Synthesis and antimicrobial activity of some new sugar-based monocyclic β-lactams

Article abstract of DOI:10.3390/90100029

The syntheses of some new sugar-based monocyclic β-lactams possessing several other functionalities in addition to the carbohydrate moiety are described. The key step was the Staudinger [2+2] cycloaddition of chiral carbohydrate Schiff base 5 with phthalimidoacetyl chloride to yield the sugar-based monocyclic β-lactam 6 as a single isomer. Treatment of protected β-lactams 6 and 8 with methylhydrazine afforded the free amino β-lactams 9 and 10. Acylation of these free amino β-lactams with benzoyl, phenoxyacetyl, cinnamoyl and phenylacetyl chloride in the presence of pyridine afforded β-lactams 11a-d and 12a-d. Some of these novel β-lactams were found to be active against Staphilococcus citrus, Klebsiella pneumoniae, Escherichia coli and Bacillus subtilis.

Full text of DOI:10.3390/90100029

Molecules 2004, 9, 29-38
molecules
ISSN 1420-3049
http://www.mdpi.org/
Synthesis and Antimicrobial Activity of Some New Sugar-Based
Monocyclic β-Lactams
1
,
1
2
A. A. Jarrahpour *, M. Shekarriz and A. Taslimi
1
Chemistry Department, College of Sciences, Shiraz University, Shiraz 71454, Iran. Tel.: +98 711 228
822, Fax: +98 711 228 0926.
Institute of Standards and Industrial Research of Iran, Shiraz 71447, Iran.
4
2
*
Author to whom correspondence should be addressed; e-mail: jarrah@susc.ac.ir;
aliasghar6683@yahoo.com
Received: 14 June 2003; in revised form: 22 August 2003 / Accepted: 7 September 2003 / Published:
1 January 2004
3
Abstract: The syntheses of some new sugar-based monocyclic β-lactams possessing
several other functionalities in addition to the carbohydrate moiety are described. The key
step was the Staudinger [2+2] cycloaddition of chiral carbohydrate Schiff base 5 with
phthalimidoacetyl chloride to yield the sugar-based monocyclic β-lactam 6 as a single
isomer. Treatment of protected β-lactams 6 and 8 with methylhydrazine afforded the free
amino β-lactams 9 and 10. Acylation of these free amino β-lactams with benzoyl,
phenoxyacetyl, cinnamoyl and phenylacetyl chloride in the presence of pyridine afforded
β-lactams 11a-d and 12a-d. Some of these novel β-lactams were found to be active
against Staphilococcus citrus, Klebsiella pneumoniae, Escherichia coli and Bacillus
subtilis.
Keywords: Monocyclic sugar-based β-lactam, galactose, Schiff base, phthalimidoacetyl
chloride, antimicrobial activity.

Molecules 2004, 9
30
Introduction
β-Lactam antibiotics have been successfully used in the treatment of infectious diseases for many
years [1]. Despite the large number of compounds containing a β-lactam moiety that have already been
synthesized and tested, there is still a need for new compounds of this kind [2] due to the increasing
resistance of bacterial strains to certain types of antibiotics [3]. A class of β-lactams known as the
monocyclic β-lactams, which includes compounds such as the nocardicins, aztreonam and carumonam,
has been described for their chemotherapeutic importance as antibiotics [4-7]. On the other hand, during
the past few years carbohydrates have received increasing attention as stereodifferentiating auxiliaries in
stereoselective syntheses [8]. Cephalosporin β-lactams containing carbohydrates have been prepared [9].
A monocyclic β-lactam containing a carbohydrate moiety resulting from the reaction of phthalimido-
acetyl chloride with the imine of D-glucosamine and cinnamaldehyde has been reported [10]. The
monocyclic β-lactam derived from the imine of the amino acid D-threonine upon treatment with
azidoacetyl chloride and triethylamine has been reported by Bose [11]. Gunda has reported the synthesis
of monocyclic β-lactams with high diastereoselectivity [12], while Palomo and his co-workers have
synthesized monocyclic β-lactams bearing carbohydrate moieties with moderate diastereoselectivity
[
13]. The recent discovery of new biologically active monocyclic β-lactam compounds displaying
activities other than the usual antibiotic one, such as Thrombin [14], Prostate Specific Antigen [15],
Human Cytomegalovirus Protease [16] and the Cholesterol Absorption inhibitors [17] is also interesting.
The presence of a carbohydrate moiety side chain in a drug may also overcome the frequently observed
water insolubility problem [18]. With all this in mind, we focused on the synthesis of some novel sugar
based monocyclic β-lactams. Of these new β-lactams, compounds 6, 7, 8, 11a-d and 12a-d were tested
against Bacillus subtilis, Klebsiella pneumoniae, Escherichia coli and Staphilococcus citrus.
Results and Discussion
D-(+)-Galactose (1) was chosen as the starting material for these studies. It was converted into
,3,4,6-tetra-O-acetyl-α-D-galactopyranosyl bromide (2) according to a reported method [19]. The
2
thermodynamically more stable α-anomer was formed. The halogen in the acylgalactosyl halide is
reactive and may be readily displaced by an azido group. Thus, treatment of 2 with sodium azide in 9:1
acetone-water for 5h at reflux temperature gave 2,3,4,6-tetra-O-acetyl-β-D-galactopyranosyl azide (3) as
a white crystalline substance in 70% yield (Scheme 1). In the case of the D-galactose derivative, the
replacement involves inversion of configuration at the anomeric site and thus the α-galactopyranosyl
halide 2 yields a β-galactosyl azide 3. The presence of the protecting acyl groups in the acylgalactosyl
halide ensures that the pyranose ring structure is retained. Heterogeneous reduction of the azide group of
3
with Raney Nickel in ethyl acetate under reflux for 2.5h resulted in a 90% yield of 2,3,4,6-tetra-O-
acetyl-β-D-galactopyranosyl amine (4). The Schiff base β-D-galactopyranosyl amino-(N-cinnamyl-
idene)-2,3,4,6-tetra-O-acetate (5) was prepared in quantitative yield by treatment of 4 with
cinnamaldehyde in refluxing dry dichloromethane in the presence of anhydrous Na SO . The β-D-
2
4

Molecules 2004, 9
31
galactopyranosyl imine 5 has a predominant E-configuration [20]. Compound 5 was next treated with
o
phthalimidoacetyl chloride in the presence of triethylamine in dry CH
2
Cl
2
at -10 C to give 1-[(2,3,4,6-
tetra-O-acetyl-β-D-galactopyranosyl)-3-phthalimido-4-styryl]azetidin-2-one (6) in 75% yield. The
o
ozonolysis of the styryl group of 6 at -78 C in dry CH Cl afforded 1-[(2,3,4,6-tetra-O-acetyl-β-D-
2
2
galactopyranosyl-3-phthalimido-4-formyl]azetidin-2-one (7) in 90% yield. Reduction of 7 using
o
LiAlH(t-OBu) in dry THF at 0 C gave 1-[(2,3,4,6-tetra-O-acetyl-β-D-galactopyranosyl)-3-phthalimido-
3
4
-hydroxymethyl]-azetidin-2-one (8) in 60% yield (Scheme 1).
Scheme 1
AcO
OH
AcO
CH OAc
2
CH OH
CH OAc
2
O
2
O
Ac O
Red P, Br2
2
O
AcO
NaN3
HO
AcO
N3
OH
H
OAc
3
HO
OAc
1
Br
2
Raney nickel
AcO
CH OAc
Ph
2
O
AcO
AcO
N
CH OAc
2
Ph CH=CHCHO
O
OAc
AcO
H
NH2
phthalimidoacetyl
chloride
OAc
5
H
4
O
O
Ph
N
O
CHO
O
N
O
1) O3
2) Me2S
CH OAc
2
CH OAc
OAc
2
OAc
N
O
N
O
OAc
O
OAc
OAc
7
OAc
6
LiAlH(t-OB4)3
O
N
O
CH OH
2
CH OAc
2
O
OAc
O
N
OAc
OAc
8
Next we turned our attention to the removal of the phthalimido protecting group. Methylhydrazine is
the most suitable deprotecting agent [21-24] among the various different methods available for the
removal of this protecting group. Indeed, a significant improvement in the yield and in the ease of
isolation of the free amino β-lactam is achieved when methylhydrazine is employed instead of
hydrazine. This is attributed to the fact that because of the decreased acidity of the by-product
N-methylphthalhydrazide (with respect to phthalhydrazide), no complex is formed with the free amine
and therefore no heating or acid treatment of the reaction mixture is required. N-methylphthalhydrazide
separates from a chloroform solution of the methylhydrazine-phthalisoimide adduct leaving the free
amine in solution. The advantages of using methylphydrazine are thus particularly noticeable when
applied to the dephthaloylation of more sensitive substrates [23].

Molecules 2004, 9
32
Compounds 6 and 8 were thus converted into the free amino β-lactams 9 and 10, respectively, by
treatment with methylhydrazine in ethanol [Scheme 2]. Finaly, treatment of these free amino
monocyclic β-lactams with benzoyl, phenoxyacetyl, phenylacetyl and cinnamoyl chloride in the
presence of pyridine afforded β-lactams 11a-d and 12a-d (Scheme 2).
Scheme 2
Ph
H N
O
Ph
2
H
CH OAc
2
R C N
OAc
OAc
O
CH OAc
CH NHNH
RCOCl
Pyridine
2
3
2
N
6
O
OAc
OAc
O
N
OAc
O
9
12
OAc
a) R =
b) R = CH O
2
c)
R =
CH2
d) R = CH=CH
O
H
CH OH
H N
CH OH
2
2
2
R C N
CH NHNH
CH OAc
2
CH OAc
2
3
2
8
O
OAc
OAc
RCOCl
Pyridine
O
OAc
N
N
O
O
OAc
OAc
OAc
10
11
a) R =
b) R = CH2O
c) R = CH2
d) R = CH=CH
Biological Screening: Antimicrobial Activity Tests.
There has been a suggestion that the bacteria may utilize a carbohydrate uptake mechanism, which
allows for a better transport of the monocyclic β-lactams across the membrane. The antibacterial
activities of compounds 6, 7, 8, 11a-d and 12a-d were tested against one strain each of a Gram +ve
bacteria (Staphylococcus citrus), a Gram -ve bacteria (Escherichia Coli), a Gram -ve containing capsule
(
Klebsiella) and a Gram +ve spore (Bacillus subtilis). From the data presented in Table 1, it is clear that
compound 6 was highly active against Staphilococcus citrus, Klebsiella pneumoniae, Escherichia coli
and Bacillus subtilis and β-Lactams 12b and 12c were moderately active against these four
microorganisms Compounds 8 and 11b were highly active against Bacillus subtilis, while compound
1
1d was moderately active against B. subtilis. Compounds 11a-d were slightly active against S. citrus.
Finally, compounds 7 and 12a were found to be inactive against all microorganisms used.

Molecules 2004, 9
Table 1. Results of antimicrobial activity tests of the synthetic monocyclic β-lactams.
33
Microorganism
K. Pneumoniae
Compound No.
B. Subtilis
E. coli
S. citrus
Ampicillin
Gentamycin
+++
+++
+++
-
+++
-
+++
+
++
-
+++
+++
+++
-
-
-
-
-
-
+++
+++
+++
-
-
-
-
-
-
+++
+++
+++
-
+
+
+
+
+
-
6
7
8
1
1
1a
1b
1c
1d
2a
2b
2c
2d
1
1
1
1
-
-
++
++
+
++
++
-
++
++
-
++
++
-
1
1
Key to symbols:
Highly active = +++ (inhibition zone>12 mm)
Moderately active = ++ (inhibition zone 9-12 mm)
Slightly active = + (inhibition zone 6-9 mm)
Inactive = - (inhibition zone<6 mm)
Acknowledgements
The authors wish to express their thanks to Research Council of Shiraz University for financial
support (Grant No.81-SC-1540-C-220).
Experimental
General
Melting points were taken in open capillaries on a Buchi 510 circulating oil melting point apparatus
and are uncorrected. Infrared spectra (KBr disks) were recorded on a Perkin Elmer 781
spectrophotomer. NMR spectra were recorded on JEOL JNM-EX 90A FT-NMR and Bruker Avance
DPX 250 MHz spectrometers. The mass spectra were recorded on a GCMS-QP 1000 EX Shimadzu
Gas Chromatography-MS apparatus. The bacterial strains used are susceptible to β-lactam antibiotics.

Molecules 2004, 9
34
Synthesis of compounds
2
,3,4,6-Tetra-O-acetyl-α-D-galactopyranosyl bromide (2). This compound was prepared according to a
literature procedure [19].
2
,3,4,6-Tetra-O-acetyl-β-D-galactopyranosyl azide (3). Sodium azide (0.47 g, 7.30 mmol) was added to
a solution of 2 (3.00 g, 7.30 mmol) in 9:1 acetone-water (30 mL) and the mixture was refluxed for 10
hours, then stirred at room temperature for 24 hours. The solvent was evaporated. The residue was taken
up in CH Cl (30 mL) and washed with H O (3 x 30 mL), dried (Na SO ) and filtered. Evaporation of
2
2
2
2
4
the solvent under reduced pressure afforded a solid. Recrystallization from ether gave 3 as white crystals
o
–1
1
(
70% yield); m.p. 91-92 C; IR (cm ): 2140 (N
3
), 1740-1750 (OCOCH
3
); H-NMR (CDCl
3
) δ (ppm):
1
H
4
.95-2.15 (12H, s, 4COCH ), 3.95-4.21 (3H, m, H CH OAc), 4.35 (1H, t, H ), 5.05-5.20 (2H, m, H
3
2,
2
3
4,
5
), 5.95(1H, d, H
6
); Anal.(C14
H
19
N
3
O
9
), Calc. C, 45.04; H, 5.13; N, 11.26; O, 38.57%, found: C,
+
5.00; H, 5.18; N, 11.24; O, 38.55%; MS, m/z (%): 373.11 (M , 100.00), 374.11 (M+1, 16.20).
2
,3,4,6-Tetra-O-acetyl-β-D-galactopyranosyl amine (4). To a solution of 3 (6.00 g, 16.07 mmol) in ethyl
acetate (50 mL) was added Raney Ni (6.00 g) and the reaction mixture was refluxed for 2.5 hours. The
resulting solid was filtered and washed with ether. Recrystallization from ether gave 4 as yellow crystals
o
–1
1
in 90% yield; m.p. 120-122 C. IR (cm ): 3400, 3330 (NH
δ (ppm): 1.95-2.15 (12H, s, 4COCH ), 3.95-4.21 (3H, m, H2, CH
NH ), 5.05-5.20 (2H, m, H4, ), 5.95 (1H, d, H ); Anal., Calc. for C14
2
), 1730-1740 (OCOCH
3
); H-NMR (CDCl
3
)
3
2
OAc), 4.35 (1H, t, H ), 4.55 (2H, d,
3
2
H
5
6
H21NO
9
: C, 48.41; H, 6.09; N,
+
4
3
.03; O, 41.46%, found: C, 48.38; H, 6.05; N, 4.01; O, 41.42%; MS, m/z (%): 347.12 (M , 100.00),
48.12 (M+1, 15.90).
β-D-galactopyranosylamino-(N-cinnamylidene)-2,3,4,6-tetra-O-acetate (5). A mixture of compound 4
3.47 g, 10.00 mmol), cinnamaldehyde (1.32 g, 10.00 mmol) and Na SO (15.00 g, 105.61 mmol) in dry
CH Cl (40 mL) was refluxed for 5 hours. The resulting mixture was filtered and washed with H O (3 x
0mL). The organic layer was separated and dried (Na SO ). Evaporation of the solvent under reduced
(
2
4
2
2
2
3
2
4
pressure gave 5 as a solid which was recrystalized from ether-hexane (1:1) (3.69g, 80.00%); m.p.110-
-1
1
1
(
(
12ºC; IR (cm ): 1740 (OCOCH
12H, s, 4COCH
1H, dd, J=8.00 and 16.00 Hz, CH=CHPh), 5.95 (1H, d, H ), 6.60 (1H, d, CH=CHPh), 7.10-7.30 (5H,
3
), 1685 (C=C), 1680 (C=N); H-NMR (CDCl
3
) δ (ppm): 1.95-2.15
3
), 4.20-4.25 (3H, m, H
2
and CH
2
OAc), 4.35 (1H, t, H
3
), 5.05-5.2 (2H, m, H
4
, H ), 5.60
5
6
m, Ph), 7.50 (1H, d, N=CH); Anal. Calc. for C23
C, 59.80; H, 6.10; N, 3.00; O, 31.10%; MS, m/z: 461.11 (M ).
H
27NO
9
: C, 59.86;H, 5.90; N, 3.04; O, 31.20%, found:
+
1
-[(2,3,4,6-tetra-O-acetyl-β-D-galactopyranosyl)-3-phthalimido-4-styryl]azetidin-2-one (6). A solution
of phthalimidoacetyl chloride (7.00 g, 35.00 mmol) in dry CH Cl (50 mL) was added dropwise to a
solution of Schiff base 5 (4.61g, 10.00 mmol) and triethylamine (1.01g, 10 mmol) in dry CH Cl (100
2
2
2
2
o
mL) at -10 C. After the addition was complete, the solution was stirred at room temperature for 24 hours

Molecules 2004, 9
35
and then washed with saturated NaHCO
The organic layer was separated, dried (Na
3
solution (2 x 70 mL), water (3 x 50 mL) and brine (70 mL).
2
SO ), filtered and evaporated. Recrystallization from
4
o
-1
hexane-ether (1:1) gave 6 in 75% yield; m.p. 95-97 C; IR (cm ): 1780 (β-lactam CO), 1760, 1735
1
(
(
phthalimido CO), 1740 (OCOCH
3H, m, H ,CH
3
); H-NMR (CDCl
3
) δ (ppm): 1.95-2.15 (12H, s, 4COCH
3
), 3.95-4.20
2
2
OAc), 4.35 (1H, t, H
3
), 4.50 (1H, dd, J=5 and J=8Hz, CH-CH=CH), 4.90 (1H, d, J=5,
CH-N(CO) ), 5.05-5.2 (2H, m, H , H ), 5.95 (1H, d, H ), 6.20 (1H, dd, J=8 and J= 16, CH-CH=CHPh),
2
4
5
6
6
C
.40 (1H, d, CH=CHPh), 7.10-7.25 (5H, m, Ph), 7.60-7.95 (4H, m, C
6
H
4
(CO) N); Anal. Calc. for
2
33
H
33
N O
2
12: C, 61.11; H, 4.97; N, 4.32; O, 29.60%, found: C, 61.02; H, 5.05; N, 4.28; O, 29.70%.
1
-[(2,3,4,6-tetra-O-acetyl-β-D-galactopyranosyl)-3-phthalimido-4-formyl]azetidin-2-one (7). A solution
o
of compound 6 (2.00 g, 3.00 mmol) in dry CH Cl (50 mL) was cooled to -78 C. A stream of ozone was
2
2
passed through the reaction mixture until a pale blue coloration was observed and then it was purged
o
with nitrogen. A solution of dimethyl sulfide (3 mL) in CH Cl (10 mL) was added dropwise at -78 C.
2
2
When the addition was complete, the cooling bath was removed and the solution was stirred at room
temperature. The reaction mixture was washed with H O (3 x 40mL) and saturated brine (2 x 30mL).
2
The organic layer was separated and dried (Na
residue was heated under high vacuum for 20 hours at 50 C to remove the benzaldehyde formed during
2
SO
4
). Evaporation of the solvent gave a syrup. The oily
o
the ozonolysis. Compound 7 was isolated as a solid and was recrystalized from ether-hexane in 97%
o
–1
yield; m.p. 88-90 C; IR (cm ): 1780 (β-lactam CO), 1765, 1730 (phthalimido CO), 1750 (CHO), 1740
1
(
4
H
C
OCOCH
3
); H-NMR (CDCl
3
) δ (ppm): 1.95-2.15 (12H, s, 4COCH
), 4.70 (1H, dd, J=5Hz, CH-CHO), 4.90 (1H, d, J=5Hz, CH-N(CO)
), 5.95 (1H, d, H ), 7.60-7.95 (4H, m, C (CO) N), 10.00 (1H, d, CHO); Anal. Calc. for
13: C, 54.36; H, 4.56; N, 4.88; O, 36.20%, found: C, 54.30; H, 5.01; N, 4.87; O, 36.27%.
3
), 3.95-4.20 (3H, m, H
2
, CH
2
OAc),
.35 (1H, t, H
, H
3
2
), 5.05-5.2 (2H, m,
4
5
6
6
H
4
2
26
H
26
2
N O
1
-[(2,3,4,6-tetra-O-acetyl-β-D-galactopyranosyl)-3-phthalimido-4-hydroxymethyl] azetidin-2-one (8):
o
To aldehyde 7 (1.73 g, 3.00 mmol) in tetrahydrofuran (80 mL) at 0 C was added lithium tri (tert-butoxy)
aluminum hydride (1.53 g, 6.00 mmol). After stirring under nitrogen for 3 hours (0 C), the mixture was
o
acidified with dilute HCl (2%) to pH 5, and 1 g of silica gel was added. The suspension was stirred for
NMR (CDCl
3
3
), 3.75 (2H, m, CH
2
OH), 3.95-4.20 (3H, m,
OH), 4.90 (1H, d,
H
2
,CH OAc), 4.35 (1H, t, H
2
2
J=5Hz, CH-N(CO) ), 5.05-5.2 (2H, m, H , H ), 5.95 (1H, d, H ), 7.60-7.95 (4H, m, C H (CO) N);
2
4
5
6
6
4
2
Anal. Calc. for C26
O, 36.18%.
H
28
2
N O13: C, 54.17; H, 4.90; N, 4.86; O, 36.08%, found: C, 54.10, H, 5.00; N, 4.80;

Molecules 2004, 9
36
General procedure for dephthaloylation of protected β-lactams 6 and 8. The monocyclic β-lactam 6
6.50 g, 10.00 mmol) (or 8) was dissolved in ethanol (25 mL) and methylhydrazine (0.046 g, 10.00
(
mmol) was added. After refluxing for 2 hours, the reaction mixture was stored overnight and then
concentrated to dryness under vacuum. The methylphthalhydrazide residue was stirred for 2 hours with
2
5 mL of 5N HCl, and filtered. The aqueous and acid extracts were combined and 3 mL of concentrated
HCl was added. After 2 hours, the aqueous solution was evaporated to give the product 1-[(2,3,4,6-tetra-
O-acetyl-β-D-galactopyranosyl)-3-amino-4-styryl] azetidin-2-one (9, 3.64g, 70.0%) (or 1-[(2,3,4,6-
tetra-O-acetyl-β-D-galactopyranosyl)-3-amino-4-hydroxymethyl]azetidin-2-one (10, 3.57g, 80.0%) as
the corresponding hydrochloride salts.
-1
+
1
(
9) m.p.: 91-92ºC (free amine); IR (cm ): 3250-3550 (br, N H ), 1780 (β-lactam), 1740 (ester); -
3
H-NMR (CDCl
3
) δ (ppm): 1.95-2.15 (12H, s, 4COCH
3
), 3.95-4.20 (3H, m, H
2
,CH OAc), 4.35 (1H, t,
2
H ), 4.50 (1H, dd, J=5 and J=8Hz, CH-CH=CH), 4.90 (1H, d, J=5Hz, CH-NH ), 5.05-5.2 (2H, m, H ,
3
2
4
H
5
), 5.95 (1H, d, H
6
), 6.20 (1H, dd, J=8 and J=16Hz, CH-CH=CHPh), 6.40 (1H, d, CH=CHPh), 7.10-
10: C, 57.91; H, 5.83; N, 5.40; O, 30.86%, found: C, 57.99;
7.25 (5H, m, Ph); Anal. Calc. for C25
H
30
2
N O
H, 5.70; N, 5.35; O, 30.91%.
-
1
(
10) m.p.: 110-112 °C (free amine); IR (cm ): 3200-3550 (NH
3
and OH), 1780 (β-lactam CO),
), 3.75 (2H, m, CH OH), 3.95-
), 4.45 (1H, br, OH), 4.70 (1H, dd, J=5Hz, CH-CH OH),
), 5.05-5.2 (2H, m, H , H ), 5.95 (1H, d, H ); Anal Calc. for C18
1
1
4
4
740(OCOCH
.20 (3H, m, H
.90 (1H, d, J=5Hz, CH-NH
3
); H-NMR (CDCl
3
) δ (ppm): 1.95-2.15 (12H, s, 4COCH
3
2
2
,CH
2
OAc), 4.35 (1H, t, H
3
2
2
4
5
6
H
26
N
2
11
O :
C, 48.43; H, 5.87; N, 6.28; O, 39.42%, found: C, 48.64; H, 5.80; N, 6.18; O, 39.50
General procedure for acylation of free-amino β-lactams 11a-d and 12a-d. Pyridine (1.50 g, 18.96
mmol) was added to a solution of 9 (3.12 g, 6.00 mmol), followed by dropwise addition of phenylacetyl
o
chloride (0.93 g 6.00 mmol) in CH
2
Cl
2
(20 mL). The solution was stirred for 2h. at 25 C. Then it was
washed with 10% HCl, 10% NaHCO
3
and water, dried (MgSO
4
), and the solvent was evaporated to give
the impure amide which was recrystallized from ethanol to give 1-[(2,3,4,6-tetra-O-acetyl-β-D-
galactopyranosyl)-3-phenylacetamido-4-styryl] azetidin-2-one ) 12c (2.00 g, 64.00%). M.p. 85-87°C. IR
–
1
1
(
cm ): 3410 (NH), 1770 (β-lactam), 1740 (ester), 1685 (amide); H-NMR (DMSO-d
6
) δ (ppm): 1.95-
2
.15 (12H, s, 4COCH ), 3.45 (2H, s, PhCH CO), 3.95-4.20 (3H, m, H , CH OAc), 4.35 (1H, t, H ), 4.50
3
2
2
2
3
(
1H, dd, J=5 and J=8Hz, CH-CH=CH), 4.90 (1H, dd, J=5 and J=8Hz, CH-NH), 5.05-5.2 (2H, m, H
), 5.95 (1H, d, H ), 6.20 (1H, dd, J=8 and J= 16Hz, CH-CH=CHPh), 6.40 (1H, d, CH=CHPh), 6.65
1H, d, J=8Hz, NH), 7.00-7.30 (10H, m, 2Ph); Anal. Calc. for C H N O : C, 62.26; H, 5.70; N, 4.40;
4
,
H
(
5
6
3
3
36
2
11
O, 27.64%, found: C, 62.20; H, 5.75; N, 4.42; O, 27.70%.
Compounds 11a-d, 12a-b and 12d were treated identically and their spectra were similar except for the
expected variations due to the amide side chains.

Molecules 2004, 9
37
Antibacterial Activity Tests
The antimicrobial activity tests were performed according to the disk diffusion method [25] using
Ampicillin and Gentamycin as the reference compounds. The sterile disks were impregnated with
different compounds (600 µg/disk). A nutrient agar medium was used and the disks were incubated at
o
3
7 C for 24 hours. After incubation, the relative susceptibility of the microorganisms to the potential
antimicrobial agent is demonstrated by a clear zone of growth inhibition around the disk. The inhibition
zones caused by the various compounds on the microorganisms were measured and the activity rated on
the basis of the size of the inhibition zone.
References
1
. (a) Grafe, U. Biochemie der Antibiotika; Spektrum Akademischer Verlag: Heidelberg, 1992; p. 359;
b) Jones, R.N.; Barry, A.L.; Thornsberry, C. J. Antimicrob.Chemother.1989, 24 (Suppl.A), 9.
(
2
3
4
. Chu, D.T.W; Plattner, J.; Katz, L. J. Med. Chem. 1996, 39, 3853.
. Page, M.I. The Chemistry of ß-Lactams; Blackie Academic & Professional: London, 1992.
. (a) Hashimoto, M.; Komori, T; Kamiya, T. J. Am. Chem. Soc., 1976, 98, 3023; (b) Aoki, H.; Sakai,
H.; Kohsaka, M.; Konomi, T.; Hosoda, J.; Kuboch, Y.; Iguch, E.; Imanka, H. J. Antibiot. 1976, 29,
4
92.
5
6
7
8
9
1
. Sykes, R.B.; Bonner, D.P. Am. J. Med. 1985, 7, 2.
. Bose, A.K.; Manhas, M.S.; Kapur, J.C.; Sharma, S.D.; Amin, S.G. J. Med. Chem. 1974, 17, 541.
. Fujisawa, T.; Shibuya, A.; Sato, D.; Shimitzu, M. Synlett 1995, 1067.
. Ricci, A. Modern Amination Methods; WILEY-VCH Verlag: Weinheim (FRG), 2000, p. 103.
. Miskolczi, I.; Sztaricskai, F.; Herczegh, P.; Bognar, R.; Koczka, I. J. Antibiot., 1985, 38, 1273.
0. (a) Barton, D.H.R.; Getau-Olesker, A.; Anaya-Mateos, J.; Cleophax, J.; Gero, S.D.; Chiaroni, A.;
Riche, C. J. Chem. Soc., Perkin Trans.1 1990, 3211; (b) Hernando, J.I.M.; Laso, N.M.; Anaya, J.;
Gero, S.D.; Grande, M. Synlett 1997, 281; (c) Anaya, J.; Gero, S.D.; Grande, M.; Hernando, J.I.M.;
Laso, N.M. Bioorg. Med. Chem. 1999, 7, 837.
1
1. Bose, A.K.; Manhas, M.S.; van der Veen, J.M.; Bari, S.S.; Wagle, D.R. Tetrahedron, 1992, 48,
4
831.
1
1
1
2. Gunda, T.E.; Sztaricskai, F. Tetrahedron, 1997, 53, 7985.
3. Palomo, C.; Oiarbide, M.; Esnal, A.; Miranda, J. I.; Linden, A. J. Org. Chem.. 1998, 63, 5838.
4. Han, W.T.; Trehan, A.K.; Wright, J.J.K.; Federici, M.E.; Seiler, S.M.; Meanwell, N.A. Bioorg. Med.
Chem Lett. 1995, 3, 1123.
1
1
1
5. Adlington, R.M.; Baldwin, J.E.; Chen. B.; Cooper, S.L.; McCoull, W.; Pritchard, G.J.; Howe, T.J.
Bioorg. Med. Chem. Lett. 1997, 7, 1689.
6. Borthwick, A.D.; Weingarten, G.; Haley, T.M.; Tomaszewski, W.; Wang, W.; Hu, Z.; Bedard, J.;
Jim, H.; Yuen, L.; Mansour, T.S. Bioorg. Med. Chem. Lett. 1998, 8, 365.
7. Palomo, C.; Aizpurua, J.M.; Gamboa, I.; Oiarbide, M. Amino Acids 1999, 16, 321.

Molecules 2004, 9
38
1
1
8. Kamel, M.M.; Naser, M.E. Die Pharmazie 1979, 34(7), 440.
9. Furniss, B.S.; Hannaford, A.J.; Smith, P.W.G.; Tartchel, A.R. Textbook of Practical Organic
th
Chemistry (Vogel’s), 5 ed.,; John Wiley & Sons, Inc.: New York 1991; p. 648.
2
2
2
2
2
2
0. Ricci, A, ref. [8], p. 106
1. Wolfe, S.; Hasan, S.K. Can. J. Chem. 1970, 48, 3572.
2. Kukolja, S.; Lammert, S.R. J. Am. Chem. Soc. 1975, 97, 5582.
3. Sheehan, J.C.; Ryan, J.J. J. Am. Chem. Soc. 1951, 73, 4367.
4. Sheehan, J.C.; Dadic, M. J. Heterocycl. Chem. 1968, 5, 779.
th
5. Finegold, S.M.; Martin, W.J. Diagnostic Microbiology, 6 ed.; Louis S.: London, 1982; p. 450.
Sample availability: Available from the authors
2004 by MDPI (http:www.mdpi.org). Reproduction is permitted for noncommercial purposes.
©