378 Journal of Natural Products, 2009, Vol. 72, No. 3
Pettit et al.
0.136 mmol), anhydrous toluene (8 mL), Ph3P (97 mg, 0.372 mmol),
and DEAD (55 µL, 0.372 mmol); reaction was complete after the
mixture was stirred at 0 °C for several hours and then at room
temperature for several hours. Workup as before and flash CC of the
residue provided depsipeptide 22 (97 mg, 84%) as an oil: Rf 0.5 (2:1
mg, 63%) as a liquid: Rf 0.4 (2:1 hexane-EtOAc); IR (CHCl3) νmax
3307, 2963, 2927, 2359, 1750, 1655, 1534, 1370, 1145 cm-1; 1H NMR
(CDCl3, 400 MHz) δ 7.87 (1H, d, J ) 6.8 Hz), 7.70 (1H, d, J ) 5.6
Hz), 7.62 (1H, d, J ) 6.4 Hz), 7.56 (1H, d, J ) 7.6 Hz), 7.34-7.28
(5H, m), 7.08 (1H, d, J ) 6.0 Hz), 7.07 (1H, d, J ) 9.2 Hz), 5.29 (1H,
q, J ) 6.8 Hz), 5.25-5.22 (4H, m), 4.58 (1H, d, J ) 11.2 Hz), 4.55
(1H, d, J ) 11.6 Hz), 4.26 (1H, dd, J1 ) 5.6 Hz, J2 ) 8.4 Hz),
4.27-4.24 (1H, m), 4.10 (1H, dd, J1 ) 7.6 Hz, J2 ) 6.8 Hz), 4.02-3.98
(2H, m), 3.94 (1H, dd, J1 ) 5.6 Hz, J2 ) 8.0 Hz), 3.90 (1H, dd, J1 )
6.6 Hz, J2 ) 8.2 Hz), 2.34-2.22 (4H, m), 2.16-2.03 (4H, m),
1.79-1.58 (3H, m), 1.42 (3H, d, J ) 6.8 Hz), 1.41 (3H, d, J ) 6.8
Hz), 1.39 (9H, s), 1.38 (3H, d, J ) 6.8 Hz), 1.35-1.19 (4H, m), 1.02
(3H, d, J ) 6.2 Hz), 0.99-0.83 (51H, m); APT (CDCl3, 400 MHz) δ
174.9, 171.6, 171.0, 170.9, 170.86, 170.8, 170.46, 170.4, 170.3, 170.0,
137.2, -128.6, -128.1, -127.9, 81.4, -76.6, -76.5, -76.0, -73.1,
72.2, -70.5, -70.0, -60.2, -59.7, -59.6, -58.7, -57.9, 41.0, -36.8,
-36.7, -30.9, -30.1, 29.7, -29.3, -29.2, -29.0, -27.9, 26.3, 26.1,
-24.4, -23.1, -21.4, -19.7, -19.4, -19.36, -19.3, -19.2, -19.1,
-19.0, -18.9, -18.8, -18.4, -18.3, -17.4, -17.1, -14.1, -13.9,
-11.8, -11.7; HRMS (APCI) m/z 1317.809 [M + H]+, (calcd for
C68H113N6O19, 1317.806).
Depsipeptide 28. Cleavage of the benzyl group from 27 (55 mg)
was carried out as described above (see 17) using 20% Pd (OH)2/C
(40 mg) in CH3OH (5 mL) under hydrogen (overnight). Purification
by flash CC gave alcohol 28 (45 mg, 88%) as an oily solid: Rf 0.6 (1:1
hexane-EtOAc); IR (CHCl3) νmax 3306, 2966, 1750, 1656, 1534,
1181,1146 cm-1; 1H NMR (CDCl3, 400 MHz) δ 7.80 (1H, d, J ) 6.0
Hz), 7.65 (1H, d, J ) 8.8 Hz), 7.63 (1H, d, J ) 7.2 Hz), 7.57 (1H, d,
J ) 6.8 Hz), 7.55 (1H, d, J ) 6.4 Hz), 7.14 (1H, d, J ) 8.4 Hz), 5.33
(1H, q, J ) 7.2 Hz), 5.25 (1H, d, J ) 2.4 Hz), 5.20-5.17 (2H, m),
5.09 (1H, d, J ) 2.8 Hz), 4.50 (1H, dd, J1 ) 7.2 Hz, J2 ) 8.4 Hz),
4.21 (1H, dd, J1 ) 5.6 Hz, J2 ) 8.4 Hz), 4.10 (1H, dd, J1 ) 7.0 Hz,
J2 ) 6.9 Hz), 4.00-3.90 (4H, m), 2.32-2.03 (8H, m), 1.74-1.64 (3H,
m), 1.35 (9H, s), 1.37-1.35 (6H, m), 1.25 (3H, d, J ) 6.8 Hz),
1.32-1.14 (4H, m), 1.03-0.84 (54H, m); HRMS (APCI) m/z 1227.757
[M + H]+ (calcd for C61H107N6O19, 1227.759).
hexane-EtOAc); [R]28 +12 (c 0.51, CHCl3); IR (CHCl3) νmax 3320,
D
1
2971, 1752, 1671, 1532, 1624 cm-1; H NMR (CDCl3, 500 MHz) δ
7.51 (1H, d, J ) 7.0 Hz), 7.44 (1H, d, J ) 6.5 Hz), 7.37-7.31 (5H,
m), 7.12 (1H, d, J ) 8.5 Hz), 7.09 (1H, d, J ) 6.5 Hz), 5.33 (1H, q,
J ) 7.0 Hz), 5.28 (1H, d, J ) 2.5 Hz), 5.24 (1H, d, J ) 2.5 Hz), 4.61
(1H, d, J ) 11.0 Hz), 4.58 (1H, d, J ) 11.0 Hz), 4.31 (1H, dd, J1 )
6.0 Hz, J2 ) 8.0 Hz), 4.25 (1H, t, J ) 7.3 Hz), 4.19 (1H, t, J ) 7.3
Hz), 4.13 (1H, t, J ) 7.5 Hz), 4.05 (1H, q, J ) 6.8 Hz), 2.36-2.01
(6H, m), 1.45 (3H, d, J ) 6.5 Hz), 1.41 (3H, d, J ) 7.5 Hz), 1.40 (9H,
s), 1.35-1.26 (4H, m), 1.05-0.91 (36H, m); 13C NMR (CDCl3, 500
MHz) δ 174.7, 171.5, 170.9, 170.8, 170.4, 170.3, 169.5, 137.3, 128.6,
128.1, 127.9, 81.4, 76.9, 76.0, 72.2, 70.4, 59.3, 59.2, 58.6, 58.1, 37.0,
36.8, 30.5, 29.9, 29.8, 29.7, 29.4, 28.0, 26.1, 19.3, 19.2, 19.1, 19.0,
18.9, 18.6, 18.5, 18.4, 17.2, 14.1, 14.0, 11.8; APT (CDCl3, 500 MHz)
δ -174.7, -171.5, -170.9, -170.8, -170.4, -170.3, -169.5, -137.3,
128.6, 128.1, 127.9, -81.4, -76.8, 76.0, -72.2, 70.4, 59.3, 59.2, 58.6,
58.1, 37.0, 36.8, 30.5, 29.9, 29.8, 29.7, 29.4, 28.0, -26.1, 19.3, 19.2,
19.1, 19.0, 18.8, 18.6, 18.5, 18.4, 17.2, 14.1, 14.0, 11.8; HRMS (APCI)
m/z 933.5759 [M + H]+ (calcd for C49H81N4O13, 933.5800).
Depsipeptide 24. Selective deprotection of tert-butyl ester 22 (50
mg, 0.054 mmol) in anhydrous CH2Cl2 (3 mL) and TFA (3.5 mL) under
argon with cooling (ice bath) was performed as described above (see
16) to yield carboxylic acid 23. The product was obtained as a solid in
almost quantitative yield and was used without further purification in
a Mitsunobu reaction as described above (see 19) with alcohol 10 (29.6
mg, 0.064 mmol), anhydrous toluene (4.5 mL), Ph3P (42 mg, 0.162
mmol), and DEAD (25.6 µL, 0.162 mmol). The mixture was stirred at
0 °C for 5 h and at room temperature overnight (reaction complete by
TLC). Flash CC led to separation of the crude product (24, 48 mg,
1
69%) as a liquid: Rf 0.5 (2:1 hexane-EtOAc); H NMR (CDCl3, 300
MHz) δ 7.71 (1H, d, J ) 5.7 Hz), 7.67 (1H, d, J ) 7.8 Hz), 7.64 (1H,
d, J ) 6.3 Hz), 7.37-7.31 (5H, m), 7.13 (1H, d, J ) 8.4 Hz), 7.09
(1H, d, J ) 6.6 Hz), 6.82 (1H, d, J ) 8.7 Hz), 5.32-5.17 (6H, m),
4.60 (2H, s), 4.54-4.40 (2H, m), 4.33-3.96 (4H, m), 2.40-2.06 (9H,
m), 1.45-1.40 (18H, m), 1.32-1.25 (6H, m), 1.07-0.90 (54H, m);
HRMS (APCI) m/z 1317.8137 [M + H]+ (calcd for C68H113N6O19,
1317.8061).
Bacillistatin 2. To a cooled (ice bath) solution of tert-butyl ester 28
(38 mg, 0.031 mmol) in anhydrous CH2Cl2 (3 mL) under argon was
added TFA (3 mL) dropwise. The solution was stirred overnight at
room temperature to complete the reaction, and the solvent was removed
(in vacuo). Toluene (3 mL × 4) and CH2Cl2 (3 mL × 4) were added
successively to the residue, the solvent being removed after each
addition, to yield hydroxy acid 29 as a syrup/solid (almost quantitative
yield) that was used without further purification. Next, to a stirred
solution of MNBA (6.8 mg, 0.02 mmol) and DMAP (6.5 mg, 0.054
mmol) in anhydrous CH2Cl2 (13 mL) under argon was added dropwise
(over 3 h from a syringe) a solution of 29 (18 mg, 0.0154 mmol) in
anhydrous CH2Cl2 (2 mL). The mixture was stirred overnight and then
cooled to 0 °C before termination of the reaction with water (2 mL).
The mixture was extracted with CH2Cl2 (30 mL × 2), and the combined
extract was washed and dried as stated previously. The solvent was
removed, and flash CC of the residue afforded bacillistatin 2 (2, 15.6
mg, 88%) as a colorless solid: Rf 0.55 (4:1 hexane-EtOAc); mp
Depsipeptide 25. To a cooled (ice bath) solution of amine 6 (0.18
g, 0.47 mmol) in anhydrous CH2Cl2 (8 mL) that was stirring under
argon were added O-benzyl-D-lactic acid (0.10 g, 0.56 mmol), EDCl
(144 mg, 0.75 mmol), HOBt (101 mg, 0.75 mmol), and NMM (0.21
mL, 1.88 mmol). The mixture was stirred overnight and then reaction
was terminated and the mixture treated as described above (see 15).
Flash CC gave product 25 (0.23 g, 91%) as a liquid: Rf 0.3 (4:1
hexane-EtOAc); [R]26D +22.5 (c 0.59, CHCl3); 1H NMR (CDCl3, 300
MHz) δ 7.36-7.31 (5H, m), 7.10 (1H, d, J ) 8.4 Hz), 6.58 (1H, d, J
) 7.5 Hz), 5.21 (1H, dd, J1 ) 3.9 Hz, J2 ) 9.3 Hz), 4.62 (1H, d, J )
12.3 Hz), 4.57 (1H, d, J ) 12.3 Hz), 4.54 (1H, dd, J1 ) 5.1 Hz, J2 )
8.4 Hz), 4.34 (1H, dd, J1 ) 5.1 Hz, J2 ) 8.4 Hz), 4.01 (1H, q, J ) 6.9
Hz), 2.28-2.14 (2H, m), 1.82-1.66 (3H, m), 1.44 (9H, s), 1.42 (3H,
d, J ) 7.2 Hz), 0.99-0.91 (18H, m); HRMS (APCI) m/z 549.3549 [M
+ H]+ (calcd for C30H49N2O7, 549.3540).
143-146 °C; [R]20 +33.9 (c 0.44, CHCl3); IR (CHCl3) νmax 3004,
D
1
2965, 1757, 1659, 1539, 1184 cm-1; H NMR (CDCl3, 500 MHz) δ
7.92 (1H, d, J ) 8.0 Hz), 7.83 (1H, d, J ) 8.0 Hz), 7.80 (1H, d, J )
5.5 Hz), 7.74 (1H, d, J ) 7.0 Hz), 7.70 (1H, d, J ) 6.5 Hz), 7.61 (1H,
d, J ) 5.5 Hz), 5.25 (1H, dd, J1 ) 7.0 Hz, J2 ) 10.0 Hz), 5.18 (1H,
d, J ) 2.5 Hz), 5.28-5.17 (3H, m), 5.11 (1H, d, J ) 2.5 Hz), 4.22
(1H, dd, J1 ) 9.5 Hz, J2 ) 8.5 Hz), 4.12 (1H, dd, J1 ) 8.0 Hz, J2 )
9.5 Hz), 2.33-2.09 (8H, m), 1.77-1.72 (3H, m), 1.45 (3H, d, J ) 7.0
Hz), 1.44 (3H, d, J ) 7.0 Hz), 1.43 (3H, d, J ) 6.5 Hz), 1.38-1.27
(4H, m), 1.08-0.87 (54H, m); APT (CDCl3, 500 MHz) δ 172.6, 172.2,
172.17, 172.13, 171.8, 171.6, 171.5, 170.8, 170.23, 170.17, 169.8,
-76.6, -73.4, -70.6, -70.5, -70.1, -60.6, -60.5, -59.6, -59.15,
-59.07, -58.4, 40.7, -36.9, -36.7, -28.7, -28.5, -28.42, -28.39,
-28.35, -28.3, 26.22, 26.19, -24.5, -23.1, -21.5, -19.9, -19.8,
-19.6, -19.45, -19.41, -19.38, -19.3, -19.23, -19.17, -19.14,
-19.12, -19.0, -17.3, -17.1, -16.9, -14.2, -11.8; HRMS (APCI)
m/z 1153.691 [M + H]+ (calcd for C61H107N6O19, 1153.686). The
synthetic specimen of bacillistatin 2 was found to be identical (TLC,
Depsipeptide 26. Selective benzyl group cleavage from 25 (230 mg)
was carried out as described above (see 21) using 20% Pd (OH)2/C
(60 mg) in CH3OH (15 mL) under hydrogen (overnight) to provide,
following flash CC, alcohol 26 (0.18 g, 95%) as a syrup/solid: Rf 0.4
(1:1 hexane-EtOAc); IR (CHCl3) νmax 3310 (br), 2967, 1739, 1661,
1
1530, 370, 1150 cm-1; H NMR (CDCl3, 300 MHz) δ 6.90 (1H, d, J
) 8.4 Hz), 6.58 (1H, d, J ) 8.7 Hz), 5.21 (1H, m), 4.56 (1H, dd, J1 )
6.3 Hz, J2 ) 9.0 Hz), 4.41 (1H, dd, J1 ) 5.0 Hz, J2 ) 8.8 Hz), 4.27
(1H, dq, J1 ) 6.6 Hz, J2 ) 6.6 Hz), 3.08 (1H, d, J ) 6.6 Hz), 2.30-2.14
(2H, m), 1.84-1.68 (3H, m), 1.46 (3H, d, J ) 7.2 Hz), 1.45 (9H, s),
1.01-0.91 (18H, m).
Depsipeptide 27. Alcohol 26 (31 mg, 0.068 mmol) was condensed
with carboxylic acid 23 (0.057 mmol) in anhydrous toluene (4.5 mL
at 0 °C under argon) under Mitsunobu reaction conditions (see 19)
with Ph3P (45 mg, 0.171 mmol) and DEAD (27 µL, 0.171 mmol). The
mixture was stirred at 0 °C for 5 h and at room temperature overnight,
by which time the reaction was complete, and the product mixture was
extracted (see 19). Separation by flash CC yielded depsipeptide 27 (35
1
1
HPLC, H NMR, 13C NMR, HRMS) with the natural product.
Compound 31. The tert-butyl group was removed from 20 (94 mg,
0.172 mmol) in anhydrous CH2Cl2 (2 mL) and TFA (3 mL) (ice bath
under argon) as described earlier (see 16) to yield carboxylic acid 30.