butynoate (0.22 mL, 2.20 mmol) and tributylphosphine
(0.10 mL, 0.40 mmol) were added. The mixture was stirred at
RT overnight, and then all volatiles were removed in vacuo to
give a dark brown syrup. The pure product was obtained by
column chromatography (9:1 petroleum spirit:ethyl acetate as
eluent) which gave the title compound (340 mg, 0.678 mmol,
86.6%) as an amber syrup. A diastereoisomeric ratio of 60:40
was estimated from the 1H NMR spectrum. 1Η ΝΜR (400
MHz) δ 0.65 (3H, d, J 7.2 Hz, major), 0.66 (3H, d, J 7.2 Hz
minor), 0.78 (3H, d, J 7.2 Hz, major), 0.80 (3H, d, J 7.2 Hz,
minor), 0.86 (3H, d, J 6.4 Hz, minor), 0.87 (3H, d, J 6.4 Hz,
major), 0.7-1.0 (4H, m), 1.28 (3H, t, J 7.2 Hz), 1.2-1.8 (5H,
m), 2.85-3.30 (4H, m), 4.17 (2H, q, J 7.2 Hz), 4.40-4.50 (1H,
m), 6.60 (1H, br s), 7.15-7.60 (10H, m); 13C ΝΜR δ major
isomer: 14.2 (CH3), 15.9 (CH3), 20.8 (CH3), 22.0 (CH3) 22.9
(CH2), 25.7 (CH) 31.2 (CH), 34.1 (CH2), 40.1 (CH2), 46.7 (CH),
47.0 (CH2), 47.2 (CH2), 60.2 (CH2), 72.9 (C), 75.2 (CH), 127.8,
127.9, 127.9, 128.0, 128.6, 128.6, 128.7, 128.7, 130.0, 130.1 (d,
Ar CH’s), 134.0 (C), 133.7 (C), 140.1 (CH), 140.8 (C), 164.4 (C),
168.8 (C), 173.6 (C); MS (ES +ve) m/z 502 (M + 1); HRMS
Calcd for C32H40NO4 (MH+) 502.2957. Found 502.2955.
1-(1R,2S,5R)-Men t h yl 3-E t h yl (1R)-1-Am in o-3-cyclo-
p en t en ed ica r b oxyla t e H yd r och lor id e Sa lt (30) a n d
1-(1R,2S,5R)-Men th yl 3-Eth yl (1S)-1-Am in o-3-cyclop en -
ten ed ica r boxyla te Hyd r och lor id e Sa lt (31). The above
mixture of cycloadducts 28 and 29 (340 mg, 0.678 mmol) was
dissolved in diethyl ether (10 mL), and then 1 N HCl (10 mL)
was added. The mixture was stirred at RT for 16 h and then
diluted with diethyl ether and 1 N HCl, and the two layers
were separated. The organic portion was extracted with 1 N
HCl, and the combined aqueous layers were evaporated in
vacuo to give the title compound (250 mg, 0.669 mmol, 94.7%),
as a clear gum. 1Η ΝΜR (CDCl3) δ 0.74 (3H, d, J 6.3 Hz), 0.80-
1.20 (9H, m), 1.29 (3H, t, J 6.9 Hz), 1.46 (2H, m), 1.60-1.90
(3H, m), 2.02 (1H, br d), 3.26 (4H, m), 4.20 (2H, q, J 6.9 Hz),
(3H, d, J 7.2 Hz), 0.86 (3H, d, J 7.2 Hz), 0.89 (3H, d, J 7.2 Hz),
1.28 (3H, t, J 6.9 Hz), 0.70-2.10 (9H, m), 2.85-3.00 (2H, m),
3.10-3.30 (2H, m), 4.19 (2H, q, J 6.9 Hz), 4.70 (1H, td, J 10.5,
4.2 Hz), 5.08 (2H, AB system, J 12.0 Hz), 5.55 (1H, br.s), 6.70
(1H, br. s), 7.30-7.40 (5H); 13C ΝΜR δ 14.2 (CH3), 15.9 (CH3),
20.7 (CH3), 21.9 (CH3), 23.0 (CH2), 26.0 (CH), 31.3 (CH), 34.1
(CH2), 40.3 (CH2), 43.5 (CH2), 44.3 (CH), 46.8 (CH), 60.4 (CH2),
64.2 (C), 66.7 (CH2), 76.0 (CH), 128.0, 128.0, 128.1, 128.5, 128.5
(Ar CH’s) 133.3 (C), 135.2 (C), 140.2 (CH), 155.0 (C), 164.1 (C),
172.7 (C); MS (CI +ve) m/z 472 (M + 1); HRMS Calcd for
C
27H38NO6 (MH+) 472.2699, Found 472.2700.
(R)-1-Am in o-3-cyclop en ten e 1,3-d ica r boxylic Acid (h y-
d r och lor id e sa lt) (2) a n d (S)-1-Am in o-3-cyclop en ten e 1,3-
d ica r boxylic Acid (h yd r och lor id e sa lt) (2). Compound 33
(30 mg, 0.064 mmol) was dissolved in diethyl ether (5 mL) and
transferred to a sealed tube, and the diethyl ether removed in
vacuo. Hydrochloric acid solution (6N, 8 mL) was added and
the tube sealed and shaken. The tube was warmed to 85 °C,
with frequent shaking until all material was in solution, and
then the mixture stirred at that temperature for 4 d. The
mixture was cooled to RT and then diluted with water (40 mL)
before it was washed with diethyl ether (2×) and evaporated
in vacuo to give a pale yellow solid. The pure product was
obtained by ion exchange chromatography which afforded (R)-2
(13 mg, 0.062 mmol, 98%) as the hydrochloride salt. (S)-2 was
produced from compound 32 (15 mg, 0.032 mmol), in the same
way giving 6 mg (0.029 mmol, 90.3%). NMR spectral data were
identical to (R)-2 and rac-2 prepared according to Schemes 2
22
and 4, respectively. (R)-2: [R]D +5.9 (c 0.6, 1 N HCl). (S)-2:
22
[R]D -5.8 (c 0.87, 1 N HCl).
Sign a l Tr a n sd u ction a t Clon ed Ra t m Glu 1a , m Glu 5a ,
a n d m Glu 2 Recep tor s in CHO Cells. CHO cells expressing
the cloned rat mGlu1a and mGlu5a receptors (a kind gift from
S. Nakanishi, Kyoto University, J apan) and the human mGlu2
receptor (cloned and expressed in house) were grown in
DMEM/Glutamax-I to which 2 mM glutamine, 46 mg/L pro-
line, and 10% dialyzed fetal calf serum were added.
Ca 2+ F lu or im etr ic Assa y for Ra t m Glu 1a a n d m Glu 5a .
CHO cells expressing the mGlu1a or mGlu5a were plated at