6796 J . Org. Chem., Vol. 62, No. 20, 1997
Fehrentz et al.
Boc-P h e-Va l-Ala -H 10: 1.5 g of peptidyl-resin (0.66 mmol),
prepared from the Weinreb amide modified MBHA by succes-
sive couplings and deprotection steps (Boc-Ala-OH, Boc-Val-
OH, and Boc-Phe-OH were coupled using BOP as activating
reagent) according to the general procedure used for SPPS,
was suspended in anhydrous THF and placed in an ice bath.
LiAlH4 (114 mg; 5 mol equiv) was added, and the reaction was
stirred for 30 min. The reaction was then hydrolyzed with a
1 M potassium hydrogenosulfate solution. The resulting
mixture was filtered, and the resin was washed twice with
dichloromethane. The solvents were recovered, diluted with
dichloromethane, and washed with a 1 M potassium hydro-
genosulfate solution and a saturated solution of sodium
hydrogenocarbonate and sodium chloride. After drying and
concentration of the solvent in vacuo, a white powder was
obtained (crude yield: 180 mg ) 71%). The aldehydic peptide
Boc-Phe-Val-Ala-H 10 was purified either by silica gel chro-
matography with an isocratic solution of AcOEt/hexane: 7/3
containing 0.1% pyridine (yield 45%) or by reversed phase
HPLC in a gradient mode from 100% A to 100% B in 50 min
of solutions A (H2O with TFA 0.1%) and B (CH3CN with TFA
0.1%) as solvent system (yield after lyophilization: 40%).
1H-NMR (CDCl3): δ 0.9 (dd, 6H, CH3), 1.35 (d, 3H, CH3),
1.4 (s, 9H, Boc), 2.3 (m, 1H, CH), 3.2 (qd, 2H, CH2), 4.1 (m,
1H, CH), 4.2 (m, 1H, CH), 4.35 (m, 1H, CH), 4.95 (m, 1H, NH),
6.45 (d, 1H, NH), 6.7 (m, 1H, NH), 7.25 (m, 5H, Ar), 9.52
(before purification: s, 1H, CHO) 9.49-9.52 (after purifica-
tion: 2s, 1 H, CHO). [M + H]+: 330.
placed in an ice-water bath. Three equivalents (2.45 mL) of
LiAlH(OtBu)3 (1 M in THF, Aldrich)) was added, and the
reaction mixture was stirred for 1 h. The reaction was then
hydrolyzed with a 1 M potassium hydrogenosulfate solution.
The resulting mixture was filtered, and the resin was washed
twice with dichloromethane. The solvent was collected, diluted
with dichloromethane, and washed with a potassium hydro-
genosulfate solution and a saturated solution of sodium
bicarbonate and sodium chloride. After drying and concentra-
tion in vacuo a white powder was obtained (crude yield: 210
1
mg ) 60%). The H NMR study revealed the presence of 25%
of the corresponding alcohol and 75% of the desired aldehydic
peptide. These compounds could be easily separated by
preparative HPLC, but with resulting partial racemization of
the R carbon of the aldehydic function.
1H NMR Ch em ica l Sh ifts (p p m ) Obser ved for th e P As
Syn th esized Both via Wein r eb Lin k er a n d via P h en yl
Ester Lin k er . The aldehydic tripeptides described were
purified by reversed phase HPLC and revealed two aldehydic
peaks.
N-r-(ter t-Bu t oxyca r b on yl)-L-p h en yla la n yl-L-va lyl-L-
a la n in a l: 1H-NMR (CDCl3): 0.9 (dd, 6Η, CH3), 1.35 (d, 3H,
CH3), 1.4 (s, 9H, Boc), 2.3 (m, 1H, CH), 3.2 (qd, 2H, CH2), 4.1
(m, 1H, CH), 4.2 (m, 1H, CH), 4.35 (m, 1H, CH), 4.95 (m, 1H,
NH), 6.45 (d, 1H, NH), 6.7 (m, 1H, NH), 7.25 (m, 5H, Ar), 9.49-
9.52 (2s, 1 H, CHO).
N-r-(ter t-Bu toxycar bon yl)-L-leu cyl-L-leu cyl-N-E-(2-ch lo-
r oben zyloxyca r bon yl)-L-lysin a l: 1H-NMR (CDCl3): 0.9 (dd,
12H, CH3), 1.5 (s, 9H, Boc), 1.8 (m, 2H, CH2), 1.95 (m, 2H,
CH2), 3.2 (m, 2H, CH2), 4.1 (m, 1H, CH), 4.3-4.5 (m, 2H, CH),
4.85 (m, 1H, NH), 5.2 (s, 2H, CH2), 5.35 (m, 1H, NH), 6.5 (d,
1H, NH), 6.9 (d, 1H, NH), 7.4 (m, 4H, Ar), 9.49-9.51 (2s, 1H,
CHO).
N-r-Acetyl-L-leu cyl-L-va lyl-N-E-(2-ch lor oben zyloxyca r -
bon yl)-L-lysin a l: 1H-NMR (DMSO-d6): 0.85 (2 dd, 12H, CH3),
1.29 (m, 2H, CH2), 1.42 (m, 4H, 2CH2), 1.57 (m, 3H, 1.5CH2),
1.74 (m, 1H, 0.5CH2), 1.83 (s, 3H, CH3CO), 1.98 (m, 1H, CH),
2.98 (m, 2H, CH2), 4.04 (m, 1H, CH), 4.18 (m, 1H, CH), 4.32
(q, 1H, CH), 5.08 (s, 2H, CH2), 7.31 (t, 1H, NH), 7.36 (m, 2H,
Ar), 7.46 (m, 2H, Ar), 7.71 (d, 1H, NH), 8.00 (d, 1H, NH), 8.30
(d, 1H, NH), 9.37 (s, 1H, CHO).
N-r-(Ben zyloxyca r bon yl)-L-va lyl-L-p h en yla la n in a l: 1H-
NMR (CDCl3): 0.9 (dd, 6H, CH3), 2.1 (m, 1H, CH), 3.15 (d,
2H, CH2), 4 (m, 1H, CH), 4.7 (m, 1H, CH), 5.1 (s, 2H, CH2),
5.3 (m, 1H, NH), 7.2 (d, 1H, NH), 7.2-7.4 (m, 10H, Ar), 9.61
(s, 1H, CHO).
P h en yl Ester Ba sed Lin k er . Syn th esis of th e F ir st
N-P r otected Am in o Acid Ester ified by th e Lin k er . p-
Hydroxybenzoic acid (4 mmol) and DIEA (8 mmol) were
dissolved in DCM (20 mL), and then 1.5 equiv of the Boc-
UNCA was added to the solution. After 1.5 h, the solvent was
concentrated in vacuo and the compound extracted with ethyl
acetate. The organic layer was washed with a 1 M solution of
potassium hydrogenosulfate and with brine. After drying over
sodium sulfate, the solvent was removed in vacuo to yield
quantitatively the desired compound 7.
Boc-Va l-O-p-p h en yl-COOH: 1H NMR (CDCl3) δ 1.07 (d,
3H, CH3), 1.11 (d, 3H, CH3), 1.54 (s, 9H, Boc), 2.3 (m, 1H, CHâ),
4.42 (m, 1H, CHR), 5.11 (d, 1H, NH), 7.61 (m, 4H, Ar), 10.73
(s, 1H, COOH). [M + H]+: 338.
Boc-P h e-O-p-p h en yl-COOH: 1H NMR (CDCl3) δ 1.47 (s,
9H, Boc), 3.25 (m, 2H, CH2â), 4.87 (m, 1H, CHR), 5.12 (d, 1H,
NH), 7.34 (m, 5H, Ar), 7.62 (m, 4H, Ar), 8.8 (s, 1H, COOH).
[M + H]+: 386.
Boc-Ala -O-p-p h en yl-COOH: 1H NMR (CDCl3) δ 1.46 (s,
9H, Boc), 1.6 (d, 3H, CH3â), 4.60 (m, 1H, CHR), 5.13 (d, 1H,
NH), 7.63 (m, 4H, Ar), 10.23 (s, 1H, COOH). [M + H]+: 310.
An ch or in g of th e F ir st Am in o Acid via th e P h en yl
Ester Lin k er . The MBHA resin (0.63 mmol/g) was acylated
with 2 equiv of compound 7 in the presence of BOP (2 equiv)
and DIEA (pH 8-9) in DCM. After 4 h, classical solid phase
workup was performed (washings), and the Kaiser test was
used to verify the completion of the reaction. The N-Boc
temporary protection was removed with TFA/DCM (40/60) by
two treatments (3 and then 27 min), and after classical
washings, the resin was ready for peptide elongation.
Typ ica l Red u ction of th e P ep tid e Lin k ed via P h en yl
Ester to th e Resin . Boc-Phe-Val-Ala-phenyl ester linked
resin (2 g, 0.818 mmol) was suspended in anhydrous THF and
Syn th esis of a n Hexa p ep tid e Con ta in in g a Red u ced
Bon d betw een Lys a n d P h e Resid u es. Ac-Leu -Leu -
Lys[CH2-NH]P h e-Asp -Ala -NH2 9. This hexapeptide was
synthesized via the two methods to yield an identical com-
pound 9 as described in Scheme 4. 1H-NMR (DMSO-d6): 0.85
(dd, 12H, CH3), 1.25 (d, 3H, CH3), 1.3 (m, 2H, CH2 âLys), 1.4-
1.7 (m, 10H, â + γLeu + γ + δLys), 1.9 (s, 3H, CH3CO), 2.8
(m, 1H, âAsp), 2.75 (m, 3H, âAsp + CH2 ꢀLys), 2.78 (m, 2H,
CH2NH), 3.09 (d, 2H, CH2 âPhe), 4 (m, 2H, CH RLys + CH
RPhe), 4.2 (m, 3H, CH RLeu + CH RLeu + CH RAla), 4.6 (q,
1H, HRAsp), 7.2-7.35 (m, 5H, Ar), 7.55 (d, 1H, NH Lys), 7.8
(d, 1H, NH), 7.85 (d, 1H, NH), 7.97 (d, 1H, NH), 8.65 (s, 1H,
NH Asp).
J O962408D