L. Ferron et al. / Tetrahedron 67 (2011) 6036e6044
6043
1H, NH, J¼6.0 Hz), 8.22 (s, 1H, NH); 13C NMR ((CD3)2O) 20.8, 23.5,
138.0, 138.5, 138.8, 139.0, 139.1, 159.3, 171.0, 171.8, 175.7, 176.0;
20
45.2, 62.6, 66.8, 126.6, 128.5, 129.1, 129.6, 137.9, 138.8, 136.8, 157.5,
[a
]
ꢀ25.0 (c 0.5, MeOH). HRMS (ESI): calcd for C32H37N4O7
D
20
169.6, 174.0; [
a]
ꢀ35.0 (c 0.5, MeOH). HRMS (ESI): calcd for
(MþHþ) 589.2662; found 589.2678.
C20H23N2O5 (MþDHþ) 371.1607; found 371.1591.
4.8. Preparation of protected octapeptide Z-(Gly-(R)-MPG)4-
4.7. Preparation of tetrapeptide Z-(Gly-(R)-MPG)2-OMe
OMe
The N-protected dipeptide (100 mg, 0.27 mmol) and the coupling
reagent HATU (1.1 equiv) were introduced in a 20 mL flask under
nitrogen. Freshly distilled THF (10 mL) was then added and the
mixture was stirred for 10 min at 0 ꢂC. Diisopropylethylamine
(0.104 mL, 2.2 equiv) was then added dropwise. After stirring at room
temperature for 6 h, HCl,H-Gly-(R)-MPG-OMe (85 mg, 1.1 equiv) and
The N-protected tetrapeptide (2.57 g, 4.36 mmol) and the coupling
reagent HATU (1.1 equiv) were introduced in a 250 mL flask under
nitrogen. Freshly distilled THF (100 mL) was then added and the
mixture was stirred for 10 min at 0 ꢂC. Diisopropylethylamine
(1.67 mL, 2.2 equiv) was then added dropwise. After stirring at room
temperaturefor8h, HCl,H-(Gly-(R)-MPG)2-OMe(2.42g,1.1equiv)and
diisopropylethylamine (0.84 mL, 1.1 equiv) were added and the mix-
ture was stirred overnight at room temperature. HATU (0.5 equiv) and
diisopropylethylamine (1.1 equiv) were added and the mixture was
stirred at room temperature for 2 days. HATU (0.5 equiv) and diiso-
propylethylamine (1.1 equiv) were one more time added and the
mixture was stirred at room temperature for 3 days. Solvent was re-
moved and dichloromethane (10 mL) was added. Organic layer was
washed with 1 M aqueous NaHCO3 (2ꢁ30 mL),10% aqueous citric acid
(2ꢁ30 mL), and water (30 mL), dried (MgSO4), and concentrated. Flash
chromatography (AcOEt/MeOH 99/1) gave pure octapeptide Z-(Gly-
(R)-MPG)4-OMe as a white powder (3.22 g, 71%). Mp 180 ꢂC (dec);
retention time 17.07 min (HPLC, ACN/H2O/TFA: 60/40/0.1); 1H NMR
(DMSO-d6) 1.67 (s, 3H, CH3), 1.70 (s, 3H, CH3), 1.73 (s, 3H, CH3), 1.80 (s,
3H, CH3), 2.25e2.28 (m, 12H, CH3), 3.51 (s, 3H, OCH3), 3.59e3.77 (m,
8H, CH2), 5.03 (s, 2H, OCH2), 7.08e7.34 (m, 21H, Ar), 7.51 (t, 1H, NH,
J¼5.6 Hz), 7.95 (t,1H, NH, J¼5.6 Hz), 8.07e8.10 (m, 5H, NH), 8.35 (s,1H,
NH); 13C NMR (MeOD) 21.0 (4C), 23.5, 24.2, 24.8, 25.0, 44.5, 45.2 (3C),
53.1, 62.9, 63.6, 64.1, 64.2, 67.9, 126.9, 127.1 (2C), 127.4, 128.9, 129.1,
diisopropylethylamine (52 mL,1.1 equiv) were added and the mixture
was stirred at room temperature overnight. Solvent was removed
and dichloromethane (10 mL) was added. The organic layer was
washed with1 M aqueous NaHCO3 (2ꢁ10 mL),10%aqueouscitric acid
(2ꢁ10 mL), and water (20 mL), dried (MgSO4), and concentrated.
Flash chromatography (AcOEt 100%) gave pure tetrapeptide Z-(Gly-
(R)-MPG)2-OMe as a white solid (130 mg, 80%). Mp 226 ꢂC; retention
time 19.04 min (HPLC, ACN/H2O/TFA: 40/60/0.1); 1H NMR (CDCl3)
1.92 (s, 3H, CH3), 1.98 (s, 3H, CH3), 2.23 (s, 3H, CH3), 2.31 (s, 3H, CH3),
3.64 (s, 3H, OCH3), 3.85e3.99 (m, 4H, CH2), 4.81 (s, 2H, OCH2), 5.67 (t,
1H, NH, J¼4.2 Hz), 6.68 (t, 1H, NH, J¼4.9 Hz), 7.08e7.31 (m, 13H, Ar),
7.71 (s,1H, NH), 8.02 (s,1H, NH); 13C NMR (CDCl3) 21.0, 21.1, 22.4, 23.4,
44.4, 45.2, 53.1, 62.9, 63.6, 67.8,126.8,127.3,128.8,129.0,129.4,129.9,
130.2, 138.1, 138.6, 138.9, 139.0 (2C), 159.0, 171.0, 171.7, 174.4, 175.3;
20
[a]
ꢀ38.4 (c 0.5, CH2Cl2); C33H38N4O7 (602.68): calcd C 65.77, H
D
6.36, N 9.30; found C 65.75, H 6.62, N 9.03; IR (cmꢀ1) 3312, 3064,
3030, 2997, 1725, 1665. Yield¼80%.
4.7.1. N-Deprotection of tetrapeptide Z-(Gly-(R)-MPG)2-OMe. To
a suspension of the protected tetrapeptide (3.72 g, 6.17 mmol) and
palladium (10% on charcoal, 521 mg, 11%) in methanol (60 mL) was
added 5e6 N HCl in iPrOH (1.6 mL) at room temperature. The flask
was purged with hydrogen and the reaction mixture was stirred at
room temperature under atmospheric pressure of hydrogen. When
the conversion was higher than 99% (HPLC), the reaction mixture
was filtered on Celite. Solvent was removed and diethylether
(20 mL) was added. Filtration and drying of the precipitate gave
pure deprotected tetrapeptide hydrochloride HCl,H-(Gly-(R)-
MPG)2-OMe as a white powder (3.11 g, quant.). Mp 180 ꢂC (dec);
retention time 3.16 min (HPLC, ACN/H2O/TFA: 50/50/0.1); 1H NMR
(MeOD) 1.84 (s, 3H, CH3), 1.92 (s, 3H, CH3), 2.27 (s, 3H, CH3), 2.30 (s,
3H, CH3), 3.61 (s, 3H, OCH3), 3.66e3.91 (m, 4H, CH2), 7.11e7.15 (m,
4H, Ar), 7.29e7.37 (m, 4H, Ar), 8.06e8.08 (m, 1H, NH); 13C NMR
(MeOD) 20.8, 23.4, 25.0, 41.7, 44.2, 53.0, 62.7, 63.9, 126.7, 127.0,
129.8, 130.0, 137.9, 138.5, 138.6, 138.8, 166.9, 170.6, 174.2, 174.8;
129.5, 129.9, 130.1, 130.2, 130.3, 137.9, 138.1, 138.6, 138.7, 138.8, 138.9,
20
139.0, 139.1, 159.1, 171.1, 171.7, 171.8, 174.4, 175.4, 175.7, 175.8; [a]
D
ꢀ6.0 (c 0.5, CH2Cl2); C57H66N8O11 (1039.18): calcd C 65.88, H 6.40, N
10.78; found C 65.85, H 6.95, N 10.73; IR (cmꢀ1) 3305, 3025, 1664.
4.8.1. C-Deprotection of octapeptide Z-(Gly-(R)-MPG)4-OMe. To
a cold (0 ꢂC) mixture of the protected octapeptide (300 mg,
0.3 mmol), water (3 mL), and iPrOH (8 mL) was added 2 N aqueous
sodium hydroxide (315
mL, 3 equiv). The reaction mixture was
stirred for 5 h at room temperature. After removal of the solvents,
methanol (10 mL) was added and the solution was stirred for 1 h at
room temperature and 5 min at 0 ꢂC. Aqueous citric acid (10%) was
added dropwise until pH¼4 at 0 ꢂC. Methanol was removed and
white precipitate was filtered and dried in a desiccator containing
P2O5 to give deprotected octapeptide Z-(Gly-(R)-MPG)4-OH as
a white powder (240 mg, 81%). Mp 220 ꢂC; retention time
10.69 min (HPLC, ACN/H2O/TFA: 60/40/0.1); 1H NMR (DMSO-d6)
1.70 (s, 3H, CH3), 1.72 (s, 3H, CH3), 1.74 (s, 3H, CH3), 1.80 (s, 3H, CH3),
2.25e2.27 (m, 12H, CH3), 3.59e3.75 (m, 8H, CH2), 5.03 (s, 2H, OCH2),
7.06e7.33 (m, 21H, Ar), 7.49 (t, 1H, NH, J¼5.6 Hz), 7.94e8.07 (m, 6H,
NH), 8.34 (s, 1H, NH), 12.64 (s, 1H, COOH); 13C NMR (DMSO-d6) 20.7,
23.2, 23.8, 24.3, 24.8, 43.3, 43.7, 44.2, 61.0, 61.7, 61.9, 62.0, 65.7,
126.1, 126.3, 127.8, 127.9, 128.5, 128.6, 128.8, 128.9, 133.5, 136.4,
136.4,136.5,137.1,138.1,138.8,138.9,156.7,168.2,168.9,169.0,172.6,
[
a]
20 ꢀ30.2 (c 0.5, MeOH); C25H33ClN4O5 (505.01): calcd C, 59.46; H,
D
6.59; N, 11.09; found: C, 59.25; H, 6.35; N, 10.87.
4.7.2. C-Deprotection of tetrapeptide Z-(Gly-(R)-MPG)2-OMe. To
a cold (0 ꢂC) mixture of the protected tetrapeptide (3.43 g,
i
5.7 mmol), water (25 mL), and PrOH (80 mL) was added 2 N
aqueous sodium hydroxide (6.87 mL, 3 equiv). The reaction mixture
was stirred for 2 h at room temperature. Solvents were removed
and water (30 mL) was added. The aqueous layer was washed with
diethylether (2ꢁ5 mL). At 0 ꢂC, 1 N HCl was added dropwise until
pH¼3. White precipitate was filtered and dried (P2O5) to give
deprotected tetrapeptide Z-(Gly-(R)-MPG)2-OH as a white powder
(2.83 g, 85%). Mp 147 ꢂC; retention time 9.93 min (HPLC, ACN/H2O/
TFA: 40/60/0.1); 1H NMR (MeOD) 1.86 (s, 3H, CH3), 1.92 (s, 3H, CH3),
2.29 (s, 3H, CH3), 2.30 (s, 3H, CH3), 3.69e3.87 (m, 4H, CH2), 5.10 (s,
2H, OCH2), 7.09e7.37 (m, 13H, Ar), 8.06 (s, 1H, NH), 8.21 (t, 1H, NH,
J¼5.3 Hz), 8.51 (s, 1H, NH); 13C NMR (MeOD) 21.1, 23.6, 24.1, 44.8,
45.4, 63.1, 63.7, 68.0, 127.0, 127.3, 128.9, 129.1, 129.6, 129.9, 130.4,
172.8, 172.9; 173.6; [
a
]
20 ꢀ20.4 (c 0.5, DMSO). HRMS (ESI): calcd for
D
C56H65N8O11 (MþHþ) 1025.4773; found 1025.4758.
4.9. Preparation of fully deprotected octapeptides HCl,H-(Gly-
(R)-MPG)4-OH
A suspension of the N-protected octapeptide (1.91 g, 1.86 mmol)
and palladium (10% on charcoal, 157 mg, 11%) in methanol (50 mL)
was purged with hydrogen, then stirred overnight at room temper-
ature under an atmospheric pressure of hydrogen. When reaction
was complete (HPLC), 5e6 N HCl in iPrOH (1.2 equiv) was added. The