3-(Hydroxy(phenyl)methyl)azetidin-2-ones obtained via catalytic asymmetric hydrogenation or by biotransformation

Article abstract of DOI:10.1016/j.tetasy.2011.03.007

The catalytic asymmetric reduction of ethyl-2-(benzamidomethyl)-3-oxo- phenylpropanoate was realized with high enantiomeric and diastereoisomeric excesses via biotransformation using whole cells of different yeasts and asymmetric hydrogenation with Ru(II)

Full text of DOI:10.1016/j.tetasy.2011.03.007

Tetrahedron: Asymmetry 22 (2011) 597–602
Contents lists available at ScienceDirect
Tetrahedron: Asymmetry
journal homepage: www.elsevier.com/locate/tetasy
3-(Hydroxy(phenyl)methyl)azetidin-2-ones obtained via catalytic
asymmetric hydrogenation or by biotransformation
Isabella Rimoldi a, , Edoardo Cesarotti a, Daniele Zerla a, Francesco Molinari b, Domenico Albanese c,
⇑
Carlo Castellano d, Raffaella Gandolfi e
a Università degli Studi di Milano, Dipartimento di Chimica Inorganica, Metallorganica e Analitica ‘‘L. Malatesta’’ e Istituto CNR-ISTM, Via Venezian 21, 20133 Milano, Italy
b Università degli Studi di Milano, DISTAM-Sez. Microbiologia Industriale, Via Celoria 2, 20133 Milano, Italy
c Università degli Studi di Milano, Dipartimento di Chimica Organica e Industriale, Via Venezian 21, 20133 Milano, Italy
d Università degli Studi di Milano, Dipartimento di Chimica Strutturale e Stereochimica Inorganica, Via Venezian 21, 20133 Milano, Italy
e Università degli Studi di Milano, Dipartimento di Scienze Molecolari Applicate ai Biosistemi, sez. Chimica Organica ‘‘A. Marchesini’’, Via Venezian 21, 20133 Milano, Italy
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 14 February 2011
Accepted 18 March 2011
The catalytic asymmetric reduction of ethyl-2-(benzamidomethyl)-3-oxo-phenylpropanoate was real-
ized with high enantiomeric and diastereoisomeric excesses via biotransformation using whole cells of
different yeasts and asymmetric hydrogenation with Ru(II) complexes prepared from different chiral
diphosphine ligands.
With these combined approaches it was possible to prepare both enantiomers of the syn-stereoisomers
in almost enantiomerically pure form; one of the enantiomers of the anti-stereoisomer was obtained in
high ee with selected yeast while the other enantiomer of the anti was prepared in low ee and de. With
three of the four epimers we were able to prepare the corresponding azetidinones.
Ó 2011 Elsevier Ltd. All rights reserved.
1. Introduction
objective for academic and industrial research. Biocatalytical
methods have been used for the preparation of enantiomerically
Over the last few years, the importance of b-amino acids has-
increased since these molecules are interesting building blocks
for the preparation of unnatural peptides and bioactive com-
pounds, more specifically, b-hydroxy-b-amino acids have proven
to be versatile precursors for drug syntheses and effective methods
for their stereoselective preparation are crucial for the synthesis of
the corresponding azetin-2-ones and consequently of new types of
b-lactam rings.1 The carbapenem family (imipenem, meropenem,
ertapenem and other derivatives) show a wide range of activities
toward many pathogens, whether Gram-positive or negative; this
activity is combined with a good resistance and a poor affinity with
b-lactamases.2 Chemocatalysis and biocatalysis are complemen-
tary approaches for the stereoselective preparation of b-hydroxy-
b-amino acids starting from the corresponding b-keto-b-amino
acids.
pure b-hydroxy-b-amino acids by using microbial whole cells bear-
ing reductases/dehydrogenases.4
The role played by a substituent at the
a-position to the
carbonyl group5 prompted us to extend the investigation to sub-
strates that are able to behave as either an electron withdrawing
or electron-donor group. In order to evaluate the influence of
factors, such as the hindrance of the substituent, the stereoelec-
tronic properties of the molecules, and the presence of a keto-eno-
lic equilibrium especially in the
a-substituted b-ketoesters, we
have focused our research toward the reduction of ethyl-2-(benz-
amidomethyl)-3-oxo-phenylpropanoate, because in the literature,
although the synthesis of the 2-aminomethyl-3-hydroxy-3-phe-
nylpropanoic acid has been reported,6 it has not been obtained
by chemo or biocatalytic asymmetric reduction.
Herein we report the preparation and characterization of
3-hydroxy(phenyl)methyl azetidin-2-ones which, to the best of
our knowledge, have not been prepared and investigated before.
The reduction of the substrate, ethyl-2-(benzamidomethyl)-
3-oxo-phenyl propanoate 4, was expected to proceed with high
enantiomeric and diastereoisomeric excess, this is the reason as
to why we have studied, in detail, Ru(II) complexes with different
chiral chelating diphosphines and different types of yeasts to
obtain all of the epimers necessary for the synthesis of all the
possible stereoisomers of 3-hydroxy(phenyl)methyl azetidin-2-
ones 10.
Ruthenium [Ru(II)] in combination with chiral diphosphines has
played a central role in asymmetric hydrogenation; Saccharomyces
cerevisieae has also played the same role in the asymmetric reduc-
tion of keto groups.3
Asymmetric catalysis utilizing metal complexes has proven to
be a powerful tool for generating pure enantiomers and a major
⇑
Corresponding author. Tel.: +39 02 503 14609; fax: +39 02 503 14615.
E-mail address: isabella.rimoldi@unimi.it (I. Rimoldi).
0957-4166/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetasy.2011.03.007

598
I. Rimoldi et al. / Tetrahedron: Asymmetry 22 (2011) 597–602
Table 1
2. Results and discussion
Biotransformation of 4 by yeasts
We first chose a phenyl group as the substituents at the
a
-posi-
Entry
Catalyst
5 (%)
ee (%)
de (%)
tion of carbonyl group since good results were obtained in the
reduction of ethyl-3-oxo-3-phenyl propanoate either with yeasts
or with Ru(II)-chiral diphosphines complexes.7,8
1
2
Pichia glucozyma CBS 5766
Kluyveromyces marxianus CBS 1553
90
100
>99 (2R,3S)
>99 (2R,3R)
70
98
The synthesis of ethyl-2-(benzamidomethyl)-3-oxo-phenylpro-
panoate 4 was carried out in good yield via a procedure modified to
that developed by Takasago Co. for the synthesis of ethyl 2-(benz-
amidomethyl)-3-oxobutanoatoate (Scheme 1).
Therefore, biocatalytic reduction gave access to enantiomerically
pure (2R,3R)-5 and (2R,3S)-5.
It is noteworthy that during the process of screening and
optimization, it was observed that a few yeasts also gave N-(3-
oxo-3-phenylpropyl)-benzamide 7 and N-(3-hydroxy-3-phenyl-
propyl)-benzamide 8 as by-products (Scheme 2).5b
O
O
O
PhCOCH2COOEt
3
When P. glucozyma CBS 5766 was used without any co-sub-
strate 7 was obtained in high yield (85–90%) and could be used
to give (S)-8 with ee >99%. Compound (S)-8 is an interesting build-
ing block in the synthesis of fluoxetine, an active principle of one of
the most diffuse antidepressive drugs.9 The formation of 7 is
caused by the spontaneous decarboxylation of the intermediate
carboxylic acid 6 produced by hydrolytic enzymes occurring in
the whole cells. Compound 7 can then be enantioselectively re-
duced by dehydrogenase(s) to (S)-8. An alternative synthesis was
carried out to confirm the structures of by-products 7 and 8.
Ketone 7 was obtained by hydrolysis of 4 catalyzed by lipase from
Candida cylindraceae and racemic alcohol 8 was obtained by
reduction with NaBH4. Compound 8 had the same configuration
of the product obtained by asymmetric hydrogenation with
[RuCl2(DMF)n(ꢀ)-tetraMe-BITIOP]; the absolute configuration was
assigned as (S) by comparison with the enantioselectivity shown
by the same catalyst on ethyl-3-oxo-3-phenyl propanoate.10
The biocatalytic reduction gave the target molecule 5 almost
enantiomerically pure; however for a detailed study of the biolog-
ical activity of the azetidinone, it was necessary to have all
epimers.
Ph
N
H
X
Ph
OEt
NHCOPh
NaH, THF, 0 ºC
4
1, X = OH
2, X = Cl
SOCl2,
CH2Cl2
Scheme 1. Synthesis of ethyl-2-(benzamidomethyl)-3-oxo-phenylpropanoate 4.
The reduction of 4 was performed using biocatalytic (whole
cells of yeasts) and chemical methods (Ru(II)/phosphine com-
plexes). A screen for the modification of 4 was carried out using
the same library of yeasts previously used for the reduction of
ethyl 2-(benzamidomethyl)-3-oxobutanoate.5b
The reduction of 4 into ethyl-2-(benzamidomethyl)-3-hydroxy-
phenylpropanoate 5 with good yields was obtained with Kluyver-
omyces marxianus CBS 1553 and Pichia glucozyma CBS 5766. We ob-
served that only two yeasts (from a screen performed using 20
yeasts) gave good yields in the reduction of 4, while twelve yeasts
were able to reduce ethyl 2-(benzamidomethyl)-3-oxobutanoate;
this is probably due to the additional steric hindrance caused by
the N-(methyl)benzamide substituent, which causes mismatching
The asymmetric reduction of the C@O bond has been realized in
many ways, but the most versatile catalysts remain those based on
Ru(II) complexes.11,12 We investigated the ligands described in Fig-
ure 1. Some of them are commercially available, such as BDPP, BIN-
AP and Tol-BINAP,13 while others have been developed in our
laboratory.5a,10,14 Some of these diphosphines are characterized
by the presence of a stereogenic sp3 carbon (Zedphos, XilylZed-
phos, Prolophos, BDPP); others by atropoisomeric chirality
(tetraMe-BITIOP, BINAP, Tol-BINAP and DIOPHEP), or by a contem-
porary presence of both elements of chirality (Isaphos).
between the two substituents at the
a-position to the carbonyl
group. The biotransformations with K. marxianus and P. glucozyma
were optimized, following a strategy described in a previous work;
the results are shown in Table 1. The type (glucose) and concentra-
tion (50 mg/mL, 280 mM) of co-substrate added for cofactor regen-
eration played a crucial role with regard to the high yields and
stereoselectivity.
K. marxianus CBS 1553 gave (2R,3R)-5 quantitatively and in
enantiomerically and diastereoisomerically pure form (entry 2),
while P. glucozyma CBS 5766 furnished the enriched diastereoiso-
mer (2R,3S)-5 (entry 1). It was possible to separate the pure stereo-
isomer (2R,3S)-5 from (2S,3S)-5 by flash chromatography.
Most ligands, i.e. those characterized by atropoisomeric chiral-
ity and those with sp3 stereogenic carbons (Table 2, entries
OH
O
O
NH
5
O
O
O
O
O
O
OH
O
NH
OH
NH
O
NH
NH
4
O
O
O
6
8
7
Scheme 2. Asymmetric reduction of 4.

I. Rimoldi et al. / Tetrahedron: Asymmetry 22 (2011) 597–602
599
OR'
PAr2
PAr2
N
R
(2R,5R)-Zedphos Ar = C6H5
(2R,5R)-XilylZedphos Ar = (3,5-(CH3)2)C6H3
(R)-Prolophos
R = R' = PPh2
O
P
O
PPh2 PPh2
(R,Sax)-Isaphos C1
R =
(S,S)-BDPP
R' = PPh2
CH3
S
PPh2
PPh2
H3C
H3C
H
H
PAr2
PAr2
O
O
O
O
PPh2
PPh2
S
CH3
Ar = Ph
(R)-BINAP
(S,S,Sax)-DIOPHEP
(-)-TetraMe-Bitiop
Ar = 4-Me-Ph (R)-Tol-BINAP
Figure 1. Chiral chelating diphosphines used in asymmetric hydrogenation of 4.
cyclization of the amino acid 9 thus obtained (Scheme 3). The
hydrolysis was carried out by heating at phenylpropanoate 5
90 °C in the presence of 10% HCl. After aqueous work-up, the b-
amino acid (2R,3R)-9 was isolated by neutralization of the hydro-
chloride salt with Et3N. The cyclization of the b-amino acid
(2R,3R)-9 to azetidinone (R,R)-10 occurred smoothly by heating
at 65 °C in the presence of 2,20-dipyridyl disulfide and PPh3 in ace-
tonitrile at 65 °C. The same procedure was used to generate azetid-
inone (S,S)-10 from (S,S)-5.
Table 2
Asymmetric hydrogenation of 4a
Entry
Ligands
eeb (%)
de (%)
1
2
3
4
5
6
7
8
(S)-BINAP
(R)-BINAP
(S)-Tol-BINAP
(ꢀ)-TetraMe-BITIOP
(+)-TetraMe-BITIOP
(S,S,Sax)-DIOPHEP
(R,R)-BDPP
35 (2S,3S)
32 (2R,3R)
58 (2S,3S)
>99(2R,3R)
>99(2S,3S)
67 (2S,3S)
50 (2R,3R)
67 (2S,3S)
68 (2R,3R)
63 (2R,3R)
65 (2R,3R)
80 (2S,3R)
70 (2S,3R)
90
93
94
97
95
85
80
88
80
84
52
50
62
(S,S)-BDPP
9
(2R,5R)-Zedphos
(2R,5R)-Xylil Zedphos
(R)-Prolophosc
(R,Rax)-Isaphos C1
(R,Sax)-Isaphos C1
10
11
12
13
c
c
a
Reaction conditions: solvent: MeOH/CH2Cl2 = 50:50; reaction time 60 h; con-
version >99%; substrate/[RuCl2(DMF)n(PP⁄)] = 100:1; substrate concentra-
tion = 0.030 M; T = 60 °C; P = 50 atm H2. The ee and de values were determined by
chiral HPLC with a Daicel Chiralpak AD (90:10 = hexane/isopropanol, flow = 0.6 mL/
min).
b
Stereochemistry of the prevailing diastereoisomer.
Conversion = 80%.
c
Scheme 3. Synthetic elaboration of phenylpropanoates to azetidinones. Reagents
and conditions: (a) 10% HCl, 90 °C; (b) Et3N, CH3CN; (c) PPh3, PySSPy, CH3CN, 65 °C,
6 h; (d) NaH, 4-Br–C6H4–CH2Br, THF–DMF, 25 °C, 20 h.
1–11) essentially only gave the syn diastereoisomers. However,
good results were obtained with (+)- and (ꢀ)-tetraMe-BITIOP.
The Ru(II) complex with (ꢀ)-tetraMe-BITIOP afforded (2R,3R)-
ethyl-2-(benzamidomethyl)-3-hydroxy-phenylpropanoate 5, the
same enantiomer obtained with K. marxianus, in 97% de and almost
enantiomerically pure (entry 4); (+)-tetraMe-BITIOP afforded the
opposite enantiomer (2S,3S)-5 (entry 5).
The ligands with mixed chirality (R,Rax)-Isaphos C1 and (R,Sax)-
Isaphos C1 mainly gave the anti-diastereoisomers (entries 12 and
13). The prevailing enantiomer was (2S,3R)-5 in both cases, the
opposite enantiomer obtained by P. glucozyma; these results
showed the predominant role of the sp3 carbon atom, which con-
trolled the whole stereochemistry of the catalytic reaction.
Phenylpropanoate (2R,3R)-5 was transformed into phenyl-
substituted azetidinones 10 via hydrolysis of 5 followed by
On the other hand, when (2R,3S)-5 was subjected to the same
protocol, azetidinone (R,R)-10 was isolated as the main product
along with minor amounts of the expected (R,S)-10. This behavior
could be tentatively explained by the better stability of the syn-
amino acid (2R,3R)-9 with respect to the anti-(2R,3S)-9.
The absolute configuration was determined by single-crystal
structure determination of N-(p-bromobenzyl)-11 which was syn-
thesized by N-alkylation of azetidinone (R,R)-10 with p-bromoben-
zyl bromide (Fig. 2). The introduction of a bromine atom inserted
into the molecule an anomalous X-ray scatterer in order to
determine the absolute configuration.

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I. Rimoldi et al. / Tetrahedron: Asymmetry 22 (2011) 597–602
6.73 (bs, 1H), 7.42–7.62 (m, aromatic), 7.71 (dd, aromatic, Jo = 1.5,
Jm = 8.0), 8.10 (dd, aromatic, Jo = 2.2, Jm = 5.1); 13C-NMR(CDCl3):
d = 14.1 (CH3), 39.2 (CH2–N), 53.7 (CH), 62.03 (CH2), 127.1–135.9
(CH aromatic), 167.9 (C@O isomer), 169.2 (C@O isomer), 194.8
(C@O aromatic). HRMS of C19H19NO4 (m/z): calcd 325,13, found
348.5 (MNa+).
4.2. General procedure for the asymmetric hydrogenation
In a Schlenk tube sealed under argon, substrate 4 was added to
the precatalyst followed by 20 mL of a choice solvent, the solution
was stirred for 30 min and then transferred to an autoclave with a
cannula.
Figure 2. A perspective view of the (R)-1-(4-bromobenzyl)-3-((R)-hydroxy-
(phenyl)methyl)azetidin-2-one 11.
The stainless steel autoclave (200 mL), equipped with tempera-
ture control and a magnetic stirrer, was purged five times with
hydrogen, after the transfer of the reaction mixture, the autoclave
was pressurized. At the end the autoclave was vented and the mix-
ture was analyzed by GC–MS, NMR spectra and HPLC.
3. Conclusion
The complementary use of biocatalysis with dehydrogenases
present in whole cells of yeasts and organometallic catalysis with
transition metal catalysts allowed for the preparation of three of
the four epimers of ethyl-2-(benzamidomethyl)-3-hydroxy-phe-
nylpropanoate 5. Compound Ru(II)-(+)tetraMe-BITIOP gave
(2S,3S)-5, Ru(II)-(ꢀ)tetraMe-BITIOP, K. marxianus CBS 1553 gave
(2R,3R)-5 and P. glucozyma CBS 5766 gave (2R,3S)-5. The phosphine
ligands bearing an axial chirality and a stereogenic sp3 carbon gave
(2S,3R)-5 but with lower stereoselectivity. The three epimers ob-
tained as enantiomerically pure molecules were transformed into
the corresponding azetininones.
4.2.1. Ethyl 2-(benzamidomethyl)-3-hydroxy-3-
phenylpropanoate syn-5
1H NMR (CDCl3): d = 1.01 (t, 3H, J = 7.0 Hz), 2.93–3.01 (m, 1H,
syn diastereoisomers), 3.61–3.69 (m, 2H), 3.98 (q, 2H, J = 7.0 Hz),
4.12–4.19 (m, 2H), 4.96 (d, 1H, J = 7.3 Hz), 6.72 (br s, 1H), 7.29–
7.53 (m, aromatic), 7.77 (dd, aromatic, Jo = 1.5 Hz, Jm = 8.0 Hz);
13C NMR (CDCl3): d = 14.0 (CH3), 37.9 (CH2–N, isomer), 53.0 (CH),
61.1 (CH2), 72.6 (CH), 126.4–132.0 (CH aromatic), 173.5 (C@O
amide). HRMS of C19H21NO4 (m/z): calcd 327,15, found 350.4
4. Experimental
(MNa+).
½
a 2D0
ꢃ
ð2R; 3RÞ ¼ þ33:2 (c 0.15, CHCl3);
½
a 2D0
ꢃ
ð2S; 3SÞ ¼
ꢀ11:0 (c 0.18, CHCl3); HPLC data: CHIRALPAK AD, eluent:
hexane/2-propanol=90:10, flow = 0.6 mL/min, k = 230 nm, rt:
(2R,3R) = 35.1 min, (2S,3S) = 37.1 min.
Catalytic reactions were performed in a 200 mL stainless steel
autoclave equipped with temperature control and a magnetic stir-
rer. Unless otherwise stated, materials were obtained from com-
mercial suppliers and used without further purification. The
ruthenium catalysts are prepared according to the well established
literature procedure.15 Compounds 1 and 2 are prepared as re-
ported in the literature.16
4.3. Microorganisms: culture conditions
Strains from official collections or from our own collection
(Microbiologia Industriale Milano) were routinely maintained on
malt extract (8 mg/mL, agar 15 mg/mL, pH 5.5). To obtain cells
for biocatalytic activity tests, the microorganisms were cultured
in 500 mL Erlenmeyer flasks containing 100 mL of medium and
incubated for 48 h at 28 °C on a reciprocal shaker (100 spm). The
yeasts were grown on malt extract with 5 mg/mL Difco yeast ex-
tract. Fresh cells from submerged cultures were centrifuged
(5000 rpm per 10 min) and washed with 0.1 M phosphate buffer
pH 7 prior the use.
1H NMR and 13C NMR spectra were recorded on a Varian
200 MHz, a Bruker DRX Avance 300 MHz equipped with a non-re-
verse probe and also or on a Bruker DRX Avance 400 MHz. GC-MS
spectra were recorded on Thermo Finningan MD 800 equipped
with GC Trace (SE 52 column: length 25 m, / int. 0.32 mm, film
0.4–0.45 lm) and MS analyses were performed by using a Thermo
Finnigan (MA, USA) LCQ Advantage system MS spectrometer with
an electrospray ionization source and an ‘Ion Trap’ mass analyzer.
The MS spectra were obtained by direct infusion of a sample solu-
tion in a mixture MeOH/H2O/AcOH 10:89:1 under ionization, ESI
positive. HPLC analysis: Merck-Hitachi L-7100 equipped with
Detector UV6000LP and Daicel Chiralcel OD or Chiralpak AD.
4.4. Bioreduction conditions
General procedure for the screening: reductions were carried
out in 10 mL screw-capped test tubes with a reaction volume of
3 mL. The cells (20 mg/mL, dry weight) suspended in 0.1 M phos-
phate buffer pH 7 with or without 5% of glucose and 1 mg/mL of
ethyl 2-(benzamidomethyl)-3-oxo-phenylpropanoate 4 as sub-
strate. The substrate 4, dissolved in DMSO, was added to the
biotransformation system to give 1 mg/mL of substrate concentra-
tion and 2% of solvent. The reactions were carried out at 28 °C
under magnetic stirring.
The production of (2S,3R)-5 on a semi-preparative scale was
carried out with P. glucozyma CBS 5766. The desired amount of
cells was suspended in different 0.1 M phosphate buffers contain-
ing glucose and 1.3 g of substrate was added to reach the desired
concentration (2 mg/mL); the suspensions obtained were magnet-
ically stirred at 28 °C.
4.1. Preparation of ethyl-2-(benzamidomethyl)-3-oxo-phenyl-
propanoate 4
To a solution of NaH (250 mg, 1.04 ꢁ 10ꢀ2 mol), ethyl-3-oxo-3-
phenyl propanoate 3 was added in anhydrous THF (50 mL) at 0 °C.
The reaction mixture was stirred for 15 min, after which N-(chloro-
methyl)benzamide 2 in THF at 0 °C was added. The reaction mix-
ture was stirred for 12 h while the temperature of the reaction
was allowed to return to room temperature, then treated with Na2-
SO4ꢂ10H2O and the reaction mixture stirred for 30 min. The mix-
ture was filtered and concentrated in vacuo. Purification of the
residue by flash chromatography on silica gel with cyclohexane/
ethyl acetate (30:70) as eluents, gave the title compound (2.77 g,
82%) as a white oil. When the compound was stored in the freezer,
it solidified. 1H NMR(CDCl3): d = 1.18 (t, 3H, J = 6.9), 3.93–3.97 (m,
2H), 4.09–4.16 (m, 2H), 4.19 (q, 2H, J = 6.9), 4.87 (dd, 1H, J =5.5),
After 48 h, the biotransformation was stopped by centrifugation
of the cellular suspension; the liquid fraction was extracted with
ethyl acetate (3 ꢁ 300 mL). The organic extracts were dried over

I. Rimoldi et al. / Tetrahedron: Asymmetry 22 (2011) 597–602
601
Na2SO4 and the solvent was removed under reduced pressure.
After flash chromatography on silica gel eluted with cyclohexane/
ethyl acetate (3:7), 751 mg of pure products were obtained.
37.1 (CH2), 58.5 (CH), 69.4 (CH), 125.6 (CH), 126.4 (CH), 127.8
(CH), 141.7 (C), 169.5 (C).
4.4.5. (S)-3-((S)-Hydroxy(phenyl)methyl)azetidin-2-one 10
The title compound has been obtained in 44% overall yield from
(S,S)-5 by following the same procedure described for (R,R)-10.
NMR data identical with those reported above for (R,R)-10.
4.4.1. Ethyl 2-(benzamidomethyl)-3-hydroxy-3-phenylpropan-
oate (2R,3S)-5
1H NMR (CDCl3): d = 1.01 (t, 3H, J = 7.0 Hz), 3.15–3.24 (m, 1H,
anti diastereoisomers), 3.61–3.69 (m, 2H), 3.98 (q, 2H), 4.12–4.18
(m, 2H), 4.95 (d, 1H, J = 7.3 Hz), 6.72 (br s, 1H), 7.29–7.53 (m, aro-
matic), 7.77 (dd, aromatic, Jo = 1.5 Hz, Jm = 8.0 Hz); 13C NMR
(CDCl3): d = 14.0 (CH3), 38.0 (CH2–N), 53.0 (CH), 61.1 (CH2), 72.6
(CH), 126.4–132.0 (CH aromatic), 173.5 (C@O amide). HRMS of
½
a 2D0
ꢃ
¼ ꢀ128:1 (c 0.6, CHCl3).
4.4.6. Attempted synthesis of (R)-3-((S)-hydroxy(phenyl)methyl)
azetidin-2-one 10
When ester (2R,3S)-5 was subjected to the same protocol de-
scribed above for the conversion of (R,R)-5 to the corresponding
azetidinone, unexpectedly (R,R)-10 was isolated in 28% overall
yield along with a 3.4:1 mixture (12% overall yield) of the title
compound with (R,R)-10 (as determined by integration of the ben-
zylic proton 1H NMR (CDCl3): (R,R)-10, 5.24 (d, 1H, J = 3.7 Hz);
(R,S)-10, 5.00 (d, 1H, J = 7.6 Hz). HPLC data: CHIRALCEL OD, eluent:
hexane/2-propanol = 90:10, flow = 0.8 mL/min, k = 215 nm, rt:
(2S,3S) = 21.5 min, (2R,3R) = 27.4 min, (2R,3S) = 30.3 min.
C
½
19H21NO4 (m/z): calcd 327,15, found 350.4 (MNa+).
a 2D0
ꢃ
ð2R; 3SÞ ¼ ꢀ11:3 (c 0.12, CHCl3). HPLC data: CHIRALPAK AD,
eluent: hexane/2-propanol = 90:10, flow=0.6 mL/min, k = 230 nm,
rt: (2S,3R) = 49.8 min, (2R,3S) = 68.8 min.
4.4.2. N-(3-Oxo-3-phenylpropyl)benzamide 7
N-(3-Oxo-3-phenylpropyl)benzamide 7 was obtained by bio-
transformation. At first, 2 mg/mL of ethyl-2-(benzamidomethyl)-
3-oxo-phenylpropanoate 4 and 10 mg/mL of lipase from Candida
cylindracea were added to a 0.1 M phosphate buffer pH 7. The bio-
transformation was carried out at 30 °C with magnetic stirring.
After 72 h, the reaction was extracted three times with ethyl ace-
tate. The collected organic phases were dried over Na2SO4 and re-
duced under vacuum. The crude extract was purified with
preparative TLC on silica gel (Kiesel 60 with fluorescent indicator)
1H NMR (CDCl3): d = 3.35 (t, 2H, J = 5.5 Hz), 3.89 (q, 2H), 6.86 (br s,
1H), 7.36–7.63 (m, aromatic), 7.76 (dd, aromatic, Jo = 1.5 Hz,
Jm = 8.0 Hz), 7.97 (dd, aromatic, Jo = 2.2 Hz, Jm = 5.1 Hz); 13C NMR
(CDCl3): d = 35.1 (CH2), 38.4 (CH2,), 127.1–136.8 (CH aromatic),
167.5 (C@O carbonyl), 199.9 (C@O, amide). HRMS of C16H15NO2
(m/z): calcd 253,11, found 253.0 (M+).
4.4.7. (R)-1-(4-bromobenzyl)-3-((R)-
hydroxy(phenyl)methyl)azetidin-2-one 11
At first, 50% NaH (0.022 mmol, 11 mg) was added to a stirred
solution of azetidinone (R,R)-10a (0.02 mmol, 37 mg) in a THF/
DMF 6:1 mixture (3.5 mL) under a nitrogen atmosphere and at
0 °C. After 5 min, 4-bromobenzyl bromide (0.022 mmol, 55 mg)
was added and the temperature was allowed to reach room tem-
perature. After 20 h a few drops of saturated aqueous NH4Cl were
added and the solvent evaporated at reduced pressure. The residue
was dissolved in CH2Cl2 (5 mL) and washed with water (1 mL). The
organic phase was dried over Na2SO4, filtered and evaporated to
dryness. The residue was purified by column chromatography (elu-
ant AcOEt/PE 6:4) to give 35 mg of the title compound, yield 51%
along with 13 mg of the azetidinone (R,R)-10a. Crystals for X-ray
diffraction were obtained by crystallization with Et2O. (R,R)-
11—1H NMR (CDCl3): d = 7.44 (d, 2H, J = 8.2 Hz), 7.33 (s, 5H), 7.07
(d, 2H, J = 8.0 Hz), 5.23 (s, 1H), 4.46 (d, 1H, J = 5.6 Hz), 4.22 (d,
1H, J = 5.6 Hz), 3.59 (br s, 1H), 3.31 (m, 1H), 3.04 (t, 1H,
J = 5.3 Hz). 13C NMR 40.3 (CH2), 45.4 (CH2), 56.9 (CH), 69.9 (CH),
122.1 (C), 125.6 (CH), 127.9 (CH), 128.6 (CH), 129.7 (CH), 131.9
(CH), 134.5 (C), 141.5 (C), 168.1(C).
4.4.3. N-(3-Hydroxy-3-phenylpropyl)benzamide 8
The N-(3-oxo-3-phenylpropyl)benzamide 7 was reduced by
NaBH4 (1:5) in ethanol. The reaction was maintained at room tem-
perature with magnetic stirring. After 12 h, water and CH2Cl2 were
added to give product 8. The organic extract was dried over Na2SO4
and concentrated in vacuum. The product was purified via prepara-
tive TLC. 1H NMR (CDCl3): d = 1.00 (q, 2H), 3.21 (br s, 1H), 3.42–3.54
(m, 1H), 3.81–3.95 (m, 1H), 4.80–4.88 (m, 1H), 6.78 (br s, 1H),
7.31–7.55 (m, aromatic), 7.76 (dd, aromatic Jo = 1.5 Hz, Jm = 8.0 Hz).
13C NMR (CDCl3): d = 37.7 (CH2), 38.8 (CH2), 73.1 (CH), 125.8–144.3
(CH aromatic), 168.1 (C@O amide). HRMS of C16H17NO2 (m/z):
calcd 255.13, found 237.0 (M+ꢀ18).
4.4.8. X-ray structure determination of (R)-1-(4-bromobenzyl)-
3-((R)-hydroxy(phenyl)methyl)azetidin-2-one 11
Crystal data.
C17H16BrNO2, M = 346.22, orthorhombic, a =
6.052(1), b = 8.895(2), c = 28.428(6) Å, U = 1530.4(5) Å3, T =
4.4.4. (R)-3-((R)-Hydroxy(phenyl)methyl)azetidin-2-one 10
A mixture of (R,R)-5 (7.12 mmol, 2.53 g) and 10% HCl (12 mL)
was heated at 90 °C for 24 h. After cooling to rt, AcOEt (15 mL)
was added and the organic phase separated. The aqueous
phase was evaporated to dryness to give 1.08 g of solid 9ꢂHCl.
The latter (6 mmol, 1.49 g) was suspended in CH3CN (20 mL) and
Et3N (6.4 mmol, 0.9 mL) was added dropwise. After stirring at rt
for 24 h, the white precipitate was filtered and washed with cold
CH3CN. The amino acid (R,R)-9 thus obtained (5 mmol, 0.98 g)
was suspended in CH3CN (100 mL), and PPh3 (6 mmol, 1.57 g)
and dipyridyl disulfide (6 mmol, 1.32 g) were added sequentially.
After heating at 72 °C for 20 h, the crude reaction mixture was
directly purified by column chromatography (Kieselgel Merck
Typ 9385–400 mesh, 60 Å; eluant CH2Cl2/MeOH 96:4) to give
0.58 g of the title compound, 65% yield, dr (NMR) 93:7. A diastereo-
isomerically pure compound was obtained through crystallization
294(2) K, space group P212121 (no. 61), Z = 4, l = (Mo-Ka)
2.690 mmꢀ1. 8889 reflections (3884 unique) were collected at
room temperature in the range 2.40 6 2h 6 29.55°, employing a
0.35 ꢁ 0.09 ꢁ 0.04 mm crystal mounted on a Bruker Apex II CCD
diffractometer. 2291 reflections (Rint = 0.0267), final R1 [wR2] val-
ues of 0.0314 [0.0667] on I > 2r(I) [all data], with GoF = 0.824 for
254 parameters.
Intensities were corrected for Lorentz-polarization effects and
empirical absorption correction (SADABS).17 The structure was solved
by direct methods (SIR-97)18 and refined on F2o with the SHELXL-9719
program (WINGX suite20) All the non-hydrogen atoms were refined
with anisotropic thermal parameters, while hydrogen atoms, lo-
cated on the
DF maps, were treated isotropically.
The choice of the correct enantiomer was confirmed by the
value of the Flack parameter [ꢀ0.010(8)]. The refinement of the
wrong enantiomer resulted in a R index of 0.0628, significantly
higher than that of the correct one.
Crystallographic data (excluding structure factors) have been
deposited with the Cambridge Crystallographic Data Centre as
(CH2Cl2–hexane). Mp 145 °C, ½a D20
ꢃ
¼ þ129:0 (c 0.6, CHCl3). 1H NMR
(CDCl3): d = 7.38–7.26 (m, 5H), 5.66 (br s, 1H), 5.24 (d, 1H,
J = 3.7 Hz), 3.63 (m, 1H), 3.50 (m, 1H), 3.24 (m, 1H). 13C NMR

602
I. Rimoldi et al. / Tetrahedron: Asymmetry 22 (2011) 597–602
7. (a) Ramos, A. S.; Ribeiro, J. B.; Vazquez, L.; Fiaux, S. B.; Leite, S. G. F.; Ramos, M.
C. K. V.; Aquino Neto, F. R.; Antunes, O. A. C. Tetrahedron Lett. 2009, 50, 7362–
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supplementary publication No. CCDC-785072. These data can be
obtained free of charge via www.ccdc.cam.ac.uk/conts/retriev-
ing.html (or from CCDC, 12 Union Road, Cambridge CB2 1EZ, UK;
fax: +44 1223 336033; e-mail: deposit@ccdc.cam.ac.uk.
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