α,α-dichloroisoxazolidinones for the synthesis and chemoselective peptide ligation of α-peptide α-ketoacids

Article abstract of DOI:10.3987/COM-10-S(E)106

In seeking to develop an iterative approach to the preparation of α-oligopeptides by the chemoselective amide-forming coupling of α-ketoacids and hydroxylamines, we have designed and synthesized novel enantiopure monomers. Key to our approach is the use of α, α-dichloroacids as masked α-ketoacids. The preparation of these monomers, their coupling with α-ketoacids, and the conversion of the α, α-dichloroacids to α-ketoacids is described. These studies provide a first step to a conceptually unique approach to peptide synthesis that does not require activating reagents or produce chemical byproducts. The Japan Institute of Heterocyclic Chemistry.

Full text of DOI:10.3987/COM-10-S(E)106

HETEROCYCLES, Vol. 82, No. 2, 2011
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HETEROCYCLES, Vol. 82, No. 2, 2011, pp. 1515 - 1525. © The Japan Institute of Heterocyclic Chemistry
Received, 14th August, 2010, Accepted, 21st October, 2010, Published online, 2nd November, 2010
DOI: 10.3987/COM-10-S(E)106
!,!-DICHLOROISOXAZOLIDINONES FOR THE SYNTHESIS AND
CHEMOSELECTIVE PEPTIDE LIGATION OF !-PEPTIDE !-
KETOACIDS
Tetsuo Narumi1 and Jeffrey W. Bode2*
Roy and Diana Vagelos Laboratories, Department of Chemistry, University of
Pennsylvania, Philadelphia, PA 19104, USA. bode@org.chem.ethz.ch
Abstract – In seeking to develop an iterative approach to the preparation of !-
oligopeptides by the chemoselective amide-forming coupling of !-ketoacids and
hydroxylamines, we have designed and synthesized novel enantiopure monomers.
Key to our approach is the use of !,!-dichloroacids as masked !-ketoacids. The
preparation of these monomers, their coupling with !-ketoacids, and the
conversion of the !,!-dichloroacids to !-ketoacids is described. These studies
provide a first step to a conceptually unique approach to peptide synthesis that
does not require activating reagents or produce chemical byproducts.
This paper is dedicated to Professor Albert Eschenmoser on the occasion of his
85th birthday.
INTRODUCTION
The chemical and biogenic synthesis of !-amino acids relies on the activation of the carboxylic acid
moiety towards coupling with an amine nucleophile.1 The success and prevalence of this paradigm has
obscured the consideration of alternative approaches to iterative peptide couplings that could have
advantages in chemical synthesis or a potential role in the prebiotic or exobiotic assembly of oligopeptide
chains. In addressing this, we have sought to develop methods for amide bond formation that are
chemically distinct from classical peptide synthesis and have recently identified the coupling of !-
1 Present address: Department of Medicinal Chemistry, Institute of Biomaterials and Bioengineering, Tokyo
Medical and Dental University, 2-3-10 Kanda surugadai, Chiyoda-ku, Tokyo 101-0062, JAPAN
2 Present address: Laboratorium für Organische Chemie, ETH–Zürich, Wolfgang Pauli Strasse 10, Zürich 8093,
Switzerland.

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HETEROCYCLES, Vol. 82, No. 2, 2011
ketoacids and hydroxylamines as a novel, chemoselective, and water-compatible approach to amide and
peptide bond formation.2
We have previously documented the application of the !-ketoacid–hydroxylamine ligation (KAHA
ligation) to the iterative synthesis of "3-oligopeptides.3 These reactions proceed in water, do not require
coupling reagents, and produce only CO2 and MeOH as reaction byproducts (Figure 1). Provided that
suitable enantiopure monomers for the preparation of !-oligopeptides could be identified, this chemistry
could offer a novel and chemically orthogonal route to iterative peptide synthesis. Importantly, such an
approach could obviate the problematic and wasteful coupling reagents typically employed in large
excess during classical peptide synthesis.4
R1
O
HN
R2
O
O
OMe
OMe
OH
+
N
H
O
O
1) H2O, no reagents
2) LiOH
R1
O
R2
O
HN
R3
O
O
OMe
OMe
OH
+
N
N
H
H
O
O
1) H2O, no reagents
2) LiOH
O
R1
O
R2
O
R3
O
OH
N
H
N
H
N
H
O
Figure 1. Iterative synthesis of "3-oligopeptides by chemoselective amidation without reagents or
byproducts
The difficulty in extending our earlier work to the iterative synthesis of !-oligopeptides lies in the
identification and synthesis of a suitable monomer class to serve as the building blocks for the amino acid
residues. These monomers must 1) cleanly afford the peptides by decarboxylative condensations with !-
ketoacids, 2) result in products that can be converted to the next !-ketoacids following the ligation, and 3)
be amenable to preparation in enantiopure form on a multigram scale. Initially, we envisioned an
extention of the isoxazolidine approach we applied towards the preparation of "-peptides, which would
require !-ketolactone monomers 2. However, such monomers are likely to be highly prone to
epimerization, as such 5-membered ring heterocycles have a tendency to exist predominantly in the
conjugated enol form.5 After considering several alternatives, we elected to pursue the use of !,!-

HETEROCYCLES, Vol. 82, No. 2, 2011
1517
dicholoroacids as masked !-ketoacid surrogates (Figure 2). In this article, we describe our preliminary
investigations into the preparation of enantiomerically enriched !,!-dichloroisoxazolidinones and their
use for the iterative synthesis of !-oligopeptides. These studies establish a synthetic entry into these
monomers, their viability in the key amide-forming ligation reaction, and the conversion of the resulting
!,!-dichloroacids into the corresponding !-ketoacids.
HN
R1
O
HN
R1
O
HN
R1
O
OMe
OMe
OMe
O
OTMS
O
O
Cl Cl
1
2
3
!3-amino acid
"-amino acid
masked "-amino acid
monomer
monomer
monomer
HN
R1
O
O
OH
2'
Figure 2. Design of monomers for iterative !-peptide synthesis
RESULTS AND DISCUSSION
Our synthetic route to the enantiomerically enriched valine-derived dichloro monomer is illustrated in
Scheme 1. After exploring several different cycloaddition routes, we selected a modification of Vasella’s
carbohydrate-derived nitrone as a viable synthesis of monomer 8.6 Simply heating a mixture of the
nitrone, prepared from isobutyl aldehyde 4 and D-Gulose-derived hydroxylamine 5,7,8 with !,!-
dichloroketene silyl acetal 69 containing a stoichiometric amount of ZnCl2 provided directly the !,!-
dichloroisoxazolidin-5-ones 7 in 44% yield with high regio- and diastereoselectivity, rather than the
expected !,!-dichloroisoxazolidine.10,11 Removal of the chiral auxiliary by perchloric acid-mediated
hydrolysis provided L-!-Val monomer 8 in 72% yield. The enantiomer D-!-Val monomer ent-8 was
prepared from L-Gulose-derived hydroxylamine by a similar sequence.

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HETEROCYCLES, Vol. 82, No. 2, 2011
Me Me
Me Me
O
O
O
O
O
H
N
H
O
H
H
MgSO4
CH2Cl2
O
Me
HO
+
O
H
N
O
O
O
Me
Me
H
O
Me
Me
Me
Me
Me
4
5
D-gulose-derived
nitrone intermediate
hydroxylamine
Cl
Cl
OTMS
OMe
D
-gulose
6
HN
O
N
O
HClO4 (aq)
ZnCl2
Me
Me
O
O
THF, rt to 80 °C
MeCN
Cl Cl
Me
Cl Cl
Me
72% yield
44% yield
8
7
L
-valine monomer
Scheme 1. Preparation of L-Valine-derived !,!-dichloroisoxazolidinone monomer
Although the condensation of the nitrone and the silyl ketene acetal could also be applied to give
isoxazolidinones corresponding to leucine, phenylalanine, and alanine-monomers, the removal of the
chiral auxiliary proved problematic. We believe that this can be addressed with optimization, but have
instead pressed on with our exploration of the chemistry of these heterocycles using the valine-derived
monomers.
With !,!-dichloro-isoxazolidinone 8 in hand, the decarboxylative amide bond formation with
phenylpyruvic acid (9) was examined (Table 1). Polar solvents such as DMF or DMSO are ideal solvents
for the coupling of O-unsubstituted hydroxylamines.2 On the other hand, non-polar or protic solvents
including aqueous media are suitable for the coupling of isoxazolidine-based monomer.3 We therefore
began our studies with a screen of solvents for the amide-formation using phenylpyruvic acid (1.5 equiv)
and 0.1 M monomer 8 at 40 °C for 12 h. Non-polar solvents such as CH2Cl2, toluene and THF were
unproductive (entries 1-3). Polar solvents, including DMF and DMSO, gave the desired amides in low
conversion along with significant amounts of unidentified products (entries 4 and 5), probably due to the
t
decomposition of the starting material. A 1:1 mixture of BuOH and water gave the desired amide 10
cleanly, albeit with low conversion (entry 6). Reaction with higher substrate concentration (0.2 M) gave
10 with improved conversion (entry 7), but further increases in concentration did not improve the
outcome (entry 8). Additional optimization of these conditions revealed that MeOH or EtOH were
superior cosolvents (entries 9–11). Alcohol solvents alone, however, did not afford amide products (entry
12).

HETEROCYCLES, Vol. 82, No. 2, 2011
1519
Table 1. Conditions for amide-forming reactions of !-ketoacids and isoxazolidinone 8
Me
Me
O
HN
O
O
O
see Table 1
Ph
OH
Me
Ph
+
O
N
OH
40 °C
H
Cl Cl
Me
Cl Cl
10
O
9
8
entry
conditions a
coversion (%) b
1
CH2Cl2 (0.1 M)
toluene (0.1 M)
THF (0.1 M)
nr
2
3
nr
nr
4
5
6
7
8
9
10
11
12
DMF (0.1 M)
DMSO (0.1 M)
27 c
25 c
12
31
26
77
75
55
trace
1:1 H2O/tBuOH (0.1 M)
1:1 H2O/tBuOH (0.2 M)
1:1 H2O/tBuOH (0.4 M)
1:1 H2O/MeOH (0.2 M)
1:1 H2O/EtOH (0.2 M)
1:1 H2O/iPrOH (0.2 M)
MeOH (0.2 M)
a All reactions were carried out using Val monomer 8 and phenylpyruvic acid 9 (1.5 equiv) at 40 °C for
12 h. b determined by 1H-NMR. C A significant amount of unidentified products was detected.
Having identified aqueous alcohol as the preferred solvent for amide-bond formation, we performed a
second level optimization of the reaction pH (Table 2). The ligations occurred over a wide pH range, with
slightly acidic oxalic acid buffers offering the best results.
Table 2. Optimization of pH for amide-forming ligations
Me
Me
O
HN
O
O
O
see Table 2
Ph
OH
Me
Ph
+
O
N
OH
40 °C
H
Cl Cl
Me
Cl Cl
10
O
9
8
entry conditions a
coversion (%) b
1
2
3
4
5
6
7
8
1:1 H2O/MeOH
77
74
68
77
66
78
80
90
22
1:1 pH 3.4 Ac buffer/MeOH
1:1 pH 4.6 Ac buffer/MeOH
1:1 pH 5.3 Ac buffer/MeOH
1:1 pH 5.3 citric buffer/MeOH
1:1 pH 6.2 citric buffer/MeOH
1:1 pH 5.3 oxalic buffer/MeOH
1:1 pH 6.1 oxalic buffer/MeOH
1:1 pH 7.4 phosphate buffer/MeOH
9
a
All reactions were carried out using Val monomer 8 and phenylpyruvic acid 9 (1.5 equiv) in 0.2 M
solution at 40 °C for 12 h. b determined by 1H-NMR.

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HETEROCYCLES, Vol. 82, No. 2, 2011
The key underlying concept to the proposed iterative synthesis of !-oligopeptides is the use of the !,!-
dicholoro carboxylic acid as a masked !-ketoacids. Although the conversions of dihaloacids and related
compounds to !-ketoacids are rare, several encouraging examples were known12 including a thorough
study from a group at Bristol-Myers Squibb that implicated the formation of an !-lactone in the
hydrolysis.13 It was unclear, however, if any of these methods would be compatible with the proximal
amide functionality and the epimerizable !-stereocenter.
Working from the BMS precedent for conversion of !,!-dichloroacids 10 to the !-ketoacids 11, we
began our investigations with base promoted hydrolysis (Table 3). Strong bases including LiOH (entry 1),
K3PO4 (entry 2), and Na2CO3 (entry 3) afforded exclusively undesired oxazole 12. Success was obtained
with weaker bases (entries 4–8) and several conditions afforded the !-ketoacids as the major product.
Although the oxazole byproduct could not be completely excluded, acceptable yields could be obtained
with potassium bases including potassium oxalate at 80 °C (entry 8).
Table 3. Conditions for conversion of !,!-dichloroacid 10 to !-ketoacid 11
Me
Me
Me
Me
Me
O
O
O
O
see Table 3
Me
+
Ph
Ph
N
N
OH
N
OH
80 °C
Ph
H
H
CO2H
Cl Cl
O
O
ent-10
ent-11
12
entry
condition a
(1.0 M)
conv. to 11
conv. to 12
(%) b
(%) b
73
1
2
3
4
5
6
7
8
LiOH
K3PO4
Na2CO3
AcONa
Na2HPO4
nd
nd
nd
43
26
44
60
62
>90
40
39
26
33
sodium citrate
K2HPO4
potassium oxalate
22
33
a All reactions were carried out using a,a-dichloroacid 10 in 1.0 M aqueous solution at 80 ºC for 3 h. b
determined by RP-HPLC.
At this juncture, we sought to address the critical questions of the stereochemical integrity of the !-
stereocenter under the basic conditions necessary for the unmasking of the !-ketoacids. In our experience,
which is consistent with the chemical literature,14 !-ketoacids are configurationally stable in organic
solvents and in the presence of aqueous acid, but are prone to racemization in the presence of base.15
They are generally stable under the ligation conditions.

HETEROCYCLES, Vol. 82, No. 2, 2011
1521
!,!-Dichloroacid 10 and its enantiomer ent-10, prepared from the L-gulose-derived chiral auxiliary, were
each hydrolyzed with 1.0 M potassium oxalate at 80 °C for 3 h and the resulting !-ketoacids isolated in
48–52% yield by preparative HPLC (Scheme 2). We were not able to assay the enantiopurity of !-
ketoacid 11 directly and therefore elected to perform a second ligation prior to analysis. Each enantiomer
of valine-derived !-ketoacid 11 was subjected to ligation with benzylhydroxylamine 13 in aqueous DMF
under unoptimized conditions and the resulting amides analyzed by SFC on a AD-H column.
Unfortunately, this study revealed significant epimerization of the valine residue, presumably due to the
basic conditions employed in the hydrolysis of the !,!-dichloroacid.
H
N
Ph
Me
Me
Me
Me
HO
O
O
O
•HCl
13
H
potassium oxalate (aq)
80 °C, 3 h
Ph
Ph
N
Ph
Ph
Ph
Ph
10
N
N
OH
aq. DMF
40 °C, 16 h
H
H
O
O
11
14
50% ee
52% isolated yield
Me
Me
Me
Me
O
O
O
potassium oxalate (aq)
80 °C, 3 h
13
H
N
ent-10
N
N
OH
aq. DMF
40 °C, 16 h
H
H
O
O
ent-11
48% isolated yield
ent-14
58% ee
Scheme 2. Investigation of epimerization during hydrolysis of !,!-dichloroacid 10
In preliminary work, we have prepared dichloroisoxazoline monomers corresponding to other amino acid
residues including L-leucine and L-phenylalanine and found that these examples are less prone to
racemization during hydrolysis, suggesting that improvements to these protocols are possible. Further
efforts to improve the hydrolysis conditions and the preparation of the monomers may provide an
alternative route to the iterative synthesis of !-peptides that does not require coupling reagents or
protecting groups and which operates under aqueous conditions
ACKNOWLEDGMENTS
T.N. was a postdoctoral fellow supported by the Japan Society for the Promotion of Science. Additional
funding for this work was generous provided fellowships to J.W.B from the Arnold and Mabel Beckman
Foundation and the David and Lucille Packard Foundation.

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HETEROCYCLES, Vol. 82, No. 2, 2011
EXPERIMENTAL
General Methods. All reactions utilizing air- or moisture-sensitive reagents were performed in dried
glassware under an atmosphere of dry nitrogen. Thin layer chromatography (TLC) was performed on
EMD precoated plates (silica gel 60 F254, Art 5715) and were visualized by fluorescence quenching
under UV light and by staining with phosphomolybdic acid and/or p-anisaldehyde, respectively. For flash
column chromatography, EMD Silica Gel 60 (230–400 Mesh) and Silica Gel 60 N (Kanto Chemical Co.,
Inc.) were employed. 1H NMR (500 MHz) and 13C NMR (125 MHz) were measured on a Bruker Avance
AVII-500 spectrometer with a CryoProbe. Chemical shifts are expressed in parts per million (ppm) with
respect to the residual solvent peak. Coupling constants are reported as Hertz (Hz), signal shapes and
splitting patterns are indicated as follows: br, broad; s, singlet; d, doublet; t, triplet; q, quartet; m,
multiplet. Infrared (IR) spectra were recorded on a SHIMADZU IR Prestige-21 FTIR-8400S and are
reported as wavenumber (cm–1). Low- and high-resolution mass spectra were recorded on a Brucker
Daltonics microTOF-2focus in the positive and negative detection mode. Optical rotations were measured
on JASCO DIP-370 polarimeter operating at the sodium D line with a 100 mm path length cell, and were
reported as follows: [!]DT (concentration (g:100 mL), solvent).
HPLC Conditions. For analytical HPLC, a Cosmosil 5C18-ARII column (4.6 x 250 mm, Nacalai Tesque,
Inc., Kyoto, Japan) was employed with a linear gradient of MeCN containing 0.1% (v/v) TFA at a flow
rate of 1 cm3 min–1 on a LaChrom Elite HTA system (Hitachi High-Technologies corporation, Ltd.,
Tokyo, Japan) and JASCO PU-2086 plus (JASCO corporation, Ltd., Tokyo, Japan), and eluting products
were detected by UV at 220 nm. Preparative HPLC was performed using a Cosmosil 5C18-AR II column
(20 x 250 mm, Nacalai Tesque, Inc.) on a JASCO PU-2087 plus and PU-2089 plus (JASCO corporation,
Ltd., Tokyo, Japan) in a suitable gradient mode of MeCN solution containing 0.1% (v/v) TFA at a flow
rate of 7-10 cm3 min–1.
(2,2-Dichloro-1-methoxyvinyloxy)trimethylsilane 6. To a suspension of activated Zn dust (784.8 mg,
12.0 mmol) in THF (10 mL) was added dropwise a solution of Cl3CCO2Me (0.953 mL, 8.0 mmol) and
TMSCl (1.23 mL, 9.6 mmol) at ambient temperature (< 40 °C). After stirring at room temperature
overnight, the reaction mixture had changed to a clear cream-colored suspension containing the excess Zn
dust. This mixture was used in the next step without further purification.
(S)-4,4-Dichloro-2-((3aR,4R,6S,6aR)-6-((R)-2,2-dimethyl-1,3-dioxolan-4-yl)-2,2-dimethyltetrahydro-
furo[3,4-d][1,3]dioxol-4-yl)-3-isopropylisoxazolidin-5-one 7. To a solution of D-gulose derived chiral
auxiliary 116 (562.6 mg, 2.0 mmol) in CH2Cl2 (10 mL) was added isobutylaldehyde (272.4 µL, 3.0 mmol)

HETEROCYCLES, Vol. 82, No. 2, 2011
1523
in the presence of MgSO4 (1 g). After stirring at rt for 3 h, the reaction mixture was filtered to remove
MgSO4. Concentration under reduced pressure gave the nitrone as a white solid, which was used in the
next step without further purification. To a solution of the nitrone in THF (10 mL) was added a solution
of !,!-dichloroketene silyl acetal 6 (0.80 M, 8.0 mmol) containing a stoichiometric amount of ZnCl2 in
THF (10 mL), from which the excess Zn dust was removed with a PTFE filter (0.45 µm). After stirring at
rt for 30 min, the mixture was refluxed for 6 h. After cooling to room temperature, the reaction mixture
was poured into ice-cold saturated aqueous NaHCO3 and extracted with EtOAc. The extract was washed
with brine and dried over MgSO4. Concentration under reduced pressure followed by flash
chromatography over silica gel with hexanes-EtOAc (9:1) gave 7 (382.7 mg, 44% yield) as a colorless
oil: [!]25D +3.74 (c 0.710, CHCl3); 1H NMR (500 MHz, CDCl3) " 5.39 (s, 1H), 4.96 (d, J = 6.0 Hz, 1H),
4.77 (dd, J = 6.0, 4.0 Hz, 1H), 4.39 (dt, J = 8.5, 6.5 Hz, 1H), 4.32 (d, J = 3.0 Hz, 1H), 4.23 (dd, J = 9.0,
7.0 Hz, 1H), 4.15 (dd, J = 8.5, 4.0 Hz, 1H), 3.76 (dd, J = 8.5, 6.5 Hz, 1H), 2.43-2.34 (m, 1H), 1.46 (s, 3H),
1.39 (s, 3H), 1.38 (s, 3H), 1.31 (s, 3H), 1.06 (d, J = 7.0 Hz, 3H), 0.882 (d, J = 6.5 Hz, 3H); 13C NMR
(125 MHz, CDCl3) " 168.5, 113.4, 109.9, 97.8, 85.6, 84.1, 80.2, 78.4, 75.4, 74.4, 65.9, 31.2, 26.5, 26.0,
25.1, 24.7, 21.0, 15.7; IR (KBr) # 2981, 1807, 1209, 1080, 979, 948, 894, 864, 848, 759 cm-1; HRMS
(ESI), m/z calcd for C18H28Cl2NO7 [M+H]+ 440.1243 and 442.1213, found 440.1245 and 442.1214.
(S)-4,4-Dichloro-3-isopropylisoxazolidin-5-one 8 (L-Val monomer). To a solution of D-gulosyl L-Val
monomer 7 (102.8 mg, 0.233 mmol) in MeCN (2.3 mL) was added 60% HClO4 (116.7 µL, 0.700 mmol),
and the mixture was stirred at rt for 1.5 h. After being diluted with EtOAc, the mixture was washed with
saturated aqueous NaHCO3 and brine, and dried over MgSO4. Concentration under reduced pressure
followed by flash chromatography hexanes-EtOAc (9:1) gave 8 (33.3 mg, 72% yield) as a colorless oil:
1
[!]25 –3.18 (c 1.27, CHCl3); H NMR (500 MHz, CDCl3) " 6.70 (br, 1H), 3.69 (dd, J = 12.3, 8.9 Hz,
D
1H), 2.01-2.20 (br, 1H), 1.24 (d, J = 6.7 Hz, 3H), 1.24 (d, J = 6.7 Hz, 1H); 13C NMR (125 MHz, CDCl3)
" 170.0, 79.5, 76.0, 28.2, 19.0; IR (KBr) # 3431, 3226, 1811, 1197, 759 cm-1; HRMS (ESI), m/z calcd for
C6H10Cl2NO2 [M+H]+ 198.0089 and 200.0059, found 198.0082 and 200.0056.
(S)-2,2-Dichloro-4-methyl-3-(2-phenylacetamido)pentanoic acid 10. To a solution of L-Val monomer
8 (61.4 mg, 0.310 mmol) in a 1:1 mixture (1.55 mL) of MeOH/oxalic buffer (pH 6.1) was added
o
phenylpyruvic acid (76.4 mg, 0.465 mmol). After being stirred at 40 C for 41 h, purification by
preparative HPLC (Gradient: 0 min, 30% MeCN in H2O; 90 min, 60% MeCN in H2O) followed by
1
lyophilization afforded compound 10 (63.4 mg, 64% yield) as colorless powder: H NMR (500 MHz,
CDCl3) " 7.40-7.28 (m, 5H), 5.97 (d, J = 10.5 Hz, 1H), 4.82 (dd, J = 10.5, 3.5 Hz, 1H), 3.82 (d, J = 16.5,
13
1H), 3.74 (d, J = 16.5, 1H), 2.37-2.27 (m, 1H), 0.956 (d, J = 7.0, 3H), 0.745 (d, J = 6.5, 3H); C NMR

1524
HETEROCYCLES, Vol. 82, No. 2, 2011
(125 MHz, CDCl3) " 173.6, 166.5, 133.4, 129.7, 129.4, 128.0, 87.1, 60.7, 43.2, 29.4, 22.0, 17.2; HRMS
(ESI), m/z calcd for C14H18Cl2NO3 [M+H]+ 318.0664 and 320.0634, found 318.0653 and 320.0630.
(S)-4-Methyl-2-oxo-3-(2-phenylacetamido)pentanoic acid 11. Dichloro-isoxazolidinone 10 (55.6 mg,
0.175 mmol) was treated with 1.0 M aqueous solution of potassium oxalate (3.5 mL) at 80 °C for 3 h.
After cooling to room temperature, purification by preparative HPLC (Gradient: 0 min, 25% MeCN in
H2O; 90 min, 55% MeCN in H2O) followed by lyophilization afforded 11 (23.8 mg, 52% yield) as a
colorless oil: 1H NMR (500 MHz, CDCl3) " 7.41-7.27 (m, 5H), 6.14 (br, 1H), 5.03 (br, 1H), 3.69 (s, 2H),
2.30-2.23 (m, 1H), 0.924 (d, J = 7.0 Hz, 3H), 0.683 (d, J = 7.0 Hz, 3H); 13C NMR (125 MHz, CDCl3) "
193.1, 173.3, 160.5, 133.5, 129.4, 129.3, 127.9, 60.0, 43.0, 29.6, 19.6, 17.0; IR (KBr) # 3388, 1724, 1647,
1531, 1215, 761 cm-1; HRMS (ESI), m/z calcd for C14H18NO4 [M+H]+ 264.1236, found 264.1239.
2-Benzyl-4-isopropyloxazole-5-carboxylic acid 12. Dichloro-isoxazolidine 10 (55.6 mg, 0.175 mmol)
1
was converted into the title compound 12 (11.2 mg, 26% yield) as colorless freeze-dried powder: H
NMR (500 MHz, CDCl3) " 7.34-7.27 (m, 5H), 4.17 (s, 2H), 3.67-3.51 (m, 1H), 1.27 (d, J = 6.5, 6H); 13C
NMR (125 MHz, CDCl3) " 165.6, 162.6, 157.6, 135.39, 134.4, 128.9, 128.8, 127.3, 34.8, 26.0, 21.3; IR
(KBr) # 3412, 1612, 758 cm-1; HRMS (ESI), m/z calcd for C14H16NO3 [M+H]+ 246.1130, found 246.1127.
This is the side product of the previous reaction.
REFERENCES
1. J. Jones, “Amino Acid and Peptide Synthesis Second edition” Oxford University Press, Inc., New
York, 2002; T. Sonke, B. Kaptein and H. E. Schoemaker, “Amino Acids, Peptides and Proteins in
Organic Chemistry. Vol. 1-Origins and Synthesis of Amino Acids,” ed. by A. B. Hughes, Wiley-
VCH Verlag GmbH & Co. KGaA, Weinheim, 2009, pp. 79-117.
2. J. W. Bode, R. M. Fox, and K. D. Boucom, Angew. Chem. Int. Ed., 2006, 45, 1248.
3. N. Carrillo, E. A. Davalos, J. A. Russak, and J. W. Bode, J. Am. Chem. Soc., 2006, 128, 1452.
4. W. C. Chan, and P. D. White, “Fmoc Solid Phase Peptide Synthesis” Oxford University Press, Inc.
New York, 2000.
5. For example, Vitamin C has a five-membered ring containing two-carbonyl groups but normally
exists predominently as a conjugated ene-diol, see: J. Clayden, N. Greeves, S. Warren and P.
Worthers, “Organic Chemistry” Oxford University Press, Inc., New York, 2001, pp. 523-545.
HO
HO
HO
H
H
H
O
O
O
O
O
O
HO
HO
HO
H
H
O
OH
stable ene-diol form of Vitamin C
O
HO
HO
OH
one unstable keto form
another unsatble keto form

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