Synthesis of an external beta-turn based on the GLDV motif of cell adhesion proteins

Article abstract of DOI:10.1016/j.tet.2004.09.037

The (3S,6S,10S)-7/5 bicyclic lactam 8, designed as an external turn constraint, was synthesised by a new stereoselective route involving Eschenmoser condensation. The cyclic peptide 35 containing the integrin recognition motif GLDV added across the amino

Full text of DOI:10.1016/j.tet.2004.09.037

Tetrahedron 61 (2005) 301–312
Synthesis of an external beta-turn based on the GLDV motif of cell
adhesion proteins*
David E. Davies,a Paul M. Doyle,b Richard D. Hillc and Douglas W. Youngc,
*
aGlaxoSmithKline, Medicines Research Centre, Gunnels Wood Road, Stevenage, Herts SG1 2NY, UK
bBioFocus Discovery Ltd, Chesterford Research Park, Saffron Walden, Essex CB10 1XL, UK
cSussex Centre for Biomolecular Design and Drug Development, Department of Chemistry, University of Sussex, Falmer,
Brighton BN1 9QJ, UK
Received 15 July 2004; revised 7 September 2004; accepted 9 September 2004
Available online 6 October 2004
Abstract—The (3S,6S,10S)-7/5 bicyclic lactam 8, designed as an external turn constraint, was synthesised by a new stereoselective route
involving Eschenmoser condensation. The cyclic peptide 35 containing the integrin recognition motif GLDV added across the amino and
carboxyl groups of the lactam external constraint 8 was prepared.
q 2004 Elsevier Ltd. All rights reserved.
1. Introduction
integrin a4b1 with vascular cell adhesion molecule-1.9
Using BTD as a model for an external b-turn, we have
synthesised the putative external turn constraints, 2, 3, 4 and
5 (where P1 and P2 are protecting groups).10
Reverse turns connect elements of protein secondary
structure and they usually occur on the surface of the
protein. They can, therefore, be important in molecular
recognition processes.2,3 Synthetic molecules which mimic
these turns, such as the classic 5/6 bicyclic lactam BTD, 1,
prepared by Nagai,4–6 have been used to replace b-turns in
various proteins without affecting their overall biological
properties.5–7 BTD, 1, has been shown to be a type II0 b-turn
mimic from the angles j2 (K1618) and f3 (K69.48) in the
X-ray crystal structure.8
A linear GLDV construct was attached to the amino group
of the template 4 but the acyl enamine nature of the template
caused problems in the final hydrolysis and cyclisation steps
which would have yielded the external GLDV turn. We
therefore decided to prepare a homochiral bicyclic system in
which the amide was not destabilised as an enamine and to
use it as a template on which to build a cyclic GLDV turn.
We have used BTD9 as a structural constraint on which to
build an external cyclic GLDV motif. This was shown
induce a type I b-turn and to inhibit the interaction of the
* See Ref. 1.
Keywords: Bicyclic lactam; Integrin; Eschenmoser olefination.
* Corresponding author. Tel.: C44 1273 678327; fax: C44 1273 677196;
e-mail: d.w.young@sussex.ac.uk
Amino acid 7/5-bicyclic lactams of type 6 have found use as
dipeptide surrogates in angiotensin-converting enzyme
0040–4020/$ - see front matter q 2004 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2004.09.037

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D. E. Davies et al. / Tetrahedron 61 (2005) 301–312
(ACE) inhibitors11 and have been used as external
constraints for tripeptide RGD turns which proved to be
inhibitors of the aVb3 integrin receptor.12 The 7/5 bicyclic
lactam 7 was synthesised using an electrochemical approach
and conformational analysis showed that the (4S,7S,10S)-
stereoisomer had families of minimum conformations with
torsion angles close to those of classical b-turns.13 We
therefore decided to develop a new and stereoselective
synthesis of the 7/5 bicyclic lactam, 8, via a route involving
Eschenmoser coupling and to use it as an external restraint
for the GLDV motif present in the integrin family14 of cell
adhesion proteins.
Figure 1. NOE experiments on compound 14.
solid sodium bicarbonate, then the only recognisable
product was the disulfide 13, obtained in 58% yield.
When the bromide was heated at reflux with the thiolactam
1218 in CH3CN, followed by addition of Ph3P and
triethylamine however, the desired product 14 was obtained
as an oil. This was shown to be the Z-isomer by the NOE
experiments summarised in Figure 1, since irradiation of the
olefinic proton, H-6, caused enhancements to all four
protons assigned to H-4 and H-8 in the 1H NMR spectrum.
The hydrogen bond between the N–H of the vinylogous
amide and its carbonyl group, as shown, presumably
accounts for this specificity. The highest yield of the
product 14 that we could obtain was 30%.
2. Results and discussion
Since sulfur contraction reactions have been reported to be
more effective when the thiolactam is trisubstituted,19 we
prepared the N-benzyl thiolactam 17 as shown in Scheme 2.
This compound had previously been prepared by
Rapoport20 using a synthesis which involved N-alkylation
of glutamic acid prior to cyclisation to avoid the danger of
racemisation. We were able to alkylate tert-butyl (2S)-
pyroglutamate 15 (prepared from commercial (2S)-pyroglu-
tamic acid by the method of Miller21) directly to the
N-benzyl derivative 16 in 64% yield using benzyl bromide
and sodium hexamethyldisilazide. Reaction with P4S10 then
gave the thiolactam 17 in 95% yield with spectroscopic data
similar to those reported by Rapoport.20 Eschenmoser
olefination using the bromide 10 and Ph3P/Et3N in
CH3CN then gave the vinylogous amide 18 as an oil in
91% yield.
As a first step in our synthesis of the bicyclic lactam 8, we
attempted to prepare the bromide 10 by the method used by
Weygand15 to obtain the corresponding ethyl ester bromide
but without success. Modification of the method by reacting
the diazoketone 916 with HBr in CHCl3, however, gave the
a-bromoketone 10 in 89% yield, as shown in Scheme 1. On
scale-up, the yield was reduced by the presence of the
a-chloroketone 1117 but this could be separated out and
converted to the a-bromoketone 10 in 85% yield by a
Finkelstein reaction using lithium bromide in acetone.
NOE experiments on the N-benzyl derivative 18, shown in
Figure 2, indicated that it had E-stereochemistry. When the
olefinic proton, H-6, was irradiated, both of the protons H-8
and one of the benzylic CH2 protons showed enhancement,
implying that, unlike the N–H analogue 14, the N-benzyl
compound 18 had E-geometry, as might be expected from
A variety of methods were used in an attempt to complete
the Eschenmoser olefination reaction between the bromo-
ketone 10 and the thiolactone 12.18 When these compounds
were heated at reflux in dichloromethane in the presence of
Scheme 1. Reagents and conditions: (i) HBr/CHCl3, room temperature, (small scale) (89%); (ii) scale-up contains some 11; (iii) LiBr/acetone/79 h/room
temperature (80%); (iv) 12/NaHCO3/CH2Cl2, reflux (58%); (v) 12/MeCN, reflux, then Ph3P/Et3N (30%).

D. E. Davies et al. / Tetrahedron 61 (2005) 301–312
303
the bicyclic compound 22 would have shortened our
synthetic route to a homochiral 7/5-bicyclic external turn
mimic but unfortunately we were unable to obtain it
consistently and in good yield. The mechanism suggested in
Scheme 3 would account for its formation. Here, incomplete
reduction to the alcohol 20, followed by lactonisation would
give the product 21 which, on final cyclisation, would give
the product 22.
The amino ester 19 was cyclised using tBuMgCl in ether at
0 8C, giving the bicyclic lactam 8 in 29% yield. This was
shown to have (3S,6S,10S) stereochemistry by the NOE
studies summarised in Figure 3b, since H-10 showed
enhancements when either H-3 or H-6 were irradiated.
This stereochemistry was confirmed by the X-ray structure
analysis, reported in our preliminary communication.1
Scheme 2. Reagents and conditions: (i) NaHMDS/PhCH2Br/THF/K78 8C,
(64%); (ii) P4S10/THF/room temperature (95%); (iii) 10/Ph3P/Et3N/
CH3CN, room temperature (91%).
Calculations of the conformational preferences of the
compound 8, reported in our preliminary communication,1
indicated that the torsional minima for compound 8 were not
compatible with it being a b-turn mimetic in its own right,
and this was in agreement with the X-ray crystal structure
analysis of the compound also reported in that paper.1
Figure 2. NOE experiments on compound 18.
In order to examine the usefulness of the compound 8 as a
constraint for an external b-turn involving the GLDV motif
attached to its amino and carboxyl groups, we hydrolysed
the trifluoroacetylamide using K2CO3 in methanol at reflux,
as shown in Scheme 3. This gave the amine 23 in 86% yield.
Synthesis of the linear GLDV tetrapeptide on the free amine
group of compound 23 was now undertaken using Fmoc
technology as shown in Scheme 4. Reaction of the amine 23
with Fmoc-valine, TBTU and DIPEA gave the product 24 in
80% yield, as shown in Scheme 4. Since it was essential to
protect the b-carboxyl group of the aspartate residue in
GLDV as the tert-butyl ester to prevent intramolecular
cyclisation reactions, we now needed to change the
orthogonality of our protecting groups. The Fmoc-tert-
butyl ester 24 was therefore deprotected using TFA to give
the acid 25 in quantitative yield. Reprotection using
diphenyldiazomethane yielded the ester 26 as an oil in
64% yield. This was now sequentially deprotected with
piperidine and reacted in turn with Fmoc-Asp(OtBu)-OH,
steric considerations in the absence of hydrogen bonding
possibilities.
In order to reduce the vinylogous amide and hydrogenolyse
the N-benzyl group in 18, it was necessary to hydrogenate
the compound at a pressure of 60–200 psi for 50 h using
10% palladium on activated carbon (75% by weight) in
methanol containing trifluoroacetic acid as shown in
Scheme 3. The resultant single diastereoisomeric product
19 was obtained as an oil in 65% yield, but NOE
experiments were inconclusive as to its stereochemistry.
Interestingly, in some of the hydrogenation experiments, a
small yield of a by-product was obtained. This proved to be
the bicyclic lactam alcohol 22. The (3S,6S,8S,10S)-stereo-
chemistry of this compound was implied from the NOE
studies shown in Figure 3a, since irradiation at H-3 caused
enhancement of H-10, and irradiation at H-10 caused
enhancements at both H-8 and H-6. The fortuitous finding of
Scheme 3. Reagents and conditions: (i) most occasions, H2/Pd–C/MeOH/TFA/60–200 psi (56%) of 19; (ii) on some occasions up to 17% of 22 were obtained;
(iii) tBuMgCl/Et2O/0 8C (29%); (iii) K2CO3/MeOH/6 (86%).

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D. E. Davies et al. / Tetrahedron 61 (2005) 301–312
Figure 3. NOE experiments on (a) compound 22, and (b) compound 8.
Fmoc-Leu-OH and Cbz-Gly-OH to give the protected
tetrapeptide 32, as summarised in Scheme 4. Hydrogeno-
lysis, cyclisation and final deprotection were then carried
out, as in Scheme 4, and the final cyclic peptide 35 was
purified by repeated reverse phase HPLC.
We have therefore prepared the homochiral external turn
constraint 8 containing three chiral centres, and have affixed
a cyclic GLDV sequence between the amino and carboxyl
functionalities resulting in the presentation of the GLDV
motif with either type VI or type II b-turn properties.
3. Experimental
3.1. General
Melting points were determined on a Kofler hot-stage
apparatus and are uncorrected. Optical rotation measure-
ments (in units of 10K1 deg cm2 gK1) were measured on a
Perkin Elmer PE241 polarimeter using a 1 dm path length
micro cell. IR spectra were recorded on Perkin Elmer 1720
and Bio-RadFTIR (FTS155)spectrometers. 1H NMRspectra
were recorded on Varian Inova 750 FT (750 MHz), and
Bruker AMX500 FT (500 MHz), WM360 FT (360 MHz),
DPX300FT(300 MHz), ACP250FT(250 MHz), AM250FT
(250 MHz) and DPX250 FT (250 MHz) instruments.
J values are reported in Hertz (Hz). 13C NMR spectra were
recorded on Bruker AMX500 FT (125.8 MHz), WM360 FT
(100.5 MHz), DPX300 FT (75.5 MHz), DPX250 FT
(62.9 MHz) and AM250 FT (62.9 MHz) instruments.
DEPT experiments were used to help assign 13C NMR
spectra where necessary. Residual solvent peaks were used as
internal reference for all NMR spectra. Microanalyses were
performed by Ms. P. McDonough (GlaxoSmithKline) and
The high-resolution NMR spectroscopic data for this
compound, reported in our preliminary communication,
were consistent with a single backbone conformation with
either type VI or type II0 b-turn properties.22
Scheme 4. Reagents and conditions: (i) TBTU/DIPEA/DMF/Fmoc-Val-OH (80%); (ii) CF3CO2H/CH2Cl2 (quant); (iii) Ph2CHN2/CH2Cl2 (64%); (iv)
piperidine/DMF (O82%); (v) TBTU/DIPEA/Fmoc-Asp(OtBu)-OH (88%); (vi) TBTU/DIPEA/DMF/Fmoc-Leu-OH (82%); (vii) TBTU/DIPEA/Cbz-Gly-OH
(81%); (viii) H2/Pd–C/MeOH/THF (92%); (ix) TBTU/DMAP/DMF; (x) CF3CO2H/TES.

D. E. Davies et al. / Tetrahedron 61 (2005) 301–312
305
Medac Ltd. Mass spectra were recorded on Kratos MS80F
and MS25 double focusing spectrometers by Dr. A. Abdul-
Sada(Sussex)andaHPEngine(5989B)instrumentoperating
in thermospray (positive ion) mode (GlaxoSmithKline).
LC-mass spectra were obtained on a HP1100 HPLC coupled
to a Micromass Platform II mass spectrometer operating in
electrospray (positive and negative ion) mode. Accurate
mass measurements were recorded by Mr. I. Davidson
(GlaxoSmithKline), by Dr. A. Abdul-Sada (Sussex) and by
the EPSRC National Mass Spectrometry Service, Swansea.
Petroleum ether refers to that fraction of hexanes of bp
60–80 8C. HPLC studies on the cyclic peptide 35 were
performed by Mr. Eric Horteuse (GlaxoSmithKline) and
preparative HPLC was carried out using a Gilson Autoprep
system and a Supelcosil LC-ABZC column.
and washed with water (75 ml). The aqueous layer was
further washed with dichloromethane (2!50 ml). The
combined organic layers were dried (MgSO4) and the
solvent was removed in vacuo to give methyl (2S)-5-bromo-
4-oxo-2-(2,2,2-trifluoroacetamido)-pentanoate 10 as a beige
solid (3.752 g, 80%) with spectra identical to those of a
sample prepared using method A.
3.1.2. Bis-[(2S)-2-oxo-4-(2,2,2-trifluoroacetamido)-
4-carboxylmethylbutyl]sulfide (13). Methyl (2S)-5-
bromo-4-oxo-2-(2,2,2-trifluoroacetamido)-pentanoate 10
(426 mg, 1.33 mmol), tert-butyl (2S)-5-thioxoproline 1218
(116 mg, 0.57 mmol) and sodium bicarbonate (223 mg,
2.66 mmol) were heated at reflux in dichloromethane
(15 ml) for 39 h. The mixture was allowed to cool to
room temperature, filtered and the solvent was removed in
vacuo. The resultant solid was purified by column
chromatography, eluting with petroleum ether–ethyl acetate
(1:1). Bis-[(2S)-2-oxo-4-(2,2,2-trifluoroacetamido)-4-
carboxylmethylbutyl]sulfide 13 was collected as a yellow/
orange solid which was recrystallised from chloroform
(194 mg, 58%); [a]2D0ZK24.47 (c 0.41, MeOH); m/z [Cve
FAB (3-NBA)] 535 ([MCNa]C) and 513 ([MCH]C); m/z
[EI] 512 ([M]C); nmax (film)/cmK1 3311 (NH) and 1709
3.1.1. Methyl (2S)-5-bromo-4-oxo-2-(2,2,2-trifluoroace-
tamido)-pentanoate (10). Method A. Methyl (2S)-5-diazo-
4-oxo-2-(2,2,2-trifluoroacetamido)-pentanoate 9 (3.6 g,
13.48 mmol) was dissolved in distilled chloroform
(100 ml) and the mixture was stirred at room temperature.
Hydrobromic acid (48%, ca. 25 ml) was added dropwise
until nitrogen evolution ceased. The mixture was washed
with de-ionised water (3!20 ml). The organic layer was
dried (MgSO4) and the solvent was removed in vacuo to
give methyl (2S)-5-bromo-4-oxo-2-(2,2,2-trifluoroaceta-
mido)-pentanoate 10 as a pale yellow solid (3.83 g, 89%);
mp 117–118 8C; [a]D30ZC3.0 (c 1.01, CH2Cl2). (Found: C
30.2; H 2.6; N 4.5. C8H9NO4BrF3 requires C 30.0; H 2.8; N
4.4%); m/z [Cve FAB (3-NBA)] 322 and 320 (1:1)
([MCH]C); nmax (KBr)/cmK1 1750 (ester), 1727 (ketone)
and 1705 (amide); dH (360 MHz, C2HCl3) 3.32 (1H, dd,
J3B,2Z4.1 Hz, J3A,3BZ18.6 Hz, H-3A), 3.50 (1H, ABX,
J2A,3Z4.1 Hz, J3B,3AZ18.6 Hz, H-3B), 3.80 (3H, s, OCH3),
3.91 (2H, s, CH2Br), 4.84 (1H, m, H-2) and 7.33 (1H, br s,
NH, exchanges in 2H2O); dC (75.5 MHz, C2HCl3) 33.5
(CH2Br), 40.9 (C-3), 48.9 (C-2), 53.5 (OCH3), 169.7 (ester)
and 200.5 (ketone).
2
(br, amide and ketone); dH (500 MHz, H6-DMSO) 3.06
(2H, dd, J3A,2Z8.0 Hz, J3A,3BZ18.0 Hz, H-3A), 3.22
(2H, dd, J3B,2Z5.2 Hz, J3B,3AZ18.0 Hz, H-3B), 3.47 (4H,
d, JZ1.3 Hz, CH2S), 3.65 (6H, s, OCH3), 4.71 (2H, ddd,
J2,3AZ8.0 Hz, J2,3BZ5.2 Hz, J2,NHZ7.6 Hz, H-2) and 9.81
2
(2H, d, JNH,2Z7.6 Hz, NH); dC (125.8 MHz, H6-DMSO)
40.56 (CH2S), 40.74 (C-3), 48.12 (C-2), 52.55 (OCH3),
1
2
114.50–116.80 (q, JC,FZ288 Hz, CF3), 156.15 (q, JC,F
37 Hz, CF3CO), 169.99 (ester) and 201.63 (ketone).
Z
3.1.3. Methyl (2S)-5-[(2S)-2-tert-butoxycarbonyl-5-
pyrrolidinylidenyl]-4-oxo-2-(2,2,2-trifluoroacetamido)-
pentanoate (14). tert-Butyl (2S)-5-thioxoproline 1218
(49 mg, 0.25 mmol) and methyl (2S)-5-bromo-4-oxo-2-
(2,2,2-trifluoroacetamido)-pentanoate 10 (156 mg,
0.488 mmol) were heated at reflux in acetonitrile (5 ml)
under nitrogen. After 1 h a solution of triphenylphosphine
(258 mg, 0.975 mmol) and triethylamine (0.135 ml,
0.975 mmol) in acetonitrile (10 ml) was slowly added at a
rate of ca. 0.5 ml/min. After addition was complete, the
mixture was heated at reflux for 1 h and the solvent was
removed in vacuo. The crude mixture was purified by
column chromatography on silica gel, eluting with
petroleum ether–ethyl acetate (2:1) to yield methyl (2S)-5-
[(2S)-2-tert-butoxycarbonyl-5-pyrrolidinylidenyl]-4-oxo-2-
(2,2,2-trifluoroacetamido)-pentanoate 14 as a yellow oil
which foamed in vacuo (30 mg, 30%); [a]2D5ZC12.27 (c 1,
CHCl3); m/z [EI] Found: 408.152141 ([M]C).
C17H23N2O6F3 requires 408.150822; m/z [Cve FAB
(3-NBA)] 409 ([MCH]C); nmax (film)/cmK1 3318 (NH),
3083 (]CH) and 1728 (ketone and amide); lmax (MeOH)/
nm 304 (3 6604); dH (360 MHz, C2HCl3) 1.47 (9H, s,
C(CH3)3), 2.05–2.32 (2H, m, H-3), 2.62–2.72 (2H, m, H-4),
2.79 (1H, dd, J8A,9Z4.2 Hz, J8A,8BZ17.1 Hz, H-8A), 3.17
(1H, dd, J8B,9Z4.2 Hz, J8B,8AZ17.1 Hz, H-8B), 3.75 (3H,
s, OCH3), 4.32 (1H, dd, J2,3BZ5.4 Hz, J2,3AZ8.5 Hz, H-2),
4.77 (1H, dt, J9,8AZJ9,8BZ4.2 Hz, J9,NHZ7.2 Hz, H-9),
5.09 (1H, s, ]CH), 8.14 (1H, d, JNH,9Z7.2 Hz, NH,
On scale-up, varying amounts of methyl (2S)-5-chloro-4-
oxo-2-(2,2,2-trifluoroacetamido)-pentanoate, 11, were
obtained together with the bromide 10. This compound
had identical spectra to a sample prepared by Dr. R. A.
August17 with characterisation as follows: mp 131–133 8C,
[a]2D3ZC55.9 (c 1.1, CHCl3). (Found C, 35.0; H. 3.1; N,
4.9. C8H9ClF3NO4 requires C, 34.9; H, 3.3; N, 5.1%); m/z
(EI) 245 and 243 (1:3) ([MKMeOH]C); nmax (KBr)/cmK1
3301 (NH), 1747 (ester), 1739 (ketone) and 1708 (amide);
dH (360 MHz, C2HCl3) 3.25 (1H, dd, J3R,2Z4.2 Hz,
J3R,3SZ18.9 Hz, H-3R), 3.46 (1H, dd, J3S,2Z4.2 Hz,
J3S,3RZ18.9 Hz, H-3S), 3.80 (3H, s, OCH3), 4.11 (2H, s,
CH2Cl), 4.85 (1H, dt, J2,3RZJ2,3SZ4.2 Hz, J2,NHZ7.8 Hz,
H-2) and 7.36 (1H, br d, NH).
Method B. Methyl (2S)-5-chloro-4-oxo-2-(2,2,2-trifluoro-
acetamido)-pentanoate 11 (3.79 g, 13.7 mmol) was dis-
solved in acetone (100 ml) with stirring at room temperature
under nitrogen. Lithium bromide (11.9 g, 137 mmol) was
added in one portion with vigorous stirring. After 19 h, a
further portion of lithium bromide (5.97 g, 68.7 mmol) was
added and the reaction mixture was vigorously stirred for a
further 60 h. The solvent was removed in vacuo to yield an
amber oil which was dissolved in dichloromethane (75 ml)
2
exchanges in H2O) and 9.78 (1H, s, NH, exchanges in

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D. E. Davies et al. / Tetrahedron 61 (2005) 301–312
2H2O); dC (125.8 MHz, C2HCl3) 25.54 (C-3), 27.87
(C(CH3)3), 31.56 (C-4), 40.55 (C-8), 49.34 (C-2), 52.86
(C-9), 60.50 (C]CH), 61.82 (OCH3), 82.53 (OC(CH3)3),
89.72 (C]CH), 111–119 (q, 1JC,FZ287 Hz, CF3), 156–157
(q, 2JC,FZ38 Hz, CF3CONH), 167.69 (ester), 170.45 (ester)
and 193.26 (ketone).
solution was stirred under nitrogen, at room temperature, for
46 h. The mixture was diluted with dichloromethane
(15 ml). A solution of triphenylphosphine (4.45 g,
17 mmol) in dichloromethane (10 ml) was added via
syringe and, after stirring for 10 min, triethylamine
(3.6 ml, 25.6 mmol) was slowly added. The mixture was
stirred under nitrogen for a further 24 h at room temperature
and the solvents were removed in vacuo to yield a yellow/
brown solid which was purified by column chromatography
on silica gel eluting with cyclohexane–ethyl acetate (2:1).
This afforded methyl (2S)-5-[(2S)-1-benzyl-2-tert-butoxy-
carbonyl-5-pyrrolidinylidenyl]-4-oxo-2-(2,2,2-trifluoroace-
tamido)-pentanoate 18 as an amber oil (3.86 g, 91%);
[a]3D2ZC122.6 (c 1.025, CHCl3); m/z [EI] Found:
498.198641 ([M]C). C24H29N2O6F3 requires 498.197772;
m/z [Cve FAB (3-NBA)] 499 ([MCH]C); nmax (film)/cmK1
3315 (NH), 3065 (NH) and 1735 (br, ester); lmax (MeOH)/
nm 305 (3 24,750); dH (300 MHz, C2HCl3) 1.41 (9H, s,
3.1.4. tert-Butyl (2S)-1-benzylpyroglutamate (16). tert-
Butyl (2S)-pyroglutamate 1521 (5.1 g, 27.6 mmol) was
dissolved in anhydrous THF (50 ml) in a flask purged
with nitrogen. The solution was cooled to K78 8C and an
approximately 1 M solution of sodium hexamethyldisila-
zide in THF (30.3 ml, 30.3 mmol) was slowly added via
syringe, over a period of 20 min. After stirring for 1 h at
K78 8C, benzyl bromide (3.2 ml, 27 mmol) was added over
3–4 min. After 2.5 h the solution was slowly allowed to
warm to room temperature and stirred overnight. The
solvent was removed in vacuo and the residue was dissolved
in ethyl acetate (50 ml). The solution was washed with
water (50 ml) and the aqueous layer was extracted with
ethyl acetate (3!25 ml). The combined organic layers were
dried (MgSO4) and the solvents were removed in vacuo to
give an oil which was purified by column chromatography
on silica gel, eluting with cyclohexane–ethyl acetate (3:2).
tert-Butyl (2S)-1-benzylpyroglutamate 16 was obtained as a
white solid (4.47 g, 59%); mp 59–61 8C, [a]2D5ZC36.87
(c 1, CHCl3) (lit. mp20 62–63 8C, lit.23 [a]2D0ZC40.3
(c 0.51, CHCl3). (Found: C, 69.5; H, 7.7; N, 4.9. C16H21NO3
requires C, 69.8; H, 7.7; N, 5.1%); m/z [electrospray] 276
([MCH]C); nmax (film)/cmK1 1740 (ester) and 1676
(lactam); dH (300 MHz, C2HCl3) 1.37 (9H, s, C(CH3)3),
1.41–2.08 (1H, m, H-3A), 2.11–2.21 (1H, m, H-3B), 2.29–
2.39 (1H, m, H-4A), 2.44–2.52 (1H, m, H-4B), 3.76 (1H, dd,
C(CH3)3), 2.06–2.26 (2H, m, H-3), 2.81 (1H, dd, J8A,9
Z
4.0 Hz, J8A,8BZ17.0 Hz, H-8A), 3.03–3.12 (1H, m, H-4A),
3.21 (1H, dd, J8B,9Z4.0 Hz, J8A,8BZ17.0 Hz, H-8B), 3.38–
3.47 (1H, m, H-4B), 3.75 (3H, s, OCH3), 3.98 (1H, dd,
J2,3BZ3.1 Hz, J2,3AZ9.1 Hz, H-2), 4.22 (1H, d, JAB
Z
15.6 Hz, PhCHAN), 4.61 (1H, d, JBAZ15.6 Hz, PhCHBN),
4.71 (1H, ddd, J9,8AZJ9,8BZ4.0 Hz, J9,NHZ7.7 Hz, H-9),
5.18 (1H, s, ]CH), 7.14–7.38 (5H, m, aromatics) and 7.99
(1H, d, JNH,9Z7.7 Hz, NH, exchanges in 2H2O); dC
(75.5 MHz, C2HCl3) 26.46 (C-3), 28.26 (C(CH3)3), 32.66
(C-4), 42.82 (C-8), 49.73 (C-2), 49.89 (NCH2Ph), 53.25
(C-9), 60.42 (C]CH), 65.30 (OCH3), 82.94 (OC(CH3)3),
90.23 (C]CH), 111–123 (q, 1JC,FZ289 Hz, CF3), 128.00–
2
129.34 and 135.03 (aromatics), 156–157 (q, JC,FZ36 Hz,
CF3CO), 167.36 (ester), 170.97 (ester) and 193.31 (ketone).
J2,3AZ3.3 Hz, J2,3BZ9.1 Hz, H-2), 3.88 (1H, d, JAB
Z
14.8 Hz, PhCHAN), 4.99 (1H, d, JBAZ14.8 Hz, PhCHBN)
and 7.13–7.29 (5H, m, aromatics); dC (75.5 MHz, C2HCl3)
23.2 (C-3), 28.3 (C(CH3)3), 30.0 (C-4), 45.9 (NCH2Ph),
59.9 (C-2), 82.6 (OC(CH3)3), 128.1, 128.9, 129.1 and 136.3
(aromatics), 171.1 (ester) and 175.5 (amide).
3.1.7. Reduction of methyl (2S)-5-[(2S)-1-benzyl-2-tert-
butoxycarbonyl-5-pyrrolidinylidenyl]-4-oxo-2-(2,2,2-
trifluoroacetamido)-pentanoate 18. Trifluoroacetic acid
(0.97 ml, 13 mmol) and 10% palladium on activated carbon
(2.30 g, 75% by weight) were added to a solution of methyl
(2S)-5-[(2S)-1-benzyl-2-tert-butoxycarbonyl-5-pyrrolidiny-
lidenyl]-4-oxo-2-(2,2,2-trifluoroacetamido)-pentanoate 18
(3.081 g, 6.18 mmol) in methanol (150 ml). The suspension
was hydrogenated at 60 psi for 50 h in a Parr pressure
reactor. The reaction vessel was repressurised as necessary.
The mixture was filtered through Celitew and the filtrate was
adjusted to pH 7 by addition of 2 M aqueous sodium
hydroxide. The solvents were removed in vacuo and the
residual oil was dissolved in dichloromethane (60 ml). The
solution was washed with water (20 ml) and the aqueous
layer was further extracted with dichloromethane
(2!20 ml). The combined organic layers were dried
(MgSO4) and the solvent was removed in vacuo to yield a
yellow oil which was purified by column chromatography
on silica gel, eluting with ethyl acetate. This provided
methyl (2S)-5-[(2S,5S)-2-tert-butoxycarbonyl-5-pyrroli-
dinyl]-2-(2,2,2-trifluoroacetamido)-pentanoate 19 as an
amber oil (1.36 g, 56%); [a]2D8ZC8.53 (c 1, CHCl3); m/z
[EI] Found: 397.194013 ([MCH]C). [C17H27N2O5F3CH]
requires 397.195032; m/z [Cve FAB (3-NBA)] 397
([MCH]C); nmax (film)/cmK1 3326 (NH) and 1724
(ester); dH (360 MHz, C2HCl3) 1.16–1.27 (2H, m, H-7),
1.45 (9H, s, C(CH3)3), 1.54–1.57 (2H, m, H-4), 1.82–2.08
(6H, m, H-3, H-6CH-8), 2.60 (O1H, br s, NH, exchanges
3.1.5. tert-Butyl (2S)-1-benzyl-5-thioxoproline (17). This
was prepared using the method of Rapoport20 from tert-
butyl 1-benzyl-(2S)-pyroglutamate 16 as a white solid (88%
yield); mp 84.8–85.9 8C (lit.20 78–79 8C). (Found: C, 65.8;
H, 7.4; N, 4.8; S, 10.9. C16H21NO2S requires C, 65.95; H,
7.3; N, 4.8; S, 11.0%); m/z [Cve electrospray] 292
([MCH]C); nmax (KBr)/cmK1 1730 (ester); dH (300 MHz,
C2HCl3) 1.43 (9H, s, C(CH3)3), 2.10–2.35 (2H, m, H-3)
3.05–3.2 (2H, m, H-4), 4.08 (1H, dd, J2,3BZ3.3 Hz, J2,3A
Z
9.3 Hz, H-2), 4.21 (1H, d, JABZ14.6 Hz, PhCHAN), 4.75
(1H, d, JBAZ14.6 Hz, PhCHBN) and 7.20–7.30 (5H, m,
aromatics); dC (75.5 MHz, C2HCl3) 23.8 (C-3), 26.8
(C(CH3)3), 42.48 (C-4), 49.45 (NCH2Ph), 65.33 (C-2),
81.87 (OC(CH3)3), 127.16–133.65 (aromatics), 168.17
(ester) and 202.59 (C]S).
3.1.6. Methyl (2S)-5-[(2S)-1-benzyl-2-tert-butoxycarbo-
nyl-5-pyrrolidinylidenyl]-4-oxo-2-(2,2,2-trifluoroaceta-
mido)-pentanoate (18). tert-Butyl (2S)-1-benzyl-5-
thioxoproline 17 (2.49 g, 8.55 mmol) and methyl (2S)-5-
bromo-4-oxo-2-(2,2,2-trifluoroacetamido)-pentanoate 10
(4.10 g, 12.8 mmol) were mixed in a flask purged with
nitrogen. Acetonitrile (25 ml) was added via syringe and the

D. E. Davies et al. / Tetrahedron 61 (2005) 301–312
307
in 2H2O), 3.01 (1H, br s, H-9), 3.62 (1H, dd, JZ4.9, 9.2 Hz,
H-5), 3.77 (3H, s, OCH3), 4.55 (1H, br s, H-2) and 7.75 (1H,
br s, NH, exchanges in 2H2O); dC (75.5 MHz, 2H6-benzene)
23.18 (C-4), 28.05 (C(CH3)3), 30.72 and 31.02 (C-6 and
C-7), 32.35 (C-3), 34.70 (C-8), 52.19 (OCH3), 53.53 (C-9),
59.49 (C-5), 60.78 (C-2), 80.92 (OC(CH3)3), 171.59 (ester)
and 174.64 (ester).
2.19 (1H, dt, J1A,2/10Z7.3 Hz, J1A,1BZ12.5 Hz, H-1A),
3.76 (1H, dd, J10,1AZ7.3 Hz, JZ16.7 Hz, H-10), 4.35 (1H,
m, J6,7BZ5.7 Hz, J6,7AZ10.5 Hz, H-6), 4.48 (1H, t, J3,2
Z
5.8 Hz, H-3) and 7.88 (1H, br s, NH exchanges in 2H2O); dC
(75.5 MHz, C2HCl3) 27.81 and 28.19 (C-1 and C-8), 28.41
(C(CH3)3), 30.51 (C-9), 33.22 (C-2), 34.51 (C-7), 54.23
(C-6), 59.75 (C-10), 61.91 (C-3), 82.25 (OC(CH3)3), 169.78
and 171.08 (2!C]O).
On one occasion repetition of the hydrogenation using
identical quantities and conditions to those outlined above,
gave a second product after purification by column
chromatography. The second compound, tert-butyl
(3S,6S,8S,10S)-8-hydroxy-6-(2,2,2-trifluoroacetylamino)-5-
oxo-octahydro-1H-pyrrolo[1,2-a]azepine-3-carboxylate 22
was collected as a yellow crystalline solid (17% yield); mp
141–143 8C; [a]2D8ZC14.57 (c 1, CHCl3). (Found: C 50.8,
H 6.1, N 7.3. C16H23N2O5F3 requires C 50.5, H 6.1, N
7.4%); m/z [Cve electrospray] 381 ([MCH]C); nmax (KBr)/
cmK1 3407 (OH), 1733 (ester) and 1656 (amide); dH
(250 MHz, C2HCl3) 1.48 (9H, s, C(CH3)3), 1.67–1.90 (3H,
m, H-2 and H-7A), 2.04–2.19 (3H, m, H-1B and H-9),
3.1.9. tert-Butyl (3S,6S,10S)-6-amino-5-oxo-octahydro-
1H-pyrrolo[1,2-a]azepine-3-carboxylate (23). A suspen-
sion of tert-butyl (3S,6S,10S)-6-(2,2,2-trifluoroacetylamino)-5-
oxo-octahydro-1H-pyrrolo[1,2-a]azepine-3-carboxylate 8
(158 mg, 0.434 mmol) and potassium carbonate (310 mg,
2.25 mmol) in methanol–water (12:1, 13 ml) was heated at
reflux for 2 h. The solvent was removed in vacuo, the
residue was dissolved in dichloromethane (30 ml) and water
(20 ml) was added. The aqueous layer was extracted with
a further two portions of dichloromethane (30 ml). The
combined organic fractions were dried (MgSO4) and the
solvent was removed in vacuo to yield t-butyl (3S,6S,10S)-
6-amino-5-oxo-octahydro-1H-pyrrolo[1,2-a]azepine-3-car-
boxylate 23 as a yellow oil (100 mg, 86%); [a]2D3ZK80.76
(c 1.01, CH2Cl2); m/z [Cve electrospray] Found:
268.178708 ([M]C). C14H24N2O3 requires 268.178693;
m/z [Cve thermospray] 269 ([MCH]C); nmax (diffuse
reflectance)/cmK1 3362 (NH), 1737 (ester) and 1657
(amide); dH (300 MHz, C2HCl3) 1.46 (9H, s, C(CH3)3),
1.55–1.80 (5H, m, H-1B, H-9B, H-8B, H-7), 1.87 (1H, br,
H-9A), 1.96–2.02 (3H, m, H-2 and H-8A), 2.20 (1H, td,
J1A,10/2Z7.0 Hz, J1A,1BZ12.0 Hz, H-1A), 2.29 (2H, br s,
NH, exchanges in 2H2O), 3.46 (1H, br s, H-6), 3.75 (1H, dd,
J10,1AZ7.0 Hz, JZ15.2 Hz, H-10) and 4.49–4.54 (1H, t,
J3,2Z5.5 Hz, H-3); dC (62.9 MHz, C2HCl3) 27.67 and 28.19
(C-1 and C-8), 28.02 (C(CH3)3), 29.70 (C-9), 33.07 (C-2),
34.58 (C-7), 55.16 (C-6), 58.69 (C-10), 61.51 (C-3), 81.26
(OC(CH3)3), 171.45 and 174.89 (2!C]O).
2.25–2.40 (2H, m, H-1A and H-7B), 2.57 (1H, br s, OH,
2
exchanges in H2O), 3.77–3.88 (1H, td, JZ7 Hz, J10,1A
Z
Z
10 Hz, H-10), 4.03–4.17 (1H, tt, J8,9B/7BZ4 Hz, J8,9A/7A
11 Hz, H-8), 4.38 (1H, dd, J6,NHZ6 Hz, J6,7Z11 Hz, H-6),
4.55 (1H, t, J3,2Z6 Hz, H-3) and 7.99 (1H, d, JNH,6Z6 Hz,
2
NH, exchanges in H2O); dC (100.5 MHz, C2HCl3) 27.77
(C-1), 27.93 (C(CH3)3), 32.66 (C-2), 38.57 (C-9), 42.52
(C-7), 49.13 (C-10), 54.64 (C-6), 61.51 (C-3), 69.68 (C-8),
1
82.05 (OC(CH3)3), 111–120 (q, JC,FZ287.8 Hz, CF3),
156.0–156.5 (q, 2JC,FZ37.4 Hz, COCF3), 168.16 (ester) and
170.40 (amide).
3.1.8. tert-Butyl (3S,6S,10S)-6-(2,2,2-trifluoroacetyla-
mino)-5-oxo-octahydro-1H-pyrrolo [1,2-a]azepine-3-
carboxylate (8). Methyl (2S)-5-[(2S,5S)-2-tert-butoxycar-
bonyl-5-pyrrolidinyl]-2-(2,2,2-trifluoroacetamido)-pen-
tanoate 19 (1.30 g, 3.29 mmol) was dissolved in anhydrous
diethyl ether (70 ml) under nitrogen. The solution was
cooled to 0 8C and tert-butyl magnesium chloride in diethyl
ether (ca. 2 M, 8.3 ml, 16.4 mmol) was added via syringe
over a period of 30 min. A white precipitate instantly
formed. The mixture was stirred at 0 8C for 2 h, the ice bath
was removed and the solution was stirred overnight at room
temperature. Saturated aqueous ammonium chloride
(40 ml) was added and the aqueous layer was diluted with
water until all of the precipitate had dissolved. The solution
was extracted with diethyl ether (2!40 ml). The combined
organic layers were dried (MgSO4) and the solvent was
removed in vacuo to give a yellow oil, which was purified
by column chromatography on silica gel, eluting with
cyclohexane–ethyl acetate (2:1). tert-Butyl (3S,6S,10S)-6-
(2,2,2-trifluoroacetylamino)-5-oxo-octahydro-1H-pyrrolo
[1,2-a]azepine-3-carboxylate 8 was collected as a yellow
crystalline solid (338 mg, 29%); mp 127–129 8C;
[a]3D2ZK20.6 (c 1, CHCl3). (Found: C 52.7; H 6.3; 7.6.
C16H23N2O4F3 requires C 52.7; H 6.4; N 7.7%); m/z [EI]
Found 365.169228 ([MCH]C). [C16H23N2O4F3CH]
requires 365.168817; m/z [Cve electrospray] 387
([MCNa]C) and 365 ([MCH]C); nmax (KBr)/cmK1 3343
(NH), 1734 (ester), 1700 and 1661 (amide); dH (300 MHz,
C2HCl3) 1.42 (9H, s, C(CH3)3), 1.52–1.82 (5H, m, H-1B,
H-9, H-8B, H-7B), 1.87–2.08 (4H, m, H-2, H-8A, H-7A),
3.1.10. tert-Butyl (3S,6S,10S)-6-[(2S)-9-fluorenylmethoxy-
carbonylamino-3-methylbutanamido]-5-oxo-octahydro-
1H-pyrrolo[1,2-a]azepine-3-carboxylate (24). tert-Butyl
(3S,6S,10S)-6-amino-5-oxo-octahydro-1H-pyrrolo[1,2-a]
azepine-3-carboxylate 23 (100 mg, 0.373 mmol) and
9-fluorenylmethoxycarbonyl-(2S)-valine (127 mg, 0.373
mmol) were dissolved in dry dimethylformamide (7 ml)
under nitrogen. 2-(1H-Benzotriazole-1-yl)-1,1,3,3-tetra-
methyluronium tetrafluoroborate (TBTU) (120 mg,
0.373 mmol) and diisopropylethylamine (0.065 ml,
0.373 mmol) were added and the solution was stirred
overnight at room temperature. The solvent was removed in
vacuo at 30 8C. The residue was dissolved in ethyl acetate
(40 ml) and washed with 5% aqueous citric acid (2!15 ml),
5% aqueous sodium hydrogen carbonate (2!15 ml) and
brine (2!15 ml). The organic layer was dried (MgSO4) and
the solvent was removed in vacuo to give a yellow oil which
was purified by column chromatography on silica gel,
eluting with methanol–dichloromethane (3:97) to give tert-
butyl (3S,6S,10S)-6-[(2S)-9-fluorenylmethoxycarbonyla-
mino-3-methylbutanamido]-5-oxo-octahydro-1H-pyrrolo
[1,2-a]azepine-3-carboxylate 24 as a white solid (176 mg,
80%); mp 129–131 8C; [a]2D5ZK24.40 (c 1.08, CHCl3);
m/z [electrospray] Found: 590.322190 ([MCH]C).
[C34H43N3O6CH] requires 590.323012; nmax (diffuse

308
D. E. Davies et al. / Tetrahedron 61 (2005) 301–312
reflectance)/cmK1 3314 (NH), 1725 (ester) and 1638
(amide); dH (300 MHz, C2HCl3) 0.85 (6H, t, Jb,b-Me
7.3 Hz, 2!b-Me), 1.38 (9H, s, C(CH3)3), 1.44–1.99 (O9H,
m, H-2, H-1B, H-9, H-8 and H-7), 2.04–2.15 (2H, m,
azepine-3-carboxylate 26 as a yellow oil (316 mg, 64%);
[a]3D0ZK32.46 (c 1, CHCl3); m/z [Cve electrospray]
Found: 717.364916 ([MCNH4]C)]. [C43H45N3O6CNH4]
requires 717.365211; m/z [Cve electrospray] 717 ([MC
NH4]C)] and 700 ([MCH]C); nmax (film)/cmK1 3309 (NH),
1720 (ester) and 1637 (amide); dH (300 MHz, C2HCl3) 0.85
Z
H-1ACH-b), 3.72 (1H, m, H-10), 4.04 (1H, dd, Ja,b
Z
7.0 Hz, Ja,NHZ8.6 Hz, H-a), 4.14 (1H, t, J6,7Z7.0 Hz,
H-6), 4.34–4.39 (3H, m, CH2CH-fluorenenyl), 4.44 (1H, t,
J3,2Z5.6 Hz, H-3), 5.53 (1H, d, JNH,aZ8.6 Hz, NH
(3H, d, Jb-MeA,bZ6.9 Hz, b-MeA), 0.88 (3H, d, Jb-MeB,b
6.8 Hz, b-MeB), 1.35–2.26 (11H, m, H-2, H-1, H-9, H-8,
H-7CH-b), 3.75 (1H, m, H-10), 4.01–4.08 (1H, q, Ja,b
Z
2
exchanges in H2O) and 7.19–7.68 (8H, m, aromatics); dC
Z
(62.9 MHz, C2HCl3) 17.58 (b-Me), 27.46 and 27.65 (C-1
and C-8), 27.92 (C(CH3)3), 31.07, 32.78 and 34.14 (C-2,
C-9 and C-7), 47.13 (C-a), 53.31 (C-b), 59.17 (C-6), 60.01
(C-10), 61.23 (C-3), 66.94 (Fmoc-CH2), 81.52 (OC(CH3)3),
119.85–127.56, 141.20 and 143.89 (aromatics) and 170.50–
170.83 (4!C]O).
7.1 Hz, H-a), 4.13–4.39 (4H, m, H-6 and CH2CH-
fluorenenyl), 4.73 (1H, t, J3,2Z5.5 Hz, H-3), 5.42 (1H, d,
JZ8.7 Hz, NH), 6.79 (1H, s, OCHAr2) and 7.18–7.69 (19H,
m, NHCaromatics); dC (75.5 MHz, C2HCl3) 18.09 and
19.63 (b-Me), 27.93 (C-1 and C-8), 30.13, 31.43, 33.27 and
34.53 (C-b, C-2, C-9 and C-7), 47.62 (C-a), 59.63 and 60.57
and 61.02 (C-6, C-10 and C-3), 67.47 (Fmoc-CH2), 78.44
(OCHAr2), 120.38–129.01 and 139.88–144.69 (aromatics)
and 156.70–171.27 (4!C]O).
3.1.11. (3S,6S,10S)-6-[(2S)-9-Fluorenylmethoxycarbon-
ylamino-3-methylbutanamido]-5-oxo-octahydro-1H-pyr-
rolo[1,2-a]azepine-3-carboxylate (25). tert-Butyl (3S,6S,
10S)-6-[(2S)-9-fluorenylmethoxycarbonylamino-3-methyl-
butanamido]-5-oxo-octahydro-1H-pyrrolo[1,2-a]azepine-3-
carboxylate 24 (375 mg, 0.634 mmol) was dissolved in
dichloromethane (12 ml). Trifluoroacetic acid (4 ml) was
added and the solution was stirred for 4 h at room
temperature, during which time the yellow solution turned
brown. The solvents were removed in vacuo to yield a
brown oil which was washed with diethyl ether (4!10 ml).
Removal of the solvents in vacuo followed by drying in
vacuo yielded (3S,6S,10S)-6-[(2S)-9-fluorenylmethoxycar-
bonylamino-3-methylbutanamido]-5-oxo-octahydro-1H-
pyrrolo[1,2-a]azepine-3-carboxylate 25 as a yellow solid
(379 mg, Oquantitative); mp 131–132 8C; [a]2D6ZK69.57
(c 0.465, CHCl3); m/z [Cve electrospray] Found:
551.287467 ([MCNH4]C). [C30H35N3O6CNH4] requires
551.286960; nmax (KBr)/cmK1 2927 (acid), 1719 (acid) and
1637 (amide); dH (250 MHz, C2HCl3/C2H3O2H) 0.88 (6H, t,
Jb-Me,bZ7.2 Hz, b-Me), 1.53–2.23 (11H, m, H-2, H-1, H-9,
H-8, H-7 and H-b), 3.80 (1H, m, H-10), 4.01 (1H, m, H-a),
4.25–4.43 (4H, m, H-6 and CH2CH-fluorenenyl), 4.55 (1H,
dd, J3,2BZ4.2 Hz, J3,2AZ6.9 Hz, H-3), 5.82 (1H, d, JZ
8 Hz, NH) and 7.21–7.72 (8H, m, aromatics); dC (75.5 MHz,
C2HCl3) 14.59 and 18.23 (b-Me), 27.67 and 27.81 (C-1 and
C-8), 30.12, 31.23, 33.34 and 34.35 (C-b, C-2, C-9 and C-7),
47.47 (C-a), 59.91 and 60.82 and 61.14 (C-6, C-10 and
C-3), 67.78 (Fmoc-CH2), 120.38–128.14, 141.61 and
144.26 (aromatics) and 157.09–170.83 (4!C]O).
3.1.13. Diphenylmethyl (3S,6S,10S)-6-[(2S)-2-amino-3-
methylbutanamido]-5-oxo-octahydro-1H-pyrrolo[1,2-
a]azepine-3-carboxylate (27). Diphenylmethyl (3S,6S,
10S)-6-[(2S)-9-fluorenylmethoxycarbonylamino-3-methyl-
butanamido]-5-oxo-octahydro-1H-pyrrolo[1,2-a]azepine-3-
carboxylate 26 (310 mg, 0.44 mmol) was dissolved in dry
dimethylformamide (6 ml). Piperidine (1.2 ml, 20% by
volume) was added and the solution was stirred at room
temperature under nitrogen for 2.5 h. The solvent was
removed in vacuo at !50 8C to yield a white solid which
was purified by column chromatography on silica gel,
eluting with a gradient of methanol–dichloromethane (1:99
to 5:95). Diphenylmethyl (3S,6S,10S)-6-[(2S)-2-amino-3-
methylbutanamido]-5-oxo-octahydro-1H-pyrrolo[1,2-a]
azepine-3-carboxylate 27 was collected as a yellow foam
(182 mg, 87%); [a]3D0ZK16.18 (c 1, CHCl3); m/z [Cve
electrospray]
Found:
478.271527
([MCH]C).
[C28H35N3O4CH] requires 478.270582; m/z [Cve electro-
spray] 955 ([2MCH]C) and 478 ([MCH]C); nmax (film)/
cmK1 3368 (NH), 1744 (ester) and 1640 (amide); dH
(300 MHz, C2HCl3) 0.79 (3H, d, Jb-MeA,bZ6.9 Hz, b-MeA),
0.92 (3H, d, Jb-MeB,bZ6.9 Hz, b-MeB), 1.40–2.03 (O9H,
m, H-2, H-1B, H-9, H-7, H-8), 2.10–2.20 (2H, m, H-b and
H-1A), 2.28 (2H, br s, NH, exchanges in 2H2O), 3.21 (1H, br
d, Ja,bZ3.9 Hz, H-a), 3.78 (1H, m, H-10), 4.41 (1H, br dd,
JZ6.5, 10.4 Hz, H-6), 4.73 (1H, dd, J3,2BZ4.0 Hz, J3,2A
Z
7.6 Hz, H-3), 6.81 (1H, s, OCHAr2), 7.19–7.94 (10H, m,
aromatics) and 8.14 (1H, d, JZ6.5 Hz, NH, exchanges in
2H2O); dC (75.5 MHz, C2HCl3) 16.87 and 19.97
(CH(CH3)2), 27.91 and 28.00 (C-1 and C-8), 31.58 (C-b),
31.65, 33.31 and 34.62 (C-2, C-9 and C-7), 53.49 (C-a),
59.55, 60.60 and 61.03 (C-6, C-10 and C-3), 78.30
(OCHAr2), 126.30–129.45, 139.92 and 140.24 (aromatics)
and 171.37–173.40 (3!C]O).
3.1.12. Diphenylmethyl (3S,6S,10S)-6-[(2S)-9-fluorenyl-
methoxycarbonylamino-3-methylbutanamido]-5-oxo-
octahydro-1H-pyrrolo[1,2-a]azepine-3-carboxylate (26).
(3S,6S,10S)-6-[(2S)-9-Fluorenylmethoxycarbonylamino-3-
methylbutanamido]-5-oxo-octahydro-1H-pyrrolo[1,2-a]
azepine-3-carboxylate 25 (379 mg, 0.711 mmol) was dis-
solved in dichloromethane (25 ml). Diphenyldiazomethane
(205 mg, 1.67 mmol)24 was added via pipette. The pale
brown solution turned red and there was some effervescence
which quickly subsided. The mixture was stirred at room
temperature overnight. The solvent was removed in vacuo
to yield a red oil which was purified by column
chromatography on silica gel, eluting with petroleum
ether–ethyl acetate (1:1) to give diphenylmethyl
(3S,6S,10S)-6-[(2S)-9-fluorenylmethoxycarbonylamino-3-
methylbutanamido]-5-oxo-octahydro-1H-pyrrolo[1,2-a]
3.1.14. Diphenylmethyl (3S,6S,10S)-6-[(2S)-2-((2S)-2-
(9H)-fluoren-9-ylmethoxycarbonylamino-4-tert-butoxy-
carbonylpropanamido)-3-methylbutanamido]-5-oxo-
octahydro-1H-pyrrolo[1,2-a]azepine-3-carboxylate (28).
Diphenylmethyl (3S,6S,10S)-6-[(2S)-2-amino-3-methylbu-
tanamido]-5-oxo-octahydro-1H-pyrrolo[1,2-a]azepine-3-
carboxylate 27 (170 mg, 0.356 mmol) and 4-tert-butyl
N-9-fluorenylmethoxycarbonyl-(2S)-aspartate (161 mg,
0.392 mmol) were dissolved in dry dimethylformamide

D. E. Davies et al. / Tetrahedron 61 (2005) 301–312
309
0
0
0
0
(10 ml). 2-(1H-Benzotriazole-1-yl)-1,1,3,3-tetramethyluro-
nium tetrafluoroborate (TBTU) (126 mg, 0.392 mmol) and
diisopropylethylamine (0.068 ml, 0.392 mmol) were added
and the solution was stirred under nitrogen at room
temperature overnight. The solvent was removed in vacuo at
!50 8C and the resulting oil was dissolved in dichloro-
methane and washed with 5% aqueous citric acid, 5% aqueous
sodium hydrogen carbonate and brine. The organic layer was
dried (MgSO4) and the solvent was removed in vacuo. The
residue was purified by column chromatography on silica gel,
eluting with methanol–dichloromethane (3:97) to yield
diphenylmethyl (3S,6S,10S)-6-[(2S)-2-((2S)-2-(9H)-fluoren-
9-ylmethoxycarbonylamino-4-tert-butoxycarbonylpropana-
mido)-3-methylbutanamido]-5-oxo-octahydro-1H-pyrrolo
[1,2-a]azepine-3-carboxylate 28 as a white powder (274 mg,
88%); [a]2D8ZK14.74 (c 1, CHCl3); m/z [Cve electrospray]
Found: 888.455720 ([MCNH4]C). [C51H58N4O9CNH4]
requires 888.454754; m/z [Cve electrospray] 909
([MCK]C), 893 ([MCNa]C) and 871 ([MCH]C); nmax
(film)/cmK1 1729 (ester) and 1646 (amide); dH (300 MHz,
C2HCl3) 0.93–0.98 (6H, t, Jb-Me,bZ6.8 Hz, b-Me, 1.46 (9H, s,
C(CH3)3), 1.54–2.26 (11H, m, H-2, H-1, H-9, H-7, H-8C
Jb B,a Z4 Hz, Jb B,b AZ17 Hz, H-b0B), 3.73 (1H, dd,
Ja ,b BZ4 Hz, Ja ,b AZ8 Hz, H-a0), 3.83 (1H, br dd, JZ8,
17 Hz, H-10), 4.23 (1H, dd, Ja,bZ6 Hz, Ja,NHZ11 Hz,
0
0
0
0
H-a), 4.40 (1H, dd, JZ7, 10 Hz, H-6), 4.76 (1H, dd, J3,2B
Z
5 Hz, J3,2AZ7 Hz, H-3), 6.84 (1H, s, OCHAr2), 7.25–7.35
(10H, m, aromatics), 7.43 (1H, d, JZ6 Hz, NH) and 8.00
(!1H, d, JZ8 Hz, NH); dC (62.9 MHz, C2HCl3) 17.62 and
19.27 (2!b-Me), 27.44 (C-1 and C-8), 28.03 (C(CH3)3),
31.04 (C-b0 and C-b), 32.79, 34.09 and 39.87 (C-2, C-9 and
C-7), 52.09 (C-a0), 53.28 (C-a), 58.26 (C-6), 59.13 (C-10),
60.53 (C-3), 77.92 (OCH(Ph)2), 81.27 (OC(CH3)3), 128.98–
128.47, 139.43 and 139.70 (aromatics) and 169.98–173.07
(5!C]O).
3.1.16. Diphenylmethyl (3S,6S,10S)-6-[(2S)-2-(((2S)-
2-((2S)-2-(9H)-fluor-9-enylmethoxycarbonylamino-4-
methylpentanamido)-4-tert-butoxycarbonylpropana-
mido)-3-methylbutanamido]-5-oxo-octahydro-1H-pyr-
rolo[1,2-a]azepine-3-carboxylate (30). Diphenylmethyl
(3S,6S,10S)-6-[(2S)-2-((2S)-2-amino-4-tert-butoxycarbo-
nylpropanamido)-3-methylbutanamido]-5-oxo-octahydro-
1H-pyrrolo[1,2-a]azepine-3-carboxylate 29 (180 mg,
0.277 mmol) and 9-fluorenylmethoxycarbonyl-(2S)-leucine
(108 mg, 0.305 mmol) were dissolved in dry dimethyl-
formamide (3 ml). 2-(1H-Benzotriazole-1-yl)-1,1,3,3-
tetramethyluronium tetrafluoroborate (TBTU) (98 mg,
0.305 mmol) and diisopropylethylamine (0.054 ml,
0.305 mmol) dissolved in dimethylformamide (1 ml) were
added and the solution was stirred under nitrogen at room
temperature overnight. The solvent was removed in vacuo
and the residue was dissolved in dichloromethane and
washed with 5% aqueous citric acid, followed by 5%
aqueous sodium hydrogen carbonate and brine. The
organic layer was dried (MgSO4) and the solvent was
removed in vacuo to give an oil which was purified by
column chromatography on silica gel, eluting with
methanol–dichloromethane (3:97). This yielded diphenyl-
methyl (3S,6S,10S)-6-[(2S)-2-(((2S)-2-((2S)-2-(9H)-fluor-9-
enylmethoxycarbonylamino-4-methylpentanamido)-4-tert-
butoxycarbonylpropanamido)-3-methylbutanamido]-5-oxo-
octahydro-1H-pyrrolo[1,2-a]azepine-3-carboxylate 30 as a
white foam (223 mg, 82%); [a]2D9ZK30.65 (c 1, CHCl3);
m/z [Cve FAB (3-NBA)] 983 ([M]C); nmax (film)/cmK1
3301 (NH), 1723 (ester) and 1637 (amide); dH (300 MHz,
C2HCl3) 0.80–0.85 (12H, m, b-MeCg00-Me), 1.32 (9H, s,
C(CH3)3), 1.40–1.99 (12H, m, H-2, H-1B, H-9, H-7, H-8,
H-b00CH-g00), 2.04–2.14 (2H, m, J1A,10Z7.1 Hz, H-1A and
0
0
0
0
H-b), 2.73 (1H, dd, Jb A,a Z6.7 Hz, Jb A,b BZ17.4 Hz,
H-b0A), 2.97 (1H, dd, Jb B,a Z4.1 Hz, Jb B,b AZ17.4 Hz,
0
0
0
0
H-b0B), 3.82 (1H, m, JZ9.4 Hz, H-10), 4.26 (1H, q, Ja,b
Z
7.2 Hz, H-a),4.32(1H, dd, J6,NHZ5.6 Hz, J6,7Z8.2 Hz, H-6),
4.42 (3H, br d, JZ7.2 Hz, CH2CH-fluorenenyl), 4.60 (1H, br
0
0
0
s, H-3), 4.80 (1H, t, Ja ,b Z5.2 Hz, H-a ), 6.12 (1H, d, JZ
8.4 Hz, NH, exchanges in 2H2O), 6.89 (1H, s, OCHAr2), 7.16
2
(1H, d, JNH,6Z5.6 Hz, NH, exchanges in H2O), 7.25–7.80
(18H, m, aromatics); dC (62.9 MHz, C2HCl3) 17.56 and 19.17
(b-Me), 27.42 (C-1 and C-8), 27.91 (C(CH3)3), 30.93 (C-b0
and C-b), 31.15, 32.76 and 37.41 (C-2, C-9 and C-7), 47.02
(C-a0), 53.27 (C-a), 58.62, 59.08 and 60.41 (C-6, C-10 and
C-3), 67.25 (FmocCH2), 76.49 (Fmoc-CH), 77.93 (OCHAr2),
81.78 (OC(CH3)3), 119.90–128.49, 139.40 and 143.76
(aromatics) and 155.97–171.44 (6!C]O).
3.1.15. Diphenylmethyl (3S,6S,10S)-6-[(2S)-2-((2S)-2-
amino-4-tert-butoxycarbonylpropanamido)-3-methylbu-
tanamido]-5-oxo-octahydro-1H-pyrrolo[1,2-a]azepine-
3-carboxylate (29). Diphenylmethyl (3S,6S,10S)-6-[(2S)-2-
((2S)-2-(9H)-fluoren-9-ylmethoxycarbonylamino-4-tert-
butoxycarbonylpropanamido)-3-methylbutanamido]-5-oxo-
octahydro-1H-pyrrolo[1,2-a]azepine-3-carboxylate 28
(250 mg, 0.287 mmol) was dissolved in dry dimethylfor-
mamide (5 ml). Piperidine (1 ml, 20% by volume) was
added and the solution was stirred at room temperature
under nitrogen for 2 h. The solvent was removed in vacuo
(!40 8C) and the residual oil was purified by column
chromatography on silica gel, eluting with a gradient of
methanol–dichloromethane (1:99–5:95). Diphenylmethyl
(3S,6S,10S)-6-[(2S)-2-((2S)-2-amino-4-tert-butoxycarbo-
nylpropanamido)-3-methylbutanamido]-5-oxo-octahydro-
1H-pyrrolo[1,2-a]azepine-3-carboxylate 29 was collected
as a white oily foam (152 mg, 82%); [a]2D8ZK42.91 (c 1,
CHCl3); m/z [Cve FAB (PEGH/NOBA)] Found:
649.364583 ([MCH]C). [C36H48N4O7CH] requires
649.360125; nmax (film)/cmK1 1727 (ester) and 1639
(amide); dH (250 MHz, C2HCl3/C2H3O2H) 0.91 (6H, t,
Jb-Me,bZ7 Hz, b-Me), 1.44 (9H, s, C(CH3)3), 1.47–2.27
(w11H, m, H-2, H-1, H-9, H-7, H-8 and H-b), 2.57 (1H, dd,
0
0
0
0
H-b), 2.58 (1H, dd, Jb A,b BZ17.0 Hz, JbA ,a Z7.0 Hz,
H-b0A), 2.74 (1H, dd, Jb B,b AZ17.0 Hz, JbB ,a Z3.7 Hz,
H-b0B), 3.70 (1H, br dd, JZ7.1, 16.8 Hz, H-10), 4.13 (1H, t,
J6,7Z6.9 Hz, H-6), 4.22–4.37 (5H, m, JZ7.0 Hz, H-a,
H-a00 and CH2CH-fluorenenyl), 4.73 (2H, br dd, H-3 and
0
0
0
0
2
H-a0), 5.35 (1H, d, JZ7.9 Hz, NH, exchanges in H2O),
6.78 (1H, s, OCHAr2), and 7.19–7.69 (21H, m, aromaticsC
3!NH, 3 protons exchange with H2O); dC (75.5 MHz,
2
C2HCl3) 18.13, 19.67, 22.25 and 23.40 (b-MeCg00-Me),
25.14 (C-g00), 27.86–27.93 (C-1, C-8 and C-b0), 28.37
(C(CH3)3), 31.50, 33.25, 34.57 and 39.41 (C-2, C-9, C-7 and
C-b), 42.29 (C-b00), 47.57, 53.60 and 53.70 (C-a, C-a0 and
C-a00), 59.04, 59.53 and 60.92 (C-3, C-10 and C-6), 67.45
(Fmoc-CH2), 78.32 (OCHAr2), 82.28 (OC(CH3)3),
120.37–128.99 and 139.90–144.35 (aromatics) and
156.56–172.59 (7!C]O).
Jb A ,a Z8 Hz, Jb A ,b BZ17 Hz, H-b0A), 2.81 (1H, dd,
0
0
0
0
0
0

310
D. E. Davies et al. / Tetrahedron 61 (2005) 301–312
3.1.17. Diphenylmethyl (3S,6S,10S)-6-[(2S)-2-(((2S)-2-
((2S)-2-amino-4-methylpentanamido)-4-tert-butoxycar-
bonylpropanamido)-3-methylbutanamido]-5-oxo-octa-
hydro-1H-pyrrolo[1,2-a]azepine-3-carboxylate (31).
Diphenylmethyl (3S,6S,10S)-6-[(2S)-2-(((2S)-2-((2S)-2-
(9H)-fluor-9-enylmethoxycarbonylamino-4-methylpentana-
mido)-4-tert-butoxycarbonylpropanamido)-3-methylbutana-
mido]-5-oxo-octahydro-1H-pyrrolo[1,2-a]azepine-3-carbox-
ylate 30 (200 mg, 0.203 mmol) was dissolved in dry
dimethylformamide (6 ml). Piperidine (20% by volume,
1.2 ml) was added and the solution was stirred at room
temperature under nitrogen for 2 h. The solvent was removed
in vacuo at 40 8C. The residual oil was purified by column
chromatography on silica gel, eluting with a gradient from
pure dichloromethane to dichloromethane–methanol (95:5).
Diphenylmethyl (3S,6S,10S)-6-[(2S)-2-(((2S)-2-((2S)-2-
amino-4-methylpentanamido)-4-tert-butoxycarbonylpropa-
namido)-3-methylbutanamido]-5-oxo-octahydro-1H-pyr-
rolo[1,2-a]azepine-3-carboxylate 31 was collected as a foam
(137 mg, 88%); [a]2D7ZK41.12 (c 0.98, CHCl3); m/z [Cve
FAB (PEGNa/NOBA)] Found: 762.445100 ([MCH]C).
[C42H59N5O8CH] requires 762.444190; m/z [Cve FAB
(3-NBA)] 762 ([MCH]C); nmax (film)/cmK1 1735 (ester)
and 1636 (amide); dH (250 MHz, C2HCl3) 0.89–0.97 (12H, m,
b-MeCg00-Me), 1.20 (9H, s, C(CH3)3), 1.51–2.25 (14H, m,
chromatography on silica gel, eluting with methanol–
dichloromethane (3:97). This gave diphenylmethyl
(3S,6S,10S)-6-[(2S)-2-((((2S)-2-(((2S)-2-(2-carbobenzyloxy-
aminoacetamido)-4-methylpentanamido)-4-tert-butoxycar-
bonylpropanamido)-3-methylbutanamido]-5-oxo-octa-
hydro-1H-pyrrolo[1,2-a]azepine-3-carboxylate 32 as a
white powder (110 mg, 81%); [a]3D0ZK29.68 (c 1,
CHCl3); nmax (film)/cmK1 3276 (NH), 1730 (ester) and
1630 (amide); m/z [Cve FAB (3-NBA)] 975 ([MCNa]C)
and 953 ([MCH]C); dH (250 MHz, 318 K, C2HCl3)
0.89–0.96 (12H, m, b-MeCg00-Me), 1.41 (9H, s,
C(CH3)3), 1.47–2.01 (O12H, m, H-2, H-1B, H-9, H-7,
H-8, H-b00CH-g00), 2.15 (2H, m, H-1A and H-b), 2.72 (1H,
0
0
0
0
0
dd, Jb A,b BZ16.6 Hz, Jb A,a Z6.8 Hz, H-b0A), 2.84 (1H,
0
0
0
0
dd, Jb B,b AZ16.6 Hz, Jb B,a Z5.7 Hz, H-b B), 3.72–3.92
(3H, m, H-10 and H-a000), 4.40 (2H, m, H-a and H-a000),
4.61–4.78 (1H, m, JZ5.4 Hz, H-6), 4.83 (2H, m, H-3 and
H-a0), 5.12 (2H, d, JZ5.4 Hz, OCH2C6H5), 5.81 (1H, br s,
2
NH, exchanges in H2O), 6.85 (1H, s, OCHAr2), 7.00 (1H,
d, JZ6.6 Hz, NH, exchanges in 2H2O), 7.30–7.38 (15H, m,
aromatics) and 7.63 (1H, d, JZ7.7 Hz, NH, exchanges in
2H2O); dC (75.5 MHz, C2HCl3) 18.37, 19.64, 23.18 and
25.22 (b-MeCg00-Me), 23.18 (C-g00), 27.84 (C-1 and C-8),
28.36 (C(CH3)3), 31.53 (C-b), 31.75 (C-b00), 33.51, 34.53,
and 38.29 (C-2, C-9 and C-7), 39.4 (C-b00), 42.30 (C-b0),
44.80 (C-a000), 52.02 (C-a000), 53.64 (C-a), 58.82, 59.27 and
60.73 (C-3, C-10 and C-6), 67.32 (OCH2Ph), 78.31
(OCHAr2), 81.71 (OC(CH3)3), 126.27–128.99, 136.92,
139.88 and 140.21 (aromatics) and 157.16–172.49
(8!C]O).
H-2, H-1, H-9, H-7, H-8, H-b00CH-g00CH-b), 2.50 (!2H, br
2
0
0
s, NH2, exchanges in0 H2O), 2.71 (1H, dd, Jb A,b B 16.9 Hz,
0
0
0
0
Jb A,a Z6.5 Hz, H-b A), 2.88 (1H, dd, Jb B,b AZ16.9 Hz,
0
00
0
0
Jb B,a Z5.2 Hz, H-b B), 3.50 (1H, m, H-a ), 3.80 (1H, br q,
JZ9.7 Hz, H-10), 4.25 (1H, dd, J6,7Z5.2 Hz, J6,NHZ8.1 Hz,
H-6), 4.40(1H, dd, JZ6.0, 10.2 Hz, H-a), 4.77(2H, brdd, H-3
and H-a0), 6.87 (1H, s, OCHAr2), 7.20 (1H, d, JZ6.0 Hz, NH,
2
3.1.19. (3S,6S,10S)-6-[((((2S)-2-(((2S)-2-(((2S)-2-(2-Ami-
noacetamido)-4-methylpentanamido)-4-tert-butoxycar-
bonylpropanamido)-3-methylbutanamido]-5-oxo-octa-
hydro-1H-pyrrolo[1,2-a]azepine-3-carboxylate (33).
Diphenylmethyl (3S,6S,10S)-6-[(2S)-2-((((2S)-2-(((2S)-2-(2-
carbobenzyloxyaminoacetamido)-4-methylpentanamido)-4-
tert-butoxycarbonylpropanamido)-3-methylbutanamido]-5-
oxo-octahydro-1H-pyrrolo[1,2-a]azepine-3-carboxylate 32
(56 mg, 0.056 mmol) was dissolved in a 1:1 mixture of
methanol and THF (6 ml). 10% Palladium onactivated carbon
(6 mg) was added and the reaction mixture was hydrogenated
at atmospheric pressure for 4 h, at room temperature. The
suspensionwasfilteredthroughfinefilterpaperandthesolvent
was removed in vacuo to yield (3S,6S,10S)-6-[((((2S)-2-
(((2S)-2-(((2S)-2-(2-aminoacetamido)-4-methylpentana-
mido)-4-tert-butoxycarbonylpropanamido)-3-methylbuta-
namido]-5-oxo-octahydro-1H-pyrrolo[1,2-a]azepine-3-car-
boxylate 33 as a white solid (38 mg, 92%); [a]3D0ZK77.62
(c 0.5, MeOH); m/z [Cve FAB (3-NBA)] 675 ([MCNa]C),
nmax (film)/cmK1 1723 (ester), 1710 (acid) 1639 (amide)
and 1545 (amide); dH (300 MHz, C2H3O2H) 0.83 (12H, t,
JZ6.8 Hz, b-MeCg00-Me), 1.33 (9H, s, C(CH3)3), 1.48–
2.07 (14H, m, H-2, H-1, H-9, H-7, H-8, H-bCH-b00C
H-g00), 2.53–2.90 (2H, m, H-b0), 3.37–3.62 (3H, m, H-10
and H-a000), 3.79 and 4.13 (2H, m, H-a and H-a00), 4.34 (2H,
m, H-6 and H-a0) and 4.63 (1H, m, H-3); dC (75.5 MHz,
C2H3O2H) 17.30, 18.59, 22.16 and 24.63 (b-MeCg00-Me),
20.81 (C-g00), 27.09 (C(CH3)3), 27.67 and 28.20 (C-1 and
C-8), 30.99 (C-b), 30.41 (C-b00), 30.75, 32.87 and 33.87
(C-2, C-9 and C-7), 36.60 (C-b0), 40.89 (C-a000), 48.27
(C-a), 52.23 (C-a00), 53.51 (C-a0), 59.33, 59.69 and 63.73
exchanges in H2O), 7.24–7.38 (11H, m, aromaticsCNH, 1
2
proton exchanges in H2O) and 8.40 (1H, d, JNH,6Z8.1 Hz,
NH, exchanges in H2O); dC (62.9 MHz, C2HCl3) 17.69,
2
19.22, 21.45 and 23.18 (b-MeCg00-Me), 24.72 (C-g00), 27.40
(C-1 and C-8), 27.94 (C(CH3)3), 30.89 (C-bCC-b0), 32.79,
34.10 and 36.92 (C-2, C-9 and C-7), 43.42 (C-b00), 53.23 and
53.38 (C-a and C-a0), 58.79, 59.10 and 60.51 (C-6, C-10 and
C-3), 77.89 (OCHAr2), 81.54 (OC(CH3)3), 126.95–128.48,
139.43 and 139.68 (aromatics) and 169.66–175.23
(6!C]O).
3.1.18. Diphenylmethyl (3S,6S,10S)-6-[(2S)-2-((((2S)-2-
(((2S)-2-(2-carbobenzyloxyaminoacetamido)-4-methyl-
pentanamido)-4-tert-butoxycarbonylpropanamido)-3-
methylbutanamido]-5-oxo-octahydro-1H-pyrrolo[1,2-a]
azepine-3-carboxylate (32). Diphenylmethyl (3S,6S,10S)-
6-[(2S)-2-(((2S)-2-((2S)-2-amino-4-methylpentanamido)-4-
tert-butoxycarbonylpropanamido)-3-methylbutanamido]-5-
oxo-octahydro-1H-pyrrolo[1,2-a]azepine-3-carboxylate 31
(109 mg, 0.143 mmol) and N-carbobenzyloxyglycine
(33 mg, 0.158 mmol) were dissolved in dry dimethylforma-
mide (5 ml). 2-(1H-Benzotriazole-1-yl)-1,1,3,3-tetramethy-
luronium tetrafluoroborate (TBTU) (51 mg, 0.158 mmol)
and diisopropylethylamine (0.028 ml, 0.158 mmol) were
added and the solution was stirred under nitrogen at room
temperature overnight. The solvent was removed in vacuo
and the residue was dissolved in dichloromethane and
washed with 5% aqueous citric acid, followed by 5%
aqueous sodium hydrogen carbonate and brine. The organic
layer was dried (MgSO4) and the solvent was removed in
vacuo to give an oil which was purified by column

D. E. Davies et al. / Tetrahedron 61 (2005) 301–312
311
(C-3, C-10 and C-6), 81.22 (OC(CH3)3) and 126.05–171.36
(6!C]O).
(2H, 2!m, H-b0), 3.62, 4.01 (2H, 2!m, H-a000), 4.35 (1H,
m, H-a), 4.45 (1H, m, H-6), 4.47 (1H, m, H-3), 4.48 (1H, m,
C-10) 4.53 (1H, m, H-a0), 4.78 (1H, m, H-a00), 7.24 (1H, d,
Ja,NHZ9.5 Hz, a-NH), 7.53 (10H0 , d, J6,NHZ5.6 Hz, 6-NH),
3.1.20. (3S,6S,10S)-3,6-Cyclo-[glycyl-(2S)-leucyl-(4-tert-
butyl (2S)-aspartyl)-(2S)-valylamino]-5-oxo-octahydro-
1H-pyrrolo[1,2-a]azepine (34). (3S,6S,10S)-6-[((((2S)-2-
(((2S)-2-(((2S)-2-(2-Aminoacetamido)-4-methylpentana-
mido)-4-tert-butoxycarbonylpropanamido)-3-methylbuta-
namido]-5-oxo-octahydro-1H-pyrrolo[1,2-a]azepine-3-car-
boxylate 33 (21 mg, 0.032 mmol) was dissolved in dry
dimethylformamide (40 ml). 2-(1H-Benzotriazole-1-yl)-
1,1,3,3-tetramethyluronium tetrafluoroborate (TBTU)
(62 mg, 0.193 mmol) and dimethylaminopyridine (28 mg,
0.225 mmol) were added and the solution was stirred under
nitrogen at room temperature for 28 h. An aliquot (1 ml)
was removed from the reaction mixture and the solvent was
removed in vacuo. FAB mass spectrometry showed no sign
of any starting material and intense ions were observed
for DMAP and the coupling reagents. [MCNa]C for
(3S,6S,10S)-3,6-cyclo-[glycyl-(2S)-leucyl-(4-tert-butyl
(2S)-aspartyl)-(2S)-valylamino]-5-oxo-octahydro-1H-pyr-
rolo[1,2-a]azepine 34 was also present at m/z 657. The
solvent was removed in vacuo (!40 8C) and the residue
was dissolved in dichloromethane and washed with 5%
aqueous sodium hydrogen carbonate (2!15 ml) and brine
(2!15 ml). The organic layer was dried (MgSO4) and the
solvent was removed in vacuo. Attempts at purification by
column chromatography and preparative HPLC were
unsuccessful and so (3S,6S,10S)-3,6-cyclo-[glycyl-(2S)-
leucyl-(4-tert-butyl (2S)-aspartyl)-(2S)-valylamino]-5-oxo-
octahydro-1H-pyrrolo[1,2-a]azepine 34 was used directly in
the next step without purification or analysis.
00
0
8.40 (1H, d, JNH,a Z9.0 Hz, a -NH), 8.58 (1H, d, JNH,a
Z
5.5 Hz, a0-NH) and 8.68 (1H, q, Ja ,NHZ5.4, 7.0 Hz,
a000-NH). ROESY data (200 ms mixing time—loosely
characterised as strong (s), medium (m) or weak (w)).
Intra-residue a/NH—Glya (m/w), Glya0 (s); Asp (m/w);
Val (s); Leu (under H2O peak. Intra-residue NH/bH-Asp
(b&b0) (m); Leu-b (s); Val (m); also intra-residue a/b cross-
peak for Val and sequential NH/NH (i, iC1) cross peaks for
Asp/Val. The 1H–1H COSY spectrum is included as
supplementary material.
000
Acknowledgements
We thank GlaxoSmithKline and the EPSRC for a CASE
studentship (to R. D. H.), Dr. A. G. Avent and Mr. C. M.
Dadswell (Sussex) and Dr. R. D. Farrant, Dr. P. N.
Sanderson and Mr. S. Richards (GlaxoSmithKline) for
NMR spectra; Mr. Eric Horteuse (GlaxoSmithKline) for
preparative HPLC; and Mr. I. Davidson (GlaxoSmithKline),
Dr. A. Abdul-Sada (Sussex) and the EPSRC National Mass
Spectrometry Service, Swansea for mass spectra.
References and notes
3.1.21. (3S,6S,10S)-3,6-Cyclo-[glycyl-(2S)-leucyl-(2S)-
aspartyl-(2S)-valylamino]-5-oxo-octahydro-1H-pyrrolo
[1,2-a]azepine (35). A solution of triethylsilane (0.15 ml)
and trifluoroacetic acid (3 ml) was added to the crude
(3S,6S,10S)-3,6-cyclo-[glycyl-(2S)-leucyl-(4-tert-butyl
(2S)-aspartyl)-(2S)-valylamino]-5-oxo-octahydro-1H-pyr-
rolo[1,2-a]azepine 34 (13 mg, 0.0198 mmol) at 0 8C. The
mixture was stirred under nitrogen at this temperature for
20 min, warmed to room temperature and stirred for a
further 2 h. The solvents were removed in vacuo to yield a
brown oil which was washed with diethyl ether (4!10 ml).
Removal of the solvents in vacuo followed by drying in
vacuo yielded a pale brown solid. The crude product was
purified using preparative HPLC using a Supelcosil LC-
ABZCPlus column (10 cm, 21.2 mm diameter). The
solvent gradient started at 100% H2O (C0.1% TFA) and
went to 95% CH3CN (C0.05% TFA) over 17.5 min, then
continued at 95% CH3CN (C0.05% TFA) for 10 min.
The retention time of the required compound was 15.5–
16.0 min. The sample was obtained as a very fine glassy
solid from freeze drying the combined fractions of multiple
HPLC runs to give (3S,6S,10S)-3,6-cyclo-[glycyl-(2S)-
leucyl-(2S)-aspartyl-(2S)-valylamino]-5-oxo-octahydro-
1H-pyrrolo[1,2-a]azepine 35; m/z [Cve electrospray] 579
([MCH]C); m/z [Kve electrospray] 577 ([MKH]C); dH
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