Chemical reactivity of 6-diazo-5-oxo-l-norleucine (DON) catalyzed by metalloporphyrins (Fe,Ru)

Article abstract of DOI:10.1016/j.tet.2010.04.080

The transfer of the metallocarbene derived from N- and O-protected 6-diazo-5-oxo-l-norleucine (DON) catalyzed by metalloporphyrins undergoes dimerization, cyclopropanation, N-H and S-H insertion reactions, respectively. An efficient and direct synthesis of 5-oxo-l-pipecolic acid from DON is described from unprotected 6-diazo-5-oxo-l-norleucine.

Full text of DOI:10.1016/j.tet.2010.04.080

Tetrahedron 66 (2010) 4462e4468
Contents lists available at ScienceDirect
Tetrahedron
journal homepage: www.elsevier.com/locate/tet
Chemical reactivity of 6-diazo-5-oxo-
metalloporphyrins (Fe,Ru)
L-norleucine (DON) catalyzed by
*
Paul Le Maux, Irène Nicolas, Soizic Chevance, Gérard Simonneaux
Ingénierie Chimique et Molécules pour le Vivant, UMR CNRS 6226, Campus de Beaulieu, Université de Rennes 1, 35042 Rennes, France
a r t i c l e i n f o
a b s t r a c t
Article history:
The transfer of the metallocarbene derived from N- and O-protected 6-diazo-5-oxo-L-norleucine (DON)
Received 15 March 2010
Received in revised form 14 April 2010
Accepted 19 April 2010
catalyzed by metalloporphyrins undergoes dimerization, cyclopropanation, NeH and SeH insertion re-
actions, respectively. An efficient and direct synthesis of 5-oxo- -pipecolic acid from DON is described
from unprotected 6-diazo-5-oxo- -norleucine.
L
L
Available online 24 April 2010
Ó 2010 Elsevier Ltd. All rights reserved.
Keywords:
6-Diazo-5-oxo-L-norleucine
Ruthenium porphyrin
Iron porphyrin
Catalytic carbene transfer
1. Introduction
cyclopropanation, NeH and SeH insertion reactions. We also report
intramolecular NeH insertion with unprotected 6-diazo-5-oxo-
L-
6-Diazo-5-oxo-
antibiotic that was isolated from Streptomyces in 1956.1 It is the
diazo analog of -glutamic acid and has been shown to interfere
L
-norleucine (commonly known as DON) is an
norleucine, which led to 5-keto-L-pipecolic acid.
L
2. Results
with a number of biological pathways, including purine, pyrimi-
dine, and protein synthesis where glutamine is used as a nitrogen
source.2 DON acts as a suicide inhibitor of glutaminase/aspar-
aginase3 and has subsequently been used in a variety of applica-
tions including labeling of the active sites of enzymes4 and possibly
as anticancer drug.2,5 Combination therapy of animal tumors with
The antibiotic 6-diazo-5oxo-L-norleucine has been synthesized
several times from the N-protected acyl chloride and diazo-
methane, but in low overall yields.10 Herein, we used a more recent
method reported by Coutts and Saint,11 in which the lithium salt of
trimethylsilyldiazomethane was reacted with pyroglutamates. In
our hands, the N- and O-protected diazo derivative was prepared in
50% yield (Scheme 1). Using a similar route, the unprotected diazo
amino acid was also prepared (Scheme 2, 52% yield) (see also
Experimental section).
L
-asparaginase and 6-diazo-5-oxo-L-norleucine, was also sug-
gested, more than thirty years ago,6 and is now starting a new
development.7
For the objective of understanding better the in vivo reactivity,
we have studied the chemical reactivity of protected DON with
metalloporphyrins as models of heme enzymes. Actually, possible
interaction of diazoketones with enzymes such as cytochrome
P450 has been previously suggested.8,9 It is also expected that
metalloporphyrins (Fe,Ru) are well suited as catalysts for in-
termolecular carbene transfer using polyfunctionnalyzed diazo
derivatives as reagents due to good selectivity. Here we report that
metallocarbene transfers derived from N- and O-protected 6-diazo-
To demonstrate the presence of possible metallocarbene in-
termediate, first cyclopropanation reaction was investigated
(Scheme 3). The results are summarized in Table 1. Thus to evaluate
the reactivity of N- and O-protected 6-diazo-5-oxo-L-norleucine
compound, its ruthenium-catalyzed decomposition was first ex-
amined in the presence of styrene in toluene at room temperature
in the presence of tetraphenylporphyrin ruthenium carbon mon-
oxide 1 (Fig. 1) as catalyst (Table 1, entry 1). The cyclopropane was
formed with 72% yield and high diastereoselectivity (trans/cis¼99/
1) with a concomitant formation of the dimer (21%), resulting from
coupling of two carbene precursors. An increased yield (85%) with
a selectivity (trans/cis¼91/9) was observed using tetraphenylpor-
phyrin iron chloride 3 instead of the ruthenium complex 1, as
catalyst. As previously reported by Woo and coll.12,13 with ethyl
5-oxo-L-norleucine (DON) catalyzed by metalloporphyrins are
possible giving intermolecular reactions such as dimerization,
* Corresponding author. Fax:þ33
2 23 23 56 37; e-mail address: gerard.
simonneaux@univ-rennes1.fr (G. Simonneaux).
0040-4020/$ e see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2010.04.080

P. Le Maux et al. / Tetrahedron 66 (2010) 4462e4468
4463
O
O
CO2Et
NHBoc
CO2H
(CH3)3SiCHN2, nBuLi
-105°C, THF
1. SOCl2/ EtOH
2. KOH
(t-BuO2C)2O, NEt3, DMAP
RT
BocN
HN
N2
HO
NH2
O
O
EtO2C
EtO2C
Scheme 1.
O
O
CO2H
NH2
CO2Et
NHFmoc
(CH3)3SiCHN2, nBuLi
piperidine
1. LHMDS, THF, -78°C
2. FmocCl, -78°C --> RT
FmocN
EtO2C
HN
N2
N2
-105°C, THF
O
O
EtO2C
Scheme 2.
Scheme 3.
diazoacetate, we took benefit of the addition of cobaltocene to in-
crease the reactivity. Actually, the absence of cobaltocene did not
give any cyclopropanation reaction, probably due to the difficulty of
reducing the ferric state to the ferrous state. As expected with 1 or
3, no enantiomeric excess is observed.
Table 1
Cyclopropanation of styrene with N2CHCO(CH2)2CH(CO2Et)NHBoc catalyzed by Ru
(CO)(TPP) (1), (ꢀ)-Ru(CO)(Halt) (2), Fe(Cl)(TPP) (3), (ꢀ)-Fe(Cl)(Halt) (4)a
Enty
Catalyst
Time
(h)
Yieldtransþcis
(%)b
Ratio
ee trans
(%)c
Dimer
(%)
trans/cis
We then investigated the cyclopropanation with chiral Halter-
man porphyrin ruthenium carbon monoxide 2 (Fig. 1) (Table 1).
With styrene, the cyclopropane was formed with 75% yield, 99/1
trans/cis ratio and 80% enantioselectivity for the trans isomer (Table
1, entry 2). We also investigated the asymmetric cyclopropanation
catalyzed by chiral Halterman porphyrin iron chloride 4 (Table 1).
As shown in Table 1, the chemical yields are higher (w95%) with
iron porphyrins than those obtained with ruthenium porphyrins 1
or 2 (72e75%) and the enantioselectivity was quite good, 80% ee. It
should be noted that other chiral metalloporphyrins derived from
1
2
1
5
5
2
2
72
75
85
95
99/1
99/1
91/9
95/5
d
80
d
80
21
18
7
2e
3
3
4d
4
<5
a
A molar ratio of 1:200:1000 for catalyst: diazo: styrene was employed at am-
bient temperature in toluene.
b
Yields were determined by isolation of cyclopropanes by column chromato-
graphy on silica gel.
c
ees were determined by chiral HPLC using a Chiralcel OD column.
At 40 ꢁC.
The other enantiomer (þ)-Ru(CO)(Halt) gave an ee¼68%.
d
e
L1
L1
N
N
M
N
N
M
N
N
N
N
L2
L2
1 ML1= RuCO
3 ML1= FeCl
2 ML1= RuCO
4 ML1= FeCl
SO3Na
L1
NaO3S
N
N
M
N
N
SO3Na
L2
NaO3S
5 ML1= RuCO
Figure 1. Catalysts used in the reactions of cyclopropanation, NeH and SeH insertions with N2CHCO(CH2)2CH(CO2Et)NHBoc.

4464
P. Le Maux et al. / Tetrahedron 66 (2010) 4462e4468
aminophenylporphyrins14 have been reported to give cyclopro-
panes with high enantiomeric excesses, when ethyl diazoacetate
was used as reagent.15,16
Since peptidyl diazomethyl ketones appeared initially to be
specific inactivators of cysteine proteinases,17,18 insertion of DON
into SeH bonds, catalyzed by metalloporphyrins, was assayed
(Scheme 4). The different results obtained with 1 or 3 as catalysts
are summarized in Table 2. Treatment of thiophenol with N-tert-
DON and its next smaller homolog 5-diazo-4-oxo-
act as suicide inhibitors of glutaminase/asparaginases and were
proposed to form an -keto ether linkage through the reaction with
L-norvaline,
a
the OH group of a threonine in the active site.3 Thus to fully char-
acterize the catalytic property of the ruthenium and iron porphyrin
complexes, an investigation of the competition between the SeH
and OeH insertions was also undertaken to see if OeH insertion
was also possible. As can be seen in Table 2 (entries 5 and 6), we
observed only SeH insertion with 2-mercaptoethanol. This is not
too surprising since we previously observed that the carbene
complexes of iron and ruthenium porphyrins are stable in water
and protic solvents.20
butoxycarbonyl-6-diazo-5-oxo-L-norleucine methyl ester catalyzed
by complex 1 and 3 gave insertion of the diazo derivative into the
SeH bond with 85 and 51% yield, respectively. To complete the
reactivity of this diazoketone, an investigation of the competition
between the cyclopropanation of alkenes and the insertion reaction
catalyzed by complex 1 or 3 was also undertaken (entries 3 and 4,
Table 2). Only the SeH insertion compound is observed by 1H NMR
when allyl sulphide is the substrate with 87 and 58% yield, re-
spectively. It should be noted that a preference for insertion has
already been observed in ruthenium-catalyzed reaction of diazo-
esters with unsaturated thiols.19
The insertion of electrophilic carbenes in the NeH bonds of
protected a-amino esters or amides is a powerful method for N-
alkylating this class of compounds.21 Since we19,22 and others23e25
previously reported NeH insertion of diazoesters catalyzed by
metalloporphyrins (Fe, Ru), NeH insertion with a diazoketone such
as DON was also investigated (Scheme 5). The results presented in
Table 2 (entries 7 and 8) show that the ruthenium complex, and to
CO2Et
O
O
O
Catalyst
S
RSH
N2
+
N2
+
R
O
O
N
H
Toluene
O
CO2Et
R
=
Ph
H2CHC CH2
Scheme 4.
a less extent the iron complex, are good catalysts for the trans-
formation of aniline into the expected products. Due to a probable
reduction of iron(III) to iron(II), addition of cobaltocene together
with complex 3 was not necessary in the reaction. However the
reaction needs 5 h to be completed compared to few minutes when
diazoacetate was used as reagent.19,24 The stereochemical con-
straints that result from the steric interaction of the axial carbene
atoms with atoms of the porphyrinato core26 may explain the de-
crease of the rate. The mechanisms of the cyclopropanation re-
action, NeH insertion and SeH insertion, catalyzed by
metalloporphyrins, have been previously discussed by us19 and
others23e25 several times, and will not be presented herein.
Next, we also report that the ruthenium complex 1 catalyzes the
stereoselective decomposition of protected DON to form olefinic
products (Scheme 6). This coupling reaction catalyzed by ruthe-
nium porphyrin to form olefins, proceeds through a metallocarbene
intermediate (detected by the chemical shift of the carbene proton
Table 2
SeH and NeH insertions with N2CHCO(CH2)2CH(CO2Et)NHBoc catalyzed by Ru(CO)
(TPP) (1) and Fe(Cl)(TPP) (3)a
Entry Catalyst Substrate
Product
Yieldb
(%)
1
2
3
4
5
6
7
8
1
3
1
3
1
3
1
3
PhSH
PhSH
PhSCH2COCH2CH2CHRNHBoc
PhSCH2COCH2CH2CHRNHBoc
H2C]CHCH2SH H2C]CHCH2SCH2COCH2CH2CHRNHBoc 87
H2C]CHCH2SH H2C]CHCH2SCH2COCH2CH2CHRNHBoc 58
HOCH2CH2SH HOCH2CH2SCH2COCH2CH2CHRNHBoc 70
HOCH2CH2SH HOCH2CH2SCH2COCH2CH2CHRNHBoc 54
85
51
PhNH2
PhNH2
Ph NH CH2COCH2CH2CHRNHBoc
Ph NH CH2COCH2CH2CHRNHBoc
71
55
a
A molar ratio of 1:200:200 for catalyst: diazo: substrate was employed at am-
bient temperature in toluene.
b
Yields were determined by isolation of insertion product by column chroma-
tography on silica gel.
H
CO2Et
O
O
O
Catalyst
N
PhNH2
N2
+
N2
+
Ph
O
O
N
H
Toluene
O
CO2Et
Scheme 5.
CO2Et
CO2Et
O
H
H
O
O
O
O
O
O
N
H
N
H
O
O
CO2Et
cis (82%)
O
Catalyst 1
N2
+
N2
+
O
N
H
Toluene
trans (18%)
O
O
H
CO2Et
H
N
N
H
O
CO2Et
H
O
Scheme 6.

P. Le Maux et al. / Tetrahedron 66 (2010) 4462e4468
4465
which appears at a very low field, 13.72 ppm by 1H NMR), giving
rise to a high cis/trans ratio of 82/18 in overall 81% yield. Attribution
of cis and trans isomer compounds was realized by NMR (HMBC,
see Experimental section). In comparison, a similar ruthenium(II)
porphyrin species has been reported to catalyze rapidly the de-
composition of ethyl diazoacetate to form diethyl maleate and
diethyl fumarate with a cis/trans ratio of 15.27 The generally ac-
cepted reaction pathway for transition metal catalyzed carbene
dimerization of diazo derivatives involves initial attack of the diazo
compound at the metal center to generate a metal carbene.28,29
Nucleophilic attack on the metal carbene by diazo compound,
followed by dissociation of the olefin from the metal complex,
completes the catalytic cycle. The origin of cis selectivity during the
different reactions giving the cyclopropane product, the SeH in-
sertion product and the NeH insertion product, respectively,
without any intramolecular reaction.
There have been a number of examples of Rh2(OAc)4-catalyzed
intramolecular NeH insertions as a key step in the synthesis of b-
lactams.40,10 Most of the intramolecular NeH insertion reactions
catalyzed by Rh2(OAc)4 involve insertion into amide bonds.40
Consequently, intermolecular reactions with diazoketones de-
rived from protected amino acids are difficult with rhodium acetate
catalysts and consequently, of rather limited use. It should also be
noted that addition of electrophilic carbenes to olefinic amino acid
derivatives is also possible but this approach is limited due to a low
diastereoselectivity.21 In contrast, carbene transfers catalyzed by
metalloporphyrins are much more selective41,42 due probably to
a lower reactivity of the metal-carbene intermediate.
coupling of a-diazo compounds has been previously interpreted on
the basis of steric and electronic effects.30e32
Finally, reactivity of unprotected DON was also tested using
ruthenium porphyrin 5 as a water-soluble catalyst (Scheme 7).
To examine whether metalloporphyrins could be used for
intramolecular NeH insertion, the diazo was completely depro-
tected. In this case, the intramolecular reaction was observed and
the cyclic product was obtained in 71% yield using 5 as catalyst. In
this case, intramolecular NeH insertion is probably much faster
than intermolecular addition of the unprotected diazo derivative to
the double bond of styrene, since no intermolecular addition was
observed, even in presence of a large excess of styrene. It should be
noted that we previously reported with a similar catalytic system19
that NeH insertion is preferred to cyclopropanation reaction when
4-aminostyrene is the substrate. The intramolecular insertion led to
Complex 5 in presence of an excess of unprotected 6-diazo-5-oxo-
norleucine in water at 25 ꢁC under an inert atmosphere gives, in
73% yield, 5-keto- -pipecolic acid, resulting from an intramolecular
L-
L
NeH insertion. In the presence of 10 equiv of styrene in water, no
cyclopropanation products are observed at room temperature and
only intramolecular NeH insertion is noted. Although an NeH
insertion was previously observed for the N-protected DON methyl
ester catalyzed by rhodium acetate,33,34 such reaction for un-
protected DON is quite surprising owing the presence of a free acid
group on the substrate. The present NeH insertion can offer a direct
5-oxo-L-pipecolic acid, which is an intermediate in the synthesis of
way to cis-5-hydroxy-
L
-pipecolic acid, a natural product, which is
the natural product, cis-5-hydroxy-L-pipecolic acid, which is pres-
present in various plants such as acacia and Rhodesian teak,33 from
ent in various plants such as Rhodesian teak, dates and acacia.33,34
a commercially available but quite expensive diazo derivative.
4. Conclusion
O
CO2H
Ru(CO)(TPPS)
In summary, the intermolecular carbene transfer of N- and
O-protected DON catalyzed by iron or ruthenium porphyrin occurs
in a highly selective manner giving cyclopropanation, NeH and
SeH insertion reactions. It should be noted that no intramolecular
NeH amide bond insertion was detected in contrast to reaction
catalyzed by rhodium acetate. However, with unprotected DON,
only intramolecular NeH insertion in the free amino group was
observed, even in presence of a large excess of alkene. Investigation
of the catalytic properties of these metalloporphyrins for the
epoxidation of olefins in water is currently underway in our
laboratory and will be reported in due course.
+
N2
N2
HN
H2O
NH2
O
CO2H
Scheme 7.
3. Discussion
The antitumor diazoketone, 6-diazo-5-oxo-
has been isolated from a natural source and can be considered as
a modified -amino acid. This glutamine analog 6-diazo-5-oxo-L-
L-norleucine (DON),
a
norleucine (DON) has been shown to bind and irreversibly in-
activate glutaminase/asparaginases from several organisms at low
concentration3 and, to a less extent, also inactivate, glutaminase
5. Experimental section
5.1. General experiments
enzymes from the
appears that the enzyme removes the diazo group of DON, pro-
ducing N2 and 5-oxo- -norleucine covalently bound to the serine 74
b
-lactamase superfamily.35 In the latter case, it
L
All reactions were performed under argon and were magneti-
cally stirred. Solvents were distilled from appropriate drying agent
prior to use: toluene from sodium and benzophenone, CH2Cl2 from
CaH2, CHCl3 from P2O5. Commercially available reagents were used
without further purification unless otherwise stated. All reactions
were monitored by TLC with Merck pre-coated aluminum foil
sheets (Silica gel 60 with fluorescent indicator UV254). Compounds
were visualized with UV light at 254 nm and 365 nm. Column
chromatographies were carried out using silica gel from Merck
(0.063e0.200 mm). 1H NMR and 13C NMR in CDCl3 were recorded
using Bruker (Advance 500dpx and 300dpx spectrometers).
UVevisible spectra were recorded on a UVIKON XL from Biotech.
The enantiomeric excess of the cyclopropane was determined on
a Varian Prostar 218 system equipped with Chiralcel columns.
The porphyrins were synthesised according to literature
methods.43 The corresponding ruthenium carbonyl complexes, Ru
(CO)(TPP) and Ru(CO)(Halt), were obtained by refluxing the
side-chain through its terminal carbon atom. Since several different
functionalities, the diazoketone unit, the amino group and the
carboxylic acid group, are in the same molecule, different reactions
in biological media are possible such as intramolecular NeH in-
sertion or intermolecular carbene transfers. A wide range of metal
catalysts derived from copper, iron, and others have been reported
to catalyze the diazo reagent decomposition21,36,37 and, conse-
quently, metalloproteins can be also considered as good candidates
to react with DON.20 Since evidences have been presented in favor
of the formation of cytochrome P450-iron-carbene complexes
during metabolism of various substrates,38,39 the catalytic de-
composition of DON have been carried out in the presence of
metalloporphyrins as models of the active site of heme proteins.
Accordingly, we herein discovered that intermolecular transfer of
N- and O-protected 6-diazo-5-oxo-L-norleucine is possible. Three
intermolecular insertion products have been identified from three

4466
P. Le Maux et al. / Tetrahedron 66 (2010) 4462e4468
porphyrins in o-dichlorobenzene with Ru3CO12 at 180 ꢁC44,45 and
eluted with pentane/ethyl acetate (4/1). Recrystallization in hot
ethyl acetate afforded 605 mg (83%) of the desired product as
the iron porphyrins were prepared as previously reported.46
a colorless crystalline solid. 1H NMR (CDCl3, ppm)
d 1.30 (t, 3H,
5.2. Preparation11 of N2CHCO (CH2)2CH(CO2Et)NHBoc (Scheme
1)
J¼7.2 Hz, CO2CH2CH3), 2.08e2.20, 2.32e2.52, 2.58e2.85 (3 m,
1Hþ1Hþ2H, CH2CH2), 4.20e4.37 (m, 3H, CO2CH2þCH), 4.47e4.72
(m, 3H, OCH2þ*CH), 7.31, 7.45 (2 t, 4H, J¼7.4 Hz, mH Ph), 7.75, 7.80
(2d, 4H, J¼7.4 Hz, oH Ph).
5.2.1. Synthesis of (S)-(þ)-ethyl-pyroglutamate. Freshly distilled
thionyl chloride (30 ml) was added to a solution of L-glutamic acid
(25.6 g, 175 mmol) in 250 ml of absolute ethanol cooled in a ice
bath. The reaction mixture was stirred at room temperature for 1 h
and heated at reflux for 0.5 h. This solution was neutralized with
KOH in ethanol. The salt KCl formed was removed by suction fil-
tration. After evaporation of the solvent, the product was purified
by distillation under reduced pressure, yielding a colorless oil,
which solidified at room temperature, 18 g (66%). 1H NMR (CDCl3,
5.3.2. Synthesis of (S)-(þ)-N-Fmoc -6-diazo-5-oxo-norleucine ethyl
ester. To a cold (ꢀ100 ꢁC) solution of trimethylsilyldiazomethane
(3.15 ml of a 2 M solution in hexane, 6.31 mmol) in THF (30 ml)
under an inert atmosphere was added n-butyllithium in hexane
(4.04 ml of a 1.6 M solution in hexane, 6.48 mmol).The reaction was
stirred for 30 min and transferred via a cannula to a solution of (S)-
(þ)-ethyl-N-Fmoc-pyroglutamate (2 g, 5.27 mmol) in THF (50 ml)
at ꢀ105 ꢁC. The mixture was stirred for a further 10 min and
quenched by addition of the cold solution to saturated ammonium
chloride 170 ml. After extraction with ethyl acetate, the residue
obtained after evaporation of solvents was purified by column
chromatography on silica gel eluted with pentane/ethyl acetate
(7/3). After evaporation of solvents, the resulting yellow oil was
dissolved in a minimum of ethyl acetate. The product recrystallizes
ppm)
7.26 (br s, 1H).
d
1.24 (t, 3H, J¼7 Hz), 2.27e2.47 (m, 4H), 4016 (q, 2H, J¼7 Hz),
5.2.2. Synthesis of (S)-(þ)-ethyl-N-tert-butoxycarbonyl pyrogluta-
mate. To a solution of (S)-(þ)-5-carbethoxy-2-pyrrolidinone (2 g,
12.7 mmol), in 20 ml CH2Cl2 were added triethylamine (1.77 ml,
12.7 mmol), di-tert-butyl dicarbonate (5.56 g, 25.4 mmol), and 4-
(dimethylamino) pyridine (1.55 g, 12.7 mmol). The solution was
stirred for 7 h at 25 ꢁC under an argon atmosphere. The solvent was
removed, and the residue was purified by column chromatography
on silica gel. Elution with dichloromethane/pentane/ether: 1/1/0.2
afforded 2.75 g (84%) of the desired compound. 1H NMR (CDCl3,
as pale yellow plates (1.22 g, 55%). 1H NMR (CDCl3, ppm)
d 1.32 (t,
3H, J¼7.2 Hz, CO2CH2CH3), 2.00e2.44 (m, 4H, COCH2CH2C*),
4.20e4.29 (m, 3H, CO2CH2þCH), 4.38e4.45(m, 3H, OCH2þ*CH),
5.29 (br s,1H, CHN2), 5.60 (br d,1H, J¼7.4 Hz, NH), 7.35, 7.44 (2 t, 4H,
J¼7.4 Hz, mH Ph), 7.63, 7.81 (2d, 4H, J¼7.2 Hz, oH Ph).
ppm)
d
1.32 (t, 3H, J¼7.0 Hz), 1.52 (s, 9H), 2.04e2.10 (m, 1H),
2.29e2.44 (m, 1H), 2.51e2.67 (m, 2H), 4.25 (q, 2H, J¼7.0 Hz), 4.60,
5.3.3. Synthesis of (S)-(þ)-6-diazo-5-oxo-norleucine (DON). (S)-
(þ)-N-Fmoc-6-diazo-5-oxo-norleucine (1 g, 2.37 mmol) was added
to piperidine (20 ml) and the mixture stirred for 2 min. The
resulting solution was poured into ice-cold water (60 ml). The re-
action was filtered and the filtrate evaporated under high vacuum
at room temperature. The pale brown residue was crystallized from
a minimum amount of water by addition of methanol at ꢀ18 ꢁC to
give 210 mg (52%) of the desired product as a pale yellow solid. 1H
4.64 (2d, 1H, J¼2.2 Hz).
5.2.3. Synthesis of (S)-(þ)-N-tert-butoxycarbonyl-6-diazo-5-oxo-
norleucine ethyl ester. To a cold (ꢀ100 ꢁC) solution of trimethylsi-
lyldiazomethane (4.2 ml of a 2 M solution in hexane, 8.4 mmol) in
THF (40 ml) under an inert atmosphere was added n-butyllithium
in hexane (5.4 ml of a 1.6 M solution in hexane, 8.6 mmol). The
reaction was stirred for 30 min and transferred via a cannula to
a solution of (S)-(þ)-5-carbethoxy-N-tert-butoxycarbonyl-2-pyr-
rolidone (7 mmol) in THF (70 ml) at ꢀ105 ꢁC. The mixture was
stirred for a further 10 min and quenched by addition of the cold
solution to saturated ammonium chloride 200 ml. After extraction
with ethyl acetate, the residue obtained after evaporation of sol-
vents was purified by column chromatography on silica gel eluted
with dichloromethane/pentane/ether: 1/1/0.2. After evaporation of
solvents, the resulting yellow oil was dissolved in a minimum of
ether. The product recrystallizes as pale yellow plates (1 g, 50%). 1H
NMR (D2O, ppm)
d
2.08e2.15 (m, 2H, CH2C*), 2.52 (br t, 2H, J¼7 Hz,
CH2CO), 3.73 (t, 1H, J¼6.2 Hz, C*H), 5.87 (br s, 1H, CHN2).
5.4. Cyclopropanation of styrene using catalyst 1, 2, 3, 4 with
N2CHCO(CH2)2 CH (CO2Et) NHBoc (Scheme 3)
In a typical experiment 1
were placed in a Schlenk tube under argon, and dissolved in 400
of toluene. With the Fe-porphyrin catalysts 3 and 4, 2 mg (10 mol)
of cobaltocene was added for the reduction of Fe. The diazo com-
pound (0.2 mmol in 100 l of toluene) was then slowly added at
mmol of catalyst and 1 mmol of styrene
ml
m
NMR (CDCl3, ppm)
(m, 1H), 2.12e2.29 (m, 1H), 2.44 (br t, 2H), 4.16e4.30 (qþm, 2H,
J¼7.0 Hz), 5.20 (br d, 1H, J¼7.6 Hz), 5.31 (br s, 1H).
d
1.26 (t, 3H, J¼7.2 Hz), 1.46 (s, 9H), 2.07e1.92
m
room temperature. After 5 h of stirring, the cyclopropanes were
purified by column chromatography on silica gel (pentane/CH2Cl2/
ether: 4/5/1) to give a mixture of cis and trans-2-phenyl-1-5-oxo-N-
5.3. Preparation11 of N2CHCO(CH2)2CH(CO2H)NH2 (DON)
(Scheme 2)
Boc-L-norleucine ethyl ester cyclopropanes. The trans/cis diaster-
eoselectivity was determined by 1H NMR. The dimer product was
then recovered after elution with CH2Cl2.
5.3.1. Synthesis of (S)-(þ)-ethyl-N-Fmoc-pyroglutamate. To a solu-
tion of (S)-(þ)-ethyl-pyroglutamate (300 mg, 1.91 mmol) in THF
(10 ml) at ꢀ78 ꢁC was added LHDMS (1.81 ml of a 1 M solution in
THF, 1.81 mmol) slowly. The resultant pale yellow mixture was
stirred at ꢀ78 ꢁC for 15 min, and slowly transferred via a cannula to
a solution of Fmoc-Cl (2.47 g, 9.55 mmol) in THF (10 ml) at ꢀ78 ꢁC.
The reaction was allowed to stir at ꢀ78 ꢁC for 2 h, after which it was
allowed to rise to room temperature. After 14 h of stirring at room
temperature, the reaction was quenched by addition of saturated
NH4Cl (2 ml), and H2O (1 ml). The solution was extracted with ethyl
acetate and washed with brine. The organic phase was dried over
MgSO4 and concentrated under reduced pressure. The crude
product was purified by column chromatography on silica gel
The ee was determined by chiral HPLC analysis using a Chiralcel
OD column: n-hexane/iPrOH¼90/10, flow rate¼0.5 mL/min,
wavelength¼250 nm:
Cyclopropanation catalyzed by (ꢀ) 2: tr¼21.12 min for the minor
isomer, tr¼25.32 min for the major isomer, 79.50% ee, [
a
]
25 ꢀ168.5
D
(c 0.27, CH2Cl2).
Cyclopropanation catalyzed by (þ) 2: tr¼20.57 min for the major
isomer, 68.20% ee, tr¼25.70 min for the minor isomer, [
a
]
25 þ128 (c
D
0.27, CH2Cl2).
Cyclopropanation catalyzed by (ꢀ) 4: tr¼20.93 min for the mi-
25
nor isomer, tr¼24.79 min for the major isomer, 79.80% ee, [
ꢀ170 (c 0.35, CH2Cl2).
a]
D

P. Le Maux et al. / Tetrahedron 66 (2010) 4462e4468
4467
1H NMR (CDCl3, ppm), trans isomer:
d
1.29 (t, J¼7.1 Hz, 3H,
Aniline (0.2 mmol) and diazo (0.2 mmol) in 100
then slowly added at room temperature. After 5 h of stirring, the
insertion product was purified by column chromatography on silica
m
l of toluene were
CO2CH2CH3), 1.38e1.42 (m, 1H, CH2 cyclopropane), 1.45 (s, 9H, tert-
butyl), 1.67e1.72 (m, 1H, CH2 cyclopropane), 1.92e1.98 (m, 1H,
CH2C*), 2.17e2.22 (m, 2H, CH2C*þCHPh cyclopropane), 2.51e2.56
(m, 1H, CH cyclopropane), 2.70e2.76 (m, 2H, CH2CO), 4.21 (q,
J¼7.1 Hz, 2H, CO2CH2CH3), 4.27e4.32 (m, 1H, *CHCO2Et), 5.13 (br d,
J¼6.7 Hz, 1H, NH), 7.11 (d, J¼7.4 Hz, 2H, oPh), 7.24 (t, J¼6.7 Hz, 1H,
pPh), 7.31 (t, J¼7.4 Hz, 2H, mPh). 13C NMR (CDCl3, ppm) and HMQC
gel (pentane/CH2Cl2/ether: 2/7/1) to give N-5-oxo-N-Boc-
L-nor-
leucine ethyl ester phenyl amine. 1H NMR (CDCl3, 300 MHz)
d 1.31
(t, J¼7.0 Hz, 3H, CO2CH2CH3), 1.46 (s, 9H, tert-butyl), 1.93e2.03 (m,
1H, CH2C*), 2.22e2.31 (m, 1H, CH2C*), 2.61e2.71 (m, 2H, CH2CO),
4.03 (s, 2H, NHCH2CO), 4.23 (q, J¼7.1 Hz, 2H, CO2CH2CH3),
4.29e4.34 (m, 1H, *CHCO2Et), 4.60 (br s, 1H, NHPh), 5.15 (br d,
J¼7 Hz, 1H, NHCO), 6.62 (d, J¼8.4 Hz, 2H, oPh), 6.71 (t, J¼7.4 Hz, 1H,
pPh), 7.23 (t, J¼8.0 Hz, 2H, mPh). 13C NMR (CDCl3, 125 MHz) and
d
14.17 CH3 (CO2Et), 19.14 (CH2 cyclopropane), 28.60 (CH2C*), 28.31
(CH3 tert-butyl), 29.07 (CH cyclopropane), 32.36 (CH Ph), 39.69
(CH2CO), 53.02 (*CH), 61.48 CH2 (CO2Et), 79.92 (C tert-butyl), 126.03
(oPh), 126.56 (pPh), 128.50 (mPh), 140.22 (CipsoPh), 155.46 (CONH),
172.39 (CO2Et), 207.58 (C]O). HRMS (ESI): calcd for C21H29NO5
(MþNa)þ: 398.19434, found: 398.1943.
HMQC
d
14.13 (CH3 (CO2Et)), 27.65 (CH2C*), 28.30 (CH3 tert-butyl),
37.47 (CH2CO), 49.59 (CH2NH), 53.42 (*CH), 61.48 (CH2 (CO2Et)),
80.00 (C tert-butyl), 112.92 (oPh), 117.55 (pPh),129.34 (mPh), 146.62
(CipsoPh), 155.77(CONH), 171.91 (CO2Et), 206.16 (C]O). HRMS
(ESI): calcd for C19H28N2O5 (MþNa)þ: 387.1896, found: 387.1896.
5.5. SeH insertion with N2CHCO(CH2)2CH(CO2Et)NHBoc
catalyzed by Ru(CO)(TPP) (1) and Fe(Cl)(TPP) (3) (Scheme 4)
5.7. Dimerization of N2CHCO(CH2)2CH(CO2Et)NHBoc catalyzed
by Ru(CO)(TPP) (1) (Scheme 6)
In a typical experiment, 1
a Schlenk tube under argon, and dissolved in 400
With the Fe-porphyrin catalysts 3 and 4, 2 mg (10
tocene was added for the reduction of Fe. Thioanisole (0.2 mmol)
and diazo (0.2 mmol) in 100 l of toluene were then slowly added
m
mol of catalyst was placed in
l of toluene.
mol) of cobal-
m
m
Ru(CO)(TPP) (0.75 mg, 1
ene in a Schlenk tube under argon. Diazo (30 mg, 0.10 mmol) in
200 l of toluene was then added slowly at room temperature and
mmol) was dissolved in 500 ml of tolu-
m
m
at room temperature. After 15 h of stirring, the insertion product
was purified by column chromatography on silica gel (pentane/
CH2Cl2/ether:1/1/0.1).
stirred for 2 h. The dimerization product was purified by column
chromatography on silica gel (pentane/CH2Cl2/ether: 0.1/0.7/0.2) to
give 22 mg (yield¼81%) of the dimer compound. In the 1H NMR
spectrum, the proportions of the cis and trans dimer compounds
were, respectively 82 and 18%. The attribution of cis and trans
5.5.1. 5-Oxo-N-Boc-
L
-norleucine ethyl ester phenylsulfide. 1H NMR
isomer compounds was realized by HMBC (CDCl3, ppm)
d
(
(
13C)
13C)
(CDCl3, ppm)
d
1.30 (t, J¼7.0 Hz, 3H, CO2CH2CH3), 1.47 (s, 9H, tert-
butyl),1.81e1.94 (m,1H, CH2C*), 2.06e2.21 (m,1H, CH2C*),2.69e2.75
(m, 2H, CH2CO), 3.72 (s, 2H, SHCH2CO), 4.21 (q, J¼7.0 Hz, 2H,
CO2CH2CH3), 4.29e4.34 (m, 1H, *CHCO2Et), 5.12 (br d, J¼7.2 Hz, 1H,
NHCO),7.28e7.35(m, 5H, Ph). 13C NMR and HMQC (CDCl3, ppm)
201.42 and
198.89 and
NMR (CDCl3, 500 MHz)
d
(1H) 6.32 (JC1H2¼7.5 Hz, O]C1eCH1]CH2, cis),
d
d
(1H) 6.79 (JC1H2¼4.9 Hz, O]C1eCH1]CH2, trans). 1H
d
1.29 (t, J¼7.0 Hz, 3H, CO2CH2CH3), 1.45 (s,
9H, tert-butyl), 1.92e2.03 (m, 1H, CH2C*), 2.19e2.27 (m, 1H,
CH2C*),2.67 (dd, J¼8.7, 8.1 Hz, 2H, CH2CO), 4.22 (q, J¼7.1 Hz, 2H,
CO2CH2CH3), 4.25e4.31 (m, 1H, *CHCO2Et), 5.12 (br d, J¼7.2 Hz, 1H,
NHCO), 6.32 (s, 2H, cis CH]CH), 6.79 (s, 2H, trans CH]CH). 13C NMR
d
14.14 (CH3 (CO2Et)), 26.82(CH2C*), 28.29 (CH3 tert-butyl), 36.41
(CH2CO),43.95 (CH2S), 52.78 (*CH), 61.52 (CH2 (CO2Et)), 79.98 (C tert-
butyl), 128.77 (pPh), 129.16 (mPh), 129.52 (oPh), 134.70 (CipsoPh),
156.40 (CONH), 172.04 (CO2Et), 204.51 (C]O). HRMS (ESI): calcd for
C19H27NO5S (MþNa)þ: 404.1508, found: 404.1509.
(CDCl3, 125 MHz)
d
14.15 (CH3 (CO2Et), 26.45(CH2C*), 28.30 (CH3
tert-butyl), 38.35 (CH2CO)), 52.84 (*CH), 61.53 (CH2 (CO2Et)), 79.95
(C tert-butyl), 135.62 (CH]CH), 155.51(CONH), 172.25 (CO2Et),
201.42 (C]O). HRMS (ESI): calcd for C26H42N2O10 (MþNa)þ:
565.27372, found: 565.2733.
5.5.2. 5-Oxo-N-Boc-
(CDCl3, ppm)
L
-norleucine ethyl ester allyl sulfide. 1H NMR
d
1.32 (t, J¼7.0 Hz, 3H, CO2CH2CH3), 1.47 (s, 9H, tert-
butyl), 1.87e1.98 (m, 1H, CH2C*), 2.14e2.21 (m, 1H, CH2C*), 2.71e2.78
(m, 2H, CH2CO), 3.14 (d, J¼7.4 Hz, 2H, CH2S), 3.22 (s, 2H, SCH2CO), 4.23
(q, J¼7.2 Hz, 2H, CO2CH2CH3), 4.29e4.34 (m,1H, *CHCO2Et), 5.14 (br d,
J¼7.2 Hz, 1H, NHCO), 5.20 (m, 2H, CH2 allyl), 5.69e5.83 (m, 1H, CH
5.8. NeH insertion of (S)-(D)-6-diazo-5-oxo-norleucine
(DON) catalyzed by Ru(CO)(TPPS) (5) (Scheme 7)
allyl). 13C NMR (CDCl3, 125 MHz) and HMQC
d 14.15 (CH3 (CO2Et)),
A solution of the diazoketone DON (34.2 mg, 200
degazed water (0.5 ml) was added dropwise for 1 h to a solution of
Ru(CO)(TPPS) (1.2 mg, 1 mol) in 0.5 ml degazed water. After 5 h of
stirring, evaporation of the water gave 21 mg (73%) of the insertion
product as a solid. 1H NMR (D2O, ppm)
1.88e2.00 (m, 3H, CH2CH2),
mmol) in
26.75(CH2C*), 28.28 (CH3 tert-butyl), 34.65 (CH2S), 36.51 (CH2CO),
52.90(*CH),61.51(CH2 (CO2Et)), 79.96 (Ctert-butyl),118.49(CH2 allyl),
132.82 (CH allyl),156.40(CONH),172.31 (CO2Et), 204.73 (C]O). HRMS
(ESI): calcd for C16H27NO5S (MþNa)þ: 368.1508, found: 368.1511.
m
d
2.21e2.23 (m, 1H, CH2CH2), 3.07, 3.27 (2d, 2H, CH2), 3.67e3.71 (m,
1H, HC*), 3.88 (br s,1H, NH). HRMS (ESI): calcd for C6H8NO3 (MꢀH)ꢀ:
5.5.3. 5-Oxo-N-Boc-
NMR (CDCl3, ppm)
L
-norleucine ethyl ester ethan-2-ol sulfide. 1H
1.29 (t, J¼7.3 Hz, 3H, CO2CH2CH3), 1.45 (s, 9H,
142.05097, found: 142.0511. [a]
25 ꢀ24 (c 0.66, D2O).
d
D
tert-butyl), 1.79e1.89 (m, 1H, CH2C*), 2.15e2.22 (m, 1H, CH2C*),
2.74e2.82 (m, 2H, CH2CO), 2.72 (t, J¼5.8 Hz, 2H, CH2S), 3.31 (s, 2H,
SCH2CO), 3.75 (t, 2H, J¼5.7 Hz, CH2OH), 4.22 (q, J¼7.2 Hz, 2H,
CO2CH2CH3), 4.24e4.29 (m, 1H, *CHCO2Et), 5.16 (br d, J¼6.1 Hz, 1H,
NHCO). HRMS (ESI): calcd for C15H27NO6S (MþNa)þ: 372.14568,
found: 372.1456.
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