Base-induced dimerization of urethane-protected amino acid N-carboxanhydrides

Article abstract of DOI:10.1021/jo951260f

tert-Butyloxycarbonyl-protected N-carboxanhydrides of amino acids dimerize in the presence of base in aprotic media to form 3,5-dialkyl-2,4-dioxo-1-pyrrolidine analogs. Depending on the nature of the base, different ratios of isomers were obtained. The reaction with lithium bis(trimethylsilyl)-amide lead to one isomer only. After deprotection of the tert-butyloxycarbonyl groups and coupling of (benzyloxycarbonyl)valine, a homogeneous product was obtained. Reduction with sodium borohydride again gave a homogeneous product. Nuclear Overhauser enhancement spectroscopy and X-ray crystallography identified the stereochemistry in positions 3 and 5 of the pyrolidine as Z. When 1, 8-diazabicyclo[5.4.0]undec-7-ene was used as the base, the condensation led to a 1:3 ratio of isomers. The major isomer was different from the one obtained with lithium bis-(trimethylsilyl)amide. The (benzyloxycarbonyl)valine derivative from this compound was obtained as a 1:1 mixture of isomers, leading to the conclusion that this condensation product was an enantiomeric mixture of the E isomers. The pure Z isomer from the lithium bis(trimethylsilyl)-amide reaction was converted to a mixture of Z and E isomers in a ratio of 1:3 when treated with 1,8-diazabizycol[5.4.0]undec-7-ene. The (benzyloxycarbonyl)valine derivative of the E isomer from this conversion was again a 1:1 mixture; therefore, the Z isomer obtained with lithium bis-(trimethylsilyl)amide was believed to have been an enantiomeric mixture. Several other examples indicated that this reaction occurred also with other tert-butyloxycarbonyl-protected N-carboxy-anhydrides.

Full text of DOI:10.1021/jo951260f

228
J . Org. Chem. 1996, 61, 228-231
Ba se-In d u ced Dim er iza tion of Ur eth a n e-P r otected Am in o Acid
N-Ca r boxa n h yd r id es
J ohann J . Leban*,† and Kimberly L. Colson‡
Divisions of Organic Chemistry and Bioanalytical Sciences, Wellcome Research Laboratories,
3030 Cornwallis Road, Research Triangle Park, North Carolina 27709
Received J uly 13, 1995X
tert-Butyloxycarbonyl-protected N-carboxanhydrides of amino acids dimerize in the presence of
base in aprotic media to form 3,5-dialkyl-2,4-dioxo-1-pyrrolidine analogs. Depending on the nature
of the base, different ratios of isomers were obtained. The reaction with lithium bis(trimethylsilyl)-
amide lead to one isomer only. After deprotection of the tert-butyloxycarbonyl groups and coupling
of (benzyloxycarbonyl)valine, a homogeneous product was obtained. Reduction with sodium
borohydride again gave a homogeneous product. Nuclear Overhauser enhancement spectroscopy
and X-ray crystallography identified the stereochemistry in positions 3 and 5 of the pyrolidine as
Z. When 1, 8-diazabicyclo[5.4.0]undec-7-ene was used as the base, the condensation led to a 1:3
ratio of isomers. The major isomer was different from the one obtained with lithium bis-
(trimethylsilyl)amide. The (benzyloxycarbonyl)valine derivative from this compound was obtained
as a 1:1 mixture of isomers, leading to the conclusion that this condensation product was an
enantiomeric mixture of the E isomers. The pure Z isomer from the lithium bis(trimethylsilyl)-
amide reaction was converted to a mixture of Z and E isomers in a ratio of 1:3 when treated with
1,8-diazabizycol[5.4.0]undec-7-ene. The (benzyloxycarbonyl)valine derivative of the E isomer from
this conversion was again a 1:1 mixture; therefore, the Z isomer obtained with lithium bis-
(trimethylsilyl)amide was believed to have been an enantiomeric mixture. Several other examples
indicated that this reaction occurred also with other tert-butyloxycarbonyl-protected N-carboxy-
anhydrides.
In tr od u ction
for some time. They can be obtained commercially and
are an attractive alternative to other activated amino acid
species. Many of these reactions require the presence of
base. We found that base can cause a dimerization of
two molecules of the amino acid to yield pyrrolidine-2,4-
diones. A few selected examples of this reaction and the
characterization of the products are presented.
N-Carboxy-R-amino acid anhydrides (NCAs), or Leuchs’
anhydrides,1 represent a unique species of amino acid
derivatives where both the carboxylate activation and the
amino group protection are achieved simultaneously.
Their sensitivity to moisture and tendency to polymeri-
zation, however, limited broader application. For these
reasons, additional protection of the R-amino group was
attempted early on.
Resu lts a n d Discu ssion
The reaction of Boc-Phe-NCA with lithium bis(trim-
ethylsilyl)amide (LHS) in THF at -78 °C yielded, after
purification, a single product with the observed mass of
a di-Boc-oxopiperazine. The 1H-NMR spectrum, however,
was not consistent with this structure. The presence of
an NH signal, one R-proton, two distinctly different
benzylic signals, and two distinct signals for the Boc
groups suggested structure 2 as shown in Scheme 1.
HPLC analysis of the crude reaction mixture indicated
one main product with the a second minor peak (9%).
When the products were separated by HPLC and ana-
lyzed by mass spectroscopy, both had the same mass. The
major isomer from the LHS reaction was obtained pure
after workup and purification in 35% yield. Most of the
nonreacted product is hydrolyzed UNCA. We avoided the
use of more base and longer reaction times to encrease
the yield, because such conditions tended to yield an
elevated amount of the second isomer. In contrast to the
reaction with LHS, the reaction with 1 equiv of 1,8-
diazabicyclo[5.4.0]undec-7-ene (DBU) yielded a 1:3 ratio
of isomers. The major isomer 3 crystallized in pure form
Tosyl-protected Gly-N-carboxanhydride was reported
first by Zaoral et al.2 Nitrophenylsulfonyl-protected Phe-
N-carboxanhydride was used by Kricheldorf and Hal-
strøm for peptide synthesis.3,4 UNCAs (Boc, Cbz, and
Fmoc) and their use in peptide synthesis were described
only recently.5 These derivatives represent valuable new
tools for peptide chemistry. The application of Boc-Xaa-
NCAs for solid-phase synthesis in the presence of tri-
ethylamine (TEA) was reported.6 Fmoc amino acid
N-carboxanhydrides were reported to esterify to p-
alkoxybenzyl alcohol resins in the presence of N-meth-
ylmorpholine.7
UNCAs have been utilized to replace isopropyl chlo-
roformate-activated N-protected amino acids for various
chemical transformations, such as reductions and nu-
cleophilic reactions.8 UNCAs are stable and can be stored
* To whom correspondence should be addressed.
† Present address: Hauptplatz 24, 2421-Kittsee, Austria.
‡ Present address: P.O. Box 4392, Chapel Hill, NC 27515.
X Abstract published in Advance ACS Abstracts, J anuary 1, 1996.
(1) Leuchs, H.; Manasse, W. Ber. Dtsch. Chem. Ges. 1906, 39, 857.
(2) Zaoral, M.; Rudinger, J . Collect. Czech. Chem. Commun. 1961,
26, 2316-2332.
(3) Kricheldorf, H. R. Angew. Chem. 1973, 85, 86-87.
(4) Halstrøm, J .; Brunfeldt, K.; Kovacs, K. Acta Chem. Scand., Ser.
B 1979, B33, 685-689.
(5) Fuller, W. D.; Cohen, M. P.; Shabankareh, M.; Blair, R. K.;
Goodmann, M.; Naider, F. R. J . Am. Chem. Soc. 1990, 112, 7414-
7416.
(7) Fuller, W. D.; Krotzer, N. J .; Swain, P. A.; Anderson, B. L.;
Comer, D.; Goodman, M. Peptides 1992: Proceedings of the 22nd
European Peptide Symposium; Escom: Leiden, The Netherlands, 1993;
pp 231-232.
(8) Fehrentz, J .-A.; Bourdel, E.; Califano, J .-C.; Chaloin, O.; Devin,
C.; Garrouste, P.; Lima-Leite, A.-C.; Llinares, M.; Rieunier, F.; Viza-
vonna, J .; Winternitz, F.; Loffet, A.; Martinez, J . Tetrahedron Lett.
1994, 35, 1557-1560.
(6) Xue, C.-B.; Naider, F. J . Org. Chem. 1993, 58, 350-355.
0022-3263/96/1961-0228$12.00/0 © 1996 American Chemical Society

Dimerization of Amino Acid N-Carboxanhydrides
J . Org. Chem., Vol. 61, No. 1, 1996 229
Sch em e 1
F igu r e 1. Summary of key NOEs observed in 5.
Sch em e 2
from methanol and was identical to the minor product
of the LHS reaction by HPLC and TLC; it also had the
1
same molecular weight and an H-NMR very similar to
2. The product had no optical rotation, and therefore,
we believed that it was a mixture of enantiomers dia-
stereomeric to 2. We made a (carbobenzyloxy)valine
derivative 6. HPLC analysis indicated the presence of
two compounds in a 1:1 ratio, which confirmed that 3
was an enantiomeric mixture. Although we have no
structural data yet to prove the E stereochemistry for
compound 3, one can assume this stereochemistry safely.
We used compound 2 and applied one-dimensional
nuclear Overhauer enhancment spectroscopy (NOE) for
the determination of the stereochemstry. Compound 2
did not yield the critical nuclear Overhauser enhance-
ment (NOE) signal between the benzylic methylene
groups that would confirm the Z stereochemistry. To
identify the stereochemistry by NOE experiments, a
(benzyloxycarbonyl)valine analog (4) was synthesized.
This product was obtained in 82% yield as a single
component on HPLC and TLC.
Attempts to determine the relative stereochemistry of
4 using NOE spectroscopy failed. No NOEs from the
protons in positions 5 or 6 were observed to the protons
in position 11 (1H NMR data not shown).
To obtain another analog for more structural informa-
tion we reduced the ketone at position 4 with sodium
borohydride (NaBH4). This reduction yielded a product
that was homogeneous by TLC and HPLC, and 5 was
obtained in 77% yield.
Ta ble 1. 1H NMR Ch em ica l Sh ift Assign m en ts of 5 a n d
th e Obser ved Nu clea r Over h a u ser En h a n cem en ts
assignment
δ
mult int
obsd NOEs
7.11 5.49 3.81
1
7.87
7.63
s
s
1
1
16
arom
4-OH
21
4
7.21 3.85 1.83 0.77 0.75
7.36-7.10
m
s
16
1
5.49
5.03
4.37
7.87 4.37
dd
d
2
1
7.21 7.11 5.49 3.81
17
5
11
6
3.85
3.81
3.06
2.80
2.24
1.84
0.77
0.75
d
m
s
dd
dd
s
1
1
2
1
1
1
3
3
7.21 5.49 4.37 2.24
7.11 3.81 2.24
6′
7.87 7.11 3.81 2.82 3.06a
18
19
19′
d
d
a
The intensity of this NOE was very small.
NOE data on 5 provided the information necessary to
assign the relative stereochemistry at positions 3 and 5.
A NOE observed between the protons on H11 (3.06 ppm)
and H6 (2.24 ppm) indicated the close spatial proximity
of these two protons. The observation of this NOE could
only be accommodated by the relative stereochemistry
as shown in Figure 1, which also displayed the key NOEs.
The H spectral data on 5 are summarized in Table 1.
The results obtained identified the product as the 3,5 Z
isomer.
We also obtained an X-ray structure of 5. The X-ray
structure confirmed the Z configuration. The configu-
ration of the chiral carbons (Figure 1) corresponds to 3R,
4S, and 5R.
If 2 was an optically pure product, we assumed that,
if it could be converted to the E isomer with DBU, it
should again yield a single (benzyloxycarbonyl)valine
derivative. We treated 2 with DBU, and again a 1:3
mixture of isomers was obtained. The major component
crystallized in pure form from methanol. The (benzyl-
oxycarbonyl)valine derivative was made, and we obtained
a product that was a 1:1 mixture of isomers and totally
identical to 6; therefore, 3 was an enantiomeric mixture.
Although compounds 4 and 5 were diastereomeric mix-
tures with the Z configuration they did not separate by
HPLC and TLC. The conditions used to get crystals for
the X-ray determination then produced only one diaste-
reomer. The fact that carbon 3 (Scheme 2) was R further
substantiated that notion.
1
We wanted to better understand the influence of
different types of bases in the condensation reaction and
their influence on the stereochemistry of the product. A
series of small scale experiments (10 µmol) were run
using Boc-Phe-NCA, with various bases and solvents. The
base was added at -78 °C, and the reaction was allowed
to warm to room temperature. The reaction mixture was

230 J . Org. Chem., Vol. 61, No. 1, 1996
Leban and Colson
Ta ble 2. Dia ster eom er ic Ra tios for 2 a n d 3 Obta in ed
fr om Rea ction of Boc-P h e-NCA w ith Va r iou s Ba ses
ated the first at the R-carbanion, and then the acylated
UNCA species, in turn, acylated the amino group of the
incoming molecule (see Scheme 1). Since the anion was
planar an enantiomeric mixture was formed. The Z
isomeric enantiomers were formed in a kinetic controlled
reaction. Contrary to the LHS reaction, the reaction with
1 equiv of DBU led to a racemic mixture of E enantiomers
in a thermodynamicly controlled reaction due to the slow
deprotonation. This proposed mechanism is corroborated
by the fact that both the Z and E condensation products
equilibrate to the same ratio of Z and E isomers when
incubated with DBU.
It would be interesting to learn if such a intermolecular
condensation could be accomplished with two different
amino acids. One would expect a clean product only if
the acidity of the R-proton of the respective UNCAs were
sufficiently different or if steric effects favor the conden-
sation of different species of UNCA. Preliminary data
indicate that such condensations indeed could be achieved
under special circumstances. Such work will be the scope
of future reports.
solvent
base (equiv)
ratio of cis/transa
THF
THF
DCM
DCM
DCM
DCM
THF
DCM
DCM
DCM
THF
THF
THF
THF
LHS (0.5)
LHS (1)
TEA (0.5)
TEA (1)
TEA (5)
TEA (10)
TEA (1)
DMAP (1)
NMM (1)
DBU (1)
DBU (0.5)
DBU (1)
DBU (1)b
DBU (1)c
3.6:1
3.7:1
1:1.1
1:1
1:1.4
1:1.4
no reaction
no reaction
no reaction
1:2.6
reaction
1:3
1:2.7
1:3
a
b
Diastereomeric content measured by HPLC. Compound 2
was incubated with DBU. c Compound 3 was incubated with DBU.
directly injected into the HPLC, and the ratio of isomeric
products obtained was estimated from the peak areas of
2 and 3. These HPLC conditions distinguished between
the cis/trans isomers but did not indicate the enantio-
meric purity of the products. The results are shown in
Table 2. Little product was obtained with 0.5 equiv of
DBU, while the reaction proceeded more with 1 equiv of
DBU. It was important to notice that the reaction did
not occur with TEA in THF but did proceed in DCM,
indicating that with the proper choice of solvent the
condensation can be avoided. Furthermore, no reaction
was observed with NMM or 4-(dimethylamino)pyridine
(DMAP). This is important because these reagents are
common when UNCAs are used as activated amino acid
species. The last three experiments in Table 2 again
show on an analytical scale the formation of a 1:3 mixture
of 2 and 3 with DBU. The interconversion of pure 2 into
a 1:3 mixture of 2 and 3 and finally the conversion of
pure 3 into a 1:3 mixture of 2 and 3 occurs in the presence
of DBU. It is worth noticing that weaker bases DBU and
TEA yielded more of the E isomers while LHS and LDA
(data for LDA not shown) resulted in more Z isomer. We
believe that the slower reaction with the weaker bases
is thermodynamicly controlled. It is also interesting to
note that TEA in THF did not give much of the product
while it did in DCM. More data will be needed to come
to a precise conclusion on the nature of the relationship
between solvent, base, and condensation product.
We were interested in whether this reaction is ap-
plicable to other Boc-amino acid NCA’s and followed the
dimerization with a few more examples. The condensa-
tion of UNCAs using LHS was not restricted to the Phe
analog and worked well with Boc-Ile-NCA and Boc-Thr-
(Bzl)-NCA, yielding single isomers in both cases. When
the R-side chain was bulky, one isomer was preferred,
while a 1:1 mixture of isomers was obtained with Boc-
Ala-NCA. No further experiments to determine the
stereochemistry of these analogs were performed. It can,
however, be expected that an enantiomeric mixture of Z
isomers is obtained with Boc-Ile-NCA and Boc-Thr(Bzl)-
NCA and the enantiomers of both E and Z in the case of
Boc-Ala-NCA. Preliminary data indicated (data not
shown) that the condensation was applicable to Cbz-Phe-
NCA, but the reaction was less clean than for the Boc-
derivative.
Con clu sion
Pyrrolidine-2,4-diones are novel amino acid derived
heterocycles obtained by base-induced condensation of
UNCAs and may be useful as a chiral auxillary or
intermediate in medicinal chemistry. The reaction can
be controlled to obtain the enantiomeric mixtures of Z
and E isomers, depending on what base is used. The
correct choice of base and solvent can eliminate this
reaction when UNCAs are used during peptide synthesis.
Exp er im en ta l Section
Urethane-protected amino acids were obtained from SNPE,
Inc., of Princeton, NJ . Other chemicals were supplied by the
Aldrich Chemical Co., Milwaukee, WI.
Str u ctu r e Deter m in a tion of 5 by P r oton NMR. 1H
NMR data on 5 were acquired on a Varian Unity-400 equipped
with a Nalorac 5 mm inverse detection triple resonance
gradient probe at 26 °C. 1H NOE spectra were acquired for
the protons resonating at 7.87, 7.63, 5.49, 4.37, 3.06, 2.80, and
2.24 ppm using a sweep width of 5000 Hz, an acquisition time
of 0.499 s, and a relaxation delay of 6 s. 1H NMR data on 4
were acquired on a Varian VXR-300 equipped with a Nalorac
5 mm quad nucleus probe at 26 °C. 1H NOE spectra of 4 were
acquired for the protons resonating at 9.04, 8.44, 5.01, 3.11,
2.78, 1.97, and 1.03 ppm using a spectral width of 4400 Hz,
an acquisition time of 0.502 s, and a relaxation delay of 8 s.
Selective irradiation to accomplish the dipolar exchange was
employed throughout the relaxation delay. All NOE experi-
ments were processed using the difference method. The 1H
chemical shift assignments were made with coupling data
obtained from a COSY experiment. The COSY experiments
were acquired using a sweep width of 5000 Hz, a relaxation
delay of 1 s, and an acquisition time of 0.205 s. The COSY
experiments were acquired using 512 points in F1 and 2048
points in F2 and processed, using a sinebell weighting, to give
a 2K by 2K matrix.
Str u ctu r e d eter m in a tion of 5 by X-r a y cr ysta llogr a -
1
p h y: C31H35N3O5; /2H2O, Mr ) 538.64; monoclinic, P21; a )
1 7.100(4) Å, b ) 5.180(2) Å, c ) 23.292(2) Å, V ) 5972(1) Å3,
Z ) 8, Dc ) 1.198 g/cm3; graphite-monochromated Cu KR
radiation λ )1.541 78 Å; reflections: total 9580, unique 9233
(Rint ) 0.149), F(000) ) 2296.00; structure solution by direct
methods9 and structure refinement using full-matrix least-
squares methods.10
The reaction mechanism of the condensation can be
explained by the increased acidity of the R-proton in the
UNCAs compared to the protected amino acid. After
deprotonation with LHS, another UNCA molecule acyl-
HP LC An a lysis. Analysis of diastereomer content was
determined by HPLC using a Waters Novapak C18 radialpak
column. The sample was eluted with 0.1% TFA/H2O (A) and
0.1% TFA/acetonitrile (B) (20-90% B over 44 min).

Dimerization of Amino Acid N-Carboxanhydrides
J . Org. Chem., Vol. 61, No. 1, 1996 231
Gen er a l P r oced u r e for Ba se-In d u ced Dim er iza tion of
UNCAs. A solution of 10 mmol of urethane-protected amino
acid-NCA in 25 mL of dry THF was cooled to -78 °C under
N2. A solution of 1 M LHS in THF (5 mL) was added slowly
via syringe. The reaction mixture was allowed to warm to
room temperature (ca. 2 h). It was then quenched with 20
mL of 1 N HCl and extracted with EtOAc. The organic phase
was dried with MgSO4 and concentrated in vacuo to yield a
thick oil. The product was purified by flash column chroma-
tography (hexane/EtOAc 2:1, unless otherwise described).
Optical rotations were measured in EtOAc (1 g/100 mL).
ter t-Bu t yl 3,5-(Z)-Dib en zyl-3-[(ter t-b u t oxyca r bon yl)-
a m in o]-2,4-d ioxo-1-p yr r olid in eca r boxyla te (2) (Der ived
fr om Boc-P h e-NCA). The product was purified by hexane/
EtOAc (3:1) to obtain 1.73 g (35%). 1H NMR (d6-DMSO) δ:
8.4 (s, 1H), 7.4-6.9 (m, 10H), 3.2 (m, 1H), 3.1-2.8 (m, 4H),
1.5 (s, 9H), 1.4 (s, 9H). Mass spectra: m/ e 517.2 (M + Na)+.
Anal. Calcd for C28H34N2O6: C, 68.00; H, 6.93; N, 5.66.
Found: C, 68.23; H, 7.12; N, 5.60. HPLC: tR ) 34.71 min.
ter t-Bu t yl 3,5-(E)-Dib en zyl-3-[(ter t-b u t oxyca r b on yl)-
a m in o]-2,4-d ioxo-1-p yr r olid in eca r boxyla te (3) (Der ived
fr om Boc-P h e-NCA). DBU was substituted for LHS. Crude
product was recrystallized from MeOH to yield 0.63 g (26%).
This is the second peak by HPLC. 1H NMR (d6-DMSO) δ: 8.5
(s, 1H), 7.4-7.0 (m, 10H), 3.3 (m, 1H), 3.2-2.9 (m, 4H), 1.45
(s, 9H), 1.4 (s, 9H). Mass spectra: m/ e 295.0 (MH - 2Boc)+.
Anal. Calcd for C28H34N2O6: C, 68.00; H, 6.93; N, 5.66.
Found: C, 67.98; H, 6.91; N, 5.74. HPLC: tR ) 35.45 min.
N-r-(Ben zyloxyca r bon yl)-L-va lin e N-(3,5-(Z)-Diben zyl-
2,4-d ioxo-3-p yr r olid in yl)a m id e (4). A mixture containing
2 (1.0 g, 2 mmol) in a solution of 10 mL of 1 M HCl/EtOAc
was stirred for 1 h, concentrated, and dried. This residue, Cbz-
Val-OH (0.51 g, 2 mmol), EDC (0.39 g, 2 mmol), HOBt (0.27
g, 2 mmol) and NMM (0.22 mL, 2 mmol) in DMF were stirred
for 16 h at 25 °C. The reaction was quenched with 5% citric
acid and extracted with EtOAc. The organic phase was
washed with 5% NaHCO3, 5% citric acid, and water, dried with
MgSO4, and concentrated. The residue was purified over silica
gel (hexane/EtOAc, 1.5:1) to yield 0.43 g of a white solid (82%).
1H NMR (d6-DMSO) δ: 9.0 (s, 1H), 8.4 (s, 1H), 7.4-7.1 (m,
13H), 6.8 (m, 2H), 5.0 (m, 2H), 4.0-3.9 (m, 2H), 3.2-2.7 (m,
4H), 2.0-1.8 (m, 2H), 1.0 (m, 1H), 0.8 (m, 6H). Mass spectra:
3.4 (m, 2H), 4.0-4.2 (m, 2H), 5.0-5.2 (m, 2H), 7.1-7.5 (m,
l5H), 7.95 (s, lH), 8.4 (m, lH), 9.0 (s, lH). Mass spectra: m/ e
550.2 (M + Na)+. Anal. Calcd from C33H31N30s: C, 70.57; H,
6.30; N, 7.96. Found: C, 68.84; H, 6.23; N, 8.32. HPLC: two
isomers (50:50) were observed [tR ) 23.12 min (A) and 24.80
min (B)].
Con ver sion of 2 to 3 a n d Syn th esis of 6. A soluton of
1.6 g of 2 (3.3 mmol) in dry THF was cooled to -78 °C under
N2, DBU (500 mL, 3.3 mmol) was added, and the reaction
mixture was stirred for 2 h. The solvent was removed in vacuo
and the residue partitioned between EtOAc and 5% citric acid.
The organic phase was washed with 5% NaHCO3, dried with
MgSO4, and concentrated to yield a white solid. Recrystalli-
zation from MeOH yielded 0.6 g (36%) of 3. 1HNMR (d6-
DMSO) δ: 1.35 (s, 9H), 1.45 (s, 9H), 2.9-3.2 (m, 4H), 3.3 (m,
1H), 6.9-7.3 (m, 10H), 8.45 (s, 1H). Mass spectra: m/ e 517.4
(M + Na)+. Anal. Calcd from C28H34N2O6: C, 68.00; H, 6.93;
N, 5.66. Found: C, 67.90; H, 6.88; N, 5.74. HPLC (3, tR
)
35.49 min).
Compound 3 from this reaction was dissolved in 10 mL of 1
M HCl in EtOAC and stirred for 1 h. The solution was
concentrated in vacuo and yielded 0.24 g of white solid. Mass
spectrum: m/ e 295.3 (MH)+. A DMF solution containing 0.407
mg, 1 mmol) of this product (0.25 g, 1 mmol), CBZ-Val-OH,
(0.25 g, 1 mmol) EDC, HOBt (0.176 g, 1.3 mmol), and NMM
(0.22 mL, 2 µmol) was stirred for 18 h at rt. The DMF was
removed in vacuo, and the residue was partitioned between
EtOAc and 5% NaHCO3. The organic phase was washed with
5% citric acid, dried with MgSO4, and concentrated to yield
0.48 g of white solid. Purification by silica gel chromatography
(5% MeOH/CH2Cl2) yielded 0.21 of a material that was
identical to 6. Mass spectra: m/ e 528.5 (MH)+. HPLC:
indicated the presence of two isomers (50:50). (tR ) 23.17 and
24.81 min). Both isomers were isolated by HPLC on a small
scale and analyzed by mass spectroscopy. Both peaks gave a
m/ e of 528.3 (MH)+.
ter t-Bu tyl 3[(ter t-bu toxycar bon yl)am in o]-3,5-dim eth yl-
2,4-d ioxo-1-p yr r olid in eca r b oxyla t e (7) (Der ived fr om
Boc-Ala -NCA). The yield was 1.14 g (23%). 1H NMR (d6-
DMSO) δ: 8.25 (s, 1H), 8.15 (s, 1H), 4.65 (m, 1H), 4.35 (m,
1H), 1.5 (s, 18H), 1.45 (m, 6H), 1.35 (m, 18H), 1.3-1.25 (m,
6H). Mass spectra: m/ e 143 (MH - 2Boc)+. Anal. Calcd for
C16H26N2O6: C, 56.11; H, 7.66; N, 8.11. Found: C, 55.90; H,
7.67; N, 8.06.
m/ e 528.4 (MH)+. Optical rotation: [R]20 ) -131.0°.
589
N-r-(Ben zyloxyca r bon yl)-L-va lin e N-(3,5-(Z)-Diben zyl-
4-h yd r oxy-2-oxo-3-p yr r olid in yl)a m id e (5). Compound 4
(0.9 g, 1.7 mmol) was dissolved in 50 mL of MeOH. An excess
of NaBH4 was added and the reaction stirred 20 min. The
reaction was concentrated, water was added, and a white solid
was filtered. Recrystallization from 30 mL of MeOH yielded
0.3 g of white solid (77%). 1H NMR (see Figure 1). Mass
spectra: m/ e 530.4 (MH)+. Anal. Calcd for C31H35N3O5(0.65
mol H2O): C, 68.80; H, 6.71; N, 7.77. Found: C, 68.84; H,
ter t-Bu tyl 3,5-Bis((R)-1-Meth ylp r op yl)-3-[(ter t-bu toxy-
ca r bon yl)a m in o]-2,4-d ioxo-1-p yr r olid in eca r boxyla te (8)
(Der ived fr om Boc-Ile-NCA). The yield was 1.06 g (50%).
The product was purified by hexane/EtOAc (3:1). 1H NMR (d6-
DMSO) δ 7.9 (s, 1H), 4.2 (m, 1H), 2.0 (m, 1H), 1.8 (m, 1H), 1.5
(s, 9H), 1.3 (s, 9H), 1.1-0.7 (m, 16H). Mass spectra: m/ e 449
(M + Na)+. Anal. Calcd for C22H38N2O6: C, 61.93; H, 8.98;
6.68; N, 7.82. [R]20 ) -113.9°.
589
N-r-(Ben zyloxyca r bon yl)-L-va lin e N-(3,5-(E)-Diben zyl-
2,4-d ioxo-3-p yr r olid in yl)a m id e (6). Compound 3 (2 g, 4.04
mmol) was dissolved in 10 mL of 1 M HCl in EtOAc and stirred
for 1 h. The solvent was removed in vacuo, and the remaining
solid was crystallized with ether yielding 1.5 g (91%) of white
solid that was used without further purification. 1H NMR (d6-
DMSO) δ: 2.7-2.8 (m, lH), 3.1-3.4 (m, 3H), 3.7-3.75 (m, lH),
7.2-7.4 (m, 10H), 9.0-9.2 (m, 3H). Mass spectra: m/ e 295.0
(MH)+.
N, 6.57. Found: C, 61.86; H, 8.93; N, 6.52. [R]20 ) -6.35°.
589
ter t-Bu tyl 3,5-Bis[(R)-1-(ben zyloxy)eth yl]-3-[(ter t-bu -
t oxyca r b on yl)a m in o]-2,4-d ioxo-1-p yr r olid in eca r b oxyl-
a te (9) (Der ived fr om Boc-Th r (Bzl)-NCA). The yield was
1.4 g (24%). 1H NMR (d6-DMSO) δ: 8.05 (s, 1H), 7.4-7.05
(m, 10H), 4.6-4.5 (m, 2H), 4.3 (m, 2H), 4.1 (m, 1H), 4.0-3.8
(m, 2H), 1.35 (s, 18H), 1.3-1.2 (m, 6H). Mass spectra: m/ e
605 (M + Na)+. Anal. Calcd for C32H42N2O8: C, 65.95; H, 7.27;
A DMF mixture consisting of the deprotected 3 (1.23 g, 3
mmol), CBZ-Val-OH (753 mg, 3 mmol), HOBt (406 mg, 3
mmol), NMM (300 µL, 3 mmol), and EDC (575 mg, 3 mmol)
was stirred at rt for 12 h. Volatiles were removed, and the
residue was dissolved in EtOAc. This was then washed with
5% citric acid and 5% NaHCO3, dried (MgSO4), and concen-
trated. This material was purified over silica gel (5% MeOH/
CH2C12) to obtain 1.11 g (70%) of 6. lH NMR (d6-DMSO) δ:
0.85-0.95 (m, 6H), 1.9-2.1 (m, lH), 2.85-3.0 (m, 2H), 3.1-
N, 4.81. Found: C, 65.22; H, 7.29; N, 4.76. [R]20 ) -20.6°.
589
Ack n ow led gm en t. We thank Tony Landavazo for
helping us to prepare this manuscript, Brad Cozart and
Brian Herring for technical assistance, and Andrew
Spaltenstein for helpful discussions. We thank the
scientists from MolecularStructures for solving the
X-ray structure of 5.
(9) Miller, R.; Gallo, S. M.; Khalak, H. G.; Weeks, C. M. J . Appl.
Crystallogr. 1994, 27, 613-621.
(10) The author has deposited atomic coordinates for 5 with the
Cambridge Crystallographic Data Centre. The coordinates can be
obtained, on request, from the Director, Cambridge Crystallographic
Data Centre, 12 Union Road, Cambridge, CB2 1EZ, UK.
J O951260F