N-Methylthio β-lactam antibacterials: Effects of the C 3/C4 ring substituents on anti-MRSA activity

Article abstract of DOI:10.1016/j.bmc.2005.08.011

N-Thiolated β-lactams are a new family of antibacterials that inhibit the growth of Staphylococcus bacteria. Unlike other β-lactam drugs, these compounds retain their full antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA) strains and operate through a different mode of action. The structural features, which give these lactams their biological activity, have not yet been completely defined. Earlier efforts in our laboratory established that the N-organothio substituent is essential for antimicrobial activity while other groups at C₃ and C₄ on the lactam ring play a more subtle role. In this present study, we investigate these effects by varying the polar and steric nature of the ring substituents at these two centers. From the data presented herein, it appears that there is a need to balance the lipophilic character of the C₃/C₄ groups to obtain an optimal anti-MRSA activity. The structure-bioactivity profiles more closely relate to the compound's ability to penetrate the bacterial cell membrane to sites of action within the cytoplasm rather than to any specific non-bonding interactions with a biological target. Based on these results, a model for the compounds' mode of action is presented.

Full text of DOI:10.1016/j.bmc.2005.08.011

Bioorganic & Medicinal Chemistry 13 (2005) 6289–6308
N-Methylthio b-lactam antibacterials: Effects of the C3/C4
ring substituents on anti-MRSA activity
Edward Turos,a,* Cristina Coates,a Jeung-Yeop Shim,a Yang Wang,a J. Michelle Leslie,a
Timothy E. Long,a G. Suresh Kumar Reddy,a Alex Ortiz,a,ꢀ Marci Culbreath,a,ꢀ
Sonja Dickey,b Daniel V. Lim,b Eduardo Alonsoc and Javier Gonzalezc
aDepartment of Chemistry, 4202 East Fowler Avenue, SCA 400, University of South Florida, Tampa, FL 33620, USA
bDepartment of Biology, University of South Florida, Tampa, FL 33620, USA
cDepartamento de Quimica Organica, Universidad de Oviedo, 37006 Oviedo, Spain
Received 7 December 2004; revised 31 March 2005; accepted 31 March 2005
Available online 26 September 2005
Abstract—N-Thiolated b-lactams are a new family of antibacterials that inhibit the growth of Staphylococcus bacteria. Unlike other
b-lactam drugs, these compounds retain their full antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA)
strains and operate through a different mode of action. The structural features, which give these lactams their biological activity,
have not yet been completely defined. Earlier efforts in our laboratory established that the N-organothio substituent is essential
for antimicrobial activity while other groups at C3 and C4 on the lactam ring play a more subtle role. In this present study, we inves-
tigate these effects by varying the polar and steric nature of the ring substituents at these two centers. From the data presented here-
in, it appears that there is a need to balance the lipophilic character of the C3/C4 groups to obtain an optimal anti-MRSA activity.
The structure–bioactivity profiles more closely relate to the compoundꢁs ability to penetrate the bacterial cell membrane to sites of
action within the cytoplasm rather than to any specific non-bonding interactions with a biological target. Based on these results, a
model for the compoundsꢁ mode of action is presented.
ꢀ 2005 Elsevier Ltd. All rights reserved.
1. Introduction
and some structurally similar analogues inhibit bacterial
growth through a mode of action that is distinctly differ-
ent from those of penicillin and other bioactive b-lac-
tams.3 These b-lactams exhibit bacteriostatic activity in
certain bacteria including S. aureus (Gram-positive)
and Neisseria gonorrhoeae (Gram-negative), and no
detectable cytotoxic effects in normal mammalian fibro-
blast cells.4 Being devoid of a hydrophilic ring function-
ality, these N-methylthio b-lactams are completely
impervious to b-lactamase cleavage, enabling them to
operate at full strength as bacteriostatic agents under
conditions that render many of the familiar b-lactam
drugs ineffective. The structural features, which impart
the molecules their antibacterial properties and resis-
tance to b-lactam hydrolysis, have not yet been assessed
in detail. We have, however, established that the N-thio
substituent is required for an antibacterial activity.5 The
substituents at C3 and C4 of the lactam ring also appear
to play a secondary role in the biological properties. One
of the first issues we addressed in the case of lactam 2 is
whether bioactivation by an electrophilic species is
required for antibacterial activity. This was considered
a possibility because in organic media, electrophilic
Recently, our laboratories have identified a new family
of N-methylthio-substituted b-lactams 1 that possess
promising antibacterial properties. Curiously, most of
this activity is directed toward Staphylococcus bacteria,
including methicillin-resistant strains of Staphylococcus
aureus (MRSA).1 The initial lead compound in these
studies, N-methylthio b-lactam 2, was first discovered
in our laboratories during routine biological screening.2
Its potent antibacterial nature was totally unexpected
since no other b-lactam compound lacking an ionizable
or acidic residue on or near the lactam nitrogen has ever
been reported to have antibacterial activity. In fact, the
structural features of lactam 2 are entirely unprecedent-
ed for a b-lactam antibiotic, with only lipophilic substit-
uents occupying positions on the four-membered ring.
Our more recent studies have indicated that lactam 2
Keywords: N-Thiolated b-lactams; MRSA; SAR; Antibiotics.
*
Corresponding author. Tel.: +1 813 974 1733; fax: +1 813 974 173;
e-mail: eturos@shell.cas.usf.edu
Undergraduate research participant.
ꢀ
0968-0896/$ - see front matter ꢀ 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2005.08.011

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E. Turos et al. / Bioorg. Med. Chem. 13 (2005) 6289–6308
reagents, such as molecular iodine, cause lactam 2 to
undergo cyclization to isopenem 3 (Scheme 1).2 The
intermediate in this reaction, bicyclic sulfonium interme-
diate II, is a powerful alkylating agent that rapidly
demethylates in the presence of nucleophiles.
O
SCH3
O
E+
S
N
N
Ph
CH3O
CH3O
H H
H H
E
Ph
2
3
+
E+
-CH3
O
SCH3
O
SCH3
N
N
CH3
S+
O
SCH3
O
R
1 R
CH3O
N
H H
I
R4
N
2 R3
Ph
Ph
CH3O
1
2
CH3O
Ph
H H
II
E
E
+
The likelihood that a similar sequence of events could be
triggered in vitro was ruled out, however, upon examin-
ing phenylethyl lactam 4, which possessed much of the
same activity against MRSA as compound 2.6 Thus,
p-unsaturation at C4 is neither a requirement for, nor
a detriment to, the antibacterial nature of N-methylthio
lactams. This was revealed further for propargylic and
alkenyl derivatives 5–7, and aryl-substituted lactams 8
and 9. 5,6 All six of the C4-varied analogues had antibac-
terial activity, with 4, 6, 8, and 9 having the strongest
growth inhibition of MRSA. This led to the realization
that other groups at C3 or C4 of the b-lactam ring could
alter biological activity, and more information was
needed to understand the effects these substituents might
have on the compoundsꢁ microbiological properties.
Scheme 1.
analogues of C4-aryl-substituted lactams 8 and 9 where-
in the functionalities, size, and orientation of the aryl
substituent are systematically altered. A model which
takes into account the experimental results is postulated
at the conclusion, to illustrate how we believe these
lactams exert their antibacterial effects.
2. Results and discussion
2.1. Synthesis and evaluation of C3-substituted analogues
O
SCH3
O
SCH3
Noting that all of the above N-methylthio b-lactams 2
and 4–9 bear a methoxy group at the C3 carbon, our first
goal was to determine if different types of substituents at
this position would affect antibacterial activity. Two
derivatives prepared in earlier studies, phthalimidyl
compound 10 and benzylthio lactam 11, were found to
be much less active against MRSA than C3 methoxy lac-
tam 2.7 On the other hand, C3-ethyl derivative 12 is
totally devoid of antibacterial activity, contrasting
sharply with the strong anti-MRSA properties of struc-
turally similar lactam 6.
N
N
CH3O
CH3O
AcO
4
5
O
SCH3
O
SCH3
N
N
CH3O
CH3O
6
O
N
SCH3
O
SCH3
7
O
SCH3
N
N
N
O
Et
BnS
O
SCH3
O
SCH3
Ph
N
N
O
Ph
Ph
Cl
10
11
CH3O
12
CH3O
Prior to initiating more detailed investigations, we
decided to examine C3-unsubstituted compounds 13–
15.8
8
9
The focus of this present study was thus to understand
the primary role of the C3 and C4 ring substituents on
the anti-MRSA activity of N-methylthio b-lactams 1.
Our overall aim was to cover a wide range of structural
variations among the C3/C4 substituents, differing in
their polarities, lipophilicities, steric requirements,
hydrogen-bonding capabilities, and stereochemistry.
The starting point for us was to consider whether the
C3 methoxy substituent is even needed or whether other
types of heteroatomic or non-heteroatomic groups can
be tolerated. We also investigated the effect of disubsti-
tution and steric crowding at C3, before focusing on the
O
SCH3
Cl
O
SCH3
O
SCH3
O
N
N
N
O
CH3
13
14
15
Derivative 13 was obtained by the [2+2]-cycloaddition9
of 2-chlorostyrene (16) with N-chlorosulfonylisocya-
nate, followed by N-thiolation with N-methylthioph-
thalimide10 and triethylamine (Scheme 2).

E. Turos et al. / Bioorg. Med. Chem. 13 (2005) 6289–6308
6291
O
O
SCH3
Cl
O
O
NH
N
NH
NH
O
Cl
O
C
NSO2Cl
[ref]9
a
a
Cl
O
CH3
17
18
16
13
b
b
Scheme 2. Reagents and condition: (a) N-methylthiophthalimide, Et3,
CH2Cl2, rt.
O
SCH3
O
O
SCH3
N
N
O
CH3
15
14
Compounds 14 and 15 were synthesized from commer-
cially available 4-acetoxyazetidinone (17) as shown in
Scheme 3. C-Allylation of 17 using allyltrimethylsilane
and BF3-etherate11 gave C4-allyl lactam 18. N-Thiola-
tion of 17 and 18 gave N-methylthio lactams 14 and
15, respectively.
Scheme 3. Reagents and conditions: (a) BF3ÆOEt2, allytrimethylsilane,
rt; (b) N-methylthiophthalimide, Et3N, CH2Cl2, rt.
OCH3
OCH3
Of these three C3-unsubstituted lactams, compounds 13
and 14 had only weak anti-MRSA activity, while allyl
derivative 15 was completely inactive. This suggests that
a polar C3 group, in combination with an appropriate
C4 substituent, may be required for bioactivity of these
lactams.
O
Cl
O
N
a
N
X
+
AcO
AcO
X
20
21a X=Cl
b X=H
22a X=Cl
b X=H
b
Thus, we then moved on to begin our studies on C3-
substituted lactams, beginning with C3-halo compounds
19a and 19b, and the azido derivative 19c.
OCH3
OCH3
O
O
N
c
N
Cl
Cl
or d
HO
X
23
24a-c
e
C3-Halogenated and azido b-lactams can normally be
prepared by the Staudinger coupling of an imine with
an a-halo or a-azidoacetyl chloride.12 However, we
found it more convenient to access these three lactams
directly from C3-hydroxy b-lactam 23, which is easily
synthesized in two steps from acetoxyacetyl chloride
(20) and N-(4-methoxyphenyl)imine 21 (Scheme 4). C3-
Chloro b-lactam 24a (X = Cl) was obtained in one step
from hydroxyl lactam 23 through the action of Ph3P
and a catalytic amount of sodium bicarbonate in reflux-
ing carbon tetrachloride. To obtain the iodo derivative
24b (X = I), hydroxy lactam 23 was first converted to
its mesylate by the reaction of its sodium salt with meth-
anesulfonyl chloride in anhydrous CH2Cl2 and then dis-
placed with NaI in DMF at 80 ꢁC. Similarly, azido
lactam 24c (X = N3) was obtained by the reaction of mes-
ylate intermediate with sodium azide in DMF. In each
case, trans-disubstituted lactams 24a–c were obtained
exclusively, as revealed by a characteristically small pro-
ton NMR coupling constant (J = 1.5–2.5 Hz) for the b-
lactam ring protons.13 These N-protected lactams were
then transformed into the N-methylthio lactams 19a–c
by N-dearylation/N-thiolation as described above.
O
O
SCH3
Cl
NH
N
Cl
f
X
X
25a-c
19a-c
Scheme 4. Reagents and conditions: (a) Et3N, CH2Cl2, rt; (b) NaOH,
MeOH, 0 ꢁC; (c) for 24a: PPh3, CCl4, NaHCO3 (catalytic), 70 ꢁC; (d)
for 24b,c: NaH, MsCl, CH2Cl2, 0 ꢁC to rt; then NaX, DMF, 80 ꢁC; (e)
(NH4)2Ce(NO)6, MeCN–H2O, 0 ꢁC; (f) N-methylthiophthalimide,
Et3N, CH2Cl2, 40 ꢁC (yields vary depending on compound; see
Section 4).
methicillin-susceptible and methicillin-resistant S. aure-
us strains by the Kirby–Bauer method of well diffu-
sion on agar plates (Fig. 1). Previously, we have
demonstrated that the growth inhibition zone sizes
for N-methylthio lactams correlate well with their
minimum inhibitory concentrations (MICs) obtained
from broth dilution experiments, and thus represent
a reliable way to gauge bioactivity within a closely
related series of analogues.4a One MSSA (ATCC
25923), nine MRSA strains (ATCC 43300), and eight
clinical isolates from a local hospital) were used for
these assays, all of which were b-lactamase producing
strains. Table 1 gives the zones of growth inhibition
2.2. Microbiological testing of C3/C4-substituted lactams
The b-lactams that were prepared in this study were
individually tested for antibacterial activity against

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E. Turos et al. / Bioorg. Med. Chem. 13 (2005) 6289–6308
the usual N-dearylation/N-methylthiolation conditions.
In the case of 26a and 26b, the N-thiolation step oc-
curred cleanly on the lactam nitrogen without affecting
the 2ꢁ amine at C3.
Lactam 9
O
SCH3
Cl
O
SCH3
Cl
N
N
NH
NH
26a
26b
MRSA
O
N
SCH3
Cl
O
N
SCH3
Cl
N
N
PenG
26c
26d
These C3 amino-substituted analogues turned out to
be significantly less potent against MRSA than meth-
oxy compound 9 or the C3-halo or azido lactams
19a–c. In fact, only the N-benzyl-substituted com-
pound 26b displayed any activity at all against
MRSAs, with average zone sizes of ꢀ11 mm diameter.
Thus, amino substituents at C3 of these N-methylthio
lactams appear to significantly diminish anti-MRSA
activity.
Figure 1. Kirby–Bauer well diffusion assay on an agar plate. The clear
regions appearing around the black wells correspond to zones where
bacterial growth is inhibited by the diffusing drug. Isolated white spots
appearing in this zone, such as that observed for penicillin G (PenG),
indicate surviving colonies of resistant bacteria.
for each compound against these microbes. For the
purpose of easier visualization, the zone data from
these assays are also plotted out graphically, as shown
in Figure 2, with the vertical bars indicating the aver-
age diameter (from 3 trials) of the growth inhibition
zones. The margin of error of these measurements is
1 mm. It is relevant that for the two reference drugs,
penicillin (Pen G) and vancomycin (Van), bioactivity
drops precipitously against the MRSA strains, and a
closer inspection of the growth inhibition zones for
these standards against the MRSAꢁs (see Fig. 1) re-
veals some speckled regions of bacterial growth indic-
ative of resistant colonies. This is not observed for
lactam 9 or any of the other N-methylthio b-lactam
analogues included in this study. On the other hand,
N-methylthio lactams 9 and 19a–c display equal effec-
tiveness against MSSA and MRSA. Of these latter
three compounds, chloro derivative 19a had the stron-
gest activity, with zones of inhibition against the
MRSA strains being more than 100% larger than
those of penicillin and equal to that of methoxy com-
pound 9. The iodo and azido-substituted lactams 19b
and 19c were appreciably weaker in activity than
either 9 or 19a. Thus, for these C3 varied analogues,
the activity follows the trend of N3 < I < Cl = OMe.
Our next series of analogues consists of a selection of
different C3-oxygenated derivatives of lactams 8 and 9,
including alkoxy and phenoxy derivatives, acyl ester,
and sulfonates, to determine if C3-alkoxy, acyloxy, or
sulfonyloxy groups differ in activity. Compounds in this
grouping bear a broad range of electronically and steri-
cally distinct functionalities that could affect lipophilici-
ty and bioactivity. Ethers 30a–d, ester 30e, and hydroxyl
lactam 30f were examined first.
O
O
SCH3
Cl
O
O
SCH3
Cl
N
N
30b
30a
O
O
SCH3
Cl
O
SCH3
Cl
N
N
H3C
O
PhO
30c
30d
Next, we examined the C3-amino-substituted analogues
26a–d, which were synthesized in four steps from hydro-
xy lactam 23, as shown in Scheme 5. Moffatt oxidation
of 23 with P2O5 in DMSO gave ketolactam 27, which
underwent reductive amination using an alkyl- or dial-
kylamine and NaBH(OAc)3 in a mixture of acetic acid
and dichloroethane, to afford amino adducts 28a–d in
good yield.14 The trans stereochemistry was established
O
O
SCH3
Cl
O
SCH3
Cl
N
N
O
HO
H3C
30e
30f
Compounds 30a–c were synthesized in 3 steps from hy-
droxy b-lactam 23 by base-induced O-alkylation with
the desired alkyl halide, affording N-aryl lactams 31a,c
1
by H NMR. These PMP-protected compounds were
then converted to N-methylthio products 26a–d under

E. Turos et al. / Bioorg. Med. Chem. 13 (2005) 6289–6308
6293
Table 1. Growth inhibition zones obtained from well diffusion experiments on agar plates
O
SCH3
N
X
R
R1
Compound
R
R1
X
ATCC
43300
USF
652
USF
653
USF
654
USF
655
USF
656
USF
657
USF
658
USF
659
9
OMe
N3
H
2-Cl
2-Cl
28
23
24
29
0
30
21
20
30
0
29
22
23
29
0
28
22
25
30
0
27
20
23
30
0
27
19
23
29
0
25
21
24
31
0
27
23
25
30
0
23
22
23
30
0
19a
19b
19c
26a
26b
26c
26d
30a
30b
30c
30d
30e
30f
33a
33b
33c
37a
37b
37c
37d
37e
37f
37g
42a
42d
43a
43b
43c
43d
43e
43f
43g
43h
43i
H
I
Cl
H
2-Cl
2-Cl
H
NHCH(CH2)4
NHCH2Ph
N(CH2CH3)2
NiBu2
OAllyl
Opropyl
OCH2OMe
OPh
H
2-Cl
2-Cl
H
11
0
10
0
12
0
10
0
10
0
10
0
11
0
10
0
10
0
H
2-Cl
2-Cl
H
0
0
0
0
0
0
0
0
0
H
2-Cl
2-Cl
22
23
15
16
18
18
15
16
21
27
26
19
28
24
27
21
21
18
26
25
26
20
26
29
24
23
22
19
23
27
23
20
18
12
13
17
10
14
16
25
24
18
18
23
18
15
17
24
28
28
18
26
25
28
20
22
17
25
26
27
21
25
34
23
27
22
20
24
29
23
18
17
13
13
14
10
13
17
24
24
18
15
24
17
15
17
20
26
26
17
28
26
27
20
19
18
28
26
28
24
27
29
23
24
21
21
23
32
27
23
22
12
12
19
11
12
16
20
23
19
15
21
17
18
17
23
24
24
17
25
22
26
18
20
15
22
26
25
18
22
27
21
24
21
19
23
27
23
20
16
11
13
18
10
13
15
23
22
17
14
23
19
14
15
20
26
26
18
26
23
26
20
21
16
23
25
23
18
21
28
24
24
24
20
22
27
23
22
14
14
12
18
10
13
17
23
21
18
16
22
18
15
16
20
26
26
17
28
25
28
21
20
17
23
25
26
19
25
29
22
25
19
21
19
28
25
22
18
13
13
19
11
14
16
20
20
18
13
20
16
15
16
19
26
25
18
28
26
29
21
18
15
22
23
26
21
25
28
25
24
20
21
22
27
23
20
18
12
14
19
12
13
14
20
20
18
15
21
19
15
15
20
24
25
16
26
24
26
20
17
16
19
24
25
18
23
28
23
20
21
19
20
26
22
21
17
12
13
18
9
24
24
16
16
15
18
14
15
21
26
25
18
23
21
24
20
21
14
18
18
21
17
20
23
22
17
17
16
18
23
20
18
11
10
10
0
H
H
2-Cl
2-Cl
H
OC(O)Me
OH
H
2-Cl
2-Cl
H
OSO2Me
OSO2Ph
OSO2Tol
OAllyl
Opropyl
OCH2OMe
OAc
H
H
H
H
H
H
2-Cl
CH3
CH3
CH3
CH3
Allyl
Propyl
Ph
2-Cl
2-Cl
2-Cl
2-Cl
OAc
OAc
2-Cl
2-Cl
OAc
–O(CH2)4–
–CH2OCH2CH2–
OMe
H
H
3-Cl
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
OMe
OMe
4-Cl
2-F
OMe
OMe
3-F
4-F
OMe
OMe
2-I
3-I
OMe
OMe
4-I
2,4-Cl
2,6-Cl
2,3,5-Cl
2-OCH3
2-CH3
2-NO2
3-NO2
4-CO2CH3
4-O2CCH=CH2
4-OH
4-Ph
3,4-CH2C6H4
4-NO2
43j
OMe
OMe
43k
43l
OMe
OMe
43m
43n
43o
43p
43q
43r
43s
43t
47a
OMe
OMe
OMe
OMe
OMe
OMe
8
OMe
–O(CH2)4–
11
12
10
14
The values indicate the average diameters in mm (of three trials) for the zone of growth inhibition observed after 24 h of incubation at 37 ꢁC. Twenty
micrograms of each test compound in DMSO solution was used. All of the microbes listed are b-lactamase producing, methicillin-resistant strains of
Staphylococcus aureus (MRSA). Those labeled as USF652-659 were obtained from a clinical testing laboratory at Lakeland Regional Medical
Center, Lakeland, FL. Error values are within 1 mm.
(Scheme 6). Phenoxy lactam 31d (R = Ph) was synthe-
sized directly from phenoxyacetyl chloride and imine
21 under the typical Staudinger coupling conditions, as
outlined in Scheme 4.15
Likewise, acyl ester 31e was prepared from hydroxyl lac-
tam 23 by base-promoted acylation. These four N-aryl
lactams were taken onto their N-methylthio lactams
30a, c, d, and e accordingly. 30b was obtained from

6294
E. Turos et al. / Bioorg. Med. Chem. 13 (2005) 6289–6308
30c = 30f > 30d. Adjusted for their differences in molec-
ular weight, the bioactivities of the lactams 30a–f show
the same trend, suggesting that these slight variations
in bioactivity may be due to slightly different lipophilic-
ity and permeability properties of the individual
compounds.
Staphylococcus aureus
MRSA
C3-Sulfonates 33a–c were also examined in continuation
of this series, and their anti-MRSA activities compared
to those of C3-methoxy lactam 8.
These three new derivatives were prepared via the reac-
tion of the sodium salt of 34 with the appropriate chlo-
rosulfonate, as shown in Scheme 7.
For these three C3 sulfonates, activity increases with
molecular weight: methyl < phenyl < p-tolyl (Fig. 4).
This trend is also observed in broth MIC values, which
range from 64 to 128 lg/mL for the MSSA (ATCC
25923) and MRSA (ATCC 43300) strains. For mesyl
compound 33a, the MICs are all around 128 lg/mL,
while for phenylsulfonyl lactam 33b, the MICs are be-
tween 64 and 128 lg/mL, and for the tosyl derivative
33c, the values decrease to around 64 lg/mL.
Figure 2. Comparison of antimicrobial activities of lactams 9 and 19a–
c with penicillin G (PenG) and vancomycin (Van) against Staphylo-
coccus aureus bacteria. The y-axis is the zone of inhibition in
millimeters. The vertical bars indicate the average diameter (from 3
trials) of the growth inhibition zones. The blue bars are for methicillin-
susceptible S. aureus (MSSA), while the red bars are for methicillin-
resistant S. aureus (MRSA). Thus, the red bars denote the average of
27 trials (9 microbes in triplicate) for each test compound.
32a (R = allyl) by hydrogenation of the allyl group, pri-
or to N-thiolation. Hydroxy lactam 30f was obtained
from 32e (R = acetyl) by base hydrolysis of the acetate
(KOH, MeOH) prior to N-thiolation.
To evaluate the effects of steric crowding at the C3 cen-
ter, several differentially configured tertiary ethers 37a–c
and esters 37d–g of 3,3-disubstituted analogues of 9 were
studied next.
As Figure 3 depicts, lactams 30a–f all showed enhanced
activity against MRSA as compared to penicillin, but an
activity lower than that of C3-methoxy lactam 9. The
relative efficacies of the C3-oxygenated lactams in this
series do vary somewhat, with 9 > 30a = 30b = 30e >
These compounds were made from keto b-lactam 27 by
introduction of the alkyl or aryl group via Grignard
reaction using the method of Buynak (Scheme 8).16
In each case, nucleophilic attack occurred exclusively
O
O
SCH3
O
O
SCH3
O
O
SCH3
N
N
N
O
S
O
S
O
S
H3C
H3C
O
O
O
33c
33a
33b
O
SCH3
O
SCH3
Cl
O
SCH3
N
N
N
Cl
Cl
CH3
CH3
CH3
O
O
CH3O
O
37a
37b
37c
O
SCH3
Cl
O
SCH3
Cl
N
N
CH3
AcO
AcO
37d
37e
O
SCH3
Cl
O
SCH3
Cl
N
N
AcO
AcO
37g
37f

E. Turos et al. / Bioorg. Med. Chem. 13 (2005) 6289–6308
6295
Scheme 5. Reagents and conditions: (a) P2O5, DMSO, rt; (b) RR0NH, NaBH(OAc)3, AcOH, ClCH2CH2Cl, rt; (c) (NH4)2Ce(NO)6, MeCN–H2O,
0 ꢁC; (d) N-methylthiophthalimide, Et3N, CH2Cl2, 40 ꢁC (yields vary depending on compound; see Section 4).
OCH3
OCH3
O
O
a
N
N
Cl
Cl
RO
HO
31a,c-e
23
b
O
SMe
O
c
N
NH
Cl
Cl
RO
RO
32a-f
30a-f
Scheme 6. Reagents and conditions: (a) for 31a,c,d: NaH, R-X,
CH2Cl2, rt; for 31e,f: AcCl, pyridine, CH2Cl2, rt; (b) (NH4)2Ce(NO3)6,
MeCN–H2O, 0 ꢁC, then for 32b: H2, Pd/C, rt; (c) N-methylthioph-
thalimide, Et3N, C6H6, 7- ꢁC; for 30f, KOH, MeOH, 0 ꢁC, then N-
methylthiophthalimide, Et3N, C6H6, 70 ꢁC (yields vary depending on
the compound; see Section 4).
Figure 3. Comparison of antimicrobial activities of lactam 9, and C3-
alkoxy and C3-acyloxy lactams 30a–f with penicillin G (PenG). The
vertical bars indicate the average diameter in mm of the growth
inhibition zones produced against nine different MRSA strains (in
triplicate).
than the more polar methoxymethyl derivative 37c.
For the second series of compounds, 37d–g, the ether
is replaced with an acetoxy ester, while the alkyl side
chain is varied. In this case, the saturated alkyl side
chain (methyl and propyl) analogues, 37d,f, are at least
33% more active than the lactams 37e,g bearing allyl
or phenyl C3-residues. For these four ester compounds,
the MIC values were determined to be between 16 and
64 lg/mL, with the order being 37f P 37d > 37g P 37e.
Thus, zone measurements and MIC values indicate the
same activity trends.
from the less hindered a-face to afford b-lactams 38d–
g. Relative stereochemistry was established by ROESY
NMR. Tertiary alcohol 38d (R = Me) was then O-al-
kylated, as described above, and carried on to N-meth-
ylthio b-lactams 37a–c. Tertiary alcohols 38d, e, and g
were O-acylated and converted to N-methylthio b-lac-
tams 37d, e, and g, respectively. b-Lactam 37f
(R = propyl) was obtained from 41e (R = allyl) by
hydrogenation of the allyl group prior to N-thiolation.
The zone data for these seven 3,3-disubstituted com-
pounds indicate that steric crowding on the ring
generally diminishes bioactivity, compared to 3-mono-
substituted lactam 9 (Fig. 5). In the series of tertiary
ethers 37a–c, in which only the ether alkyl moiety is var-
ied, the propyl and allyl ethers 37a, b are more active
Finally, to conclude the study of C3-derivatives, we
examined spirocyclic lactams 42a and 42b, in which
the C3 ether is contained within a conformationally
restricted ring.17 Compound 42a had slightly
more activity than its isomer 42b, but was 25% less

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E. Turos et al. / Bioorg. Med. Chem. 13 (2005) 6289–6308
OCH3
OCH3
b
OCH3
OCH3
OCH3
O
O
O
O
O
O
a
a
N
N
N
N
N
Cl
Cl
Cl
R
R
RO
HO
R'O
HO
27
39a-c
40d,e,g
38d,e,g
34
35a-c
b
c
O
SCH3
O
SMe
O
O
N
c
N
NH
NH
Cl
d
Cl
R
R
R'O
RO
RO
R'O
37a-g
41a-g
36a-c
33a-c
Scheme 8. Reagents and conditions: (a) RMgBr, THF, NH4Cl,
ꢁ78 ꢁC; (b) for 39–c, NaH, R0-X, TBAl, CH2Cl2, rt; for 40d,e,g,
NaH, AcCl, CH2Cl2, rt; (c) (NH4)2Ce(NO)6. MeCN–H2O, 0 ꢁC; then
fpr 41f; 41c, H2, Pd/C, EtOAc, rt; (d) N-methylthiophthalimide, Et3N,
C6H6, 70 ꢁC (yields vary depending on compound; see Section 4).
Scheme 7. Reagents and conditions: (a) NaH, R-X, CH2Cl2, rt; (b)
(NH4)2Ce(NO3)6, MeCN–H2O, 0 ꢁC; (c) N-methylthiophthalimide,
Et3N, C6H6, 70 ꢁC (yields vary depending on compound; see
Section 4).
similar antibacterial activity to the C4-acetylenic lead
compound 2.4 We have also observed previously that
lipophilic substituents at certain locations on the aryl
ring of 43 (X = CH3) seemed to enhance in vitro activi-
ties, while more polar groups (X = CN) diminished the
activity.5
Figure 4. Comparison of antimicrobial activities of lactam 8 and C3-
sulfonate lactams 33a–c with penicillin G (PenG). The vertical bars
indicate the average diameter in mm of the growth inhibition zones
produced against nine different MRSA strains (in triplicate).
active than the 3ꢁ open-ring analogues 37a and 37b
(Fig. 6).
O
SCH3
O
SCH3
N
N
O
O
42a
42b
To summarize these findings thus far, it appears that C3-
alkoxy and acyloxy substituents provide for the best
anti-MRSA activity.
We then turned our attention to the effects of the C4 ring
substituents on bioactivity. For this study, we chose to
look specifically at C4 aryl-substituted lactams 43, based
upon the observation that the phenyl lactam 8 (X = H in
43) and chlorophenyl lactam 9 (X = ortho-Cl in 43) had

E. Turos et al. / Bioorg. Med. Chem. 13 (2005) 6289–6308
6297
For this, a focused library of different C4 aryl-substitut-
ed b-lactams, 43a–t, was prepared, as illustrated in
Scheme 9. To complement this set, previously reported
compounds 43c–e, m, q, and r were also included.
Figure 5. Comparison of antimicrobial activities of lactam 9 and C3-
tertiary ether and ester lactams 37a–g with penicillin G (PenG). The
vertical bars indicate the average diameter in mm of the growth
inhibition zones produced against nine different MRSA strains (in
triplicate).
Staudinger coupling of N-(4-methoxyphenyl)imines 44
with methoxyacetyl chloride in the presence of three
equivalents of triethylamine exclusively afforded the
1
cis-disubstituted b-lactams 45, as corroborated by H
NMR. These adducts were converted to N-methylthio
lactams 43, as previously described.
Figure 7 shows that the location of the variable ring sub-
stituent had a discernable effect on activity, with the
ortho substituents displaying the most potent activities,
followed by the groups at the para then meta positions
(9 > 43e > 43d). This agrees with our previous studies
of aryl-fluorinated N-methylthio b-lactams, which found
that those analogues having at least one fluorine ortho to
the lactam ring displayed the highest potencies, and
within the monofluoro series, 43a > 43c > 43b.4a
Figure 6. Comparison of antimicrobial activities of lactam 8, 37a–b,
and C3-spirocyclic lactams 42a–b with penicillin G (PenG). The
vertical bars indicate the average diameter in mm of the growth
inhibition zones produced against nine different MRSA strains (in
triplicate).
Thus, our next objective was to investigate further the
influence of the C4 aryl ring substituent and the position
of the X moiety on the aryl ring on anti-MRSA activity.
Second, activity among the different halo derivatives is
largely independent of which halogen is present on the
Scheme 9. Reagents and conditions: (a) p-anisidine, CSA, CH2Cl2; (b) CH3OCH2COCl, iPr2EtN, PhMe, 0 ꢁ to rt; (c) (NH4)2Ce(NO3)6, MeCN–H2O,
i
0ꢁ; (d) N-methylthiophthalimide, Pr2EtN, C6H6, 70 ꢁC (yields vary depending on compound; see Section 4).

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E. Turos et al. / Bioorg. Med. Chem. 13 (2005) 6289–6308
Figure 7. Comparison of antimicrobial activities of lactam 9 and C4-
monohaloaryl lactams 43a–h with penicillin G (PenG). The vertical
bars indicate the average diameter in mm of the growth inhibition
zones produced against nine different MRSA strains (in triplicate).
Figure 9. Comparison of antimicrobial activities of lactam 9 and C4-
monosubstituted aryl lactams 43l–t with penicillin G (PenG). The
vertical bars indicate the average diameter in mm of the growth
inhibition zones produced against nine different MRSA strains (in
triplicate).
show a greatly reduced bioactivity compared to the C4-
phenyl or chlorophenyl analogues 8 and 9, respectively.
aryl ring (compare the pair of ortho-halo compounds 9
to 43a and 43f, and meta-halo lactams 43b to 43d and
43g in Fig. 7), and the number of halogens on the aryl
ring (see lactams 43i–k in Fig. 8).
Spiro compounds 47a–c17 displayed variable deviations
in activity versus C4 phenyl derivative 42a. In effect,
what these studies indicate is that various types of unsat-
urated and saturated side chains can occupy the C4
center without dramatically affecting bioactivity, but
ortho-substituted aryl ring compounds generally offer
better antimicrobial properties (see Fig. 10).
Additionally, other monosubstituted aryl analogues
43i–t were studied to compare the effects of different
electron-donating or electron-withdrawing groups on
in vitro activity (Fig. 9). What we observe is that
replacement of the ortho-chloro substituent of lactam 9
for other ortho groups, such as methoxy, methyl, or ni-
tro, diminishes activity somewhat, although zone sizes
generally remain larger than for the meta or para-substi-
tuted derivatives 43o–t. Curiously, reversal of the ester
functionality at the para position (see lactams 43o and
43r) does lead to significant reduction of anti-MRSA
activity, while conversion of the acryloyl ester 43o to
the corresponding phenol 43p does not alter activity.
Fluorenyl lactam 43s and biphenyl lactam 43t, likewise,
`
It is also interesting to consider vis-a-vis the cis and trans
stereoisomers of C3-acetoxy N-methylthio b-lactams 48a
and 48b, which we were able to prepare independently
by isolating the trans b-lactam adduct from the cis/trans
mixture of N-PMP lactams by recrystallization in
methanol, and carrying both on to the N-thiolated
compounds. These two diastereomers show similar bio-
activity with zone sizes in the mid-20 mm range,
although the trans isomer was found to be about 10%
more active than the cis lactam.
Likewise, we had the opportunity to examine the effect
of absolute stereochemistry on bioactivity through an
independent asymmetric synthesis of enantiomeric lac-
tams, (ꢁ)-50a and (+)-50a (Scheme 10). For this, we
employed Lipase PS-30 to selectively deacylate the
Figure 8. Comparison of antimicrobial activities of lactam 9 and C4-
dihaloaryl and trihaloaryl lactams 43i–k with penicillin G (PenG). The
vertical bars indicate the average diameter in mm of the growth
inhibition zones produced against nine different MRSA strains (in
triplicate).

E. Turos et al. / Bioorg. Med. Chem. 13 (2005) 6289–6308
6299
3. Conclusions
The data in this study reveal some interesting character-
istics about the structural requirements of the C3 and C4
ring substituents of N-methylthio b-lactams. First, lipo-
philicity within these groups does appear to be required
for anti-MRSA activity, although C3-alkoxy or acyloxy
side chains afford the best bioactivity. Differences in the
in vitro behavior for all the different analogues may be
more closely related to their differences in diffusability
through the bacterial membrane, rather than to any spe-
cific binding interaction with a biological target. This
lends considerable support to the suggestion that the
mode of action is as proposed in Scheme 11.
We postulate that the lactams react covalently with
their biological target by transfer of the sulfur side
chain upon passing through the bacterial cell mem-
brane. Compounds which are too polar to get through
the membrane would be expected to have a lower bac-
teriostatic activity, while those with too much lipophil-
icity may be sequestered in the membrane or internal
organelles, lowering the effective concentration in the
cytoplasm. The finding that neither relative nor abso-
lute chirality of the N-methylthio lactam affects anti-
microbial properties suggests that the lactam may
not experience significant non-bonding interactions
with its biological target prior to transferring the sul-
fenyl side chain. This suggests the model shown in
Scheme 1 in which the nucleophile attacks the mole-
cule, without precoordination, directly on the sulfur
center.
Figure 10. Comparison of antimicrobial activities of C3-spirocyclic
lactams 42a and 47a–c with penicillin G (PenG). The vertical bars
indicate the average diameter in mm of the growth inhibition zones
produced against nine different MRSA strains (in triplicate).
C3-acetoxy group of the 3S,4R-enantiomer from the
mixture of racemic C3-acetoxy lactams, ( )-31e.18 The
resulting enantiomerically pure compounds, (ꢁ)-alcohol
49 and recovered (+)-acetate 31e, were then converted
independently to the N-methylthio lactams (ꢁ)-50a
and (+)-50a, respectively.
Both antipodes of 50a displayed equal antimicrobial
activity against the MRSA isolates, giving identical
zones of growth inhibition on agar plates, as well as
equivalent minimum inhibitory concentration (MIC)
values in broth dilution experiments. Thus, from these
examinations, neither relative nor absolute stereochem-
istry of the N-thiolated lactams seems to be a factor in
anti-MRSA activity.
Further work is ongoing to identify the biological tar-
get(s) of the lactams in bacterial cells and to determine,
in more detail, the effects these compounds have on the
primary cellular processes in bacteria.
OCH3
OCH3
OCH3
O
O
O
N
N
N
Lipase PS-30
+
acetone
AcO
AcO
HO
phosphate buffer
pH 7.2
(-)-49
(3S,4R)
(+)-31e
(3R,4S)
(+/-)-31e
O
O
SCH3
N
SCH3
N
+
AcO
AcO
(-)-50a
(3S,4R)
(+)-50a
(3R,4S)
Scheme 10.

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E. Turos et al. / Bioorg. Med. Chem. 13 (2005) 6289–6308
Nu
by washing with ice-cold MeOH. The product 22a
was isolated 10.6 g (44%), as a white solid, mp 130–
132 ꢁC. H NMR (250 MHz, CDCl3): d 7.43 (1H, d,
J = 8.9 Hz), 7.32–7.23 (5H, m), 6.83 (2H, d,
J = 8.9 Hz), 6.16 (1H, d, J = 5.0 Hz), 5.78 (1H, d,
J = 5.0 Hz), 3.76 (3H, s), 1.76 (3H, s); 13C NMR
(63 MHz, CDCl3): d 168.6, 161.3, 156.6, 133.8, 130.1,
129.9, 129.7, 128.6, 126.7, 118.5, 114.4, 75.4, 58.1,
55.3, 19.8.
1
O
S
O
CH3
Nu-
N
N
Nu-SCH3
+
R
1 R
R3
R R3
1 R
2 R4
2 R4
1
Scheme 11.
4.4. ( )-(3R,4S)-3-Acetoxy-N-(4-methoxyphenyl)-4-phenyl-
azetidin-2-one (22b)
4. Experimental
White solid in 56% yield mp 138–140 ꢁC. 1H NMR
(250 MHz, CDCl3): d 7.32–7.20 (7H, m), 6.73 (2H, d,
J = 8.9 Hz), 5.87 (1H, d, J = 4.8 Hz), 5.27 (1H, d,
J = 5.0 Hz), 3.89 (3H, s), 1.60 (3H, s); 13C NMR
(63 MHz, CDCl3): d 168.6, 158.1, 142.6, 133.8, 130.1,
129.9, 129.7, 128.6, 126.7, 118.5, 114.9, 114.4, 75.4,
58.1, 55.3, 19.8.
All reagents were purchased from Sigma–Aldrich Chem-
ical Company and used without further purification.
Solvents were obtained from Fisher Scientific Company.
Thin-layer chromatography (TLC) was carried out
using EM Reagent plates with a fluorescence indicator
(SiO2-60, F-254). Products were purified by flash chro-
matography using J.T. Baker flash chromatography sil-
ica gel (40 lm). NMR spectra were recorded in CDCl3
unless otherwise noted. 13C NMR spectra were proton
broad-band decoupled.
4.5. Synthesis of ( )-(3R,4S)-4-(2-chlorophenyl)-3-hydroxy-
N-(4-methoxyphenyl)azetidin-2-one (23)
4.1. Synthesis of 2-chlorophenyl-N-(4-methoxyphenyl)-
imine (21a)
To a solution of 22 (1.23 g, 3.56 mmol) in 30 mL ace-
tone was added a solution of KOH in 10 mL MeOH
at 0 ꢁC. The hydrolysis was complete after the addi-
tion of KOH/MeOH, as indicated by TLC. The reac-
tion was quenched by adding an equal volume of
water upon which the product precipitated out of
solution. The product was filtered and dried to give
a white solid, 1.07 g (99%) of 23, mp 183–184 ꢁC.
To a solution of p-anisidine (7.27 g, 59.0 mmol) in
50 mL CH2Cl2 were added o-chlorobenzaldehyde
(6.64 g, 47.2 mmol), and a catalytic amount of camphor-
sulfonic acid. The resultant mixture was stirred until
TLC indicated the disappearance of starting materials.
The solvent was removed under reduced pressure, and
the crude material was purified by recrystallization from
ice-cold MeOH to yield 11.41 g (79%) of imine 21a as a
yellow solid. mp 62–63 ꢁC. 1H NMR (250 MHz,
CDCl3): d 8.95 (1H, s), 8.27–8.23 (1H, m), 7.45–7.36
(3H, m), 7.30 (2H, d, J = 8.9 Hz), 6.96 (2H, d,
J = 8.9 Hz), 3.86 (3H, s); 13C NMR (63 MHz, CDCl3):
d 156.5, 154.6, 144.5, 135.7, 133.3, 131.7, 129.8, 128.3,
127.0, 122.4, 114.3, 55.4.
1H NMR (250 MHz, CDCl3):
d
7.48 (1H, d,
J = 7.5 Hz), 7.33–7.22 (5H, m), 6.84 (2H, d,
J = 8.9 Hz), 5.63 (1H, d, J = 5.1 Hz), 5.33 (1H, d,
J = 5.1 Hz), 4.88 (1H, br s), 3.78 (3H, s); 13C NMR
(63 MHz, DMSO-d6): d 166.6, 156.1, 133.1, 132.9,
130.9, 129.7, 129.6, 128.9, 127.3, 118.6, 114.9, 77.2,
60.0, 55.6.
4.6. ( )-(3R,4S)-3-Hydroxy-4-phenyl-N-(4-methoxyphenyl)-
azetidin-2-one (34)
4.2. N-(4-Methoxyphenyl)phenyl-imine (21b)
White solid in 99% yield, mp 183–184 ꢁC. 1H NMR
(250 MHz, CDCl3): d 7.49 (1H, d, J = 7.5 Hz), 7.32–
7.22 (5H, m), 6.83 (2H, d, J = 8.9 Hz), 5.62 (1H, d,
J = 5.1 Hz), 5.33 (1H, d, J = 5.1 Hz), 3.77 (3H, s); 13C
NMR (63 MHz, DMSO-d6): d 166.6, 156.1, 133.1,
132.9, 130.9, 129.7, 129.6, 128.9, 127.3, 118.6, 114.9,
114.2, 77.2, 60.0, 55.6.
White solid in 85% yield. mp 58–60 ꢁC. 1H NMR
(250 MHz, CDCl3): d 8.42 (1H, s), 7.85–7.82 (2H, m),
7.40 (3H, t, J = 3.2 Hz), 7.20–7.15 (2H, m), 6.87 (2H,
dd, J = 2.0, 7.8 Hz), 3.77 (3H, s); 13C NMR (63 MHz,
CDCl3): d 156.5, 155.2, 144.2, 132.2.7, 130.3, 128.7,
122.9, 122.4, 114.8, 114.3, 55.9.
4.3. Synthesis of ( )-(3R,4S)-3-acetoxy-4-(2-chlorophenyl)-
N-(4-methoxyphenyl)azetidin-2-one (22a)
4.7. Synthesis of ( )-(3S,4S)-3-chloro-4-(2-chlorophenyl)-
N-(4-methoxyphenyl)azetidin-2-one (24a)
Imine21a (17.3 g, 70.4 mmol) was dissolved in 200 mL
of freshly distilled CH2Cl2. The solution was cooled
to approximately 5 ꢁC in an ice bath. Triethylamine
(3 equiv, 21.4 g, 211.4 mmol) was added, followed by
acetoxyacetyl chloride (20) (1.2 equiv, 11.5 g,
84.4 mmol) dissolved in 20 mL CH2Cl2. The reaction
mixture was stirred until no further change in TLC
was observed for 1 h. The solvent was removed under
reduced pressure and the crude material was purified
To a solution of b-lactam 23 (318 mg, 1 mmol) in 15 mL
CCl4 added triphenylphosphine (524 mg, 2 mmol) and a
catalytic amount (1 mg) of NaHCO3 and the mixture
was refluxed for 20 h. The solvent was removed under
reduced pressure and the crude material was purified
by column chromatography using (3:7 EtOAc/hexanes)
to give 302 mg (91%) of 24a as a solid. 1H NMR
(250 MHz, CDCl3): d 7.40 (1H, d, J = 7.5 Hz), 7.10–
7.30 (5H, m), 6.80 (2H, d, J = 7.5 Hz), 5.40 (1H, d,

E. Turos et al. / Bioorg. Med. Chem. 13 (2005) 6289–6308
6301
J = 1.5 Hz), 4.60 (1H, d, J = 2.0 Hz), 3.70 (3H, s); 13C
NMR (63 MHz, CDCl3): d 160.0, 156.7, 137.1, 133.8,
133.5, 132.5, 130.3, 128.7, 128.5, 128.4, 127.5, 126.9,
118.8, 114.5, 62.5, 55.4.
4.11. Synthesis of ( )-(3S,4S)-4-(2-chlorophenyl)-3-cyclo-
pentylamino-N-(4-methoxyphenyl)azetidin-2-one (28a)
Cyclopentylamine (35.6 mg, 0.33 mmol) and compound
27 (100 mg, 0.33 mmol) were mixed in ClCH2CH2Cl
(1.32 mL) and then treated with sodium triacetoxyboro-
hydride (98.6 mg, 0.465 mmol) and AcOH (19.8 mg,
0.33 mmol). The mixture was stirred at rt under a N2
atmosphere for 1 h until the reactants were consumed
completely. The reaction mixture was quenched by add-
ing 1 N NaOH, and the product was extracted with
ether. The ether extract was washed with brine and dried
with MgSO4. The solvent was evaporated to give 28a in
4.8. Synthesis of ( )-(3S,4S)-4-(2-chlorophenyl)-3-iodo-
N-(4-methoxy-phenyl)azetidin-2-one (24b)
To a solution of b-lactam 23 (795 mg, 2.5 mmol) in
20 mL of dry CH2Cl2 was added NaH (60% suspension
in mineral oil, 125 mg, 5 mmol) and the mixture was
stirred for 15 min. Methanesulfonyl chloride (342 mg,
3 mmol) was then added dropwise to the solution,
and the resulting solution was stirred at rt for
30 min. The solution was washed with brine (3·
25 mL), the organic layer was dried with anhydrous
MgSO4, filtered, and evaporated, and the residue was
washed with cold MeOH to give 830 mg as a white sol-
id in 82% yield. To the solution of above compound
(405 mg, 1 mmol) dry DMF was added NaI (447 mg,
3 mmol) and the resulting solution was heated to
80 ꢁC for 24 h. After cooling to rt, the solution was
concentrated under vacuum. The crude compound
was dissolved in EtOAc and washed with water (3·
20 mL), and the organic layer was dried with MgSO4,
1
88% yield. H NMR (250 MHz, CDCl3): d 7.30–7.19
(4H, m), 6.90 (2H, d, J = 8.8 Hz), 6.69 (2H, d,
J = 8.8 Hz), 4.97 (2H, s), 3.78 (1H, br s), 3.69 (3H, s),
1.71 (4H, m), 1.33 (4H, m); 13C NMR (63 MHz,
CDCl3): d 165.8, 157.9, 140.2, 133.2, 130.5, 130.4,
129.5, 129.2, 127.6, 127.4, 119.5, 114.7, 55.4, 50.0,
48.4, 42.8, 33.3, 33.2, 27.9, 27.8.
An analogous procedure was used to prepare com-
pounds 28b–d.
4.12. ( )-(3S,4S)-3-Benzylamino-4-(2-chlorophenyl)-N-(4-
methoxyphenyl)azetidin-2-one (28b)
1
filtered, and evaporated to give 24b in 81% yield. H
NMR (250 MHz, CDCl3): d 7.50 (1H, d, J = 1.0 Hz),
7.25–7.50 (5H, m), 6.83–6.89 (2H, m), 5.57 (1H, d,
J = 2.5 Hz), 4.74 (1H, d, J = 2.0 Hz), 3.80 (3H, s);
13C NMR (63 MHz, CDCl3): d 162.1, 156.7, 133.5,
130.6, 130.2, 130.0, 129.7, 129.7, 129.6, 126.8, 126.5,
118.8, 118.4, 118.2, 114.0, 62.0, 21.0.
1
95% yield. H NMR (250 MHz, CDCl3): d 7.27–7.12
(9H, m), 6.93 (1H, d, J = 8.8 Hz), 6.70 (1H, d,
J = 8.8 Hz), 4.96 (2H, s), 4.23 (2H, d, J = 6.0 Hz), 3.69
(3H, s); 13C NMR (63 MHz, CDCl3): d 164.0, 161.7,
159.4, 137.8, 134.3, 134.1, 133.9, 133.8, 130.4, 129.4,
129.0, 128.4, 128.2, 127.9, 127.4, 127.1, 119.0, 114.6,
55.7, 53.4, 51.7, 43.5.
4.9. ( )-(3S,4S)-3-Azido-N-(4-methoxyphenyl)azetidin-2-
one (24c)
4.13. ( )-(3S,4S)-4-(2-Chlorophenyl)-3-diethylamino-N-
(4-methoxyphenyl)azetidin-2-one (28c)
Brown solid, 81% yield. 1H NMR (250 MHz, CDCl3): d
7.24–7.10 (4H, m), 6.80–6.74 (2H, m), 5.24 (1H, d,
J = 1.8 Hz), 4.44 (1H, d, J = 1.8 Hz), 3.70 (3H, s); 13C
NMR (63 MHz CDCl3): d 164.2, 156.2, 142.5, 133.5,
132.2, 128.9, 128.7, 128.2, 127.9, 126.8,121.9, 121.5,
114.2, 113.8, 60.8, 52.2.
90% yield. 1H NMR (250 MHz, CDCl3): d 7.31–7.10 (4H,
m), 6.95 (2H, d,J = 8.8 Hz), 6.65 (2H, d, J = 8.8 Hz), 5.00
(2H, s), 3.67 (3H, s), 3.18 (2H, q, J = 7.0 Hz), 3.09 (2H, q,
J = 7.1 Hz), 1.14 (3H, t, J = 7.0 Hz), 0.60 (3H, t,
J = 7.1 Hz); 13C NMR (63 MHz, CDCl3) d 165.9, 164.2,
160.7, 134.2, 132.2, 131.1, 129.7, 129.4, 127.5, 114.5,
55.8, 49.3, 42.3, 37.8, 14.0, 12.2.
4.10. Synthesis of ( )-(4S)-4-(2-chlorophenyl)-N-(4-meth-
oxyphenyl)-3-oxoazetidin-2-one (27)
Phosphorous pentoxide (0.82 g, 2.88 mmol, 0.7 equiv)
was added to 15 mL of dry DMSO. The resultant mix-
ture was stirred for 5 min, followed by the addition of
23 (1.25 g, 4.12 mmol, 1.0 equiv). The reaction was
monitored by H NMR and showed complete conver-
sion after stirring for 1 h. The reaction mixture was
4.14. ( )-(3S,4S)-3-(2-Chlorophenyl)-3-diisobutylamino-
N-(4-methoxyphenyl)azetidin-2-one (28d)
98% yield. 1H NMR (250 MHz, CDCl3): d 7.31–7.20 (4H,
m), 6.96 (2H, d, J = 8.8 Hz), 6.65 (2H, d, J = 8.8 Hz), 4.98
(2H, s), 3.68 (3H, s), 2.90 (4H, t, J = 6.6 Hz), 1.90 (1H, m),
1.45 (1H, m), 0.76 (6H, d, J = 6.6 Hz), 0.35 (6H, d,
J = 6.6 Hz); 13C NMR (63 MHz, CDCl3): d 163.1,
156.2, 136.9, 135.2, 134.2, 134.1, 130.2, 129.4 127.5,
122.3, 122.2, 114.2, 114.0, 71.9, 60.0, 56.2, 55.0, 41.6,
28.3, 26.8, 20.6, 20.5, 20.3, 20.2.
1
poured into
a
cold solution of sat. NaHCO3
(100 mL) and extracted with CH2Cl2 (3· 50 mL). The
combined organic layers were washed with brine
(100 mL) and dried over MgSO4. The solvent was re-
moved under reduced pressure to yield a yellow solid,
1.16 g (93%) of 27; mp 127–130 ꢁC. 1H NMR
(250 MHz, CDCl3): d 7.49–7.41 (3H, m), 7.35–7.17
(3H, m), 6.89 (2H, d, J = 9.0 Hz), 6.07 (1H, s), 3.78
(3H, s); 13C NMR (63 MHz, CDCl3): d 189.5, 159.8,
157.9, 133.3, 130.5, 130.4, 129.5, 129.2, 127.6, 119.5,
114.7, 71.9, 55.4.
4.15. Synthesis of ( )-(3R,4S)-3-Allyloxy-4-(2-chlorophenyl)-
N-(4-methoxyphenyl)azetidin-2-one (31a)
To a solution of b-lactam 23 (1.0 g, 3.3 mmol) in 25 mL
of dry CH2Cl2 was added NaH (60% suspension in

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E. Turos et al. / Bioorg. Med. Chem. 13 (2005) 6289–6308
mineral oil, 0.26 g, 6.6 mmol) and the mixture was
stirred for 15 min. Allyl bromide (0.79 g, 6.6 mmol)
was then added, along with 5 mg TBAI (tetrabutylam-
monium iodide). The mixture was refluxed for 24 h or
until the TLC indicated the disappearance of the start-
ing material. The reaction was quenched with a 5% solu-
tion of NH4Cl and extracted (3· 25 mL) with CH2Cl2.
The combined organic layers were washed with brine
and dried over anhydrous Na2SO4. The crude material
was purified by column chromatography on silica gel
(1:9 EtOAc/hexanes) to give 0.92 g (85%) of 31a as a
7.26 (7H, m), 6.85 (2H, d, J = 8.7 Hz), 5.94 (1H, d,
J = 5.1 Hz), 5.82 (1H, d, J = 5.1 Hz), 3.75 (3H, s), 3.02
(3H, s); 13C NMR (63 MHz, CDCl3): d 163.2, 157.3,
132.5, 131.8, 130.2, 129.8, 129.3, 129.1, 128.5, 128.1,
119.4, 114.9, 102.1, 79.8, 61.9, 55.9, 39.2.
An analogous procedure was used to prepare sulfonates
35b and 35c.
4.19. ( )-(3R,4S)-3-Benzenesulfonyl-N-(4-methoxyphenyl)-
4-phenylazetidin-2-one (35b)
1
yellow oil. H NMR (250 MHz, CDCl3): d 7.43 (1H,
d,J = 1.4 Hz), 7.40–7.20 (5H, m), 6.80 (2H, d,
J = 9.0 Hz), 5.61 (1H, d, J = 4.7 Hz), 5.09 (2H, d,
J = 5.9 Hz), 5.02 (1H, d, J = 4.9 Hz), 3.92 (2H, d,
J = 5.5 Hz), 3.73 (3H, s); 13C NMR (63 MHz, CDCl3):
d 163.8, 156.3, 133.2, 133.0, 131.2, 130.3, 129.4, 129.0,
126.9, 118.6, 118.1, 114.3, 82.5, 71.9, 58.9, 55.4.
White solid, mp 162–165 ꢁC, 93% yield. 1H NMR
(250 MHz, CDCl3): d 8.05 (2H, d, J = 4.0 Hz), 7.75–
7.60 (5H, m), 7.50–7.18 (5H, m), 6.79 (2H, d,
J = 8.6 Hz), 5.90 (1H, d, J = 5.0 Hz), 5.77 (1H, d,
J = 5.0 Hz), 3.74 (3H, s); 13C NMR (63 MHz, CDCl3):
d 160.7, 157.6, 136.2, 134.5, 133.2, 132.1, 129.6, 129.2,
128.5, 128.2, 127.8 (2C), 127.1 (2C), 119.3 (2C), 114.8
(2C), 79.6, 62.1, 55.8.
4.16. ( )-(3R,4S)-4-(2-Chlorophenyl)-3-methoxymethoxy-
N-(4-methoxyphenyl)azetidin-2-one (31c)
4.20. ( )-(3R,4S)-N-(4-Methoxyphenyl)-4-phenyl-3-(4-tol-
uenesulfonyl)azetidin-2-one (35c)
1H NMR (250 MHz, CDCl3): d 7.42 (1H, d, J = 7.8 Hz),
7.26 (5H, m), 6.80 (2H, d, J = 8.8 Hz), 5.64 (1H, d,
J = 5.0 Hz), 5.21 (1H, d, J = 5.0 Hz) 4.56 (2H, s), 3.74
(3H, s), 3.19 (3H, s); 13C NMR (63 MHz, CDCl3): d
163.9, 156.4, 133.3, 131.5, 130.4, 129.5, 126.9, 118.6,
114.4, 96.6, 80.4, 58.9, 55.7, 55.4.
White solid, mp 148–151 ꢁC, 75% yield. 1H NMR
(250 MHz, CDCl3): d 7.46 (2H, d, J = 7.9 Hz), 7.36–
7.28 (8H, m), 6.77 (2H, d, J = 8.5 Hz), 5.78 (1H, d,
J = 4.9 Hz), 5.28 (1H, d, J = 4.9 Hz), 3.73 (3H, s), 2.43
(3H, s); 13C NMR (63 MHz, CDCl3) d 161.2, 157.6,
147.2, 138.8, 133.4, 131.9, 130.2 (2C), 129.3, 128.5,
128.2, 127.8 (2C), 127.4 (2C), 119.1 (2C), 114.9 (2C),
79.6, 58.1, 55.8, 22.1.
4.17. Synthesis of 3-acetoxy-N-(4-methoxyphenyl)-4-phenyl-
azetidin-2-one (31e)
To a stirred solution of benzaldehyde N-(4-methoxy-
phenyl)imine (5.31 g, 25.2 mmol) and triethylamine
(7.64 g, 75.5 mmol) was added a solution of acetoxyace-
tyl chloride (20) (5.15 g, 37.7 mmol) in CH2Cl2 dropwise
over 10 min. The resultant mixture was stirred at rt until
TLC indicated the disappearance of starting material.
The solvent was removed under reduced pressure, and
the crude material was purified by washing with ice-cold
MeOH to give 6.89 g (89%)of 31e in as white solid, mp
4.21. Synthesis of ( )-(3R,4S)-4-(2-chlorophenyl)-3-hy-
droxy-N-(4-methoxyphenyl)-3-methylazetidin-2-one (38d)
To a solution of 27 (0.496 g, 1.65 mmol) in 9 mL of
anhydrous THF was added methylmagnesium bromide
(0.549 mL, 1.05 mmol, 1.0 equiv) at ꢁ40 ꢁC. The resul-
tant solution was stirred for 1 h. The reaction mixture
was quenched by adding an equal volume of 5% ammo-
nium chloride at ꢁ40 ꢁC. The mixture was warmed to
room temperature and extracted with EtOAc (3·
50 mL). The combined organic layers were dried with
MgSO4, and the solvent was removed under reduced
pressure to yield a yellow solid, 0.383 g (73%) of 38d.
No further purification was necessary. 1H NMR
(250 MHz, CDCl3): d 7.47 (1H, d, J = 7.2 Hz), 7.29–
7.19 (5H, m), 6.83 (2H, d, J = 8.9 Hz), 5.40 (1H, s),
3.76 (3H, s), 1.83 (3H, s); 13C NMR (63 MHz, CDCl3):
d 168.2, 156.4, 133.5, 132.0, 130.4, 129.7, 129.3, 128.1,
126.9, 118.7, 114.4, 83.9, 66.1, 55.4, 21.9.
1
153–155 ꢁC. H NMR (250 MHz, CDCl3): d 7.34–7.30
(5H, t), 7.28 (2H, d, J = 8.9 Hz), 6.79 (2H, d,
J = 8.9 Hz), 5.92 (1H, d, J = 4.8 Hz), 5.33 (1H, d,
J = 4.8 Hz), 3.74 (3H, s), 1.66 (3H, s); 13C NMR
(63 MHz, CDCl3): d 169.1, 161.2, 156.5, 132.2, 130.2,
128.7, 128.4, 127.8, 118.7, 61.3, 55.3, 19.7.
4.18. Synthesis of ( )-(3R,4S)-N-(4-methoxyphenyl)-3-
methylsulfonyl-4-phenylazetidin-2-one (35a)
Compound 34 (269 mg, 1.00 mmol) was dissolved in dry
CH2Cl2 (15 mL) and 60 mg (1.50 mmol) of NaH (60% in
mineral oil, unwashed) was added. After stirring for
30 min at room temperature to the resultant solution
methanesulfonyl chloride (115 mg, 1.00 mmol) was then
added dropwise. The resultant solution was then stirred
at rt for 30 min. The solution was washed with brine (3·
15 mL). The organic layer was dried with anhydrous
MgSO4, filtered, and evaporated, and the residue was
washed with cold MeOH to give 267 mg (77%) of 35a
as a white solid in; mp 158–160 ꢁC. 1H NMR
(250 MHz, CDCl3): d 7.48 (2H, d, J = 7.8 Hz), 7.39–
An analogous procedure was used to prepare com-
pounds 38e,g.
4.22. ( )-(3R,4S)-4-(2-Chlorophenyl)-3-hydroxy-N-(4-
methoxyphenyl)-3-(2-propenyl)azetidin-2-one (38e)
1
Orange oil, 84% yield. H NMR (250 MHz, CDCl3): d
7.47 (1H, d, J = 7.3 Hz), 7.33–7.23 (5H, m), 6.83 (2H,
d, J = 8.9 Hz), 5.99–5.93 (1H, m), 5.51 (1H, s), 5.39–
5.24 (2H, m), 3.77 (3H, s), 2.90 (2H, d, J = 7.4 Hz);

E. Turos et al. / Bioorg. Med. Chem. 13 (2005) 6289–6308
6303
13C NMR (63 MHz, CDCl3): d 166.9, 156.4, 133.4,
131.8, 130.9, 130.4, 129.8, 129.4, 128.4, 127.0, 120.8,
118.7, 114.4, 85.6, 63.2, 55.4, 40.1.
reduced pressure to yield a yellow oil, 0.275 g (85%)
of 40d. No further purification was necessary. 1H
NMR (250 MHz, CDCl3): d 7.42 (1H, d, J = 8.3 Hz),
7.29–7.12 (5H, m), 6.82 (2H, d, J = 8.9 Hz), 5.47 (1H,
s), 3.75 (3H, s), 1.94 (3H, s), 1.60 (H, s); 13C NMR
(63 MHz, CDCl3): d 168.3, 164.4, 156.4, 134.0, 130.8,
130.4, 129.4, 129.3, 129.2, 126.4, 118.5, 114.3, 86.5,
64.8, 55.3, 20.2, 19.8.
4.23. ( )-(3R,4S)-4-(2-Chlorophenyl)-3-hydroxy-N-(4-
methoxyphenyl)-3-phenylazetidin-2-one (38g)
Yellow solid, 79% yield. 1H NMR (250 MHz, CDCl3): d
7.67 (2H, dd, J = 7.6, 2.0 Hz), 7.50–7.29 (10H, m), 6.85
(2H, d, J = 9.2 Hz), 5.68 (1H, s), 3.78 (3H, s); 13C NMR
(63 MHz, CDCl3): d 166.6, 156.6, 138.2, 133.8, 131.4,
129.9, 129.8, 128.6, 126.9, 125.5, 118.9, 114.4, 87.3,
67.2, 55.4.
An analogous procedure was used to prepare com-
pounds 40e,g.
4.27. ( )-(3R,4S)-3-Acetoxy-4-(2-chlorophenyl)-N-(4-
methoxyphenyl)-3-(2-propenyl)azetidin-2-one (40e)
4.24. Synthesis of ( )-(3R,4S)-3-allyloxy-4-(2-chlorophe-
nyl)-N-(4-methoxyphenyl)-3-methylazetidin-2-one (39a)
1
Orange oil, 99% yield. H NMR (250 MHz, CDCl3): d
7.42 (1H, d, J = 7.6 Hz), 7.23 (4H, d, J = 5.0 Hz), 7.17
(1H, d, J = 7.5 Hz), 6.81 (2H, d, J = 8.9 Hz), 5.93 (1H,
m), 5.56 (1H, s), 5.29 (2H, m), 3.74 (3H, s), 3.09 (2H,
d, J = 7.3 Hz); 13C NMR (63 MHz, CDCl3): d 168.3,
163.5, 156.5, 134.0, 130.8, 130.3, 129.9, 129.5, 126.4,
121.1, 118.6, 114.4, 88.4, 61.8, 55.4, 37.5, 20.2.
To a solution of b-lactam 38d (0.10 g, 0.32 mmol) in
5 mL of dry CH2Cl2 was added NaH (60% suspension
in mineral oil, 0.018 g, 0.48 mmol) and the mixture was
stirred for 15 min. Allyl bromide (0.076 g, 0.62 mmol)
was then added, along with 5 mg TBAI (tetrabutylam-
monium iodide). The mixture was refluxed for 24 h or
until the TLC indicated the disappearance of the start-
ing material. The reaction was quenched with a 5%
solution of NH4Cl and extracted with CH2Cl2 (3·
5 mL). The combined organic layers were washed with
brine and dried over anhydrous Na2SO4. The crude
material was purified by column chromatography on
silica gel (1:9 EtOAc/hexanes) to give 0.07 g (87%) of
39a as a yellow semisolid. 1H NMR (250 MHz,
CDCl3): d 7.42 (1 H, d, J = 7.7 Hz), 7.29–7.13 (5H,
m), 6.80 (2H, d, J = 8.8 Hz), 5.46–5.35 (1H, m), 5.32
(1H, s), 4.86–4.79 (2H, m), 3.90–3.85 (2H, m), 3.73
(3H, s), 1.81 (3H, s); 13C NMR (63 MHz, CDCl3): d
166.0, 156.3, 133.6, 133.3, 132.1, 130.6, 129.4, 129.1,
128.7, 126.6, 118.6, 116.0, 114.3, 88.5, 66.9, 65.4,
55.3, 19.4.
4.28. ( )-(3R,4S)-3-Acetoxy-4-(2-chlorophenyl)-N-(4-
methoxyphenyl)-3-phenylazetidin-2-one (40g)
1
Yellow oil, 85% yield. H NMR (250 MHz, CDCl3): d
7.81 (2H, d, J = 8.9 Hz), 7.46–7.29 (9H, m), 6.83 (1H,
d, J = 8.9 Hz), 6.15 (1H, s), 3.76 (3H, s), 1.68 (3H, s);
13C NMR (63 MHz, CDCl3): d 167.8, 162.6, 156.6,
134.7, 130.8, 129.8, 129.7, 129.2, 128.9, 128.5, 126.9,
126.4, 118.8, 114.4, 89.7, 63.5, 55.4, 20.3.
4.29. Synthesis of ( )-(3S,4S)-3-ahloro-4-(2-chlorophe-
nyl)-azetidin-2-one (25a)
To a solution of lactam 24a (0.335 g, 1.00 mmol) in
CH3CN (2.2 mL) at rt was added dropwise a solution
of ceric ammonium nitrate (1.64 g, 3.00 mmol) in
1.5 mL water. The reaction was stirred for 5 min and
then diluted with EtOAc (20 mL). The resultant solution
was washed sequentially with water (20 mL), 5% NaH-
CO3 (2· 20 mL), 5% NaHSO3 (2· 20 mL) and brine
(20 mL). The organic layer was dried with anhydrous
MgSO4, filtered, and evaporated, and the residue was
chromatographed on silica gel (1:2 EtOAc/hexanes) to
give 0.172 g (75%) of 25a as a yellow oil. 1H NMR
(250 MHz, CDCl3): d 7.10–7.50 (m, 4H), 5.10 (1H, d,
J = 1.5 Hz), 4.49 (1H, d, J = 2.0 Hz); 13C NMR
(63 MHz, CDCl3): d 164.7, 134.7, 132.9, 129.9, 129.8,
127.3, 126.2, 63.4, 59.7.
4.25. ( )-(3R,4S)-4-(2-Chlorophenyl)-3-methoxymeth-
oxy-N-(4-methoxyphenyl)-3-methylazetidin-2-one (39c)
1H NMR (250 MHz, CDCl3): d 7.43 (1H, d, J = 7.5 Hz),
7.30–7.17 (5H, m), 6.85–6.79 (2H, m), 5.30 (1H, s), 4.59
(2H, s), 3.75 (3H, s), 3.08 (3H, s), 1.84 (3H, s); 13C NMR
(63 MHz, CDCl3): d 167.2, 156.3, 133.1, 131.3, 130.5,
129.4, 126.7, 118.4, 114.2, 96.6, 81.4, 58.7, 55.6, 55.4,
19.2.
4.26. Synthesis of ( )-(3R,4S)-3-acetoxy-4-(2-chlorophe-
nyl)-N-(4-methoxyphenyl)-3-methylazetidin-2-one (40d)
To a solution of 38d (0.287 g, 0.903 mmol, 2.0 equiv) in
5 mL of CH2Cl2 was added NaH (0.072 g, 3.01 mmol).
The resultant suspension was stirred until the bubbling
stopped, and then acetyl chloride (0.077 mL,
1.08 mmol, 1.2 equiv) was added via syringe. The reac-
tion was followed by TLC and showed that the reac-
tion was complete immediately after adding the acetyl
chloride. The reaction was quenched by adding a 1%
solution of NaHCO3 (5 mL) and extracted with EtOAc
(3· 20 mL). The combined organic layers were dried
over MgSO4, and the solvent was removed under
An analogous procedure was used to prepare com-
pounds 25b–c, 29a–d, 32a–c, 36a–c, and 41a–g.
4.30. ( )-(3S,4S)-4-(2-Chlorophenyl)-3-iodoazetidin-2-
one (25b)
Yellow oil in 72% yield; 1H NMR (250 MHz, CDCl3): d
7.50 (1H, m), 7.10–7.30 (4H, m), 6.50 (1H, br s),
5.62(1H, d, J = 2.5 Hz), 5.26 (1H, d, J = 2.0 Hz); 13C
NMR (63 MHz, CDCl3): d 166.0, 135.9, 132.9, 129.8,
127.3, 126.8, 125.9, 59.6, 21.8.

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E. Turos et al. / Bioorg. Med. Chem. 13 (2005) 6289–6308
4.31. ( )-(3S,4S)-3-Azido-4-(2-chlorophenyl)azetidin-2-
one (25c)
J = 4.5 Hz) 4.91 (1H, d, J = 4.5 Hz), 3.47–3.38 (1H,
m), 3.25–3.19 (1H, m), 1.33–1.24 (2H, m), 0.54 (3H, t,
J = 7.4 Hz); 13C NMR (63 MHz, CDCl3): d 168.6,
133.9, 133.0, 129.2, 129.0, 128.4, 126.8, 85.8, 73.1,
56.1, 22.6, 10.0.
1
Yellow oil, H NMR (250 MHz, CDCl3): d 7.38–7.19
(4H, m), 6.20 (1H, br s), 4.90 (1H, d, J = 1.8 Hz), 4.33
(1H, d, J = 1.8 Hz); 13C NMR (63 MHz, CDCl3) d
172.1, 142.3, 132.2, 128.4, 128.2, 128.0, 126.7, 64.2, 44.3.
4.38. ( )-(3R,4S)-4-(2-Chlorophenyl)-3-methoxymeth-
oxy-azetidin-2-one (32c)
4.32. ( )-(3S,4S)-4-(2-Chlorophenyl)-3-propylaminoazeti-
din-2-one (29a)
1H NMR (250 MHz, CDCl3): d 7.48–7.26 (4H, m), 6.26
(1H, br s), 5.29 (1H, d, J = 5.0 Hz), 5.17 (1H, d,
J = 5.0 Hz), 4.55 (2H, s), 3.21 (3H, s); 13C NMR
(63 MHz, CDCl3): d 169.3, 133.3, 129.9, 128.7, 128.5,
127.7, 126.8, 96.1, 84.2, 57.3, 50.2.
1H NMR (250 MHz, CDCl3): d 7.88 (1H, br s), 7.31–7.12
(4H, m), 4.73 (1H, d, J = 6.5 Hz), 4.49 (1H, d, J = 6.4 Hz),
3.83 (1H, br s), 1.84 (4H, m), 1.43 (4H, m); 13C NMR
(63 MHz, CDCl3): d 179.5, 140.4, 133.2, 130.4, 129.5,
129.2, 127.6, 55.4, 42.7, 42.5, 33.3, 33.2, 27.9, 27.6.
4.39. ( )-(3R,4S)-3-Methylsulfonyl-4-phenylazetidin-2-
one (36a)
4.33. ( )-(3S,4S)-3-Benzylamino-4-(2-chlorophenyl)azeti-
din-2-one (29b)
Yellow oil in 90% yield. 1H NMR (250 MHz, CDCl3): d
7.36–7.19 (5H, m), 6.58 (1H, br s), 5.71 (1H, d,
J = 4.9 Hz), 4.99 (1H, d, J = 4.9 Hz), 2.68 (3H, s); 13C
NMR (63 MHz, CDCl3): d 164.7, 134.2, 129.6, 129.1,
128.7, 128.5, 127.8, 81.7, 58.2, 39.2.
1
86% yield. H NMR (250 MHz, CDCl3): d 7.78 (1H, br
s), 7.33–7.16 (9H, m), 4.53 (1H, d, J = 6.3 Hz), 4.42 (1H,
d, J = 6.1 Hz); 13C NMR (63MHz, CDCl3): d 187.6,
160.0, 137.0, 134.7, 130.4, 130.3, 130.1, 130.0, 129.7,
129.6, 129.2, 128.3, 127.6, 55.7, 51.8, 43.6.
4.40. ( )-(3R,4S)-3-Benzenesulfonyl-4-phenylazetidin-
2-one (36b)
4.34. ( )-(3S,4S)-4-(2-Chlorophenyl)-3-diethylaminoaz-
etidin-2-one (29c)
1
Yellow oil, 88% yield. H NMR (250 MHz, CDCl3): d
8.05 (1H, d, J = 8.0 Hz), 7.78–7.63 (6H, m), 7.48–7.20
(3H, m), 6.42 (1H, br s), 5.83 (1H, d,J = 4.8 Hz), 5.41
(1H, d, J = 4.8 Hz); 13C NMR (63 MHz, CDCl3): d
164.2, 136.0, 134.5, 134.3, 129.6, 129.2, 128.9, 128.5,
128.2, 128.1, 127.3, 127.1, 116.5, 81.5, 58.4.
1
80% yield. H NMR (250 MHz, CDCl3): d 7.62 (1H, br
s), 7.28–7.19 (4H, m), 4.75 (1H, d, J = 5.2 Hz), 4.49 (1H,
d, J = 5.2 Hz), 3.18 (2H, q, J = 7.1 Hz), 3.09 (2H, q,
J = 7.2 Hz), 1.15 (6H, t, J = 7.1 Hz); 13C NMR
(63 MHz, CDCl3): d 165.9, 134.2, 132.2, 131.1, 129.7,
129.4, 127.5, 55.8, 49.3, 42.3, 37.8, 14.0, 12.2.
4.41. ( )-(3R,4S)-4-Phenyl-3-(4-toluenesulfonyl)azetidin-
2-one (36c)
4.35. ( )-(3S,4S)-4-(2-Chlorophenyl)-3-diisobutylamino-
azetidin-2-one (29d)
1
Yellow oil, 85% yield. H NMR (250 MHz, CDCl3): d
7.45–7.18 (9H, m), 6.41 (1H, br s), 5.69 (1H, d,
J = 2.4 Hz), 4.95 (1H, d, J = 4.7 Hz), 2.41 (3H, s); 13C
NMR (63 MHz, CDCl3): d 164.3, 136.1, 134.6, 134.4,
130.0, 129.3, 129.0, 128.5, 128.3, 128.2, 126.6 (2C),
125.1 (2C), 81.6, 58.5.
1
78% yield. H NMR (250 MHz, CDCl3): d 7.31–7.20
(4H, m), 6.96 (2H, d, J = 8.8 Hz), 6.65 (2H, d,
J = 8.8 Hz), 4.98 (2H, s), 3.68 (3H, s), 2.90 (4H, t,
J = 6.6 Hz), 1.90 (1H, m), 1.45 (1H, m), 0.76 (6H, d,
J = 6.6 Hz), 0.35 (6H, d, J = 6.6 Hz); 13C NMR
(63 MHz, CDCl3): d 163.1, 136.9, 135.2, 134.1, 130.2,
129.4 127.5, 71.9, 56.2, 55.0, 41.6, 28.3, 26.8, 20.6,
20.5, 20.3, 20.2.
4.42. ( )-(3R,4S)-3-Allyloxy-4-(2-chlorophenyl)-3-
methylazetidin-2-one (41a)
1H NMR (250 MHz, CDCl3): d 7.48–7.26 (4H, m),
6.25 (1H, br s), 5.39–5.35 (1H, m), 4.97 (1H, s),
4.86–4.78 (2H, m), 3.82–3.79 (2H, m), 1.80 (3H, s);
13C NMR (63 MHz, CDCl3): d 170.5, 134.5, 133.7,
133.1, 129.2, 128.8, 127.8, 126.6, 116.0, 90.8, 66.8,
62.6, 20.1.
4.36. ( )-(3R,4S)-3-Allyloxy-4-(2-chlorophenyl)-azetidin-
2-one (32a)
Brown semi-solid 69% yield; 1H NMR (250 MHz,
CDCl3): d 7.50–7.26 (4H, m), 6.25 (1H, br s), 5.70–
5.55 (1H, m), 5.26 (1H, d, J = 4.6 Hz) 5.11–4.99 (2H,
m), 4.97 (1H, d, J = 4.6 Hz), 3.93–3.89 (2H, m); 13C
NMR (63 MHz, CDCl3): d 168.7, 133.8, 133.2, 129.1,
128.4, 126.9, 117.8, 84.6, 71.8, 56.0.
4.43. ( )-(3R,4S)-4-(2-Chlorophenyl)-3-methyl-3-propyl-
oxyazetidin-2-one (41b)
1H NMR (250 MHz, CDCl3): d 7.46–7.27 (4H, m),
6.65 (1H, br s), 4.93 (1H, s), 3.25–3.15 (2H, m),
1.74 (3H, s), 1.18–1.04 (2H, m), 0.47 (3H, t,
J = 7.4 Hz); 13C NMR (63 MHz, CDCl3): d 168.2,
133.2, 130.3, 129.0, 127.1, 126.5, 126.2, 89.8, 63.2,
22.5, 19.2, 10.2.
4.37. ( )-(3R,4S)-4-(2-Chlorophenyl)-3-propoxyazetidin-
2-one (32b)
1H NMR (250 MHz, CDCl3): d 7.49 (1H, d, J = 1.9 Hz),
7.46–7.27 (3H, m), 6.30 (1H, br s), 5.26 (1H, d,

E. Turos et al. / Bioorg. Med. Chem. 13 (2005) 6289–6308
6305
4.44. ( )-(3R,4S)-4-(2-Chlorophenyl)-3-methoxymeth-
oxy-3-methylazetidin-2-one (41c)
J = 2.0 Hz), 2.50 (3H, s); 13C NMR (63 MHz, CDCl3):
d 166.0, 131.2, 130.4, 128.5, 127.2, 66.2, 62.5, 20.6.
1H NMR (250 MHz, CDCl3): d 7.44–7.22 (4H, m), 6.71
(1H, br s), 4.93 (1H, s), 4.52 (2H, s), 3.05 (3H, s), 1.80
(3H, s); 13C NMR (63 MHz, CDCl3): d 170.2, 134.7,
133.1, 129.2, 128.9, 127.8, 126.7, 93.0, 89.8, 62.6, 55.6,
20.3.
An analogous procedure was used to prepare com-
pounds 19b–c, 26a–d, 30a–c, 33a–c, 37a–g, 43a–b, 43f–
j, 43l, 43n, and 43q–t.
4.50. ( )-(3S,4S)-4-(2-Chlorophenyl)-3-iodo-N-methylthi-
oazetidin-2-one (19b)
4.45. ( )-(3R,4S)-3-Acetoxy-4-(2-chlorophenyl)-3-meth-
yl-azetidin-2-one (41d)
1
Brown oil, 86% yield H NMR (250 MHz, CDCl3) d
(1H, d, J = 3.5 Hz), 7.39–7.36 (2H, m), 7.10–7.20
(1H, m), 5.60 (1H, d, J = 2.5 Hz), 5.30 (1H, d,
J = 2.5 Hz), 2.58 (3H, s); 13C NMR (63 MHz, CDCl3):
d 168.6, 134.2, 133.8, 130.2, 127.5, 127.1, 126.7, 66.4,
21.4, 20.3.
1
Brown oil, 94% yield. H NMR (250 MHz, CDCl3): d
7.49–7.18 (4H, m), 5.00 (1H, s), 1.86 (3H, s), 1.54
(3H, s); 13C NMR (63 MHz, CDCl3):
168.4, 133.2, 133.1, 129.0, 128.8, 126.4, 88.1, 62.2,
29.6, 20.1.
d 169.4,
4.51. ( )-(3S,4S)-3-Azido-3-(2-chlorophenyl)-N-meth-
ylthioazetidin-2-one (19c)
4.46. ( )-(3R,4S)-3-Acetoxy-4-(2-chlorophenyl)-3-(2-pro-
pen-yl)azetidin-2-one (41e)
Yellow solid, 83% yield. 1H NMR (250 MHz, CDCl3): d
7.35–7.19 (4H, m), 5.74 (1H, d, J = 1.8 Hz), 4.95 (1H, d,
J = 1.8 Hz), 2.37 (3H, s); 13C NMR (63 MHz, CDCl3): d
168.2, 142.5, 133.5, 128.9, 128.7, 128.2, 128.0, 60.8, 52.2,
20.8.
1
Brown oil, 35% yield. H NMR (250 MHz, CDCl3): d
7.51–7.37 (1H, m), 7.36–7.23 (3H, m), 7.02 (1H, br s),
5.99–5.92 (1H, m), 5.37–5.25 (2H, m), 5.13 (1H, d,
J = 1.5 Hz), 2.99 (2H, d, J = 7.3 Hz), 1.58 (3H, s); 13C
NMR (63 MHz, CDCl3): d 168.2, 163.4, 156.4, 133.9,
130.8, 129.9, 129.4, 126.4, 120.9, 118.5, 114.3, 88.3,
55.3, 37.5, 20.1.
4.52. ( )-(3S,4S)-N-Methylthio-3-propylaminoazetidin-2-
one (26a)
1
4.47. ( )-(3R,4S)-3-Acetoxy-4-(2-chlorophenyl)-3-propy-
lazetidin-2-one (41f)
68% yield. H NMR (250 MHz, CDCl3): d 7.33–7.16
(4H, m), 4.51 (1H, d, J = 6.4 Hz), 3.84 (1H, br s), 2.41
(3H, s), 1.85 (4H, m), 1.47 (4H, m); 13C NMR
(63 MHz, CDCl3): d 189.5, 140.2, 133.2, 130.4, 129.5,
127.6, 127.4, 48.8, 47.7, 42.7, 33.3, 33.2, 27.8, 27.7, 20.2.
1
Brown oil, 91% yield. H NMR (250 MHz, CDCl3): d
7.48–7.18 (4H, m), 6.65 (1H, s), 5.01 (1H, s), 2.35–2.11
(2H, m), 1.72–1.50 (2H, m), 1.58 (3H, s), 1.57 (3H, s),
0.98 (3H, t, J = 7.3 Hz); 13C NMR (63 MHz, CDCl3):
d 169.1, 168.6, 133.2, 133.1, 129.1, 128.9, 126.4, 115.9,
90.0, 60.6, 36.1, 20.0, 16.6, 14.1.
4.53. ( )-(3S,4S)-3-Benzylamino-N-methylthioazetidin-2-
one (26b)
1
64% yield. H NMR (250 MHz, CDCl3): d 7.25–7.05
4.48. ( )-(3R,4S)-3-Acetoxy-4-(2-chlorophenyl)-3-phenyl-
azetidin-2-one (41g)
(6H, m), 6.61 (3H, m), 4.53 (1H, d, J = 6.3 Hz), 4.42
(1H, d, J = 6.1 Hz), 2.41 (3H, s); 13C NMR (63 MHz,
CDCl3): d 187.5, 160.0, 137.0, 134.7, 130.4, 130.3,
130.1, 130.0, 129.7, 129.6, 129.2, 128.3, 127.6, 55.7,
51.8, 43.6, 19.4.
1
Brown oil, 95% yield. H NMR (250 MHz, CDCl3): d
7.75 (2H, d, J = 7.1 Hz), 7.36–7.23 (3H, m), 7.02 (1H,
br s), 5.99–5.92 (1H, m), 5.37–5.25 (2H, m), 5.13 (1H,
d, J = 1.5 Hz), 2.99 (2H, d, J = 7.3 Hz), 1.58 (3H, s);
13C NMR (63 MHz, CDCl3): d 168.0, 167.7, 134.8,
133.9, 133.1, 129.4, 129.2, 128.9, 128.8, 128.5, 126.4,
126.0, 91.6, 61.9, 20.3.
4.54. ( )-(3S,4S)-3-Diethylamino-N-methylthioazetidin-
2-one (26c)
1
54% yield. H NMR (250 MHz, CDCl3): d 7.62 (1H, br
s), 7.28–7.19 (4H, m), 4.75 (1H, d, J = 5.2 Hz), 4.49 (1H,
d, J = 5.2 Hz), 3.18 (2H, q, J = 7.1 Hz), 3.09 (2H, q,
J = 7.2 Hz), 2.41 (3H, s), 1.15 (6H, t, J = 7.1 Hz); 13C
NMR (63 MHz, CDCl3): d 175.2, 134.5, 129.2, 128.9,
128.7, 128.2, 127.8, 60.8, 54.2, 48.6, 26.2, 25.8, 20.8,
11.4, 11.3.
4.49. Synthesis of ( )-(3S,4S)-3-chloro-4-(2-chlorophe-
nyl)-N-methylthioazetidin-2-one (19a)
To a solution of 25a (0.027 g, 0.13 mmol) in benzene
were
added
N-methylthiophthalimide
(0.025 g,
0.133 mmol) and 1 drop of triethylamine. The mixture
was refluxed overnight and washed with 1% KOH.
The organic layer was dried over Na2SO4 and the sol-
vent was removed under reduced pressure. The crude
material was purified by column chromatography (1:4
EtOAc/hexanes) to yield 0.021 g (71%) of 19a as a
brown oil. 1H NMR (250 MHz, CDCl3) d 7.20–7.40
(4H, m), 5.10 (1H, d, J = 1.5 Hz), 4.60 (1H, d,
4.55. ( )-(3S,4S)-3-Diisobutylamino-N-methylthioazeti-
din-2-one (26d)
1
48% yield. H NMR (250 MHz, CDCl3): d 7.28–7.13
(4H, m), 4.48 (2H, d, J = 6.2 Hz), 3.59 (2H, d,
J = 7.5 Hz), 3.13 (2H, d, J = 7.5 Hz), 1.99 (1H, m) 1.96
(1H, m), 0.81 (12H, m); 13C NMR (63 MHz, CDCl3):

6306
E. Turos et al. / Bioorg. Med. Chem. 13 (2005) 6289–6308
d 173.1, 140.2, 136.9, 130.2, 129.4 127.5, 127.2, 49.9,
48.2, 41.6, 41.3, 39.2, 27.3, 26.8, 23.5, 23.4, 20.2.
4.62. ( )-(3R,4S)-3-Allyloxy-4-(2-chlorophenyl)-3-methyl-
N-methylthioazetidin-2-one (37a)
1
4.56. ( )-(3R,4S)-3-Allyloxy-4-(2-chlorophenyl)-N-meth-
ylthioazetidin-2-one (30a)
26% yield. H NMR (250 MHz, CDCl3): d 7.40–7.26
(4H, m), 5.41–5.34 (1H, m), 5.05 (1H, s), 4.84–4.77
(2H, m), 3.83–3.75 (2H, m), 2.48 (3H, s), 1.73 (3H, s);
13C NMR (63 MHz, CDCl3): d 169.7, 133.8, 129.7,
129.5, 128.9, 126.9, 116.4, 85.6, 69.5, 67.1, 21.5, 19.1.
1
25% yield. H NMR (250 MHz, CDCl3): d 7.39 (1H, d,
J = 5.8 Hz), 7.38–7.27 (3H, m), 5.62–5.51 (1H, m), 5.34
(1H, d, J = 4.8 Hz), 5.06 (2H, d, J = 11.6 Hz), 4.99
(1H, d, J = 4.8 Hz), 3.93–3.78 (2H, m), 2.45 (3H, s);
13C NMR (63 MHz, CDCl3): d 170.4, 133.7, 132.8,
131.6, 129.5, 129.4, 129.1, 126.7, 118.0, 84.4, 71.6,
62.8, 21.7.
4.63. ( )-(3R,4S)-4-(2-Chlorophenyl)-3-methyl-N-methyl-
thio-3-propoxyazetidin-2-one (37b)
1H NMR (250 MHz, CDCl3): d 7.36–7.28 (4H, m), 4.51
(1H, s), 3.33–3.29 (1H, m), 3.04–3.01 (1H, m), 2.40 (3H,
s), 1.61 (3H, s), 1.23–1.14 (2H, m), 0.59 (3H, t,
J = 7.4 Hz); 13C NMR (63 MHz, CDCl3): d 168.8,
133.4, 130.4, 129.2, 127.3, 126.9, 126.4, 88.5, 64.1,
23.0, 19.5, 16.1, 10.2.
4.57. ( )-(3R,4S)-4-(2-Chlorophenyl)-N-methylthio-3-prop-
oxyazetidin-2-one (30b)
1H NMR (250 MHz, CDCl3): d 7.34 (1H, d, J = 3.6 Hz),
7.27–7.24 (3H, m), 5.32 (1H, d, J = 4.8 Hz), 4.91 (1H, d,
J = 4.8 Hz), 3.38–3.32 (1H, m), 3.09–3.04 (1H, m), 2.42
(3H, s), 1.27–1.19 (2H, m), 0.49 (3H, t, J = 7.5 Hz); 13C
NMR (63 MHz, CDCl3): d 170.5, 134.6, 133.8, 131.6,
129.4, 129.1, 126.6, 85.6, 72.9, 63.0, 22.4, 21.8, 10.0.
4.64. ( )-(3R,4S)-4-(2-Chlorophenyl)-3-methoxymethoxy-
3-methyl-N-methylthioazetidin-2-one (37c)
1H NMR (250 MHz, CDCl3): d 7.36-7-.18 (4H, m), 4.98
(1H, s), 4.46 (2H, s), 3.01 (3H, s), 2.43 (3H, s), 1.72 (3H,
s); 13C NMR (63 MHz, CDCl3): d 170.1, 134.2, 130.4,
128.6, 128.4, 127.9, 126.9, 96.3, 85.1, 57.8, 51.2, 20.0,
16.2.
4.58. ( )-(3R,4S)-4-(2-Chlorophenyl)-3-methoxymethoxy-
N-methylthioazetidin-2-one (30c)
1H NMR (250 MHz, CDCl3): d 7.39 (1H, d, J = 5.2 Hz),
7.38–7.26 (3H, m), 5.37 (1H, d, J = 5.1 Hz), 5.19 (1H, d,
J = 5.1 Hz), 4.51 (1H, d, J = 6.6 Hz), 3.14 (3H, s), 2.47
(3H, s); 13C NMR (63 MHz, CDCl3): d 170.6, 134.6,
131.8, 129.5, 129.4, 128.8, 126.8, 96.3, 82.2, 62.8, 55.7,
21.7.
4.65. ( )-(3R,4S)-3-Acetoxy-4-(2-chlorophenyl)-3-methyl-
N-methylthioazetidin-2-one (37d)
Yellow oil in 40% yield. 1H NMR (250 MHz, CDCl3): d
7.39–7.38 (1H, m), 7.26 (3H, s), 5.15 (1H, s), 2.49 (3H,
s), 1.85 (3H, s), 1.57 (3H, s); 13C NMR (63 MHz,
CDCl3): d 171.1, 168.1, 134.3, 131.2, 129.5, 129.4,
128.9, 126.3, 88.1, 68.9, 21.3, 20.1, 19.5.
4.59. ( )-(3R,4S)-3-Methylsulfonyl-N-methylthio-4-phenyl-
azetidin-2-one (33a)
Yellow solid in 72% yield. 1H NMR (250 MHz, CDCl3):
d 7.47–7.26 (5H, m), 5.91 (1H, d, J = 5.3 Hz), 5.54 (1H,
d, J = 5.3 Hz), 2.96 (3H, s), 2.53 (3H, s); 13C NMR
(63 MHz): d 165.7, 133.2, 129.4, 129.0, 128.7, 127.4,
126.0, 79.9, 60.9, 38.0, 20.9.
4.66. ( )-(3R,4S)-3-Acetoxy-4-(2-chlorophenyl)-N-meth-
ylthio-3-(2-propenyl)azetidin-2-one (37e)
1
Yellow oil, 33% yield. H NMR (250 MHz, CDCl3): d
7.40–7.37 (1H, m), 7.26 (3H, br s), 5.93–5.27 (2H, m),
5.30 (1H, s), 3.03-2-.96 (2H, m), 2.48 (3H, s), 1.58
(3H, s); 13C NMR (63 MHz, CDCl3): d 170.0, 168.1,
134.3, 131.2, 129.5, 129.1 126.3, 121.5, 89.9, 65.3, 37.2,
21.8, 20.1.
4.60. ( )-(3R,4S)-3-Benzenesulfonyl-N-methylthio-4-
phenylazetidin-2-one (33b)
1
Yellow oil, 55% yield. H NMR (250 MHz, CDCl3): d
7.64–7.57 (3H, m), 7.45–7.39 (2H, m), 7.30–7.21 (5H,
m), 5.85 (1H, d, J = 5.1 Hz), 5.46 (1H, d, J = 5.1 Hz),
2.43 (3H, s); 13C NMR (63 MHz, CDCl3): d 165.2,
134.5, 133.5, 133.2, 129.2, 128.9, 128.7, 128.2, 128.0,
127.8, 127.6, 126.7, 125.8, 80.0, 60.8, 20.8.
4.67. ( )-(3R,4S)-3-Acetoxy-4-(2-chlorophenyl)-N-methyl-
thio-3-propylazetidin-2-one (37f)
1
Yellow oil, 33% yield. H NMR (250 MHz, CDCl3): d
7.38 (1H, m), 7.27 (3H, m), 5.19 (1H, s), 2.50 (3H, s),
2.33–2.10 (2H, m), 1.72–1.46 (2H, m), 1.58 (3H, s),
0.99 (3H, t, J = 7.3 Hz); 13C NMR (63 MHz, CDCl3):
d 170.7, 168.3, 134.3, 131.2, 129.5, 91.0, 67.0, 35.6,
21.5, 20.1, 16.7, 14.1.
4.61. ( )-(3R,4S)-N-(Methylthio)-4-phenyl-3-(4-toluene-
sulfonyl)azetidin-2-one (33c)
Yellow solid, mp 85–87 ꢁC, 74% yield. 1H NMR
(250 MHz, CDCl3): d 7.41–7.26 (5H, m), 7.19–7.15
(4H, m), 5.74 (1H, d,J = 5.0 Hz), 4.88 (1H, d,
J = 5.0 Hz), 2.40 (3H, s), 2.35 (3H, s); 13C NMR
(63 MHz, CDCl3): d 166.4, 145.3, 132.4, 132.0, 129.8,
129.2, 129.0, 128.5, 127.8 (2C), 126.8 (2C), 80.6, 66.0,
22.1, 21.7.
4.68. ( )-(3R,4S)-3-Acetoxy-4-(2-chlorophenyl)-N-methyl-
thio-3-phenylazetidin-2-one (37g)
1
Yellow oil, 40% yield. H NMR (250 MHz, CDCl3): d
7.73 (2H, d, J = 7.6 Hz), 7.44–7.32 (7H, m), 5.87 (1H,
s), 2.51 (3H, s), 1.65 (3H, s); 13C NMR (63 MHz,

E. Turos et al. / Bioorg. Med. Chem. 13 (2005) 6289–6308
6307
CDCl3): d 169.3, 167.6, 135.3, 134.1, 131.2, 128.9,
129.7, 129.1, 129.0, 128.6, 126.8, 126.3, 91.8, 67.2,
21.8, 20.3.
4.75. ( )-(3R,4S)-4-(2,6-Dichlorophenyl)-3-methoxy-N-
methylthioazetidin-2-one (43j)
1
Pale yellow solid; mp 77–80 ꢁC; H NMR (250 MHz,
CDCl3): d 7.23 (1H, t, J = 8.1 Hz), 5.72 (1H, d,
J = 5.3 Hz), 4.90 (1H, d, J = 5.1 Hz), 3.30 (3H, s), 2.44
(3H, s); 13C NMR (63 MHz, CDCl3): d 168.3, 131.1,
130.1, 128.8, 88.0, 62.9, 59.2, 21.6.
4.69. ( )-(3R,4S)-4-(3-Chlorophenyl)-3-methoxyazetidin-
2-one (43a)
White solid; mp 110–111 ꢁC; 1H NMR (250 MHz,
CDCl3) d 7.30–7.20 (4H, m), 6.58 (1H, brs ), 4.76 (1H,
d, J = 4.5 Hz), 4.69–4.67 (1H, m), 3.13 (3H, s); 13C
NMR (63 MHz, CDCl3): d 168.1, 137.9, 129.6, 128.5,
127.7, 125.8, 86.7, 58.3, 57.6.
4.76. ( )-(3R,4S)-3-Methoxy-4-(2-methoxyphenyl)azeti-
din-2-one (43l)
White solid; mp 120–123 ꢁC; 1H NMR (250 MHz,
CDCl3): d 7.27-07.24 (1H, app m), 7.190-7.15 (1H, m),
6.95 (1H, t, J = 7.5 Hz), 6.86 (1H, d, J = 8.2 Hz), 5.27
(1H, d, J = 4.9 Hz), 4.72 (1H, d, J = 4.9 Hz), 3.80 (3H,
s), 3.12 (3H, s), 2.37 (3H, s); 13C NMR (63 MHz,
CDCl3): d 170.9, 157.7, 129.5, 128.5, 121.5, 120.4,
110.3, 86.5, 60.2, 58.5, 55.4, 21.8.
4.70. ( )-(3R,4S)-4-(4-Chlorophenyl)-3-methoxy-N-methyl-
thioazetidin-2-one (43b)
White crystals; mp 62–66 ꢁC; 1H NMR (250 MHz,
CDCl3): d 7.33 (2H, d, J = 8.5 Hz), 7.25 (2H, d,
J = 8.5 Hz), 4.73 (2H, s), 3.13 (3H, s), 2.31 (3H, s); 13C
NMR (63 MHz, CDCl3): d 170.2, 134.9, 130.2, 128.6,
86.5, 65.5, 58.4, 22.1.
4.77. ( )-(3R,4S)-3-Methoxy-N-methylthio-4-(2-nitro-
phenyl)-azetidin-2-one (43n)
4.71. ( )-(3R,4S)-4-(2-Iodophenyl)-3-methoxy-N-methyl-
thioazetidin-2-one (43f)
1
Yellow solid; H NMR (250 MHz, CDCl3): d 8.12 (1H,
d, J = 8.2 Hz), 7.66–7.63 (1H, m), 7.50–7.41 (2H, m),
5.49 (1H, d, J = 5.1 Hz), 4.93 (1H, d, J = 5.2 Hz), 3.23
(3H, s), 2.43 (3H, s).
White crystals; mp 62–65 ꢁC; 1H NMR (250 MHz,
CDCl3): d 7.80 (1H, d, J = 7.8 Hz), 7.30 (1H, t,
J = 7.5 Hz), 7.15 (1H, t, J = 8.0 Hz), 7.00 (1H, t,
J = 7.5 Hz), 5.09 (1H, d, J = 5.0 Hz), 4.80 (1H, d,
J = 4.8 Hz), 3.82 (3H, s), 2.41 (3H, s); 13C NMR
(63 MHz, CDCl3): d 170.7, 139.9, 136.2, 130.6, 129.3,
128.6, 99.8, 87.1, 70.4, 59.6, 22.2.
4.78. ( )-(3R,4S)-3-Methoxy-N-methylthio-4-(4-prope-
nyl-oxyphenyl)azetidin-2-one (43q)
White solid; mp 87–88 ꢁC; 1H NMR (250 MHz, CDCl3):
d 7.29 (2H, d, J = 8.4 Hz), 7.08 (2H, d, J = 8.4 Hz), 6.50
(1H, d, J = 17.3 Hz), 6.21 (1H, dd, J = 17.1, 10.3 Hz),
5.92 (1H, d, J = 10.4 Hz), 4.75 (1H, d, J = 4.8 Hz),
4.69 (1H, d, J = 4.8 Hz), 3.05 (3H, s), 2.26 (3H, s); 13C
NMR (63 MHz, CDCl3): d 170.3, 164.2, 150.9, 132.9,
131.1, 129.9, 127.7, 121.4, 86.5, 65.5, 58.3, 22.0.
4.72. ( )-(3R,4S)-4-(3-Iodophenyl)-3-methoxy-N-methyl-
thioazetidin-2-one (43g)
White crystals; mp 97–99 ꢁC; 1H NMR (250 MHz,
CDCl3): d 7.65 (2H, m), 7.27 (1H, d, J = 7.7 Hz), 7.08
(1H, t, J = 8.0 Hz), 4.73–4.67 (2H, AB m), 3.13 (3H,
s), 2.32 (3H, s); 13C NMR (63 MHz, CDCl3): d 170.6,
138.4, 138.2, 136.4, 130.5, 128.5, 94.5, 87.0, 65.8, 58.3,
22.6.
4.79. ( )-(3R,4S)-4-(4-Hydroxyphenyl)-3-methoxy-N-
methylthioazetidin-2-one (43r)
White solid; mp 119–123 ꢁC; 1H NMR (250 MHz,
CDCl3): d 7.18 (2H, d, J = 9.0 Hz), 6.81 (2H, d,
J = 9.0 Hz), 5.73 (1H, br s), 4.69 (2H, app s), 3.11
(3H, s), 2.28 (3H, s); 13C NMR (63 MHz, CDCl3): d
170.9, 168.5, 130.4, 124.9, 115.4, 65.9, 58.3, 22.1.
4.73. ( )-(3R,4S)-4-(4-Iodophenyl)-3-methoxy-N-methyl-
thioazetidin-2-one (43h)
White solid; mp 102–105 ꢁC; 1H NMR (250 MHz,
CDCl3): d 7.65 (2H, d, J = 8.3 Hz), 7.03 (2H, t,
J = 8.3 Hz), 4.70 (2H, app s), 3.10 (3H, s), 2.29 (3H,
s); 13C NMR (63 MHz, CDCl3): d 170.1, 137.5, 133.2,
130.7, 94.9, 86.4, 65.6, 58.4, 22.1.
4.80. ( )-(3R,4S)-4-Biphenyl-3-methoxy-N-methylthioaz-
etidin-2-one (43s)
1
Light yellow solid; H NMR (250 MHz, CDCl3): d 7.65
(4H, d, J = 6.5 Hz), 7.43–7.34 (5H, m), 4.85 (1H, d,
J = 4.3 Hz), 4.81 (1H, d, J = 4.4 Hz), 3.12 (3H, s), 2.33
(3H, s); 13C NMR (63 MHz, CDCl3): d 169.5, 142.3,
141.6, 135.8, 129.1, 128.2, 127.5, 127.4, 87.5, 59.1, 58.3.
4.74. ( )-(3R,4S)-4-(2,4-Dichlorophenyl)-3-methoxy-N-
methylthioazetidin-2-one (43i)
Yellow crystals; mp 102–105 ꢁC; 1H NMR
(250 MHz, CDCl3): d 7.37 (s, 1H), 7.28 (1H, d,
J = 8.3 Hz), 7.16 (1H, t, J = 8.2 Hz), 5.24 (1H, d,
J = 4.6 Hz), 4.97 (1H, d, J = 4.6 Hz), 3.19 (3H, s),
2.40 (3H, s); 13C NMR (63 MHz, CDCl3): d 170.5,
135.3, 134.9, 130.6, 130.3, 129.9, 127.7, 87.1, 62.6,
59.4, 22.2.
4.81. ( )-(3R,4S)-4-Fluorenyl-3-methoxy-N-methylthioaz-
etidin-2-one (43t)
1
Colorless oil; H NMR (250 MHz, CDCl3): d 7.75 (2H,
d, J = 7.5 Hz), 7.50 (2H, d, J = 6.6 Hz), 7.34–7.24 (3H,

6308
E. Turos et al. / Bioorg. Med. Chem. 13 (2005) 6289–6308
m), 4.85 (1H, d, J = 4.9 Hz), 4.77 (1H, d, J = 4.9 Hz),
3.88 (2H, s), 3.14 (3H, s), 2.33 (3H, s).
are grateful to Dr. Josefa Anaya and Professor Manuel
Grande, Departamento de Quimica Organica, Universi-
dad de Salamanca, Spain, for providing us with a sam-
ple of compound 12.
4.82. Testing of antimicrobial susceptibilities (Kirby–Bauer
well diffusion)
Staphylococcus aureus (ATCC 25923) and MRSA (ATCC
43300 and 33591) were purchased from ATCC sources.
Eight additional strains of MRSA were obtained from
Lakeland Regional Medical Center (Lakeland, FL).
References and notes
1. Turos, E.; Konaklieva, M. I.; Ren, R. X. F.; Shi, H.;
Gonzalez, J.; Dickey, S.; Lim, D. Tetrahedron 2000, 56,
5571.
2. (a) Ren, X.-F.; Konaklieva, M. I.; Shi, H.; Dickey, S.;
Lim, D. V.; Gonzalez, J.; Turos, E. J. Org. Chem. 1998,
63, 8898; (b) Ren, X.-F.; Konaklieva, M. I.; Turos, E.
J. Org. Chem. 1995, 60, 4980.
3. Chemistry and Biology of b-Lactam Antibiotics; Morin, R.
B., Gorman, M., Eds.; Academic Press: New York, 1982;
Vols. 1–3.
4. (a) Long, T. E.; Turos, E.; Konaklieva, M. I.; Blum, A. L.;
Amry, A.; Baker, E. A.; Suwandi, L. S.; McCain, M. D.;
Rahman, M. F.; Sonja Dickey, S.; Lim, D. V. Bioorg.
Med. Chem. 2003, 11, 1859; (b) Kazi, A.; Hill, R.; Long, T.
E.; Kuhn, D. J.; Turos, E.; Dou, Q. P. Biochem.
Pharmacol. 2004, 67, 365.
5. Turos, E.; Long, T. E.; Konaklieva, M. I.; Coates, C.;
Shim, J.-Y.; Dickey, S.; Lim, D. V.; Cannons, A. Bioorg.
Med. Chem. Lett. 2002, 12, 2229.
6. Coates, C.; Long, T. E.; Turos, E.; Dickey, S.; Lim, D. V.
Bioorg. Med. Chem. 2003, 11, 193–196.
7. Konaklieva, M. I., Ph.. Dissertation, SUNY at Buffalo,
1997.
8. Coates, C.; Ph.D. Dissertation, University of South
Florida, 2004.
4.83. Culture preparation
From a freezer stock in tryptic soy broth (Difco Labora-
tories, Detroit, MI) and 20% glycerol, a culture of each
microorganism was transferred with a sterile Dacron
swab to Trypticaseꢂ Soy Agar (TSA) plates (Becton–
Dickinson Laboratories, Cockeysville, MD), streaked
for isolation, and incubated at 37 ꢁC for 24 h. A 108
standardized cell count suspension was then made in
sterile phosphate-buffered saline (pH 7.2) and swabbed
across fresh TSA plates.
4.84. Antimicrobiological testing
Prior to swabbing with the culture solution, 20 lL of a
1 mg/mL stock solution of the test lactam compound
in dimethylsulfoxide (DMSO) was added to a 6-mm
diameter well bored into the agar. The plates were
swabbed uniformly with the test microbe compound
above and then incubated for 24 h at 37 ꢁC. The antimi-
crobial susceptibilities were determined by measuring
the zones of growth inhibition around each well.
9. Mickel, S. Aldrichim. Acta 1985, 18, 2259.
10. Woulfe, S. R.; Iwagami, H.; Miller, M. J. Tetrahedron
Lett. 1985, 26, 3891.
11. Kraus, G. A.; Neuenschwander, K. J. Chem. Soc. Chem.
Commun. 1982, 134.
4.85. Determination of minimum inhibitory
concentrations
12. Bose, A. K.; Anjaneyulu, B.; Bhattacharya, S. K.;
Manhas, M. S. Tetrahedron 1967, 23, 4769.
13. Ren, X. F.; Turos, E. J. Org. Chem. 1994, 59, 5858.
14. Abdel-Magid, A. F.; Carson, K. G.; Harris, B. D.;
Maryanoff, C. A.; Shah, R. D. J. Org. Chem. 1996, 61,
3849.
15. Long, T. E., Ph.D. Dissertation, University of South
Florida, 2003.
16. Buynak, Y. D.; Borate, H. B.; Lamb, G. W.; Khasnis, D.
D.; Hustig, C.; Jsum, H.; Siriwardone, U. A. J. Org. Chem
1993, 58, 1325.
The minimum inhibitory concentration (MIC) values of
the lactams were determined for S. aureus and MRSA
by serial dilution, according to NCCLS protocols.19
The test medium was prepared in 24-well plates (Costar
3524, Cambridge, MA) by adding the test drug in
DMSO to Mueller–Hinton II agar (Becton–Dickinson
Laboratories, Cockeysville, MD) to bring the total vol-
ume in each well to 1.0 mL. Starting with an initial well
concentration of 256 lg of drug/mL, each sequential
dilution contained half the concentration of the drug.
The medium was allowed to solidify at room tempera-
ture for 24 h before inoculation with the bacteria. Using
a sterilized inoculating loop, a small amount of each
standardized Staphylococcus strain cultured on TSA
plates for 24 h was transferred into sterile test tubes con-
taining 5 mL TSA broth and incubated at 37 ꢁC for
24 h. One microliter of each culture was then applied
to the appropriate well of Mueller–Hinton agar and
incubated at 37 ꢁC overnight. After 24 h, the MICs were
determined as the lowest concentration of the drug
where bacterial growth was visibly inhibited.
17. These compounds were prepared by N-lithiation and N-
thiolation of the N-protio lactam precursor 45 (Alonso,
E., Ph.D. Dissertation, Universidad de Oviedo, Spain,
2002).
O
SCH3
O
Li
R
O
H
R
N
N
N
a
b
X
X
X
R
Y
Y
Y
47a-e
45
46
18. Carr, J. A.; Al-Azemi, T. F.; Long, T. E.; Shim, J.-Y.;
Coates, C. M.; Turos, E.; Bisht, K. S. Tetrahedron 2003,
59, 9147.
19. NCCLS (National Committee for Clinical Laboratory
Standards) Methods for Dilution of Antimicrobial Suscep-
tibility Tests for Bacteria that Grow Aerobically. NCCLS
Document M7-A4, Vol. 17, No. 2, 1997.
Acknowledgments
Funding for this research was generously provided by
the National Institutes of Health (R01 AI51351). We