Fulleroproline DeriVatiVes and Peptides
.97), 601 (-1.45, 1.31), 549 (-0.79, 0.66), 428 (-3.87, 3.32), 396
J. Am. Chem. Soc., Vol. 118, No. 17, 1996 4079
1
mmol) of paraformaldehyde. The mixture was heated to reflux for 1
h, then brought to ambient temperature, poured on top of a chromato-
graphic column, and eluted with toluene and then toluene/ethyl acetate,
95:5. The fractions containing unreacted C60 were concentrated,
affording 120 mg (40%) of pure material. The fractions containing
5b were concentrated to ca. 50 mL and added to a solution of 700 mg
(2.88, -2.62), 343 (-5.38, 3.65), 319 (2.82, -2.96); Anal. Calcd for
82 4
C H23NO
: C, 90.68; H, 2.13; N, 1.29. Found: C, 89.51; H, 2.02; N,
1
.12.
Fmoc-Fpr-L-Ala-OEt (8a and 8b). To a solution of 27.8 mg
0.0256 mmol) of 6 in 2 mL of CH Cl was added 2 mL of TFA, and
(
2
2
the mixture was stirred at ambient temperature for 4 h. The solvent
was removed in Vacuo, and the residue was washed several times with
methanol in a centrifuge tube and finally dried under reduced pressure.
Carboxylic acid 7 was obtained in nearly quantitative yield: IR (KBr)
(1.16 mmol) of (Fmoc-L-Ala)
2
O and 127 µL (1.16 mmol) of NMM in
40 mL of CH Cl . The mixture was stirred at room temperature for 3
2
2
days, and then the solvent was removed in Vacuo. Diastereomeric
dipeptides 9a and 9b were separated by flash column chromatography
(eluant: toluene/ethyl acetate, 98:2).
Fmoc-L-Ala-D-Fpr-OEt (9a): 57 mg (12%); IR (KBr) (cm-1) 3420,
1742, 1724, 1667, 1503, 1449, 1426, 1191, 758, 528; H NMR (400
3
MHz, CDCl ) δ 7.80-7.30 (m, 8H), 6.62 (s, 1H, cis); 6.54 (s, 1H,
-
1
1
(
cm ) 3435, 1718, 1638, 1450, 1424, 1222, 1151, 757, 576, 528; H
NMR (250 MHz, DMSO/CDCl , 5:2) δ 7.86-7.80 (m, 4H), 7.42-
.36 (m, 4H), 6.19 (s, 1H), 5.75-5.42 (m, 2H), 4.86-4.40 (m, 3H);
3
1
7
13
3
C NMR (62.5 MHz, CDCl ) δ 170.99, 154.57, 154.43, 150.72, 146.99,
1
1
1
1
1
4
46.89, 146.21, 145.94, 145.88, 145.84, 145.67, 145.60, 145.26, 145.12,
45.10, 145.03, 145.00, 144.90, 144.83, 144.79, 144.42, 144.14, 144.01,
43.87, 143.51, 143.42, 142.68, 142.55, 142.24, 142.19, 141.93, 141.78,
41.66, 141.59, 141.48, 141.25, 140.84, 139.73, 139.64, 139.09, 136.31,
trans), 6.15 (d, J ) 11.7 Hz, 1H, cis), 5.90 (d, J ) 8.0 Hz, 1H), 5.83
(d, J ) 11.0 Hz, 1H, trans), 5.60 (d, J ) 11.0 Hz, 1H, trans), 5.52 (d,
J ) 11.7 Hz, 1H, cis), 5.10-5.00 (m, 1H), 4.53-4.25 (m, 5H), 1.74
(d, J ) 7.3 Hz, 3H), 1.27 (t, J ) 6.6 Hz, 3H); 13C NMR (62.5 MHz,
35.70, 134.72, 127.69, 127.08, 125.14, 120.03, 67.65, 58.51, 57.90,
3
CDCl ) δ 172.26, 169.51, 155.61, 153.93, 153.52, 151.95, 151.89,
+
6.75; MALDI-MS C78
H
15NO
4
(MW ) 1029), m/z 1075 [(M + 2Na) ].
149.60, 147.56, 147.45, 147.06, 146.50, 146.40, 146.22, 146.15, 145.76,
145.62, 145.53, 145.44, 145.37, 144.70, 144.60, 144.54, 144.47, 144.37,
144.33, 143.87, 143.82, 143.64, 143.10, 142.76, 142.72, 142.23, 142.18,
142.09, 141.82, 141.32, 140.41, 140.34, 139.70, 136.07, 127.76, 127.11,
125.17, 120.02, 70.65, 69.56, 68.97, 67.20, 62.30, 58.89, 48.77, 47.17,
To a suspension of 26.4 mg (0.0256 mmol) of 7 in 5 mL of CH
Cl
2 2
were added 5.2 mg (0.0384 mmol) of HOAt and 7.4 mg (0.0384 mmol)
of EDC‚HCl, and the mixture was stirred at room temperature for 15
min. A solution of 4.3 mg (0.0282 mmol) of HCl‚H-L-Ala-OEt and 3
µL (0.0282 mmol) of NMM in 0.5 mL of CH
2
Cl
2
was added and the
19.99, 14.20; MALDI-MS C83
H
24
N
2
O
5
(MW ) 1128), m/z 1151 [(M
+ Na) ], 1129 [(M + H) ]; UV-vis (CHCl ) λmax (ꢀ) 428 (3080), 312
(37 600), 301 (39 600), 256 (128 800); CD (CHCl
+
+
solution stirred for 30 min. The solvent was removed under reduced
pressure and the mixture of dipeptides 8a and 8b purified by flash
column chromatography (eluant: toluene/ethyl acetate, 98:2). The
fractions containing the dipeptide were concentrated, and the two
diastereoisomers were separated by HPLC (C18, see Instrumentation).
3
-
3
3
T
) λmax (θ x 10 )
690 (-1.71), 654 (2.07), 595 (1.77), 544 (1.34), 477 (1.28), 428 (4.55),
397 (-3.72), 347 (2.13).
Fmoc-L-Ala-L-Fpr-OEt (9b): 57 mg (12%); IR (KBr) (cm-1) 3417,
-
1
1
Fmoc-D-Fpr-L-Ala-OEt (8a): 7 mg (24%); IR (KBr) (cm ) 3405,
1742, 1728, 1670, 1501, 1449, 1424, 1194, 758, 528; H NMR (400
1
1
718, 1688, 1509, 1221, 1151, 757, 576, 528; H NMR (250 MHz,
3
MHz, CDCl ) δ 7.82-7.30 (m, 8H), 6.66 (s, 1H, trans), 6.28 (s, 1H,
CDCl
5
(
3
) δ 7.78-7.69 (m, 4H), 7.43-7.29 (m, 4H), 6.20 (broad s, 1H),
.72 (d, J ) 11.33 Hz, 1H), 5.59 (d, J ) 11.33 Hz, 1H), 4.80-4.67
m, 3H), 4.44 (t, J ) 6.21 Hz, 1H), 4.28-4.20 (q, J ) 7.31 Hz, 2H),
cis), 6.02 (d, J ) 11.7 Hz, 1H, cis), 5.94 (d, J ) 8.0 Hz, 1H, trans),
5.77 (d, 1H, cis), 5.76 (s, 2H, trans), 5.56 (d, J ) 11.7 Hz, 1H, cis),
5.15-5.10 (m, 1H, trans), 4.98-4.90 (m, 1H, cis), 4.53-4.25 (m, 5H),
1
3
13
1
.32 (d, J ) 6.94 Hz, 3H), 1.30 (t, J ) 7.31 Hz, 3H); C NMR (62.5
) δ 172.36, 168.78, 154.59, 154.17, 152.63, 150.34, 147.54,
47.40, 146.42, 146.38, 146.34, 146.16, 146.12, 146.09, 145.83, 145.63,
1.72 (d, J ) 7.3 Hz, 3H), 1.27 (t, J ) 6.6 Hz, 3H); C NMR (62.5
MHz, CDCl
3
3
MHz, CDCl ) δ 171.90, 169.54, 155.64, 153.79, 153.31, 152.00, 149.74,
1
1
1
1
1
1
6
147.56, 147.44, 146.48, 146.38, 146.35, 146.21, 146.14, 145.75, 145.59,
145.55, 145.45, 145.37, 145.32, 144.69, 144.58, 144.51, 144.45, 144.35,
144.29, 143.88, 143.70, 143.08, 142.78, 142.73, 142.69, 142.22, 142.17,
142.10, 142.07, 142.03, 141.83, 141.79, 141.31, 140.29, 139.68, 136.63,
135.94, 135.57, 135.26, 127.77, 127.11, 125.17, 120.03, 70.53, 69.87,
68.76, 67.27, 62.30, 58.83, 48.59, 47.13, 18.47, 14.18; MALDI-MS
45.60, 145.57, 145.52, 145.40, 145.37, 145.33, 144.98, 144.65, 144.58,
44.55, 144.39, 144.33, 143.70, 143.64, 143.14, 142.73, 142.65, 142.33,
42.28, 142.21, 142.19, 142.10, 142.04, 142.01, 141.95, 141.83, 141.73,
41.47, 141.40, 140.35, 140.23, 140.12, 139.66, 137.91, 136.74, 135.38,
34.99, 127.93, 127.89, 127.28, 127.24, 125.04, 120.18, 72.44, 70.22,
+
+
9.67, 68.17, 61.88, 58.89, 48.56, 47.30, 18.12, 14.17; MALDI-MS
C
83
H
24
N
2
O
5
(MW ) 1128), m/z 1166 [(M + K) ], 1151 [(M + Na) ],
+
+
C
H
83 24
N
2
O
5
(MW ) 1128), m/z 1151 [(M + Na) ]; UV-vis (CHCl
3
)
1129 [(M + H) ]; UV-vis (CHCl
3
) λmax (ꢀ) 428 (3380), 311 (34 800),
-3
λ
max (ꢀ) 696 (319), 429 (3070), 312 (34 300), 301 (36 800), 257
301 (36 800), 256 (118 200); CD (CHCl
653 (-1.70), 596 (-1.42), 548 (-0.74), 474 (-1.03), 428 (-4.63),
396 (2.58), 341 (-3.71). Anal. Calcd for C83 : C, 88.29; H,
2.14; N, 2.48. Found: C, 85.65; H, 1.83; N, 2.40.
c-(L-Ala-D-Fpr) (10a). A solution of 17 mg (0.015 mmol) of 9a
and 0.9 mL of piperidine in 6 mL of CH Cl was stirred at room
3
) λmax (θ
T
× 10 ) 690 (1.88),
-
3
(115 300); CD (CHCl
3
) λmax (θ
T
× 10 ) 695 (-0.96), 658 (3.12), 605
(
3
2.38), 515 (-0.86), 428 (3.78), 396 (-3.12), 384 (-2.36), 343 (6.80),
24 2 5
H N O
19 (-5.84).
Fmoc-L-Fpr-L-Ala-OEt (8b): 6 mg (21%); IR (KBr) (cm-1) 3423,
1
1
718, 1689, 1635, 1510, 1222, 1151, 757, 576, 528; H NMR (250
2
2
MHz, CDCl
3
) δ 7.77-7.69 (m, 4H), 7.43-7.27 (m, 4H), 6.05 (broad
temperature for 30 min, and then the solvent was removed in Vacuo.
s, 1H), 5.73 (d, J ) 11.70 Hz, 1H), 5.57 (d, J ) 11.70 Hz, 1H), 4.90-
The residue was purified by flash chromatography (eluant: toluene/
-1
4
2
.66 (m, 3H), 4.41 (t, J ) 6.21 Hz, 1H), 4.15-4.07 (q, J ) 6.94 Hz,
H), 1.50 (d, J ) 7.31 Hz, 3H), 1.21 (t, J ) 6.94 Hz, 3H); C NMR
) δ 172.09, 168.96, 154.99, 154.05, 152.53, 150.28,
2-propanol, 95:5), affording 5.7 mg (43%) of 10a: IR (KBr) cm 3435,
1
3
1
3
3378, 3255, 1678, 1431, 575, 528; H NMR (200 MHz, CDCl /DMSO,
(62.5 MHz, CDCl
3
5:1) δ 8.67 (s, 1H), 6.09 (d, J ) 12.20 Hz, 1H), 5.85 (s, 1H), 4.76 (d,
1
1
1
1
1
1
1
1
47.50, 147.40, 146.35, 146.33, 146.16, 146.09, 145.58, 145.56, 145.52,
45.37, 145.27, 144.90, 144.63, 144.54, 144.51, 144.40, 143.74, 143.60,
43.19, 143.07, 142.70, 142.67, 142.57, 142.29, 142.21, 142.10, 142.04,
41.98, 141.91, 141.80, 141.48, 141.38, 140.31, 140.17, 140.04, 139.76,
38.00, 136.88, 135.36, 134.88, 127.99, 127.94, 127.32, 127.27, 125.03,
J ) 12.20 Hz, 1H), 4.39 (m, 1H), 1.66 (d, J ) 6.93 Hz, 3H); H NMR
R
â
NOE (200 MHz, CDCl
3
/DMSO, 5:1) H( CH-Fpr):{H( CH
3
-Ala)}, 2.0;
(MW:
â
R
H( CH
3
-Ala):{H( CH-Fpr)}, absent; MALDI-MS C66
860), m/z 861 [(M + H) ]; UV-vis (CHCl
(3260), 311 (31 000), 255 (89 500); CD (CHCl
(1.71), 481 (-1.26), 428 (12.28), 398 (-5.79), 364 (-3.31), 326
(-4.22), 311 (3.30), 303 (-1.62), 287 (9.59); λcalcd 538, 516, 455, 4.04,
341, 315, 275, 253, 242.
H
88
N
2
O
2
+
3
) λmax (ꢀ) 688 (234), 428
) λmax (θ
-
3
3
T
× 10 ) 574
20.14, 72.36, 70.50, 69.65, 68.16, 61.70, 58.81, 48.30, 47.30, 18.57,
4.16; MALDI-MS C83
+
H N O
24 2 5
(MW ) 1128), m/z 1151 [(M + Na) ];
UV-vis (CHCl
3
) λmax (ꢀ) 696 (391), 429 (3620), 308 (38 000), 300
-
3
(
6
(
41 000), 257 (127 900); CD (CHCl
3
) λmax (θ
T
× 10 ) 695 (0.85),
58 (-2.14), 603 (-2.00), 515 (0.58), 428 (-4.71), 394 (0.98), 383
: C,
c-(L-Ala-L-Fpr) (10b).The synthesis was carried out as described
for compound 10a, starting from 17 mg (0.015 mmol) of 9b. Derivative
-
1
1.26), 343 (-7.12), 317 (3.79). Anal. Calcd for C83
H
24
N
2
O
5
10b: 4.5 mg (35%); IR (KBr) (cm ) 3435, 3386, 3259, 1686, 1439,
1
8
8.29; H, 2.14; N, 2.48. Found: C, 86.32; H, 1.96; N, 2.24.
Fmoc-L-Ala-Fpr-OEt (9a and 9b). A solution of H-Gly-OEt,
prepared by treating 120 mg (0.86 mmol) of HCl‚H-Gly-OEt with 95
µl (0.86 mmol) of NMM in 5 mL of CH Cl at room temperature, was
added to a solution of 300 mg (0.416 mmol) of C60 and 50 mg (1.67
575, 528; H NMR (200 MHz, CDCl /DMSO, 5:1) δ 8.73 (s, 1H),
3
6.15 (d, J ) 12.10 Hz, 1H), 5.95 (s, 1H), 4.85 (d, J ) 12.10 Hz, 1H),
1
4.54 (m, 1H), 1.75 (d, J ) 6.89 Hz, 3H); H NMR NOE (200 MHz,
R
R
R
2
2
CDCl
3
/DMSO, 5:1) H( CH-Fpr):{H( CH-Ala)}, 2.3; H( CH-Ala):{H-
(MW: 860), m/z 861 [(M
R
( CH-Fpr)}, absent; MALDI-MS C66
H
N O
8 2 2