8550 J . Org. Chem., Vol. 63, No. 23, 1998
Notes
g, 85%): 1H NMR (DMSO-d6) δ 7.8-7.0 (m, 13H), 4.1-3.9 (m,
8H), 3.0 (m, 5H), 2.8 (m, 1H), 1.14 (s, 18H); 31P NMR (DMSO-
d6) δ -5.95 (s); 13C NMR (DMSO-d6) δ 205.9, 174.4, 156.7, 149.4,
144.6, 141.5, 136.3, 131.1, 128.4, 127.9, 126.1, 120.9, 120.3, 67.1,
66.4, 56.6, 47.5, 46.8, 36.7, 29, 27.7; FAB MS (>0, G-T) m/z
with 5% KHSO4 (4 × 50 mL), dried over Na2SO4, and concen-
trated to dryness in vacuo. Purification by silica gel column
chromatography using a stepwise gradient of MeOH (0-5%) in
CH2Cl2 afforded compound 4b (300 mg, 86%) as an oil: 1H NMR
(DMSO-d6) δ 8.3-6.9 (m, 24 H), 4.49 (m, 1H), 4.2-4.0 (m, 9H),
3.7-3.5 (m, 4H), 3.1-2.9 (m, 6H), 2.7 (m, 2H), 1.5 (m, 1H), 1.4
(m, 2H), 1.14 (s, 18H), 0.83 (dd, 6H); 31P NMR (DMSO-d6) δ
-5.99 (s); FAB MS (0<, G-T) m/z 1145 (M + H)+, 145 ((CH3)3-
CC(O)S(CH2)2)+, 120 (PhCH2CHdNH2)+, 86 ((CH3)2CHCH2CHd
NH2)+, 30 (CH2dNH2)+; FAB MS (<0, G-T) m/z 1143 (M - H)-.
ter t-bu tylSATE P h osp h otr iester Der iva tive of Leu -En -
k ep h a lin a m id e (1). Boc Clea va ge Con d ition s. The Boc
peptide phosphotriester 4a (75 mg, 0.07 mmol) was treated with
a solution of TFA/CH2Cl2 (0.8 mL, 50:50, v/v) and anisole (0.04
mL, 0.37 mmol). After 15 min, addition of Et2O led to a
formation of a white precipitate. The precipitate was collected
by filtration, washed several times with Et2O, and purified by
silica gel column chromatography using a stepwise gradient of
MeOH (0-15%) in CH2Cl2 to afford the trifluoroacetate salt of
the target compound 1 as a white solid (65 mg, 83%): 1H NMR
(DMSO-d6) δ 8.8 (m, 1H), 8.18 (m, 1H), 8.1-7.9 (m, 5H), 7.3-
7.1 (m, 10H), 6.90 (s, 1H), 4.5 (m, 1H), 4.13 (m, 5H), 4.05 (m,
1H), 3.8-3.6 (m, 4H), 3.13 (t, 4H, J ) 6.3 Hz), 3.1-2.7 (m, 4H),
1.55 (m, 1H), 1.44 (m, 2H), 1.17 (s, 18H), 0.84 (dd, 6H); 31P NMR
(DMSO-d6) δ -6.03 (s); FAB MS (0<, G-T) m/z 923 (M + H)+,
793 (M - Leu-NH2)+, 589 (M - Gly-Phe-Leu-NH2)+, 145 ((CH3)3-
CC(O)S(CH2)2)+, 120 (PhCH2CHdNH2)+, 86 ((CH3)2CHCH2CHd
NH2)+, 30 (CH2dNH2)+; FAB MS (<0, G-T) m/z 1035 (M + TFA
- H)-, 921 (M - H)-. HRMS: calcd for C42H64N6O11PS2
923.3812, found 923.3828.
F m oc Clea va ge Con d ition s. The Fmoc peptide phospho-
triester 4b (200 mg, 0.17 mmol) was treated with a solution of
2% DBU in CH2Cl2 (2.5 mL). After 15 min, addition of Et2O led
to a formation of white precipitate. The precipitate was collected
by filtration, washed several times with Et2O, and purified by
chromatography using a stepwise gradient of MeOH (0-15%)
in CH2Cl2 to afford the target compound 1 as a white solid (133
mg, 82%): 1H NMR (DMSO-d6) δ 8.2 (m, 1H), 8.1-8.0 (m, 2H),
7.3-7.0 (m, 10H), 6.98 (s, 2H), 4.47 (m, 1H), 4.13 (m, 5H), 3.8-
3.6 (m, 4H), 3.4 (m, 1H, partially obscured by water), 3.12 (t,
4H, J ) 6.2 Hz), 3.0-2.5 (m, 4H), 2.0 (m, 2H), 1.55 (m, 1H),
1.46 (m, 2H), 1.07 (s, 18H), 0.84 (dd, 6H); 31P NMR (DMSO-d6)
δ -5.94 (s); FAB MS (0<, G-T) m/z 923 (M + H)+, 793 (M -
Leu-NH2)+, 589 (M - Gly-Phe-Leu-NH2)+, 145 ((CH3)3CC(O)S-
(CH2)2)+, 120 (PhCH2CHdNH2)+, 86 ((CH3)2CHCH2CHdNH2)+;
FAB MS (<0, G-T) m/z 921 (M - H)-. HRMS: calcd for
C42H64N6O11PS2 923.3812, found 923.3834.
772 (M + H)+; FAB MS (<0, G-T) m/z 770 (M - H)-; [R]20
)
D
-3 (c 1.0, DMSO); HPLC tR 28.3 min. Anal. Calcd for C38H46
-
NO10PS2: C, 59.12; H, 6.01; N, 1.81. Found: C, 59.12; H, 5.99;
N, 1.92.
O-[bis(S-p iva loyl-2-th ioeth yl)]-L-p h osp h otyr osin e (2c).
From 2a . Compound 2a (100 mg, 0.15 mmol) was treated with
a solution of trifluoroacetic acid (TFA) in CH2Cl2 (5 mL, 50:50,
v/v) and stirred for 10 min. The solution was evaporated under
reduced pressure and coevaporated several times with hexane.
The residue was purified by silica gel column chromatography
using a stepwise gradient of MeOH (0-40%) to afford 2c as a
white solid (62 mg, 87%): 1H NMR (DMSO-d6) δ 7.27 (d, 2H, J
) 8.5 Hz), 7.10 (d, 2H, J ) 8.2 Hz), 4.13 (q, 4H, J ) 13.4 and 6.8
Hz), 3.4 (m, 1H, partially obscured by water), 3.1 (m, 5H), 2.8
(m, 1H), 1.14 (s, 18H); 31P NMR (DMSO-d6) δ -5.96 (s); 13C NMR
(DMSO-d6) δ 205.9, 170.1, 149.6, 135.6, 131.7, 120.5, 67.1, 56.3,
46.9, 37.1, 28.9, 27.7; FAB MS (>0, G-T) m/z 642 (M + G +
H)+, 550 (M + H)+, 145 ((CH3)3CC(O)S(CH2)2)+; FAB MS (<0,
G-T) m/z 1097 (2M - H)-, 640 (M + G - H)-, 548 (M - H)-;
[R]20 ) -7 (c 1.0, DMSO); HPLC tR 22.1 min. Anal. Calcd for
D
C
23H36NPO8S2: C, 50.25; H, 6.60; N, 2.55. Found: C, 50.31; H,
6.85; N, 2.66.
From 2b. Compound 2b (70 mg, 0.09 mmol) was treated with
a solution of 2% 1,8-diazabicyclo[5.4.0]undec-7-ene (1.5-S) (DBU)
in CH2Cl2 (3.5 mL) and stirred for 10 min. The solution was
evaporated under reduced pressure. The residue was purified
by column chromatography using a stepwise gradient of MeOH
(0-40%) in CH2Cl2 to give 2c as a white solid (40 mg, 80%).
The physicochemical properties were identical to those obtained
from 2a in Boc cleavage conditions.
P ep tid e Der iva tive 4a . The tetrapeptide Boc-Gly-Gly-Phe-
Leu-NH2 3 (150 mg, 0.30 mmol) was added to a solution of TFA/
CH2Cl2 (2 mL, 50:50, v/v) and stirred for 30 min. The solution
was evaporated under reduced pressure and coevaporated
several times with hexane. The residue was dissolved in CH2-
Cl2 (4 mL) and then DIEA (0.26 mL, 1.52 mmol), BOP reagent
(135 mg, 0.37 mmol), and compound 2a (200 mg, 0.31 mmol)
were successively added. The solution was stirred at room
temperature for 2 h, and the solvent was evaporated under
reduced pressure. The oily residue was dissolved in EtOAc (50
mL), washed with an aqueous solution of 5% KHSO4 (4 × 50
mL), dried over Na2SO4, and concentrated to dryness in vacuo.
Purification by silica gel column chromatography using a step-
wise gradient of MeOH (0-5%) in CH2Cl2 afforded compound
4a (264 mg, 84%) as an oil: 1H NMR (DMSO-d6) δ 8.2 (m, 1H),
8.1 (m, 2H), 7.9 (m, 1H), 7.2-6.9 (m, 12H), 4.5 (m, 1H), 4.1 (m,
6H), 3.7-3.6 (m, 4H), 3.12 (t, 4H, J ) 6.3 Hz), 3.0-2.7 (m, 4H),
1.55 (m, 1H), 1.45 (m, 2H), 1.26 (s, 9H), 1.16 (s, 18H), 0.84 (dd,
6H); 31P NMR (DMSO-d6) δ -6.0 (s); FAB MS (0<, G-T) m/z
1023 (M + H)+, 145 ((CH3)3CC(O)S(CH2)2)+, 120 (PhCH2CHd
NH2)+; FAB MS (<0, G-T) m/z 1021 (M - H)-.
P ep tid e Der iva tive 4b. The tetrapeptide Boc-Gly-Gly-Phe-
Leu-NH2 3 (150 mg, 0.30 mmol) was added to a solution of TFA/
CH2Cl2 (2 mL, 50:50, v/v) and stirred for 30 min. The solution
was evaporated under reduced pressure and coevaporated
several times with hexane. The residue was dissolved in CH2-
Cl2 (4 mL), and then DIEA (0.26 mL, 1.52 mmol), BOP reagent
(135 mg, 0.37 mmol), and compound 2b (254 mg, 0.33 mmol)
were successively added. The solution was stirred at room
temperature for 2 h and then evaporated under reduced pres-
sure. The oily residue was dissolved in EtOAc (50 mL), washed
Ack n ow led gm en t. These investigations were sup-
ported by grants from CNRS and “Agence Nationale de
Recherches sur le SIDA” (ANRS, France). One of us
(C.M.) is particularly grateful to ANRS for a postdoc-
toral fellowship.
Su p p or tin g In for m a tion Ava ila ble: The detailed syn-
thesis of 3, a full listing of NMR data accompanied by
subjective peak assignments, and procedures used for the
stability study of SATE phosphotriester structures 2a ,b and
the preliminary enzymatic stability of 1 (4 pages). This
material is contained in libraries on microfiche, immediately
follows this article in the microfilm version of the journal, and
can be ordered from the ACS; see any current masthead page
for ordering information.
J O980437D