Table Of Content1
Synthesis and Use of Pseudopeptides Derived
from 1,2,4=Oxadiazole-, 1,3,4=0xadiazole-,
and 1,2,4=Triazole=based Dipeptidomimetics
Kristina Luthman, Susanna Borg, and Uli Hacksell
1. Introduction
This chapter focuses on the tsostertc replacement of peptide bonds with three
different types of heterocycltc rmg systems (I); 1,2,4-oxadtazole (2), 1,3,4-
oxadtazole (3), and 1,2,4-trtazole rmgs (4-6). The rmg systemsa re stmtlar m
size and shape but show variations m aromattc, electrostatic, and hydrogen
bondmg properties These varrattons provide opportunittes to study properties
of importance for amtde bond mimicry. The derivatives are synthestzed from
protected natural ammo acids, and the reactton condttrons have been chosen so
that the enanttopurtty IS retained during the reaction sequences.T wo series of
mtmettcs will be described, one in which the carboxyhc acid functronality 1s
directly attached to the heterocychc rmg (2) and one series with a methylene
group inserted between the rmg and the carboxylic acid group (7). Since we
have focused on the design and synthesis of Phe-Gly mtmetrcs, the synthetic
examples described here start from t-phenylalanme. However, we have used
the same synthetic scheme also for other ammo acids and notes will be given
when other dertvattves require differences m reaction condtttons (I) The use
of the dipepttdomtmetrcs as burldmg blocks in pseudopepttde synthesis will
also be described (7). These synthesesa re performed on solid phase using Boc-
chemtstry. Also, the deprotectton and purtficatton of the pseudopeptides by
reversed phase HPLC wtll be dtscussed.
From Methods m Molecular Medune. Vol 23 Pepfdomfmebcs Protocols
Edlted by W M Kazmrerskl OHumana Press Inc , Totowa, NJ
1
2 Luthman, Borg, and Hacksell
2. Materials
2.1. Overview
‘H and 13C NMR sp ectra were recorded at 270 and 67.5 MHz with tetra-
methylsilane as the internal standard. Thin layer chromatography (TLC) was
carried out on aluminum sheets precoated with silica gel 60 F,,, (0.2 mm,
E. Merck); spots were visualized with UV detection and by spraying with a
2% ethanol solution of ninhydrin, followed by heating. Column chroma-
tography was performed on silica using Kieselgel 60 (230400 mesh, E.
Merck). Solvents were in some cases dried prior to use. THF and diethyl
ether were distilled from sodium/benzophenone, and dichloromethane was
dried over MgSO+
2. I. 1, HPLC Columns, Conditions, and Necessary Reagents
1. Analyses of the optical purity of 1,2,4-oxadiazole and 1,3,4-oxadiazole deriva-
tives: J. T. Baker Chiralcel OD-H column, Detection wavelength: 254 or 2 11 nm.
Ambient temperature. Mobile phase: Hexane/2-propanol/diethylamine (90: 10:O. 1
(2 and 5/ or 95:5:0.1 /9/). Flow rate: 0.5 mL/min.
Reagents
1. Ethyl 2-chloro-2-(hydroximino)acetate.
2. Diethyl ether.
3. Liquid or gaseous ammonia (NH,).
4. Hexane.
5. N,hr-Dicyclohexylcarbodiimide (DCC).
6. tert-Butyloxycarbonyl(Boc)-L-phenylalanine.
7. Dichloromethane.
8. Pyridine.
9. Pentane.
10. Ethyl acetate.
Il. NaHCO,.
12. Aqueous HCl (pH 2).
13. Aqueous NaCl (Brine).
14. MgSO,.
15. Methanol (MeOH).
16. tert-Butyldiphenylchlorosilane.
17. 3-Hydroxypropionitrile.
18. Triethylamine.
19. 4-Dimethylaminopyridine (DMAP).
20. Tetrahydrofuran (THF).
2 1. Citric acid.
22. Hydroxylamine hydrochloride.
23. K2C03,
24. Ethanol.
Synthesis and Use of Peptides 3
25. Na$O,,
26. Chloroform (CHCI,).
27. 1 M Tetrabutylammonium fluoride in THF.
28. Acetic acid.
29. Acetone.
30. Na$r,O,.2 H,O.
3 1. H,SO, (97%).
2. Analyses of the optical purity of 1,2,4-triazole derivatives: J. T. Baker Chiralpak
AD column. Detection wavelength: 225 nm. Temperature: 30°C. Mobile phase:
Hexane/2-propanol/diethylamine (90: 1O:O.1 /14/ or 80:20:0.1 /Z 71). Flow rate:
0.5 or 1 mlimin.
Reagents
1. Boc-L-phenylalanine methyl ester.
2. Methanol (MeOH).
3. Hydrazine hydrate.
4. Chloroform.
5. Triethylamine.
6. Tetrahydrofuran (THF).
7. Ethyl oxalyl chloride.
8. Diethyl ether.
9. NaHCOs.
10. NaHSO,.
I I. MgSO,.
12. Hexane.
13. Pyridine.
14. Thionyl chloride (SOCI,).
15. Toluene.
16. Dichloromethane.
17. Ethyl acetate.
IS. N-Methylmorpholine (NMM).
19. Methyl malonyl chloride.
20. Citric acid.
2 I. Aqueous NaCl (Brine).
3. Purification of pseudopeptides with reversed-phase semi-preparative HPLC:
Waters pBondapak C-18 column, 25 x 100 mm. Detection wavelength: 254 nm.
Ambient temperature. Mobile phase: A gradient of 0 to 60% acetonitrile in water/
I%TFA in 60 min. Flow rate: 5 mL/min.
Reagents
1. Ethyl thiooxamate.
2. Ethanol.
3. Hydrazine hydrate.
4. Activated charcoal.
5. Dichloromethane.
6. Celite.
4 Luthman, Borg, and Ha&se/l
7 Pentane
8 Ethyl chloroformate
9 Boc-t,-phenylalanme
10 Triethylamine.
11 Tetrahydrofuran (THF)
12 NJ-Dtmethylformamtde (DMF)
13 Xylenes (mixture of isomers)
14 Chloroform
15 Methanol (MeOH)
16 Ethyl acetate
17 Ethyl cyanoacetate
18. Gaseous HCl
19 Dtethyl ether
4 Analyses of pseudopepttdes wtth reversed-phase HPLC Waters pBondapak C-
18 column, 8 x 100 mm Detectton wavelength 254 nm Ambtent temperature
Mobile phase A gradient of 0 to 60% acetomtrtle m water/l%TFA in 30 mm
Flow rate 1 mL/mm
Reagents
1 4-Methylbenzhydrylamme (MBHA) resin
2 KOH
3 Ethanol.
4 Methanol
5 Hf-ton exchange resm (Dowex’ 50 W x 8)
6 Drethyl ether
7. MgS04.
8 Dtchloromethane
9 NJ-Dtmethylformamtde (DMF)
10 Benzene
11 NaHCOs
12 Aqueous HCl (I M)
13 Trtfluoroacetlc actd (TFA)
14 2-( lH-Benzotrtazol- 1-yl)- 1,1,3,3-tetramethyluromum hexafluorophosphate
(HBTU)
15 1 -Hydroxybenzotrtazole hydrate (HOBt).
16 N-Ethyl-N,N-dnsopropylamme (DIPEA)
17 2-( lH-Benzotrtazol-1 -yl)-1 ,1,3,3-tetramethyluromum tetrafluoroborate
(TBTU)
18 Ethyl chloroformate
19 N-methylmorpholme (NMM)
20 Iodotrtmethylstlane (TMSI)
2 1 Thtoamsole
Synthesis and Use of Peptldes 5
N/OH
pyrldine
(Boc-L-Phe)aO + Et0 - BocHN
2 A
0 COOEt
1 2
Figure 1
TBDPSO
(Boc-L-Phe)20 +
3 N’OH BocHN
TBAF
Jones’
BocHN BocHN
oxidation
6
Figure 2
22 1,2-Ethanedtol
23 Trlfluoromethanesulfomc acid (TFMSA)
3. Methods
3.1. Preparation of 1,2,4-Oxadiazoles (Figs. 1 and 2)
1,2,4-Oxadlazole dlpeptldomlmetlcs (see Note 1) can be obtained from
amldoxlmes and Boc-protected ammo acid anhydrides (see Note 2) (8-10). Sym-
metric anhydrldes are synthesized from Boc-protected amino acids by treatment
with N,N-dlcyclohexylcarbodnmlde (DCC) (see Note 3) (II). The amidoxlme (12)
and the anhydnd? are reacted in pyndme, initially forming an O-acylamldoxlme
(see Note 4). This intermediate is not isolated but directly heated to reflux to
achieve the cychzatlon (23) ( see Note 5) The use of a symmetric anhydride as
activating group requires two equivalents of the protected ammo acid, however, it
IS possible to recover one equivalent from the reaction mixture.
6 Luthman, Borg, and Hacksell
3 I. 7 Ethyl Z-Amino-2-(hydrox/mlno)Acetate I (14)
1 Dissolve ethyl 2-chloro-2-(hydroxlmmo)acetate (1 0 g, 6 6 mmol) m anhydrous
diethyl ether Cool the solution to 0 “C
2 Pass gaseous NH, through the solution for approx 3 mm (see Note 6) Filter off
the precipitated ammomum chloride and concentrate the filtrate to an 011 Solldlfy
by addition of hexane Filter off the precipitate to obtain approx 660 mg (76%) of
1 Mp 75-76°C (8)
3 1 2. Ethyl 5-[(S)-[(tert-Buty/oxycarbonyl)Amino]-2-Phenylethyl]-
7,2,4-Oxadlazole-3-Carboxylate 2 (Fig. 7)
DCC (239 mg, 1 16 mmol) to a solution of Boc-L-phenylalanme (6 14 mg, 2 32 mmol)
m dry dlchloromethane (30 mL) at room temperature Stir the reaction mixture at
0°C for 1 h Filter off the precipitated N,N’-dlcyclohexylurea (DCU) and concen-
trate the filtrate In vucuo
Redissolve the crude symmetric anhydride m pyrldme Add dropwise a solution
of 1 (102 mg, 0 77 mmol) m pyrldme
Heat the mixture to reflux for 2 h (TLC, pentaneiethyl acetate, 3.2) Add water
when the reaction IS complete, and evaporate the solvent (see Note 7) Partition
the residue between dlchloromethane and water and wash the organic layer with
saturated aqueous NaHCO,, aqueous HCl (pH = 2), brine, and water
Pun@ the crude product (see Note 8) by column chromatography using pentane/ethyl
acetate (8 5 1 5) as eluent Recrystalhzatlon from dlethyl etherkexane affords 200 mg
(72 %) of pure 2 Mp 75-76°C; [a]o = -29 4” (c 1 0, MeOH), tR 17 2 mm (see Sub-
beading 2.), ‘H NMR (CDC&) 6 7 3G7 00 (m, 5H), 5 39 (m, 1H ), 5 17 (br d, IH), 4 5 1
(q, J = 7 2,2H), 3.27 (AB system, 2H), 1 44 (t, 3H), 1 40 (s, 9H), ‘)C NMR (CDCl,) S
1809,1618,1575,1547,1346,1292,1289,1276,808,632,495,399,282,141
3. I .3. Preparation of Amidoxlmes
from Nitrrles (3-tert-Butyld~phenyls~lyloxypropanamide Ox/me 3
Amldoxlmes can be synthesized from mtrlles by treatment with hydroxy-
lamme In this specific case we use 3-hydroxyproplomtnle as starting material,
and the reaction proceeds m two steps. In the first step, the alcohol functlonal-
lty IS protected as a sllyl ether (151, and m the second step the mtrlle IS con-
verted mto the amldoxlme (see Note 9)
1 Add tert-butyldlphenylchlorosdane (18 5 mL, 73 mmol) to a mixture of 3-hydroxy-
proplomtrile (2 46 mL, 36 mmol), trlethylamme (12 7 mL, 91 mmol), and
dlmethylammopyridme (DMAP) (178 mg, 1 5 mmol) m dry THF (75 mL) under
a nitrogen atmosphere
2 Stir the mixture at room temperature for 3 5 h (TLC, dlethyl ether/pentane, 1 3)
Filter off the salts and evaporate the solvent uz V~CUO Redissolve the residue m
dlchloromethane. Extract with saturated aqueous NaHC03, saturated aqueous clt-
rlc acid, and brine
Synthesrs and Use of Peptides 7
3. Purify the crude product (see Note 8) by column chromatography using
dlchloromethane/pentane (2.3) as eluent Concentrate the appropriate fractions
to obtain a solid residue Recrystalllzatlon from dlchloromethane/pentane
affords 8 6 g (76%) of pure 3-tert-butyldlphenylsllyloxyproplomtrlle Mp
50-51°C (15), 1HNMR (CDCI,) 6 7.70-7 35 (m, lOH), 3 84 (t,J= 6 3,2H),
2 53 (t, 2H), 1 08 (s, 9H), 13CN MR (CDC13) 6 135 5,132 7,129 9,127 8, 117 9,
59 0, 26.7, 21 4, 19.1
4 Add a solution of hydroxylamme hydrochloride (3.8 g, 55 mmol) and K&O,
(3.8 g, 27 mmol) in water to a solution of 3-tert-butyldlphenylsllyloxy-
proplonitrlle (8 5 g, 27 mmol) m ethanol (see Note 10)
5 Stir the mixture for 23 h at room temperature Evaporate the volatlles In vacw
and add dlethyl ether to the residue Filter off the solids and dry the filtrate with
Na,SO, Filter off the drying agent and concentrate the filtrate zn vucuo
The residue can be crystallized by addition of dlethyl ether/pentane However,
recrystalllzatlon can be difficult (see Note 11) You will obtain 5 4 g (57%) of
pure 3 Mp 83-84”C, ‘H NMR (CDCl,) 6 7 70-7 25 (m, lOH), 5.04 (br s, 2H),
3 88 (t, .I=5 6,2H), 2 37 (t, 2H), 1 06 (s, 9H), 13CN MR (CDCl,) 6 153 9, 135 5,
1329, 1300, 1279,624,336,268, 19 1
3.1 4. 2-[5-[(S)- 7-(tert-Butyloxycarbonylammo-2-Pheny/ethyl]-
1,2,4-Oxadlazol-3-yl]Acetic Acid 6 (Fig. 2)
Prepare the symmetric anhydride as described m (Subheading 3.1.2.) using 1 43 g
(5 4 mmol) of Boc-L-phenylalanme and 558 mg (2 7 mmol) of DCC Dissolve the
crude anhydride m pyndme.
Add dropwlse a solution of 3 (926 mg; 2.7 mmol) m pyrldme at room tempera-
ture. Heat the mixture to reflux for 3 h (TLC, dichloromethane/pentane, 4 1)
Add water and evaporate the solvent zn V~CUO( see Note 7). Redissolve the resl-
due in dlethyl ether Extract with water, 10% aqueous citric acid, and saturated
aqueous NaHC03
Purify the crude product (see Note 8) by column chromatography using
dlchloromethane/pentane (4 1) as eluent to obtain 940 mg (63%) ofpure 5-([(S)-l-
tert-butyloxycarbonylam~no-2-phenylethyl]-3-(2-~e~~-butyld~phenyls~lyloxyethyl)-
1,2,4-oxadlazole 4 as an 011. [a],,=-9 4” (c 1.2, CHCI,), ‘H NMR (CDCl,) 6
7.65-7.00 (m, 15H), 5 29 (m, IH), 5.06 (m, IH), 4.00 (t, J= 5.4, 2H), 3 29-3 08
(m, 2H), 2 96 (t, 2H), 1 40 (s, 9H), 1 01 (s, 9H); 13C NMR (CDClJ F 178 3,
168 2, 154.7, 135 5, 135.0, 133.4, 129.7, 129 2, 128.6, 127 7, 127.2, 80.4, 60.8,
49 3,40.0, 29 5,28 2, 26 7, 19.1
Dissolve 4 (285 mg, 0 49 mmol) m dry THF (50 mL) Keep the solution under
nitrogen atmosphere Add a mixture of tetrabutylammomum fluoride (810 &;
0 80 mmol, 1 M solution in THF) and 30 pL of acetic acid m THF (see Note 12).
Stir the reactlon mixture for 2 h at room temperature (TLC, dlethyl ether), Evapo-
rate the solvent and redissolve the residue in dlethyl ether. Extract with 10%
aqueous citric acid, saturated aqueous NaHC03, and brine
8 Luthman, Borg, and Hacksell
7 Put@ the crude product (see Note 8) by column chromatography usmg first dlethyl
ether/pentane ( 1.2) and then dlethyl ether as eluents Concentration of appropriate frac-
tlons and recrystalhzatlon m dlethyl etheripentane should afford 125 mg (75%) of pure
5-([(S)-l-tert-butyl-oxycarbonylam~no-2-phenylethyl]-3-(2-hydroxyethyl)-1,2,4-
oxadiazole (5). Mp 84-85”C; [a],=-20.8” (c 0 9, CHC13), tR 30.9 mm (2 11 nm) (see
Subheading 2.), ‘H NMR (CDCl,) 6 7 30-7 00 (m, 5H), 5 3 (br, lH), 5 1 (br, lH),
3 95 (t, J= 5 9,2H), 3 31-3 14 (m, 2H), 2 96 (t, 2H), 2 30 (t, IH), 141 (s, 9H), 13C
NMR (CDC13) 6 178 8, 168.5, 154 9, 135 0, 129 2, 128 7, 127 4, 80 6, 59.4, 49 4,
39 8,29 2,28 2
8 Dissolve purltied 5 (50 mg, 0 15 mmol) m acetone and chill the solution to 0°C
Add Jones’ reagent (26) (457 mL, 0 3 1 mmol, 0 67 M) (see Note 13)
9 Stir the solution for 12 h (TLC, dlethyl ether) Filter off the sohds and evaporate the
solvent Redissolve the residue m dlethyl ether and extract wtth saturated aqueous
NaHC03. Acid@ the aqueous phase with 1 Maqueous HCl and extract with dtethyl ether
Recrystallize the crude product (see Note 8) from chloroform to obtain 34 mg (64%)
ofpure 6 Mp 163 5-164.5”C; [aID=- 5’ (c 1 1, CHCI,), ‘H NMR (acetone-d,) 6
7 25-7 15 (m, 5H), 6 70 (br d, lH), 5 22-5 10 (m, IH), 3 78 (s, 2H), 3 29 (dd, J= 13 7,
6 0, lH), 3 16 (dd, J= 9 2, IH), 1 28 (s, 9H), 13CN MR (acetone-d,) 6 181 0, 169 4,
166 3,156 2,137 8, 130 5, 129 4, 127 9,80.0,50 8,39 8,32 6,28 7
3.2. Preparation of 1,3,#-Oxadiazoles (Figs. 3 and 4)
1,3,4-Oxadlazole dipeptldomlmetics (see Note 14) can be obtained by
dehydration of the correspondmg dlacylhydrazmes (I 7) The dlacylhydrazmes are
obtained from ammo acid hydrazldes, which are conveniently synthesized from
amino acid esters and hydrazme (Z&19) ( see Note 15) Boc-protected ammo acid
esters are either commercially available or can be synthesized from Boc-protected
ammo acids by treatment with DCC/DMAP and the appropriate alcohol (20) (see
Note 16). The esters are reacted with hydrazme hydrate to produce the hydrazldes
that are treated with an acid chloride (or another activated carboxyhc acid denva-
tlve) to obtain the diacylhydrazmes (22). The dehydration condltlons should be
mild and compatible with the properties of the ammo acid protecting group (see
Note 17) Therefore, we use a mixture of thlonyl chloride (SOCI,) and pyrldme at
0°C This leads to the formation of a 1,2,3,4-oxathladmzole-S-oxide intermedlate,
which can be isolated or dn-ectly heated m a high-bollmg solvent to form the 1,3,4-
oxadlazole derivative by thermal ehmmatlon of sulfur dioxide (22)
3.2.1. Preparation of Amino Aad Hydrazides
([(S)-2-[(tert-Butyloxycarbonyl)Amlno]-3-Phenylpropanoyl]
Hydrazine, 7) (Fig. 3)
1 Dissolve Boc-L-phenylalanme methyl ester (14 g, 0 05 mol) m methanol (see
Note 18) For preparation of ammo acid esters, see Note 16 and ref. 20
Synthesis and Use of Peptldes 9
1 DCClEtOH
Boc-L-Phe c
2 Hydrame BocHN
0
ClCOCOOEt
Et3N
1 SOCl2, pyrldine
2. Toluene, reflux
BocHN BocHN
N-N
9 8
Ftgure 3
7
+ “TCOOMe Et3N *
BocHN
0
1. SOCI,, pyndme
w
2. Toluene, reflux BocHN
N-N
11
Figure 4
2 Add hydrazme hydrate (7 3 mL; 0.15 mol).
3 Stir at room temperature for I8 h (TLC, chloroform/MeOH, 9.1)
4 Put the reaction flask m the refrigerator for 1 h to allow the hydraztde to pre-
cipitate (see Note 19) The precipitate is filtered off to afford 11.5 g (82%) of
pure 7 Mp 120-122°C (see Note 20) [a]n = +9 7” (c 1 01, MeOH); ‘H NMR
(CDCls) 6 7 70 (br s, lH), 7 3c7.15 (m, 5H), 5 32 (br d, lH), 4 36 (dd, IH),
3.8 1 (br s, 2H), 3 02 (AB system, 2H), 1.39 (s, 9H), 13C NMR (CDC13) 6 17 1 9,
155 2, 136 4, 129 2, 128 8, 127 1, 80.5,54 8,38 4,28 3
10 Luthman, Borg, and Hacksell
3.2.2 Preparation of Diacylhydrazines (1 -(Ethoxyoxalyl)-2-[(S)-2-
[(tert-Butyloxycarbonyl)-Amino]-3-Phenylpropanoyl]
Hydrazme, 8) (Fig. 3)
Dtssolve 7 (1 3 g, 4 65 mol) and triethylamme (777 $; 5 58 mmol) m dry THF
under nitrogen atmosphere Chtll the solutton to -30°C
Add dropwtse ethyl oxalyl chloride (624 mL, 5 58 mmol) dissolved m 5 mL of
THF, and let the reaction stu for 20 h The temperature should be gradually
mcreased to room temperature (TLC, chloroform/MeOH, 9 1)
Filter off the precipitated trtethylammomum hydrochlortde and concentrate the
filtrate zn vucuo Partmon the residue between dtethyl ether and water and extract
the organic layer with saturated aqueous NaHCO, and 1 M NaHSO,
Crystallize the crude product (see Note 8) m dtethyl etherihexane to obtain 1 0 g
(60%) of pure 8 (see Note 21). Mp 88-89°C [cY.]~= -21.7” (c 1 01, MeOH), ‘H
NMR (CDC13) 6 9 20 (br s, 2H), 7 35-7 20 (m, 5H), 5 07 (br d, 1 H), 4 5 1 (m,
lH), 4 37 (q, .I= 7 0, 2H), 3 18 (dd, lH), 3 05 (dd, J= -14 0, 6.0, lH), 1.39 (s,
9H), 1 38 (t, 3H), 13C NMR (CDC13) 6 169.0, 158 7, 155 7, 152.9, 136 0, 129 3,
128 7, 127 1, 80 8,65 6,54 3,37 9,28 3, 13 9
3.2.3 Ethyl 5-[(S)-[(tert-Butyloxycarbonyl)Amlno]-2-Phenylethyl]-
1,3,4-Oxadiazole-2-Carboxylate 9 (Fig. 3)
Dissolve 8 (1 .O g, 2 64 mmol) m dry dtethyl ether (75 mL) under nitrogen atmo-
sphere Chill the solutron to O’C (see Note 22)
Add pyrtdme (550 mL, 6 85 mmol) and thereafter SOCl* (250 mL, 3 43 mmol)
dissolved in drethyl ether (see Note 23)
Let the reactton stir for 2 h at 0°C Filter off the precipitated pyndunum salt as quickly
and carefully as possible (see Note 24) Concentrate the filtrate WIv acua without heating
Redtssolve the residue m dry toluene and heat the solutton to reflux for 2 h (TLC,
chloroform/MeOH/hexane, 9.1 1)
Evaporate the solvent zn vucuo and purify the product by column chromatogra-
phy with dtchloromethane/ethyl acetate (19 1) as eluent Recrystalhzatron from
drchloromethane/ pentane affords 590 mg (62%) of pure 9 Mp 122-123”C,
[a],, = -3 1 9” (c 10, MeOH), tR 45 7 mm (see Subheading 2.); ‘H NMR (CDCl,) 6
73>705(m,5H),536(m, lH),509(brd, lH),451 (q,J=72,2H),332(dd, lH),
3 22 (dd, lH), 1 45 (t, 3H), 1 40 (s, 9H), 13C NMR (CDCl,) 6 168 5, 156 9, 154 6,
1541,134.7,1292,1288,1274,808,636,486,396,282,140
3.2 4 Methyl 2-~5-[(S)-l-(tert-5utyloxycarbonylamino-2-Phenylethyl]-
I, 3,4-Oxadlazol-2-yl]Ace ta te 11 (Fig. 4)
1 Dissolve 7 (433 mg; 1.6 mmol) (Subheading 3.2.1.) m dry THF under nitrogen
atmosphere. Add N-methylmorpholme (NMM) (255 mL; 2 3 mmol) and chill the
solution to -30°C (see Note 25)
2 Add methyl malonyl chloride (248 pL, 2.3 mmol) and let the reaction stir for 4 h whtle
the temperature 1sa llowed to increase to room temperature (TLC, ethyl acetate)
