Synthesis and antibacterial activity of oxazolidinones containing pyridine substituted with heteroaromatic ring

Article abstract of DOI:10.1016/j.bmc.2004.08.025

Oxazolidinone derivatives containing pyridine substituted with heteroaromatic ring were synthesized and exhibited potent in vitro and in vivo antibacterial activities against many antibiotic-resistant microbial strains. A series of oxazolidinone derivatives, which morpholino group of linezolid was replaced with heteroaromatic ring substituted pyridine moiety, were newly synthesized, and their substituted effects on in vitro and in vivo antibacterial activities were evaluated against four problematic gram-positive strains including drug resistant strains and two gram-negative strains. Most compounds exhibited the enhanced in vitro activities with 4-16-fold and three compounds exerted more than 2-fold increased in vivo efficacies than linezolid.

Full text of DOI:10.1016/j.bmc.2004.08.025

Bioorganic & Medicinal Chemistry 12 (2004) 5909–5915
Synthesis and antibacterial activity of oxazolidinones containing
pyridine substituted with heteroaromatic ring
Yeong Woo Jo,a,b Weon Bin Im,b Jae Keol Rhee,b Mi Ja Shim,c
Won Bae Kimb and Eung Chil Choia,*
aCollege of Pharmacy and Research Institute of Pharmaceutical Sciences, Seoul National University, Seoul 151-742, Korea
bDong-A Pharmaceutical Co., Ltd, Research Laboratories, Yongin, Kyunggi 449-905, Korea
cDepartment of Life Science, The University of Seoul, Seoul 130-743, Korea
Received 29 July 2004; revised 18 August 2004; accepted 18 August 2004
Available online 11 September 2004
Abstract—A series of oxazolidinone derivatives, which morpholino group of linezolid was replaced with heteroaromatic ring substi-
tuted pyridine moiety, were newly synthesized, and their substituted effects on in vitro and in vivo antibacterial activities were eval-
uated against four problematic gram-positive strains including drug resistant strains and two gram-negative strains. Most
compounds exhibited the enhanced in vitro activities with 4–16-fold and three compounds exerted more than 2-fold increased
in vivo efficacies than linezolid.
ꢀ 2004 Elsevier Ltd. All rights reserved.
O
O
1. Introduction
O
O
O
N
O
N
N
H
N
H
N
The rapidly increasing incidence of multiple drug-resis-
tant gram-positive bacteria requires an urgent discovery
of novel active agents against these pathogens.1,2 The
oxazolidinones, a new class of synthetic antimicrobial
agent, are active against a variety of clinically important
susceptible and resistant gram-positive organisms such
as methicillin-resistant Staphylococcus aureus (MRSA),
vancomycin-resistant Enterococcus faecium (VRE), and
penicillin-resistant Streptococcus pneumoniae (PRSP).3–5
These compounds have a unique mechanism of inhibit-
ing bacterial protein synthesis at an early phase of trans-
lation by binding selectively to the central loop of
domain of 23S rRNA of 50S ribosomal subunit and
show no cross resistance with other classes of protein
synthesis inhibitor6–8 (Fig. 1).
F
O
O
Dup-721
Linezolid
Figure 1. Structures of oxazolidinone antibacterial agents.
lid-resistant isolates have been reported,12,13 and the
development of resistance urgently required to explore
a new oxazolidinone series with greater potency and
broader spectra of activity by further modification of
the basic oxazolidinone skeleton. Many groups have
reported the synthesis and the activity of novel oxazoli-
dinones,14–20 particularly some analogues of nitrogen–
carbon-linked oxazolidinone showed expanded spectrum
of activity and improved activity against H. influenzae.21
On this line, we designed some derivatives by introducing
heteroaromatic rings on the concept that the electron
affluence in the flat heteroaromatic ring structures could
induce maximum interaction with the binding site. In the
course of our efforts to improve spectrum and activity, we
recently found that several oxazolidinone derivatives
containing pyridine substituted with heteroaromatic ring
exhibited the enhanced antibacterial activities against
many antibiotic resistant strains compared to linezolid.
We herein report the synthesis, in vitro biological activi-
ties, and preliminary in vivo efficacies.
Linezolid was approved for the treatment of multi-drug
resistance gram-positive infections such as nosocomial
and community-acquired pneumonia and skin infec-
tions,9,10 since Dup-721 was introduced as the first
candidate agent of oxazolidinones.11 Recently, linezo-
Keywords: Oxazolidinone; Heteroaromatic ring substitution; Pyridine;
Antibacterial activity; In vivo efficacy.
*
Corresponding author. Tel.: +82 2 880 7874; fax: +82 2 886 5802;
e-mail: ecchoi@snu.ac.kr
0968-0896/$ - see front matter ꢀ 2004Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2004.08.025

5910
Y. W. Jo et al. / Bioorg. Med. Chem. 12(2004) 5909–5915
2. Chemistry
a
Br
Br
NC
Br
N
N
Synthesis of oxazolidinone derivatives is shown in
Scheme 1. The Stille coupling reactions22 of 123 with
the appropriate heteroaromatic ring containing pyridine
bromides (2a–l) were carried out in solvent N-methyl-2-
pyrrolidone with lithium chloride and palladium cata-
lyst, and the products were afforded to 20–50% yield
after chromatographic purification on silica gel (chloro-
form/methanol). Compounds 2a–c and 2i–l were
prepared from 2,5-dibromopyridine and necessary
reagents. The compounds 2d,f, and 2g were synthesized
with introduction of tetrazole moiety followed by methyl-
ation as shown in Scheme 2. The compound 2e was
obtained by reaction of compound 5 first with hydroxyl-
amine and subsequently with acetic anhydride as shown
in Scheme 3. The compound 2h was prepared as
described in Scheme 4. Briefly, the compound 5 was
treated with methylmagnesium bromide to produce the
compound 7 and that was further reacted with DMF
dimethylacetal and hydrazine to give compound 9. Sub-
sequent methylation of 9 afforded the compound 2h
(Scheme 4).
5
4
b
N
HO
N
c
O
Br
Br
N
N
H2N
N
6
2e
Scheme 3. Reagents and conditions: (a) NaCN, CuCN, DMF; (b)
NH2OH, NaHCO3, EtOH, reflux, 2h, 93%; (c) Ac2O, reflux, 2h, 29%.
3. Results and discussion
Twelve new oxazolidinone derivatives, bearing hetero-
aromatic ring substituted pyridine, were synthesized
and evaluated for in vitro antibacterial activity against
several clinically isolated strains and in vivo efficacy
using standard lethal-systematic infection model in
O
O
O
O
a
H
N
(H3C)3Sn
N
Br
R
H
N
R
N
+
N
N
F
F
O
O
1
2
c
3
N
N
N
N
N
N
O
N
N
N
N
N
N
N
R =
N
N
N
N
H
d
N
f
a
e
b
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
j
i
k
l
g
h
Scheme 1. Synthesis of oxazolidinone derivatives. Reagents and conditions: (a) LiCl, Pd(PPh3)2Cl2, NMP, 120ꢁC, 2h, 50%.
N
N
a
b
Br
Br
Br
NC
Br
N
N
H
N
N
N
2d
5
4
c
N
N
N
N
N
N
N
+
Br
Br
N
N
N
2f
2g
Scheme 2. Reagents and conditions: (a) NaCN, CuCN, DMF, 150ꢁC, 3h, 65%; (b) NaN3, NH4Cl, DMF, 120ꢁC, 4h; (c) MeI, KOH, DMF, rt, 24h.

Y. W. Jo et al. / Bioorg. Med. Chem. 12(2004) 5909–5915
5911
O
b
a
Br
Br
NC
Br
Br
N
N
N
4
7
5
c
O
e
d
Br
Br
Br
N
HN
N
N
N
N
N
N
2h
9
8
Scheme 4. Reagents and conditions: (a) NaCN, CuCN, DMF; (b) MeMgBr, THF, À20ꢁC, 2h; (c) DMF dimethylacetal, reflux, 15h; (d) hydrazine
hydrate, EtOH, reflux, 1h, 71%; (e) MeI, KOH, DMF, 0ꢁC, 1h, 72%.
mice. The in vitro activity was assessed by minimum
inhibitory concentration (MIC50) values expressed in
lg/mL utilizing standard agar dilution method and the
in vivo efficacy was evaluated by determination of effec-
tive dose (ED50) expressed in mg/kg.
pounds substituted with triazole (3b), oxadiazole (3e),
and tetrazole (3f) showed high in vitro and in vivo activ-
ities. Among these derivatives, the tetrazole substituted
oxazolidinone compound 3f was found to be the most
active compound with MICs of 0.39, 0.78, 0.39, 0.2,
1.56, and 3.13lg/mL against MSSA, MRSA, VRE,
PRSP, M. catarrhalis, and H. influenzae, respectively,
and ED50 of 3.4mg/kg. These superior antibacterial
activities might be interpreted that the increased hydro-
phobic interaction of these heteroaromatic ring moieties
in the oxazolidinone structure with 23S rRNA binding
pocket might endow a stronger intrinsic binding attrac-
tion for the site of action as suggested by Gandhi
et al.24 although their activities vary with the structures
and position or numbers of heteroaromatic ring.
In order to procure highly potent oxazolidinone deriva-
tives with a long half-life and elucidate the SAR based
on the mode of action, we substituted oxazolidinone at
the 4-position of phenyl ring of its mother nucleus. In a
series of substitutions with hetero rings such as pyridine,
pyrimidine, and pyridazine, the pyridine derivatives
substituted at the 4-position of phenyl ring showed
potent antibacterial activities than the pyrimidine and
pyridazine derivatives (data not shown). Further substi-
tutions using heteroaromatic ring were added to pyridine
ring and their in vitro and in vivo activities were com-
pared to those of linezolid. The results of the MIC50
and ED50 are presented in Table 1. These substituted
compounds with heteroaromatic ring at pyridine (3a–l)
exhibited the enhanced activity with 4–16-fold against
six strains compared to linezolid. Especially, the com-
In comparison with linezolid, especially, compound 3f
showed potential activity against M. catarrhlis and
H. influenzae (MIC: 0.78 and 3.13lg/mL, respectively)
where linezolid was practically inactive (MIC: 6.25 and
12.5lg/mL, respectively). However, the compounds
substituted with diazole (3c,h, and 3j) exhibited low
Table 1. In vitro and in vivo activities of oxazolidinone derivatives
g
ED50 (mg/kg)
Compound
MIC50 (lg/mL)
MSSAa (30)
MRSAb (20)
VREc (21)
PRSPd (32)
M. cate (35)
H. inff (35)
3a
3b
0.39
0.39
1.56
—
0.39
0.39
1.56
>25
0.2
0.2
0.2
0.2
—
1.56
1.56
—
6.25
6.25
>25
—
12.0
3.9
—
3c
3d
0.39
25
—
—
—
3e
0.39
0.39
0.78
—
0.78
0.78
0.78
0.78
0.78
0.39
0.39
0.78
3.13
0.39
0.2
0.2
0.2
—
1.56
1.56
—
6.25
3.13
6.25
50
3.9
3.4
6.7
11.1
3f
3g
0.2
3h
3i
0.39
0.39
0.2
—
—
—
0.78
0.39
0.39
—
1.56
1.56
1.56
—
12.5
3.13
6.25
3.13
12.5
>80
3j
3k
0.1
0.1
—
14.9
6.1
8.1
8.0
0.2
3l
Linezolid
0.39
1.56
3.13
1.56
6.25
a Methicillin-susceptible S. aureus.
b Methicillin-resistant S. aureus.
c Vancomycin-resistant enterococci.
d Penicillin-resistant Streptococcus pneumoniae.
e Moraxella catarrhalis.
f Haemophilus influenzae.
g Mouse was infected intraperitoneally with mucin suspension of S. aureus Smith and test compounds were administered orally 1h after infection.
ED50 was the amount of antibiotic in milligrams per kilogram of body weight required to cure 50% of the infected mice.

5912
Y. W. Jo et al. / Bioorg. Med. Chem. 12(2004) 5909–5915
activity against H. influenzae with MICs of >12.5lg/mL
and low in vivo activity even though the antibacterial
activity against gram-positive bacteria was nearly the
same as that of 3f. In case of tetrazole substitution,
the effects of substituted forms of tetrazole ring and
the position of methyl group in tetrazole ring on their
antibacterial activity were briefly investigated. Antibac-
terial activities of the different-types of tetrazole ring
structures substituted with methyl group like 3f, 3g,
and 3k, 3l maintained the biological activity. However,
the replacement of the methyl moiety with free acidic
portion as found in 3d resulted in complete loss of anti-
bacterial activity against gram-positive bacteria. Thus,
the presence of methyl group and its orientation can
affect activity in these structures. Furthermore, com-
pound 3f showed a long in vivo half-life of 12.4h in rats
compared to 0.9h of linezolid, and it could increase
administration intervals in clinical use. This might be
correlated with the higher in vivo activity than linezolid.
and the residue was subjected to column chromato-
graphy to give 3a–l.
4.2. (S)-[N-3-(4-(2-(1-methyl-5-tetrazolyl)-pyridin-5-yl)-
3-fluorophenyl)-2-oxo-5-oxazolidinyl]methyl acetamide
(3f)
This was obtained from the 2-(1-methyl-5-tetrazolyl)-5-
bromopyridine (2f) (510mg, 2.12mmol) and intermedi-
ate 1 (1.17g, 2.82mmol); yield 530mg (1.28mmol,
45%), mp 217–219ꢁC. 1H NMR (CDCl3): d 8.89 (s,
1H, pyridine), 8.29 (d, 1H), 8.00 (m, 1H), 7.61 (dd,
1H, ArH), 7.46 (t, 1H), 7.26 (dd, 1H), 6.12 (t, 1H,
–NH–), 4.80 (m, 1H, –CH–), 4.45 (s, 3H, –NCH3),
4.07 (t, 1H), 3.83 (dd, 1H), 3.67 (m, 2H), 2.02 (s, 3H,
–COCH3). IR (KBr, cmÀ1): 3430, 3036, 2987, 2953,
1743, 1677, 1226. HRMS C19H18FN7O3, calcd (M):
411.1455; meas.: 411.1460 (Table 2).
4.3. 2-(1,2,3,4-Tetrazolyl)-5-bromopyridine (2k)
In conclusion, a series of heteroaromatic ring substi-
tuted pyridinyl oxazolidinone antibacterial agents were
discovered with in vitro activity against clinically rele-
vant resistant gram-positive organisms, M. catarrhlis
and H. influenzae and with a long half-life. Diverse
substituted forms of heteroaromatic ring are tolerated
on the pyridine and the presence or orientation of
methyl group in the heteroaromatic ring affected the
antibacterial activity. Three of the most potent com-
pounds, 3b,e, and 3f, were active against a variety of
clinically relevant resistant gram-positive organisms
and M. catarrhlis and H. influenzae, and also showed
the higher in vivo efficacy with longer half-lives than
linezolid.
To the solution of 2,5-dibromopyridine (1.0g,
4.22mmol) in N-methyl-2-pyrrolidone (10mL), 1,2,3,4-
tetrazole (0.5g, 7.14mmol) along with 1.75g of potas-
sium carbonate was added. The reaction mixture was
heated at 100ꢁC for 3h with stirring. After completion
of the reaction, the reaction mixture was added to water
and extracted with ethyl acetate. The organic layer thus
obtained was dehydrated, filtered, and concentrated and
the concentrate was subjected to column chromatogra-
phy to give 2k (0.8g, 3.54mmol, 84%). TLC (ethyl ace-
tate/hexane: 1:4, Rf: 0.4), 1H NMR (DMSO-d6): d 10.17
(s, 1H), 8.80 (d, 1H), 8.40 (dd, 1H), 8.00 (d, 1H).
4.4. 2-Cyano-5-bromopyridine (5)
2,5-Dibromopyridine (10.0g, 42.21mmol) was dissolved
in dimethylformamide (100mL) and copper cyanide
(3.0g, 33.50mmol) and sodium cyanide (1.7g,
34.69mmol) were added to this solution. The reaction
mixture was heated to reflux for 3h. After completion
of the reaction, the reaction mixture was added to the
water and extracted with ethyl acetate. The organic layer
obtained was dehydrated, filtered, and concentrated.
The crude product was triturated with n-hexane to af-
ford 5 (5g, 27.32mmol, 65%) after filtration. TLC (ethyl
4. Experimental
MICs were determined by the twofold serial agar dilu-
tion method with Mueller–Hinton agar.25 In vivo experi-
ments were also conducted according to the previously
reported method.26 The melting points were determined
1
on Mettler Toledo FP62 apparatus. H NMR spectra
were recorded on Varian 400MHz spectrophoto-
meter by CDCl3 or DMSO-d6 and TMS as an internal
standard (chemical shift in d ppm). IR spectra were
measured on Nicolet Magna 550 Series II IR spectro-
photometer. The mass spectra were recorded on
a LC/GC–MS/MS, High Resolution Tandem Mass
Spectrometer. The results were within 0.4% of the
theoretical values.
1
acetate/hexane: 1:8, Rf: 0.5), H NMR (CDCl3): d 8.76
(s, 1H), 7.96 (dd, 1H), 7.57 (dd, 1H).
4.5. 2-(5-1,2,3,4-Tetrazolyl)-5-bromopyridine (2d)
To the solution of 5 (4.5g, 24.59mmol) in dimethylform-
amide (50mL) at room temperature were added ammo-
nium chloride (4.2g, 78.52mmol) and sodium azide
(2.4g, 36.92mmol). After stirring at 120 ꢁC for 4h, the
reaction mixture was added to ice water and controlled
to pH2 with a 6N HCl solution. After additional stir-
ring for 1h at room temperature, the reaction mixture
was extracted with ethyl acetate. The organic layer thus
collected was dehydrated, filtered, and concentrated to
give crude 2d, which was used for next step without fur-
4.1. Synthesis
General procedure for the synthesis of oxazolidinone
derivatives, (3a–l): To the solution of intermediate 1 in
N-methyl-2-pyrrolidone at room temperature was added
side chain (2a–l), lithium chloride and dichlorobistriphen-
ylphosphine palladium (II). The reaction mixture was
kept at 120ꢁC for 4h with stirring and water was added
thereto. The organic layer obtained after extraction with
ethyl acetate was dehydrated, filtered, and concentrated
1
ther purification. TLC (CHCl3/MeOH: 5:1, Rf: 0.2), H

Table 2. Characterization data of target compounds
3a
3b
Yield: 35%; mp: 228ꢁC; 1H NMR (DMSO-d6): d 8.70 (s, 1H), 8.58 (s, 1H), 8.21 (m, 2H), 7.99 (m, 2H), 7.69 (m, 2H), 7.48 (dd, 1H), 7.14 (s, 1H), 4.77 (m, 1H), 4.19 (t, 1H), 3.78
(dd, 1H), 3.43 (t, 2H), 1.83 (s, 3H); HRMS C20H18FN5 O3, calcd (M): 395.1394; meas.: 395.1363
Yield: 32%; mp: 224–225ꢁC; 1H NMR (DMSO-d6): d 9.41 (s, 1H, triazole), 8.90 (d, 1H, pyridine), 8.72 (s, 1H), 8.33 (s, 1H, triazole), 8.29–8.25 (m, 2H, –NH–, pyridine), 7.99
(d, 1H), 7.76–7.62 (m, 2H, ArH), 7.46 (dd, 1H), 4.76 (m, 1H, –CH–), 4.17 (t, 1H), 3.79 (dd, 1H), 3.43 (t, 2H), 1.83 (s, 3H, –OCH3); IR (KBr, cmÀ1): 3347, 3123, 2988, 2928
1739, 1417, 987; HRMS C19H17FN6O3, calcd (M): 396.1346; meas.: 396.1350
Yield: 40%; 1H NMR (DMSO-d6): d 8.74 (s, 1H), 8.21 (m, 2H), 7.74 (m, 2H), 7.68 (m, 2H), 7.48 (dd, 1H), 6.94 (dd, 1H), 4.76 (m, 1H), 4.18 (t, 1H), 3.80 (dd, 1H), 3.43 (t, 2H),
2.25 (s, 3H), 1.87 (s, 3H); HRMS C21H20FN5O3, calcd (M): 409.1550; meas.: 409.1512
Yield: 35%; mp: 236ꢁC; 1H NMR (DMSO-d6 ): d 8.97 (s, 1H, pyridine), 8.29 (m, 3H), 7.72 (t, 1H, ArH), 7.65 (dd, 1H), 7.47 (dd, 1H), 4.78 (m, 1H, –CH–), 4.18 (t, 1H), 3.81 (dd, 1H),
3.44 (dd, 2H), 1.83 (s, 3H, –OCH3); IR (KBr, cmÀ1): 3333, 2973, 2887, 2727, 1754, 1624, 1395, 842; HRMS C18H16FN7 O3, calcd (M): 397.1299; meas.: 397.1304
Yield: 36%; mp: 215ꢁC; 1H NMR (CDCl3): d 8.85 (s, 1H, pyridine), 8.09 (d, 1H), 7.97 (m, 1H), 7.58 (dd, 1H, ArH), 7.45 (t, 1H), 7.26 (dd, 1H), 6.50 (t, 1H, –NH–), 4.81 (m, 1H, –CH–),
4.10 (t, 1H), 3.84 (dd, 1H), 3.68 (m, 2H), 2.67 (s, 3H, oxadiazole), 2.01 (s, 3H, –OCH3); IR (KBr, cmÀ1): 3306, 3096, 1774, 1761, 1407, 1221; HRMS C20H18FN5O4, calcd (M): 411.1343;
meas.: 411.1340
3c
3d
3e
3f
Yield: 50%; mp: 217–219ꢁC; 1H NMR (CDCl3): d 8.89 (s, 1H, pyridine), 8.29 (d, 1H), 8.00 (m, 1H), 7.61 (dd, 1H, ArH), 7.46 (t, 1H), 7.26 (dd, 1H), 6.12 (t, 1H, –NH–), 4.80 (m,
1H, –CH–), 4.45(s, 3H, –NCH3), 4.07 (t, 1H), 3.83 (dd, 1H), 3.67 (m, 2H), 2.02 (s, 3H, –COCH3); IR (KBr, cmÀ1): 3430, 3036, 2987, 2953, 1743, 1677, 1226, HRMS C19H18FN7O3,
calcd (M): 411.1455; meas.: 411.1460
3g
3j
Yield: 28%; 1H NMR (DMSO-d6): d 8.92 (s, 1H), 8.35 (m, 3H), 7.79 (m, 1H), 7.70 (dd, 1H), 7.46 (dd, 1H), 4.78 (m, 1H), 4.44 (s, 3H), 4.14 (t, 1H), 3.84 (dd, 1H), 3.43
(t, 2H), 1.83 (s, 3H); HRMS C19H18FN7O8, calcd (M): 411.1455; meas.: 411.1486
Yield: 35%; 1H NMR (DMSO-d6): d 8.88 (s, 1H), 8.76 (s, 1H), 8.28 (d, 1H), 8.21 (d, 1H), 8.01 (s, 1H), 7.70 (m, 2H), 7.51 (d, 1H), 4.76 (m, 1H), 4.17 (t, 1H), 3.79 (m, 1H),
3.42 (t, 2H), 1.83 (s, 3H); HRMS C19H17FN6O3, calcd (M): 396.1346; meas.: 396.1332
3k
Yield: 20%; mp: 210ꢁC; 1H NMR (DMSO-d6): 10.22 (s, 1H, tetrazole), 8.83 (s, 1H, pyridine), 8.39 (dd, 1H, pyridine), 8.30 (t, 1H, –NH–), 8.15 (d, 1H, pyridine),
7.76–7.64 (m, 2H, ArH), 7.47 (dd, 1H), 4.78 (m, 1H, –CH–), 4.16 (t, 1H), 3.80 (dd, 1H), 3.43 (t, 2H),1.83 (s, 3H, –OCH3); IR (KBr, cmÀ1): 3408, 3113, 2927, 1740, 1667,
1225; HRMS: C18H16FN7O3, calcd (M): 397.1299; meas.: 397.1307
Yield: 37%; 1H NMR (DMSO-d6): d 8.83 (s, 1H), 8.34 (d, 1H), 8.24 (t, 1H, NH), 8.07 (d, 1H), 7.77 (t, 1H), 7.65 (dd, 1H), 7.48 (dd, 1H), 4.77 (m, 1H), 4.18 (t, 1H),
3.80 (dd, 1H), 3.43 (t, 2H), 2.82 (s, 3H), 1.83 (s, 3H); HRMS C19H18FN7O3, calcd (M): 411.1455; meas.: 411.1442
3l

5914
Y. W. Jo et al. / Bioorg. Med. Chem. 12(2004) 5909–5915
NMR (DMSO-d6): d 8.91 (br s, 1H), 8.31 (br s, 1H), 8.13
(br, 1H).
4.10. 2-(3-Pyrazolyl)-5-bromopyridine (9)
2-Acetyl-5-bromopyridine (7) (2g, 10.00mmol) was ref-
luxed in dimethylformamide dimethylacetal for 15h.
After evaporating excess solvent, the resulting residue
(crude 8) was dissolved in ethanol and hydrazine hy-
drate (5mL, 99.88mmol) was added thereto. The mix-
ture was refluxed for 1h. The crude product obtained
from the ordinary work-up was further purified by col-
umn chromatography to give the desired product 9. It
was purified by column chromatography to produce
the desired product 9 (1.6g, 7.14mmol, 71%). TLC
4.6. 2-(1-Methyl-5-tetrazolyl)-5-bromopyridine (2f) and
2-(2-methyl-5-tetrazolyl)-5-bromopyridine (2g)
To the solution of crude 2d (6.0g, 26.55mmol) in
dimethylformamide (60mL) were added iodomethane
(15g, 106.18mmol) and potassium hydroxide (3.7g,
66.37mmol). After stirring for 24h at room tempera-
ture, work-up as described before followed by column
chromatography gave the polar desired product 2f
(1.0g, 4.17mmol) along with the less polar 2g (2.2g,
9.16mmol) as a by-product.
1
(ethyl acetate/hexane: 1:4, Rf: 0.1), H NMR (CDCl3):
d 11.56 (s, 1H, NH), 8.68 (d, 1H), 7.83 (dd, 1H), 7.70
(d, 1H), 7.66 (s, 1H), 6.81 (d, 1H).
Compound 2f: TLC (ethyl acetate/hexane: 1:4, Rf: 0.3),
1H NMR (CDCl3): d 8.80 (d, 1H), 8.11 (d, 1H), 7.96
(dd, 1H), 4.42 (s, 3H from CH3 on tetrazole ring).
4.11. 2-(1-Methyl-3-pyrazolyl)-5-bromopyridine (2h)
To the solution of 9 (300mg, 1.34mmol) in dimethylform-
amide were added iodomethane (760mg, 5.36mmol) and
potassium hydroxide (230mg, 4.10mmol). After stirring
for 1h at 0ꢁC, work-up as described before and purifica-
tion by column chromatography gave the desired product
2h (230mg, 0.97mmol, 72%). TLC (ethyl acetate/hexane:
Compound 2g: TLC (ethyl acetate/hexane: 1:4, Rf: 0.5),
1H NMR (CDCl3): d 8.77 (d, 1H), 8.25 (d, 1H), 8.03 (dd,
1H), 4.48 (s, 3H, from CH3 on tetrazole ring).
Compounds 2f and 2g were easily distinguished
from each other by comparison of the chemical shifts
of CH3 on tetrazole ring and their polarities on
TLC.27,28
1
1:1, Rf: 0.4), H NMR (CDCl3): d 8.63 (t, 1H), 7.78 (m,
2H), 7.38 (d, 1H), 6.81 (d, 1H), 3.94(s, 3H).
4.12. In vitro activity
4.7. 2-(2-Amino-1-oxime)-5-bromopyridine (6)
The MICs were determined using an agar dilution method
by NCCLS guidelines. An overnight culture was inocu-
lated with an inoculating device onto Muller–Hinton
agar containing serial twofold dilutions of the test com-
pounds. Organisms were incubated at 37ꢁC for 18–24h.
The MIC of a compound was defined as the lowest con-
centration that visibly inhibited growth.
To the solution of 5 (2.0g, 10.93mmol) in EtOH (20mL)
were added hydroxylamine (3.8g, 54.68mmol) and so-
dium hydrogen carbonate (4.6g, 54.76mmol) and the
reaction mixture was heated to reflux for 2h. The ordin-
ary work-up was conducted to give crude 6 (2.2g,
10.18mmol, 93%). TLC (ethyl acetate/hexane: 1:1, Rf:
1
0.2), H NMR (DMSO-d6): d 10.02 (s, 1H), 8.65 (d,
1H), 8.01 (dd, 1H), 7.78 (dd, 1H).
4.13. In vivo efficacy
Four-week-old male outbred ICR mice (18–20g) were
given an intraperitoneal injection of S. aureus Smith
(2.9 · 107 CFU/mice) suspended in 5% mucin. Eight
mice per group were used. Test compounds were orally
administered 1h after bacterial infection. Mice were ob-
served for mortality over seven days. The total number
of survivors at each dose was used to calculate the effec-
tive dose that protected 50% of the infected mice from
death (ED50). ED50 and 95% confidence limits determi-
nations were performed by Probit analysis within each
test.
4.8. 2-(5-Methyl-1,2,4-oxadiazol-3-yl)-5-bromopyridine
(2e)
Compound 6 (8.6g, 39.81mmol) was dissolved in acetic
anhydride (250mL) and the solution was heated to re-
flux for 2h. Excess acetic anhydride was removed under
reduced pressure and the resulting residue was purified
on column chromatography to yield 2e (2.8g,
11.66mmol, 29%). TLC (ethyl acetate/hexane: 1:2, Rf:
0.7), 1H NMR (CDCl3): d 8.80 (dd, 1H), 7.96 (dd,
2H), 2.67 (s, 3H).
4.9. 2-Acetyl-5-bromopyridine (7)
Acknowledgements
To the solution of 5 (1g, 5.46mmol) in THF (50mL) at
À20ꢁC was added drop by drop methyl magnesiumbro-
mide (54mL). And the reaction mixture was stirred for
2h and warmed up to room temperature slowly. The or-
dinary work-up was conducted to give crude 7 (0.5g,
2.50mmol), which was used without further purification.
TLC (ethyl acetate/hexane: 1:4, Rf: 0.6), 1H NMR
(CDCl3): d 8.70 (dd, 1H), 7.92 (dd, 2H), 2.67 (s,
3H).
This work was supported by the Korea Ministry of
Health and Welfare.
References and notes
1. Norrby, R. Exp. Opin. Pharmacother. 2001, 2, 293.
2. Tomasz, A. N. Engl. J. Med. 1994, 330, 1247.

Y. W. Jo et al. / Bioorg. Med. Chem. 12(2004) 5909–5915
5915
3. Perry, C. M.; Javis, B. Drugs 2001, 61, 525.
4. Stevens, D. L.; Herr, D.; Lampiris, H.; Hunt, J. L.; Batts,
D. H.; Hafkin, B. Clin. Infect. Dis. 2002, 34, 1481.
5. Moellering, R. C. Ann. Intern. Med. 1999, 130, 155.
6. Aoki, H.; Ke, L.; Poppe, S. M.; Poel, T. J.; Weaver, E. A.;
Gadwood, R. C.; Thomas, R. C.; Shinabarger, D. L.;
Gonaza, M. C. Antimicrob. Agents Chemother. 2002, 46,
1080.
18. Yu, D.; Huiyuan, G. Bioorg. Med. Chem. Lett. 2002, 12,
857.
19. Selvakumar, N.; Srinivas, D.; Khera, M. K.; Kumar, M.
S.; Mamidi, R. N.; Sarnaik, H.; Charavaryamath, C.;
Rao, B. S.; Raheem, M. A.; Das, J.; Iqbal, J.; Rajagopa-
lan, R. J. Med. Chem. 2002, 45, 3953.
20. Cui, Y.; Yang, Y.; Chen, K.; Ji, R.; Zhang, S. Bioorg.
Med. Chem. Lett. 2003, 13, 2311.
7. Kloss, P.; Xiong, L.; Mankin, A. S.; Shinabarger, D. L.;
Mankin, A. S. J. Mol. Biol. 1999, 294, 93.
8. Bobkova, E. V.; Yan, Y. P.; Jordan, D. B.; Kurilla, M. G.;
Pompliano, D. L.; Jordan, D. B. J. Biol. Chem. 2003, 278,
9802.
9. Bassey, P. Drug Market Dev. 2000, 11, 392.
10. Diekema, D. J.; Jones, R. N. The Lancet 2001, 358, 1975.
11. Slee, A. M.; Wuonola, M. A.; McRipley, R. J.; Zajac, I.;
Zawada, M. J.; Bartholomew, P. T.; Gregory, W. A.;
Forbes, M. Antimicrob. Agents Chemother. 1987, 31, 1791.
12. Gonzales, R. D.; Schreckenberger, P. C.; Graham, M. B.;
Kelkar, S.; DenBesten, K.; Quinn, J. P. The Lancet 2001,
357, 1179.
21. Genin, M. J.; Allwine, D. A.; Anderson, D. J.; Barbachyn,
M. R.; Emmert, D. E.; Garmon, S. A.; Graber, D. R.;
Grega, K. C.; Hester, J. B.; Hutchinson, D. K.; Morris, J.;
Reischer, R. J.; Ford, C. W.; Zurenko, G. E.; Hamel, J.
C.; Schaadt, R. D.; Stapert, D.; Yagi, B. H. J. Med. Chem.
2000, 43, 953.
22. Stille, J. K. Angew. Chem., Int. Ed. Engl. 1986, 124,
508.
23. Barbachyn, M. R.; Brickner, S. J. U.S. Patent 5,565,571,
1996.
24. Gandhi, N.; Srivastava, B. K.; Lohray, V. B.; Lohray, B.
B. Tetrahedron Lett. 2004, 45, 6269.
25. National Committee for Clinical Laboratory Standards
(NCCLS). Methods for Dilution Antimicrobial Susceptibil-
ity Tests for Bacteria that Grow Aerobically, 4th ed.;
Approved standard: NCCLS Document M7-A4; National
Committee for Clinical Laboratory Standards: Villanova,
PA, 1997.
13. Tsiodras, S.; Gold, H. S.; Sakoulas, G.; Eliopoulas, G. M.;
Wennerstein, C.; Venkaturaman, L.; Moellering, R. C.,
Jr.; Ferro, M. J. The Lancet 2001, 358, 207.
14. Huiyuan, G.; Yu, D. Bioorg. Med. Chem. Lett. 2002, 12,
857.
15. Weidner-Wells, M. A.; Boggs, C. M.; Foleno, B. D.; Wira,
E.; Bush, K.; Goldschmidt, R. M.; Hlasta, D. J. Bioorg.
Med. Chem. Lett. 2001, 11, 1829.
26. Cleland, R.; Squires, E. In Antibiotics in Laboratory
Medicine; 3rd ed.; Loria, V., Ed., Williams Wilkins:
Baltimore, 1991; pp 747–762.
16. Tucker, J. A.; Allwine, D. A.; Grega, K. C.; Barbachyn,
M. R.; Klock, J. L.; Adamski, J. L.; Brickner, S. J.;
Hutchinson, D. K.; Ford, C. W.; Zurenko, G. E.;
Conradi, R. A.; Burton, P. S.; Jensen, R. M. J. Med.
Chem. 1998, 41, 3727.
17. Tokuyama, R.; Takahashi, Y.; Tomita, Y.; Suzuki, T.;
Yoshida, T.; Iwasaki, N.; Kado, N.; Okezaki, E.; Nagata,
O. Chem. Pharm. Bull. 2001, 49, 347.
27. Tanaka, A.; Terasawa, T.; Hagihara, H.; Sakuma, Y.;
Ishibe, N.; Sawada, M.; Tanake, H. Med. Chem. 1998, 41,
2390.
28. Street, L. J.; Baker, R.; Davey, W. B.; Guiblin, A. R.;
Jelley, R. A.; Reeve, A. J.; Routledge, H.; Sternfeld, F.;
Beer, M. S.; Middlemiss, D. N.; Noble, A. J.; Stanton, J.
A.; Scholey, K.; Hargreaves, R. J.; Sohal, B.; Granham,
M. I.; Matssa, V. G. J. Med. Chem. 1995, 38, 1799.