Synthesis and structure-Activity relationships of novel lincomycin derivatives. Part 2. Synthesis of 7(S)-7-deoxy-7-(4-morpholinocarbonylphenylthio)lincomycin and its 3-dimensional analysis with rRNA

Article abstract of DOI:10.1038/ja.2015.125

Lincomycin derivatives, which possess a hetero ring at the C-7 position via sulfur atom, were synthesized by three types of reactions: (1) Mitsunobu reaction of 2,3,4-Tris-O-(trimethylsiliyl)lincomycin (1) with the corresponding thiol, (2) S N 2 reaction

Full text of DOI:10.1038/ja.2015.125

The Journal of Antibiotics (2015), 1–12
2015 Japan Antibiotics Research Association All rights reserved 0021-8820/15
www.nature.com/ja
&
ORIGINAL ARTICLE
Synthesis and structure–activity relationships
of novel lincomycin derivatives.
Part 2. Synthesis of 7(S)-7-deoxy-7-(4-morpholino
carbonylphenylthio)lincomycin and its 3-dimensional
analysis with rRNA
Yoshinari Wakiyama, Ko Kumura, Eijiro Umemura, Satomi Masaki, Kazutaka Ueda, Takashi Watanabe,
Mikio Yamamoto, Yoko Hirai and Keiichi Ajito
Lincomycin derivatives, which possess a hetero ring at the C-7 position via sulfur atom, were synthesized by three types of
reactions: (1) Mitsunobu reaction of 2,3,4-tris-O-(trimethylsiliyl)lincomycin (1) with the corresponding thiol, (2) SN2 reaction of
7-O-methanesulfonyl-2,3,4-tris-O-(trimethylsiliyl)lincomycin (2) with the corresponding thiol and (3) Pd-catalyzed cross-coupling
reaction of 7-deoxy-7-epi-7-mercaptolincomycin (35) with the corresponding aryl halides. As a result, compound 28 had potent
antibacterial activities against major pathogens, which caused respiratory infections, even compared with clindamycin. On the
other hand, compound 38 showed most potent activities against a variety of Streptococcus pneumoniae with erm gene.
The Journal of Antibiotics advance online publication, 16 December 2015; doi:10.1038/ja.2015.125
INTRODUCTION
interactions by a hydrogen bonding between the peptidyl transferase
Macrolide antibiotics possess a broad spectrum of antibacterial cavity (A2058Ec, A2059Ec and G2520Ec) and hydroxyl groups at the
activities against Streptococcus pneumoniae, Streptococcus pyogenes, sugar portion of CLDM.4,6 This observation suggests that it is difficult
Haemophillus influenzae, Moraxella catarrhalis, Mycoplasma to improve antibacterial activity by chemical modification at the sugar
pneumoniae, Neisseria gonorrhoeae and so on. Macrolide antibiotics moiety. In fact, 2-deoxylincomycin18 was reported to have weaker
have been used in clinical sites over many years. Recently, resistant antibacterial activities even when compared with LCM.
bacteria, especially S. pneumoniae with erm gene, have markedly
increased,1–3 which are causing serious problems in the field of broth of Streptomyces lincolnensis. CLDM was synthesized by the
bacterial respiratory infections. chemical modification of LCM (Figure 1), and antimicrobial activities
LCM was isolated as a secondary metabolite from the fermentation
Macrolide antibiotics inhibit chain elongation of bacterial protein and pharmacokinetics of CLDM were improved in comparison with
by binding to 23S ribosomal RNA,4–7 and consequently inhibit those of LCM. Furthermore, chemical modifications at the C-7
bacterial protein synthesis. However, clarithromycin8 and position of LCM were investigated by Hoeksema et al.,19 Magerlein
azithromycin9 are not effective enough against resistant bacteria such et al.,20,21 Sinkula et al.,22 Birkenmeyer et al.,17 Lewis et al.,23 Bannister
as S. pneumoniae with erm gene (Figure 1, Table 1). On the other et al.,18,24–30 Sztaricskai and Ōmura et al.31 and so on. As a result,
hand, telithromycin (TEL)10 and some of our novel macrolide some of the LCM derivatives possessing a substituent at the C-7
derivatives11 synthesized from 16-membered macrolide are effective position via sulfur atom with 7(S)-configuration had potent anti-
against resistant S. pneumoniae with erm gene.
bacterial activities when compared with LCM. But no LCM derivatives
TEL, however, has possibility to cause serious liver damage,12 and it were effective against resistant bacteria of S. pneumoniae or S. pyogenes
is scarcely used in Japan. No oral antibiotic, which is effective against with erm gene that caused problems in clinical sites.
resistant bacteria of S. pneumoniae and does not have any problems in
safety or taste, has been launched so far.
Lincomycin (LCM)13–16 and clindamycin (CLDM)17 inhibit 7(S)-configuration. Among them, 7(S)-7-(6-amino-benzothiazol-2-
On the other hand, we had already reported LCM derivatives
possessing a hetero ring at the 7 position via sulfur atom32–34 with
bacterial protein synthesis similar to macrolide antibiotics. X-ray yl-thio)-7-deoxylincomycin,
7(S)-7-deoxy-7-(4-methoxycarbonyl-
crystallographic analysis indicates that there are several major phenylthio)lincomycin, 7(S)-7-(5-amino-1,3,4-thiadiazol-2-yl-thio)-
Pharmaceutical Research Center, Meiji Seika Pharma Co., Ltd., Yokohama, Japan
Correspondence: Dr K Ajito, Pharmaceutical Research Center, Meiji Seika Pharma Co., Ltd., 760 Morooka-cho, Kohoku-ku, Yokohama 222-8567, Japan.
E-mail: keiichi.ajito@meiji.com
Received 10 October 2015; revised 2 November 2015; accepted 12 November 2015

Synthesis and structure–activity relationships of novel LCM derivatives
Y Wakiyama et al
2
Me
O
N
Me
HO
Me
Me
Me
OH
Me
HO
Me
OMe
Me
Me
OH
Me
OH
Me
O Me
R2
R1
O
NMe2
Me
NMe2
Me
Me
O
HO
HO
O
O
O
O
O
N
HN
Me
Me
Me
O
O
O
O
OMe
Me
OMe
HO
HO
SMe
Me
Me
Me
Me
OH
Me
OH
O
O
OH
LCM: R1 = OH; R2 = H
CLDM: R1 = H; R2 = Cl
Clarithromycin (CAM)
Figure 1 Chemical structures of the representative macrolides, lincomycin (LCM) and clindamycin (CLDM).
Azithromycin (AZM)
these intermediates to synthesize a variety of derivatives in this article,
which had the same 7(S)-configuration as CLDM. Compound 2 is a
very useful intermediate to synthesize LCM derivatives, and it was first
synthesized by us with one step from compound 1. An SN2 reaction of
2 under basic condition with the corresponding thiol gave desired
molecules 3 and 4. Other desired molecules 5–8 were directly
synthesized from 1.
Compound 3 was treated with methoxymethylcarbonyl chloride
and Et3N, followed by deprotection of TMS (trimethylsilyl) groups
under the 1 N HCl condition provided 9 in 99% yield with two steps.
On the other hand, TMS groups of ester compounds (4–8) were
deprotected under the 1 N HCl condition to give the corresponding
analogs (10–14) in good-to-excellent yields. Amido analogs 15–17 and
18 were prepared from the corresponding ester compounds (10 and
14) by aminolysis, respectively. Carboxylic acid derivatives 19–22 were
prepared by hydrolysis of the ester precursors (10–13) under the 1 N
NaOH condition and then followed by condensation with a variety of
amines to give the corresponding amido analogs (23–32).
On the other hand, compound 33 was prepared from the
methanesulfonate (2) with KSAc under the heat (80 °C) condition.
Both TMS groups and an acetyl group were successively removed
under the 1 N HCl condition and then the sodium methoxide
condition to give a key intermediate, compound 35. Pd-catalyzed
cross-coupling reactions35 of 35 with an aryl bromide or an aryl
iodide, which are more powerful than Mitsunobu reactions of 1 or
SN2 reactions of 2 in our research, were used to synthesize further
novel derivatives. Consequently, we synthesized a variety of LCM
analogs 36–45.
Table 1 Antibacterial activities of the representative macrolides,
lincomycin (LCM) and clindamycin (CLDM)
Test organisma
Characteristicsb
CAM
AZM
LCM
CLDM
Streptococcus pneumo- Susceptible
niae DP1 type I
0.03
0.06
1
0.06
S. pneumoniae-2
S. pneumoniae-3
S. pneumoniae-4
S. pneumoniae-5
S. pneumoniae-6
Susceptible
Susceptible
0.03
0.03
1
0.12
0.06
0.015 0.03
0.25
ermAM methylase (c) 4128 4128 4128 4128
ermAM methylase (c) 4128 4128 4128 4128
ermAM methylase (c) 4128 4128 4128 4128
+ mefE
S. pneumoniae-7
S. pneumoniae-8
S. pneumoniae-9
Streptococcus
pyogenes Cook
S. pyogenes-2
S. pyogenes-3
Haemophilus
ermAM methylase (i) 4128 4128
ermAM methylase (i) 4128 4128
128
128
1
128
128
mefE efflux
0.5
0.5
0.12
0.06
Susceptible
0.015 0.06
0.12
ermAM methylase (c) 4128 4128 4128
128
0.12
16
mefE efflux
8
2
8
0.25
8
Susceptible
0.25
influenzae-1
H. influenzae-2
H. influenzae-3
H. influenzae-4
Susceptible
Susceptible
⊿acr
4
8
1
2
16
16
4
8
16
1
0.5
0.5
Abbreviations: AZM, azithromycin; CAM, clarithromycin.
MIC (μg ml− 1).
aAll strains except standard organisms were clinically isolated.
b(c): constitutive; (i): inducible.
7-deoxylincomycin and 7(S)-7-deoxy-7-(5-phenyl-1,3,4-thiadiazol-2-
yl-thio)lincomycin had potent activities against resistant bacteria of
S. pneumoniae and S. pyogenes with erm gene when compared with
LCM or CLDM.
In this article, we report synthesis of novel LCM derivatives
and their potent antibacterial activities against resistant bacteria of
S. pneumoniae and S. pyogenes with erm gene. We also discuss SARs of
these molecules focusing on substituted phenylthio groups at the 7-
position of LCM with 7(S)-configuration.
SAR analysis of LCM derivatives possessing a substituted phenyl
moiety at the C-7 position via sulfur atom with 7(S)-configuration
Antibacterial activities of LCM derivatives possessing a substituted
phenyl moiety at the C-7 position via sulfur atom are shown in
Table 2. In our medicinal chemistry research, we have already
had SAR information suggestion that a carbonyl group or an ester
function was important to enhance antibacterial activities in the case
of 7-thio-phenyl derivatives such as compound 46 (Figure 2). Thus,
we first replaced the thiadiazolyl group in our selected compound 47
with a phenyl group. As a result, 9 generally showed stronger
antibacterial activities than 47.
RESULTS AND DISCUSSION
Synthesis of 7(S)-7-deoxy-7-(substituted-arylthio)lincomycin
derivatives
Compound 36, which possessed
a propyl group at the
Synthesis of 7(S)-7-deoxy-7-(substituted-arylthio)lincomycin deriva- methoxymethylcarbonylamino group of 9, had the strongest activities
tives is shown in Scheme 1. Because we had utilized compounds 1 and against resistant Streptococcus species with erm gene, but exhibited
2 as key intermediates in our drug discovery program, we also applied comparable activities to 47 against H. influenzae. Next, we were
The Journal of Antibiotics

Synthesis and structure–activity relationships of novel LCM derivatives
Y Wakiyama et al
3
interested in changing the direction of an amide bond, and we CONCLUSION
At the beginning of our LCM analogs research program, we were
interested in LCM derivatives possessing a hetero ring at the
C-7 position via sulfur atom with 7(S)-configuration. We synthesized
them by two reactions; (1) Mitsunobu reaction of 2,3,4-tris-O-
(trimethylsilyl)lincomycin (1) with the corresponding thiol and (2)
SN2 reaction of 7-O-methanesulfonyl-2,3,4-tris-O-(trimethylsilyl)lin-
comycin (2) with the corresponding thiol. These synthetic procedures,
however, had limitation in preparation of various LCM-7-thio-aryl
analogs in order to investigate their SAR. So, we have developed a
novel synthetic route for a variety of 7-thio-modified LCM derivatives
by the application of Pd-catalyzed cross-coupling reaction35 of
7-deoxy-7-epi-7-mercaptolincomycin (35) with an aryl bromide or
an aryl iodide. This methodology was very useful to synthesize a
various 7-thio-modified LCM analogs.
We first synthesized and biologically evaluated 7(S)-7-deoxy-7-
thiophenyl analogs possessing either the NHCO-type or the CONH-
type bond at the C-7 substituent. As a result, compound 28 possessing
the morpholine ring had potent antibacterial activities against major
pathogens that caused respiratory infections, even when compared
with CLDM. A substitution introduced on the benzene ring of 28,
however, did not enhance antibacterial activities. Furthermore,
conversion of the phenyl group of 28 to other hetero rings also
decreased antibacterial activities. Finally, compounds 37 and 38
showed the strongest antibacterial activities against S. pneumoniae
and S. pyogenes with erm gene, but, antibacterial activities against
H. influenzae of these analogs were not improved compared with those
of CLDM.
constructed a CONH-type bond instead of a NHCO-type to provide
15. This compound had similar activities as 9. To accumulate the
SAR information around the C-7 position, we substituted the
methyl group in 15, respectively, with a larger group such as a
cyclopropyl, cyclohexyl, adamantyl and pyridin-3-yl group, and
compounds 23–26 were prepared. Modification of the R moiety on
7(S)-7-sulfur-Ph-CONHR derivatives could not improve the
antibacterial activities of 15.
SAR analysis of LCM derivatives possessing a 4-(N, N-disubstituted-
carbamoyl)phenyl group at the C-7 position via sulfur atom with 7
(S)-configuration
Conversions of the methylamino group of 15 to other dialkylamino
groups were accomplished and antibacterial activities of the resulting
compounds are shown in Table 3. Compound 16 also showed almost
the same antibacterial spectrum as that of 15. A variety of substituted
amino functional groups (pyrrolidinyl, piperidinyl, morpholinyl,
1,4-oxazepanyl, 1-methylpiperazinyl group) were constructed to
improve antibacterial activities. Consequently, the morpholinyl
derivative (28) had strong activities against major pathogens which
caused respiratory infections, that is, S. pneumoniae, S. pyogenes and
H. influenzae. Compounds 37 and 38 possessing a substituent in the
pyrrolidine ring were prepared. Although compounds 37 and 38
exhibited potent activities against S. pneumoniae and S. pyogenes with
erm gene and/or mef gene, they showed weaker activities against
H. influenzae than 28. On the other hand, the tertiary amino analog
(40) generally showed comparable antibacterial activities to 28.
To investigate the possibilities of novel semi-synthetic LCM
antibiotics, alternative modifications of LCM analogs possessing the
7-thiophenyl group or 7-thiothiadiazolyl group are now in progress.
On the basis of information in this article, we will continuously
explore novel chemical modifications focusing on clinically promising
LCM derivatives that exhibit potent antibacterial activities against
resistant S. pneumoniae, S. pyogenes with erm gene and H. influenzae.
SAR analysis of LCM derivatives possessing a morpholinyl-
carbonylphenyl group at the C-7 position via sulfur atom with
7(S)-configuration
Next, we consequently introduced several kinds of substituents on the
phenyl group of 28, and the antibacterial activities of compounds 41 to
44 are shown in Table 4. Introducing a substituent on the phenyl
group of 28 did not improve its antibacterial activities, even though it
was an electron-withdrawing group or an electron-donating group.
EXPERIMENTAL PROCEDURE
General methods
1H NMR spectra were measured with a BRUKER Ascend 400 NMR spectro-
meter (BRUKER, Coventry, UK) for 400 MHz, JEOL JNM-GSX 400 NMR
spectrometer for 400 MHz or a Varian Gemini 300 NMR spectrometer for
300 MHz in CDCl3 or CD3OD. TMS (0 p.p.m.) in CDCl3 or CD3OD was used
as internal reference standard. Mass spectra (MS) were obtained on a JEOL
JMS-700 mass spectrometer or Agilent Technologies 6530-Q-TOF LC/MS mass
spectrometer (Agilent Technologies, Santa Clara, CA, USA). The optical
rotations were recorded with Jasco P-2300 digital polarimeter (Jasco, Tokyo,
Japan). Column chromatography was performed with silica gel (Wakogel
C200) or Diaion HP-20 (Mitsubishi Chemical, Tokyo, Japan). Preparative thin-
layer chromatography (preparative TLC) was performed with silica gel (Merck:
TLC plates Silica gel 60 F254). All organic extracts were dried over anhydrous
MgSO4, and the solvent was removed with a rotary evaporator under reduced
pressure.
SAR analysis of LCM derivatives possessing a morpholin-1-yl-
carbonylaryl moiety at the C-7 position via sulfur atom with
7(S)-configuration
Antibacterial activities of LCM derivatives possessing a morpholin-1-
yl-carbonylaryl moiety are shown in Table 5. Conversion of the
benzene ring to other hetero rings did not enhance antibacterial
activities of 28.
Docking simulation of the key compound 28
Finally, we investigated three-dimensional analysis4,6–7 of 28 and the
peptidyl transferase, and the result is shown in Figure 3. (Docking
simulation was calculated by data on bacteria, Haloarcula marismortui
(Hm).) The analysis indicated that an oxygen atom of a carbonyl
7-O-Methanesulfonyl-2,3,4-tris-O-(trimethylsilyl)lincomycin (2). To a solution
group in the C-7 side chain of 28 has a hydrogen bonding with of 2,3,4-tris-O-(trimethylsilyl)lincomycin (1) (4.0 g, 6.42 mmol) in CHCl3
(20 ml) were added Et3N (2.24 ml, 16.1 mmol), methanesulfonyl chloride
(0.99 ml, 12.8 mmol) and stirred at room temperature for 3 h. The mixture
was added to CHCl3 (60 ml), washed with saturated aqueous (aq) NaHCO3
(50 ml), dried over MgSO4 and concentrated under reduced pressure. The title
U2620Hm (Docking simulation was calculated by data on bacteria,
Haloarcula marismortui (Hm).) (U2585Ec) (The numbers in
parenthesis are expressed as the case of Escherichia coli (Ec).) on 23S
rRNA (ribosomal RNA). Furthermore, an ethylene part of the
compound was obtained as a colorless solid (4.50 g, quantitative). ESI-MS
morpholine ring in 28 was analyzed and determined to have a
hydrophobic interaction of CH-p stacking with uracil (cytosine) ring
of U2621Hm (C2586Ec) on 23S rRNA.
(m/z) 701 (M+H)+ as C28H60N2O8S2Si3; 1H NMR (400 MHz, CDCl3) δ 0.13
(s, 9 H), 0.14 (s, 9 H), 0.17 (s, 9 H), 0.89 (br t, J = 6.9 Hz, 3 H), 1.21–1.36
(m, 4 H), 1.40 (d, J = 6.6 Hz, 3 H), 1.79–1.89 (m, 1 H), 1.92–2.09 (m, 3 H),
The Journal of Antibiotics

Synthesis and structure–activity relationships of novel LCM derivatives
Y Wakiyama et al
4
2.11 (s, 3 H), 2.40 (s, 3 H), 2.99 (dd, J = 10.7, 3.7 Hz, 1 H), 3.09 (s, 3 H), (m/z) 586 (M+H)+ as C27H43N3O7S2; TOF-ESI-HR-MS (M+H)+ calcd
3.14–3.21 (m, 1 H), 3.52 (dd, J = 9.5, 2.4 Hz, 1 H), 3.75 (br d, J = 2.4 Hz, 1 H),
3.90 (d, J =9.7 Hz, 1 H), 4.15 (dd, J = 9.5, 5.6 Hz, 1 H), 4.70–4.78 (m, 1 H),
5.09–5.15 (m, 1 H), 5.16 (d, J = 5.6 Hz, 1 H), 7.61 (d, J = 10.7 Hz, 1 H).
for C27H43N3O7S2: 586.2621, found: 586.2621; 1H NMR (400 MHz, CD3OD)
δ 0.87–0.97 (m, 3 H), 1.27 (d, J =7.0 Hz, 3 H), 1.29–1.40 (m, 4 H), 1.79–1.89
(m, 1 H), 1.92–2.02 (m, 1 H), 2.02–2.09 (m, 1 H), 2.06 (s, 3 H), 2.10–2.21
(m, 1 H), 2.36 (s, 3 H), 2.97 (dd, J = 10.6, 4.5 Hz, 1 H), 3.22 (dd, J = 7.9,
6.0 Hz, 1 H), 3.48 (s, 3 H), 3.59 (dd, J = 10.3, 3.5 Hz, 1 H), 3.70–3.82 (m, 2 H),
4.03 (s, 2 H), 4.11 (dd, J = 10.3, 5.5 Hz, 1 H), 4.30–4.43 (m, 2 H), 5.28
(d, J = 5.5 Hz, 1 H), 7.39–7.46 (m, 2 H), 7.58–7.66 (m, 2 H).
7(S)-7-(4-Aminophenylthio)-7-deoxy-2,3,4-tris-O-(trimethylsilyl)lincomycin (3).
To a solution of compound 2 (5.62 g, 8.02 mmol) in DMF (50 ml) were added
K2CO3 (3.33 g, 24.1 mmol), 4-aminobenzenethiol (2.01 g, 16.1 mmol) and
stirred at 100 °C for 4.5 h. The mixture was diluted with 1 N HCl
(100 ml)-MeOH (50 ml), reacted at room temperature for 45 min and then 7(S)-7-Deoxy-7-(4-methoxycarbonylphenylthio)lincomycin (10). To a solution
concentrated under reduced pressure. The resulting residue was dissolved by of compound 2 (5.63 g, 8.0 mmol) in DMF (20 ml) were added K2CO3 (3.33 g,
water, washed with Et2O. The mixture was added to the saturated aq NaHCO3,
24.1 mmol), methyl 4-mercaptobenzoate (2.70 g, 16.1 mmol), stirred at 80 °C
then extracted with EtOAc, washed with water, dried over MgSO4 and for 1 h and concentrated under reduced pressure. The resulting residue
concentrated under reduced pressure. The resulting residue was purified by (compound 4) in MeOH (20 ml) was added to 1 N HCl (80 ml), stirred at
silica gel column chromatography (CHCl3/MeOH/28% aq NH4OH= 20/1/0.1) room temperature for 1 h and concentrated under reduced pressure. The
to obtain the 7(S)-7-(4-aminophenylthio)-7-deoxylincomycin as a colorless solid resulting residue was dissolved by water, washed with Et2O. The mixture was
(2.77 g, 67%). [α]D +142.0° (c 0.51, MeOH); ESI-MS (m/z) 514 (M+H)+ as
C24H39N3O5S2; TOF-ESI-HR-MS (M+H)+ calcd for C24H39N3O5S2: 514.2409,
found: 514.2411; 1H NMR (400 MHz, CD3OD) δ 0.89–0.98 (m, 3 H), 1.20 (d,
J = 7.1 Hz, 3 H), 1.30–1.41 (m, 4 H), 1.80–1.90 (m, 1 H), 1.92–2.00 (m, 1 H),
2.04–2.21 (m, 2 H), 2.17 (s, 3 H), 2.34 (s, 3 H), 2.98 (dd, J =10.6, 4.6 Hz, 1 H),
3.24 (dd, J = 8.2, 5.6 Hz, 1 H), 3.53 (dq, J = 7.1, 2.8 Hz, 1 H), 3.60 (dd, J = 10.3,
3.3 Hz, 1 H), 3.68–3.72 (m, 1 H), 4.10 (dd, J = 10.3, 5.6 Hz, 1 H), 4.25 (dd,
J = 9.9, 2.8 Hz, 1 H), 4.38 (br d, J = 9.9 Hz, 1 H), 5.27 (d, J = 5.6 Hz, 1 H), 6.62–
6.68 (m, 2 H), 7.20–7.26 (m, 2 H).
added to the saturated aq NaHCO3, then extracted with EtOAc, washed with
water, dried over MgSO4 and concentrated under reduced pressure.
The resulting residue was purified by silica gel column chromatography
(CHCl3/MeOH/28% aq NH4OH= 20/1/0.1) to obtain the title compound as
28
24
a colorless solid (3.19 g, 71%). [α]D +84.6° (c 0.97, MeOH); ESI-MS (m/z)
557 (M+H)+ as C26H40N2O7S2; TOF-ESI-HR-MS (M+H)+ calcd for
C26H40N2O7S2: 557.2355, found: 557.2359; 1H NMR (400 MHz, CD3OD)
δ 0.88–0.96 (m, 3 H), 1.29–1.37 (m, 4 H), 1.40 (d, J = 6.8 Hz, 3 H), 1.79–1.91
(m, 1 H), 1.84 (s, 3 H), 1.96–2.05 (m, 1 H), 2.05–2.12 (m, 1 H), 2.12–2.25
(m, 1 H), 2.40 (s, 3 H), 3.00 (dd, J = 10.6, 4.8 Hz, 1 H), 3.24 (dd, J = 8.2,
5.7 Hz, 1 H), 3.58 (dd, J = 10.2, 3.2 Hz, 1 H), 3.78 (br dd, J = 3.2, 0.7 Hz,1 H),
3.89 (s, 3 H), 4.03 (dq, J = 6.8, 2.8 Hz,1 H), 4.10 (dd, J = 10.2, 5.6 Hz, 1 H),
4.37 (br dd, J = 9.7, 0.7 Hz, 1 H), 4.52 (dd, J = 9.7, 2.8 Hz, 1 H), 5.24
(d, J = 5.6 Hz, 1 H), 7.42–7.49 (m, 2 H), 7.90–7.97 (m, 2 H).
To a solution of 7(S)-7-(4-aminophenylthio)-7-deoxylincomycin (2.0 g, 3.9
mmol) in pyridine (20 ml) were added trimethylchlorosilane (2.0 ml, 15.7
mmol), hexamethyldisilazane (2.1 ml, 16.0 mmol) and stirred at room
temperature for 20 h, then it was concentrated under reduced pressure. The
residue was diluted with water, then extracted with EtOAc, washed with water
and concentrated under reduced pressure. The resulting residue was purified by
silica gel column chromatography (hexane/EtOAc= 2/1) to obtain the title
compound as a colorless solid (2.77 mg, 97%). [α]D28 +106.6° (c 1.15, CHCl3);
ESI-MS (m/z) 730 (M+H)+ as C33H63N3O5S2Si3; TOF-ESI-HR-MS (M+H)+
calcd for C33H63N3O5S2Si3: 730.3595, found: 730.3583; 1H NMR (400 MHz,
CDCl3) δ 0.13 (s, 9 H), 0.14 (s, 9 H), 0.19 (m, 9 H), 0.84–0.95 (m, 3 H),
1.12 (d, J = 6.8 Hz, 3 H), 1.20–1.39 (m, 4 H), 1.78–1.89 (m, 1 H), 1.91–2.11
(m, 3 H), 2.21 (s, 3 H), 2.44 (s, 3 H), 2.98 (dd, J =10.8, 4.0 Hz, 1 H), 3.15–3.25
(m, 1 H), 3.59–3.79 (m, 5 H), 4.12–4.22 (m, 2 H), 4.55–4.65 (m, 1 H),
5.28 (d, J = 5.6 Hz, 1 H), 6.55–6.63 (m, 2 H), 7.12–7.22 (m, 2 H), 7.63
(d, J = 10.6 Hz, 1 H).
7(S)-7-Deoxy-7-(5-methoxycarbonylpyridin-2-ylthio)lincomycin (11). To
a
solution of compound 1 (200 mg, 0.32 mmol) in THF (3 ml) at 0 °C were
added triphenylphosphine (84.2 mg, 0.32 mmol), diisopropylazodicarboxylate
(0.065 ml, 0.32 mmol), methyl 6-mercaptonicotinate (36.2 mg, 0.21 mmol),
stirred at room temperature for 7 h and concentrated under reduced pressure.
The resulting residue (compound 5) in MeOH (3 ml) was added to 1 N HCl
(3 ml), stirred at room temperature for 40 min and concentrated under reduced
pressure. The resulting residue was dissolved by water, washed with Et2O. The
mixture was added to NaHCO3 (150 mg), then extracted with EtOAc, washed
with water, dried over MgSO4 and concentrated under reduced pressure.
The resulting residue was purified by preparative TLC (CHCl3/MeOH/28% aq
NH4OH= 9/2/0.2) to obtain the title compound as a colorless solid (91 mg,
7(S)-7-Deoxy-7-(4-methoxyacetamidophenylthio)lincomycin (9). To a solution
of compound 3 (100 mg, 0.14 mmol) in THF (1 ml) were added Et3N
(0.058 ml, 0.42 mmol), 2-methoxyacetyl chloride (0.019 ml, 0.21 mmol) and
stirred at room temperature for 3 h. The mixture was diluted with 1 N HCl
(2.6 ml)-MeOH (1.3 ml), reacted at room temperature for 40 min, and then
concentrated under reduced pressure. The resulting residue was dissolved by
water, washed with Et2O. The mixture was added to NaHCO3 (70 mg), then
extracted with EtOAc, washed with water, dried over MgSO4 and concentrated
under reduced pressure. The resulting residue was purified by preparative
TLC (CHCl3/MeOH/28% aq NH4OH= 9/2/0.2) to obtain the title compound
76%). [α]D +71.2° (c 0.25, MeOH); ESI-MS (m/z) 558 (M+H)+ as
28
C25H39N3O7S2; TOF-ESI-HR-MS (M+H)+ calcd for C25H39N3O7S2:
558.2308, found: 558.2301; 1H NMR (400 MHz, CD3OD)
δ 0.86–0.99
(m, 3 H), 1.29–1.40 (m, 4 H), 1.48 (d, J = 6.8 Hz, 3 H), 1.79 (s, 3 H),
1.80–1.90 (m, 1 H), 1.97–2.11 (m, 2 H), 2.12–2.26 (m, 1 H), 2.36 (s, 3 H), 2.99
(dd, J = 10.5, 5.0 Hz, 1 H), 3.25 (dd, J = 8.4, 6.0 Hz, 1 H), 3.55 (dd, J = 10.3,
3.2 Hz, 1 H), 3.77 (br dd, J = 3.2, 0.6 Hz, 1 H), 3.91 (s, 3 H), 4.09 (dd, J = 10.3,
5.6 Hz, 1 H), 4.33 (br dd, J = 9.7, 0.6 Hz, 1 H), 4.45 (dq, J = 6.8, 3.2 Hz,1 H),
4.52 (dd, J = 9.7, 3.2 Hz, 1 H), 5.22 (d, J = 5.6 Hz, 1 H), 7.39 (dd, J = 8.4,
0.8 Hz, 1 H), 8.10 (dd, J = 8.4, 2.2 Hz, 1 H), 8.96 (dd, J = 2.2, 0.8 Hz, 1 H).
28
as a colorless solid (80 mg, 99%). [α]D +109.3° (c 2.16, MeOH); ESI-MS
Scheme 1 Synthesis of 7(S)-7-arylthio-7-deoxylincomycin derivatives. Conditions and Results: (a) methanesulfonyl chloride, Et3N, CHCl3, r.t., 3 h,
quant; (b) (3): (1) 4-aminobenzenethiol, K2CO3, DMF, 100 °C, 4.5 h, (2) 1 N HCl, MeOH, r.t., 45 min, 67%, (3) trimethylchlorosilane, hexamethyldisilazane,
pyridine, r.t., 20 h, 97% (3). Because a part of TMS groups was removed during SN2 reaction with aminobenzenethiol, total deprotection and total re-
protection by TMS groups were performed. (4): methyl 4-mercaptobenzoate, K2CO3, DMF, 80 °C, 1 h, not isolated (4); (c) 2-methoxyacetyl chloride, Et3N,
THF, 3 h; (d) 1 N HCl, MeOH, r.t., 40 min, two steps 99%; (e) triphenylphosphine, diethylazodicarboxylate or diisopropylazodicarboxylate, the corresponding
HS-Ar, THF or toluene, 0 °C to r.t., 7–16 h, not isolated (5–8); (f) 1 N HCl, MeOH, r.t., 1 h, three steps 71% from 2 to 10, two steps 76% from 1 to 11, two
steps 74% from 1 to 12, two steps 66% from 1 to 13, two steps 45% from 1 to 14; (g) the corresponding amine, MeOH or EtOH, reflux, 18% (15), 16%
(16), 33% (17), 64% (18); (h) 1 N or 5 N NaOH, MeOH or EtOH, r.t., quant (19), 81% (20), not isolated (21), not isolated (22); (i) the corresponding
amine, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide·HCl or N,N’-dicyclohexylcarbodiimide, 1-hydroxybenzotriazole, DMF, r.t. to 60 °C, 1–48 h, Et3N
(21 and 22 only), 45% (23), 25% (24), 55% (25), 29% (26), 63% (27), 63% (28), 69% (29), 70% (30), two steps 6% from 12 to 31, two steps 7%
from 13 to 32; (j) KSAc, DMF, 60 °C, 4 h, two steps 88% from 2 to 33; (k) 2 N HCl, MeOH, r.t., 10 min, 97%; (l) sodium methoxide, MeOH, r.t., 20 min,
94%; (m) Ar–Br or Ar–I, Xantphos, Pd2(dba)3, N,N-diisopropylethylamine, 1,4-dioxane, reflux, 2–21 h, 76% (36), 79% (37), 82% (38), 73% (39), 75%
(40), 82% (41), 81% (42), 80% (43), 78% (44), 76% (45). quant, quantitative; r.t., room temperature; TMS, trimethylsilyl.
The Journal of Antibiotics

Synthesis and structure–activity relationships of novel LCM derivatives
Y Wakiyama et al
5
Me
Me
Me
Me
O
Me
O
Me
O
HN
HO
Me
O
Me
OH
O
SAr
O
SAr
O
OMs
O
N
N
N
N
HN
HN
HN
Me
Me
Me
Me
a
b
f
TMSO
SMe
TMSO
SMe
SMe
TMSO
SMe
TMSO
OTMS
TMSO
OTMS
HO
OH
TMSO
OTMS
e
1
5 - 8
2
3 - 8
10 - 14
Ar
O
O
4
10
5
11
3
NH2
OMe
N
OMe
N
N
O
N
S
N
N
7
13
6
12
8
14
OEt
OEt
S
OEt
NH2
O
O
OMe
HN
O
CO2H
Me
A
Me
Me
O
Me
O
HN
HO
Me
O
HN
HO
Me
S
SAr
S
N
HN
HO
N
N
Me
Me
Me
O
c, d
O
O
g
h
10 - 14
SMe
3
10 or 14
SMe
SMe
HO
OH
HO
OH
HO
OH
19 - 22
15 - 18
17
9
Ar
A
O
N
O
N
21
15
19
NHMe
N
N
NH2
O
N
S
N
O
22
16
18
20
N
S
NMe2
N
O
O
R
R
H
H
O
A
N
23
24
N
N
Me
Me
A
O
Me
O Me
N
30
H
N
H
N
S
S
25
27
29
26
28
N
N
N
N
HN
HO
HN
HO
Me
Me
O
O
i
31
32
19 - 22
N
N
N
O
SMe
SMe
N
N
NH2
HO
23 - 29
OH
HO
30 - 32
OH
NMe
Me
S
O
N
O
N
Me
Me
Me
Ar
O
Me
O
SAc
O
SAc
O
36
N
37
OMe
38
N
N
HN
HN
HO
j
k
Me
TMSO
Me
2
O
NMe2
OMe
NO2
SMe
SMe
F
O
N
O
TMSO
OTMS
HO
OH
N
39
40
43
41
33
34
N
O
O
O
F
Me
O
Me
Me
m
O
N
O
O
O
HN
HO
Me
O
HN
HO
Me
SH
SAr
44
42
45
N
N
N
N
l
Me
Me
O
O
O
O
SMe
SMe
O
S
HO
OH
HO
36 - 45
OH
N
35
O
The Journal of Antibiotics

Synthesis and structure–activity relationships of novel LCM derivatives
Y Wakiyama et al
6
Table 2 Antibacterial activities of 7(S)-7-deoxy-7-(4-substituted-phenylthio)-LCM derivatives
MIC (μg ml− 1).
aAll strains except standard organisms were clinically isolated.
b(c): constitutive; (i): inducible.
Grey shading strains are target strains.
O
O
N
OMe
N
O
H
N
N
N
Me
Me
Me
OMe
S
O Me
O Me
O Me
O
S
S
S
N
N
N
HN
HN
HN
Me
Me
Me
O
O
O
HO
SMe
HO
SMe
HO
SMe
HO
OH
HO
OH
HO
OH
46
47
48
Figure 2 Basic molecule 46 and alternative series of our novel LCM derivatives 47 and 48. LCM, lincomycin.
7(S)-7-(4-Amino-5-ethoxycarbonylpyrimidin-2-ylthio)-7-deoxylincomycin (12).
Compound 1 (1.87 g, 3.0 mmol), triphenylphosphine (1.18 g, 6.86 mmol),
diethylazodicarboxylate (0.71 ml, 390 mmol), ethyl 4-amino-2-mercaptopyri-
midine-5-carboxylate (894 mg, 4.49 mmol) and toluene (24 ml) were treated
according to the similar procedure as described for the preparation of 11 to
(m, 1 H), 2.36 (s, 3 H), 2.98 (dd, J = 10.4, 5.1 Hz, 1 H), 3.24 (dd, J = 8.5,
6.1 Hz, 1 H), 3.55 (dd, J = 10.3, 3.3 Hz, 1 H), 3.77–3.81 (m, 1 H), 4.10
(dd, J = 10.3, 5.6 Hz, 1 H), 4.29–4.40 (m, 4 H), 4.58 (dd, J =9.8, 3.2 Hz, 1 H),
5.24 (d, J = 5.6 Hz, 1 H), 8.21 (s, 1 H).
29
afford 12 (1.3 g, 74%) as a colorless solid. [α]D +43.3° (c 6.21, MeOH);
7(S)-7-Deoxy-7-(5-ethoxycarbonyl-1,3,4-thiadiazol-2-ylthio)lincomycin (14).
Compound 1 (950 mg, 1.52 mmol), triphenylphosphine (550 mg, 2.10 mmol),
diethylazodicarboxylate (0.5 ml, 2.74 mmol), ethyl 5-mercapto-1,3,4-thiadia-
zole-2-carboxylate (141 mg, 0.75 mmol) and toluene (20 ml) were treated
according to the similar procedure as described for the preparation of 11 to
ESI-MS (m/z) 588 (M+H)+ as C25H41N5O7S2; TOF-ESI-HR-MS (M+H)+ calcd
for C25H41N5O7S2: 588.2526, found: 588.2519; 1H NMR (400 MHz, CD3OD)
δ 0.87–0.96 (m, 3 H), 1.26–1.38 (m, 4 H), 1.36 (t, J = 7.1 Hz, 3 H), 1.49
(d, J = 6.8 Hz, 3 H), 1.77–1.87 (m, 1 H), 1.87 (s, 3 H), 1.95–2.10 (m, 2 H),
2.10–2.26 (m, 1 H), 2.36 (s, 3 H), 2.98 (dd, J =10.5, 5.1 Hz, 1 H), 3.22
(dd, J =8.5, 6.1 Hz, 1 H), 3.57 (dd, J = 10.2, 3.4 Hz, 1 H), 3.77–3.82 (m, 1 H),
4.11 (dd, J = 10.2, 5.6 Hz, 1 H), 4.29–4.40 (m, 4 H), 4.51 (dd, J = 9.7,
3.2 Hz, 1 H), 5.24 (d, J = 5.6 Hz, 1 H), 8.58 (s, 1 H).
29
afford 14 (345.3 mg, 45%) as a colorless solid. [α]D +90.7° (c 0.63, EtOH);
ESI-MS (m/z) 579 (M+H)+ as C23H38N4O7S3; TOF-ESI-HR-MS (M+H)+ calcd
for C23H38N4O7S3: 579.1981, found: 579.1976; 1H NMR (400 MHz, CD3OD)
δ 0.87–0.97 (m, 3 H), 1.28–1.38 (m, 4 H), 1.41 (t, J = 7.1 Hz, 3 H), 1.57
(d, J = 7.0 Hz, 3 H), 1.78–1.90 (m, 1 H), 1.94 (s, 3 H), 1.97–2.12 (m, 2 H),
2.13–2.28 (m, 1 H), 2.38 (s, 3 H), 3.01 (dd, J = 10.4, 5.1 Hz, 1 H), 3.26
(dd, J = 8.6, 6.1 Hz, 1 H), 3.55 (dd, J = 10.2, 3.2 Hz, 1 H), 3.81 (br dd, J = 3.2,
0.8 Hz, 1 H), 4.10 (dd, J = 10.2, 5.6 Hz, 1 H), 4.41 (br dd, J = 9.8, 0.8 Hz, 1 H),
4.47 (q, J = 7.1 Hz, 2 H), 4.53 (dq, J = 7.0, 3.2 Hz, 1 H), 4.62 (dd, J = 9.8,
3.2 Hz, 1 H), 5.25 (d, J = 5.6 Hz, 1 H).
7(S)-7-Deoxy-7-(5-ethoxycarbonylthiazol-2-ylthio)lincomycin (13). Compound
1 (930 mg, 1.49 mmol), triphenylphosphine (600 mg, 2.29 mmol), diethylazo-
dicarboxylate (0.4 ml, 2.20 mmol), ethyl 2-mercaptothiazole-5-carboxylate
(350 mg, 1.85 mmol) and toluene (15 ml) were treated according to the similar
procedure as described for the preparation of 11 to afford 13 (569.2 mg, 66%)
as a colorless solid. [α]D28 +85.7° (c 0.32, MeOH); ESI-MS (m/z) 578 (M+H)+
as C24H39N3O7S3; TOF-ESI-HR-MS (M+H)+ calcd for C24H39N3O7S3:
578.2028, found: 578.2023; 1H NMR (400 MHz, CD3OD)
δ 0.88–0.96
7(S)-7-Deoxy-7-(4-methylcarbamoylphenylthio)lincomycin (15). To a solution
of compound 10 (100 mg, 0.18 mmol) in 30% methylamine methanol solution
(m, 3 H), 1.29–1.39 (m, 4 H), 1.35 (t, J =7.1 Hz, 3 H), 1.52 (d, J = 6.8 Hz,
3 H), 1.78–1.89 (m, 1 H), 1.94 (s, 3 H), 1.96–2.10 (m, 2 H), 2.13–2.27 (1.2 ml) was refluxed 20 h and concentrated under reduced pressure. The
The Journal of Antibiotics

Synthesis and structure–activity relationships of novel LCM derivatives
Y Wakiyama et al
7
Table 3 Antibacterial activities of 7(S)-7-deoxy-7-(4-(N,N-disubstituted-carbamoyl)phenylthio)-LCM derivatives
Abbreviations: LCM, lincomycin; ND, not determined.
MIC (μg ml− 1).
aAll strains except standard organisms were clinically isolated.
b(c): constitutive; (i): inducible.
Grey shading strains are target strains.
Table 4 Antibacterial activities by substituent effects on the phenyl group of 28
MIC (μg ml− 1).
aAll strains except standard organisms were clinically isolated.
b(c): constitutive; (i): inducible.
Grey shading strains are target strains.
resulting residue was purified by preparative TLC (CHCl3/MeOH/28% aq 1.87 (s, 3 H), 1.97–2.06 (m, 1 H), 2.08–2.25 (m, 2 H), 2.43 (s, 3 H), 2.90
NH4OH= 9/2/0.2) to obtain the title compound as a colorless solid (18.0 mg, (s, 3 H), 3.06 (dd, J = 10.6, 4.8 Hz, 1 H), 3.27 (dd, J = 7.9, 5.4 Hz, 1 H), 3.58
18%). [α]D +80.5° (c 0.65, MeOH); ESI-MS (m/z) 556 (M+H)+ as (dd, J = 10.2, 3.2 Hz, 1 H), 3.75–3.80 (m, 1 H), 3.98 (dq, J = 6.8, 2.8 Hz, 1 H),
33
C26H41N3O6S2; TOF-ESI-HR-MS (M+H)+ calcd for C26H41N3O6S2: 4.10 (dd, J = 10.2, 5.5 Hz, 1 H), 4.37 (br dd, J = 9.7, 0.6 Hz, 1 H), 4.51
556.2515, found: 556.2515; 1H NMR (400 MHz, CD3OD)
δ
0.88–0.97 (dd, J = 9.7, 2.8 Hz, 1 H), 5.25 (d, J = 5.5 Hz, 1 H), 7.42–7.48 (m, 2 H),
(m, 3 H), 1.30–1.41 (m, 4 H), 1.37 (d, J = 6.8 Hz, 3 H), 1.80–1.95 (m, 1 H), 7.72–7.78 (m, 2 H).
The Journal of Antibiotics

Synthesis and structure–activity relationships of novel LCM derivatives
Y Wakiyama et al
8
Table 5 Antibacterial activities of 7(S)-7-deoxy-7-((morpholin-1-yl-carbonyl)arylthio)-LCM derivatives
Abbreviations: LCM, lincomycin; ND, not determined.
MIC (μg ml− 1).
aAll strains except standard organisms were clinically isolated.
b(c): constitutive; (i): inducible.
Grey shading strains are target strains.
Figure 3 Three-dimensional analysis of 28 and the peptidyl transferase.
7(S)-7-Deoxy-7-(4-dimethylcarbamoylphenylthio)lincomycin (16). Compound (m, 1 H), 2.08–2.26 (m, 2 H), 2.44 (s, 3 H), 3.00 (s, 3 H), 3.05 (dd, J =10.6,
10 (300 mg, 0.54 mmol) and 2 M dimethylamine methanol solution (20 ml)
4.8 Hz, 1 H), 3.08 (s, 3 H), 3.28 (dd, J = 8.1, 5.5 Hz, 1 H), 3.58 (dd, J = 10.2,
3.2 Hz, 1 H), 3.75–3.80 (m, 1 H), 3.97 (dq, J = 6.9, 2.7 Hz, 1 H), 4.10 (dd,
J = 10.2, 5.5 Hz, 1 H), 4.35 (br dd, J = 9.7, 0.5 Hz, 1 H), 4.49 (dd, J = 9.7,
2.7 Hz, 1 H), 5.26 (d, J =5.5 Hz, 1 H), 7.36–7.42 (m, 2 H), 7.44–7.50 (m, 2 H).
were treated according to the similar procedure as described for the preparation
31
of 15 to afford 16 (48.6 mg, 16%) as a colorless solid. [α]D +90.9°
(c 1.12, MeOH); ESI-MS (m/z) 570 (M+H)+ as C27H43N3O6S2;
TOF-ESI-HR-MS (M+H)+ calcd for C27H43N3O6S2: 570.2672, found:
570.2681; 1H NMR (400 MHz, CD3OD) δ 0.89–0.96 (m, 3 H), 1.29–1.42
(m, 4 H), 1.36 (d, J = 6.9 Hz, 3 H), 1.82–1.90 (m, 1 H), 1.92 (s, 3 H), 1.97–2.07
7(S)-7-Deoxy-7-(4-pyrrolidinocarbonylphenylthio)lincomycin (17). A solution
of compound 10 (100 mg, 0.18 mmol) and pyrrolidine (0.95 ml) were treated
The Journal of Antibiotics

Synthesis and structure–activity relationships of novel LCM derivatives
Y Wakiyama et al
9
according to the similar procedure as described for the preparation of 15 to reduced pressure. The resulting residue was purified by preparative TLC
30
afford 17 (35.0 mg, 33%) as a colorless solid. [α]D +64.2° (c 0.24, MeOH); (CHCl3/MeOH/28% aq NH4OH= 9/2/0.2) to obtain the title compound as a
ESI-MS (m/z) 596 (M+H)+ as C29H45N3O6S2; TOF-ESI-HR-MS (M+H)+ calcd colorless solid (48.0 mg, 45%). [α]D +84.0° (c 1.77, MeOH); ESI-MS (m/z)
29
for C29H45N3O6S2: 596.2828, found: 596.2825; 1H NMR (400 MHz, CD3OD) 582 (M+H)+ as C28H43N3O6S2; TOF-ESI-HR-MS (M+H)+ calcd for
δ 0.90–0.97 (m, 3 H), 1.30–1.43 (m, 4 H), 1.36 (d, J = 6.8 Hz, 3 H), 1.85–2.02 C28H43N3O6S2: 582.2672, found: 582.2669; 1H NMR (400 MHz, CD3OD)
(m, 5 H), 1.91 (s, 3 H), 2.03–2.12 (m, 1 H), 2.17–2.32 (m, 2 H), 2.54 (s, 3 H), δ 0.59–0.66 (m, 2 H), 0.75–0.84 (m, 2 H), 0.89–0.98 (m, 3 H), 1.29–1.42
3.22–3.30 (m, 1 H), 3.35–3.42 (m, 1 H), 3.47 (t, J = 6.6 Hz, 2 H), 3.55–3.61 (m, 4 H), 1.37 (d, J = 6.8 Hz, 3 H), 1.80–1.92 (m, 1 H), 1.87 (s, 3 H), 1.96–2.05
(m, 3 H), 3.79 (br dd, J = 3.1, 0.5 Hz, 1 H), 3.96 (dq, J = 6.8, 2.6 Hz, 1 H), 4.10 (m, 1 H), 2.06–2.25 (m, 2 H), 2.41 (s, 3 H), 2.83 (tt, J = 7.4, 3.8 Hz, 1 H),
(dd, J = 10.2, 5.6 Hz, 1 H), 4.38 (br dd, J = 9.7, 0.5 Hz, 1 H), 4.53 (dd, J = 9.7, 3.03 (dd, J = 10.5, 4.8 Hz, 1 H), 3.26 (dd, J = 8.1, 5.6 Hz, 1 H), 3.58
2.6 Hz, 1 H), 5.25 (d, J = 5.6 Hz, 1 H), 7.43–7.51 (m, 4 H).
(dd, J = 10.2, 3.2 Hz, 1 H), 3.77 (br dd, J = 3.2, 0.6 Hz, 1 H), 3.98
(dq, J =6.8, 2.8 Hz, 1 H), 4.10 (dd, J = 10.2, 5.6 Hz, 1 H), 4.37 (br dd,
J = 9.7, 0.6 Hz, 1 H), 4.50 (dd, J = 9.7, 2.8 Hz, 1 H), 5.25 (d, J = 5.6 Hz, 1 H),
7.40–7.47 (m, 2 H), 7.70–7.78 (m, 2 H).
7(S)-7-Deoxy-7-(5-morpholinocarbonyl-1,3,4-thidiazol-2-ylthio)lincomycin
(18). To a solution of compound 14 (50 mg, 0.09 mmol) in EtOH (1 ml) was
added morpholine (0.1 ml) and refluxed for 3 h and concentrated under
reduced pressure. The resulting residue was purified by preparative TLC
(CHCl3/MeOH/28% aq NH4OH= 9/2/0.2) to obtain the title compound as a
7(S)-7-(4-Cyclohexylcarbamoylphenylthio)-7-deoxylincomycin (24). Compound
19 (100 mg, 0.18 mmol) and cyclohexylamine (0.031 ml, 0.28 mmol) were
treated according to the similar procedure as described for the preparation of
31
colorless solid (34.5 mg, 64%). [α]D +73.4° (c 0.92, MeOH); ESI-MS (m/z)
620 (M+H)+ as C25H41N5O7S3; TOF-ESI-HR-MS (M+H)+ calcd for
C25H41N5O7S3: 620.2246, found: 620.2239; 1H NMR (400 MHz, CD3OD)
δ 0.88–0.97 (m, 3 H), 1.27–1.40 (m, 4 H), 1.57 (d, J = 6.8 Hz, 3 H), 1.79–1.90
(m, 1 H), 1.96 (s, 3 H), 1.97–2.11 (m, 2 H), 2.14–2.27 (m, 1 H), 2.38 (s, 3 H),
30
23 to afford 24 (29.0 mg, 25%) as a colorless solid. [α]D +105.0° (c 1.91,
CHCl3); ESI-MS (m/z) 624 (M+H)+ as C31H49N3O6S2; TOF-ESI-HR-MS
(M+H)+ calcd for C31H49N3O6S2: 624.3141, found: 624.3149; 1H NMR
(400 MHz, CD3OD) δ 0.89–0.97 (m, 3 H), 1.16–1.47 (m, 9 H), 1.37
(d, J =6.9 Hz, 3 H), 1.64–1.73 (m, 1 H), 1.77–1.97 (m, 5 H), 1.89 (s, 3 H),
1.97–2.06 (m, 1 H), 2.07–2.26 (m, 2 H), 2.41 (s, 3 H), 3.02 (dd, J = 10.6,
4.8 Hz, 1 H), 3.26 (dd, J = 8.2, 5.6 Hz, 1 H), 3.58 (dd, J = 10.2, 3.3 Hz, 1 H),
3.77 (br dd, J = 3.3, 0.6 Hz, 1 H), 3.79–3.90 (m, 1 H), 3.98 (dq, J = 6.9, 2.7 Hz,
1 H), 4.10 (dd, J = 10.2, 5.6 Hz, 1 H), 4.37 (br dd, J = 9.7, 0.6 Hz, 1 H),
4.50 (dd, J = 9.7, 2.7 Hz, 1 H), 5.25 (d, J = 5.6 Hz, 1 H), 7.41–7.47 (m, 2 H),
7.72–7.78 (m, 2 H).
3.00 (dd, J =10.5, 5.1 Hz,
1 H), 3.25 (dd, J = 8.5, 6.1 Hz, 1 H),
3.55 (dd, J = 10.3, 3.2 Hz, 1 H), 3.72–3.83 (m, 7 H), 4.10 (dd, J = 10.3, 5.6 Hz,
1 H), 4.24 (br t, J = 4.7 Hz, 2 H), 4.39 (dd, J =9.8, 0.7 Hz, 1 H), 4.47
(dq, J = 6.8, 3.2 Hz,
(d, J =5.6 Hz, 1 H).
1 H), 4.61 (dd, J =9.8, 3.2 Hz, 1 H), 5.25
7(S)-7-(4-Carboxylphenylthio)-7-deoxylincomycin (19). To
a solution of
compound 10 (1.84 g, 3.3 mmol) in MeOH (20 ml) was added 1 M aq NaOH
(5 ml) and stirred at room temperature for 19 h. The mixture was diluted with
1 N HCl (5 ml) and concentrated under reduced pressure. The resulting residue
was purified by Diaion HP-20 (Mitsubishi Chemical) column chromatography
to obtain the title compound as a colorless solid (1.78 g, quant). [α]D28 +161.2°
(c 0.34, DMF); ESI-MS (m/z) 543 (M+H)+ as C25H38N2O7S2; TOF-ESI-HR-MS
(M+H)+ calcd for C25H38N2O7S2: 543.2199, found: 543.2194; 1H NMR (400
MHz, CD3OD) δ 0.89–0.97 (m, 3 H), 1.29–1.45 (m, 4 H), 1.38 (d, J = 6.8 Hz,
3 H), 1.87 (s, 3 H), 1.92–2.03 (m, 1 H), 2.03–2.13 (m, 1 H), 2.17–2.29
(m, 1 H), 2.30–2.38 (m, 1 H), 2.57 (s, 3 H), 3.36 (dd, J = 10.3, 5.2 Hz, 1 H),
3.41 (dd, J = 9.0, 6.1 Hz, 1 H), 3.58 (dd, J = 10.2, 3.3 Hz, 1 H), 3.80
7(S)-7-(4-(Adamant-1-yl)carbamoylphenylthio)-7-deoxylincomycin
(25).
Compound 19 (100 mg, 0.18 mmol) and 1-adamantylamine (0.04 mg, 0.28
mmol) were treated according to the similar procedure as described for the
preparation of 23 to afford 25 (68.6 mg, 55%) as a colorless solid. [α]D31 +74.5°
(c 1.95, MeOH); ESI-MS (m/z) 676 (M+H)+ as C35H53N3O6S2; TOF-ESI-HR-
MS (M+H)+ calcd for C35H53N3O6S2: 676.3454, found: 676.3453; 1H NMR
(400 MHz, CD3OD) δ 0.87–0.98 (m, 3 H), 1.29–1.42 (m, 4 H), 1.35
(d, J =6.8 Hz, 3 H), 1.68–1.80 (m, 6 H), 1.81–1.95 (m, 1 H), 1.90 (s, 3 H),
1.96–2.05 (m, 1 H), 2.06–2.24 (m, 11 H), 2.42 (s, 3 H), 3.04 (dd, J = 10.5,
4.9 Hz, 1 H), 3.27 (dd, J = 8.1, 5.6 Hz, 1 H), 3.59 (dd, J = 10.2, 3.3 Hz, 1 H),
3.76–3.79 (m, 1 H), 3.97 (dq, J =6.8, 2.7 Hz, 1 H), 4.10 (dd, J = 10.2, 5.6 Hz,
1 H), 4.36 (br dd, J = 9.8, 0.5 Hz, 1 H), 4.50 (dd, J = 9.8, 2.7 Hz, 1 H), 5.26
(d, J = 5.6 Hz, 1 H), 7.39–7.45 (m, 2 H), 7.66–7.72 (m, 2 H).
(br dd, J =3.3, 0.8 Hz,
(dd, J = 10.2, 5.6 Hz,
1
H), 3.98 (dq, J = 6.8, 2.7 Hz,
1
1
H), 4.09
H), 4.55
1
H), 4.40 (br dd, J = 9.8, 0.8 Hz,
(dd, J = 9.8, 2.7 Hz, 1 H), 5.25 (d, J = 5.6 Hz, 1 H), 7.38–7.44 (m, 2 H),
7.88–7.95 (m, 2 H).
7(S)-7-Deoxy-7-(4-(pyridin-3-yl)carbamoylphenylthio)lincomycin
(26).
For the qualified analytical purpose, the above colorless solid was further
purified by reverse-phase column chromatography (Biotage SNAP Ultra C18,
25 µm, room temperature, 12.0 ml min− 1, H2O/MeOH= 100/0–0/100) to
obtain the highly purified title compound as a colorless solid.
Compound 19 (40.5 mg, 0.075 mmol) and 3-aminopyridine (10.5 mg, 0.11
mmol) were treated according to the similar procedure as described for the
preparation of 23 to afford 26 (13.5 mg, 29%) as a colorless solid. [α]D30 +31.1°
(c 0.10, MeOH); ESI-MS (m/z) 619 (M+H)+ as C30H42N4O6S2; TOF-ESI-HR-
MS (M+H)+ calcd for C30H42N4O6S2: 619.2624, found: 619.2623; 1H NMR
(400 MHz, CD3OD) δ 0.93 (br t, J = 6.9 Hz, 3 H), 1.26–1.45 (m, 4 H), 1.41
7(S)-7-(5-Carboxylpyridin-2-ylthio)-7-deoxylincomycin (20). Compound 11
(621.8 mg, 1.12 mmol), 1 M aq NaOH (6.2 ml) and MeOH (6.2 ml) were
treated according to the similar procedure as described for the preparation of (d, J =6.8 Hz, 3 H), 1.85–1.98 (m, 1 H), 1.88 (s, 3 H), 2.01–2.11 (m, 1 H),
29
2.15–2.28 (m, 2 H), 2.50 (s, 3 H), 3.14–3.24 (m, 1 H), 3.30–3.38 (m, 1 H), 3.59
(dd, J = 10.2, 3.2 Hz, 1 H), 3.80 (br d, J = 3.2 Hz,1 H), 4.04 (dq, J = 6.8, 2.7 Hz,
19 to afford 20 (488.1 mg, 81%) as a colorless solid. [α]D +90.4° (c 0.30,
DMF); ESI-MS (m/z) 544 (M+H)+ as C24H37N3O7S2; TOF-ESI-HR-MS
(M+H)+ calcd for C24H37N3O7S2: 544.2151, found: 544.2151; 1H NMR 1 H), 4.11 (dd, J = 10.2, 5.6 Hz, 1 H), 4.41 (br d, J = 9.8 Hz, 1 H), 4.57
(400 MHz, CD3OD) δ 0.88–0.98 (t, J = 7.1 Hz, 3 H), 1.25–1.50 (m, 4 H), (dd, J = 9.8, 2.7 Hz, 1 H), 5.27 (d, J =5.6 Hz, 1 H), 7.45 (br ddd, J = 8.3, 4.8,
1.46 (d, J = 7.0 Hz, 3 H), 1.83 (s, 3 H), 1.96–2.16 (m, 2 H), 2.20–2.33 (m, 1 H), 0.7 Hz, 1 H), 7.48–7.55 (m, 2 H), 7.88–7.96 (m, 2 H), 8.25 (ddd, J = 8.3,
2.38–2.47 (m, 1 H), 2.62 (s, 3 H), 3.45–3.55 (m, 2 H), 3.56 (dd, J = 10.2, 2.4, 1.4 Hz, 1 H), 8.31 (br dd, J = 4.8, 1.4 Hz, 1 H), 8.86–8.91 (m, 1 H).
3.2 Hz, 1 H), 3.80–3.83 (m, 1 H), 4.10 (dd, J = 10.2, 5.6 Hz, 1 H), 4.32–4.42
7(S)-7-Deoxy-7-(4-piperidinocarbonylphenylthio)lincomycin (27). Compound
(m, 2 H), 4.53 (dd, J = 9.7, 3.2 Hz, 1 H), 5.24 (d, J = 5.6 Hz, 1 H), 7.30–7.35
19 (100 mg, 0.18 mmol) and piperidine (0.027 ml, 0.28 mmol) were treated
(m, 1 H), 8.09 (dd, J = 8.4, 2.1 Hz, 1 H), 8.92–8.97 (m, 1 H).
according to the similar procedure as described for the preparation of 23 to
32
7(S)-7-(4-Cyclopropylcarbamoylphenylthio)-7-deoxylincomycin (23). To
a
afford 27 (71 mg, 63%) as a colorless solid. [α]D +65.4° (c 0.18, MeOH);
solution of compound 19 (100 mg, 0.18 mmol) in DMF (1 ml) were added ESI-MS (m/z) 610 (M+H)+ as C30H47N3O6S2; TOF-ESI-HR-MS (M+H)+ calcd
1-hydroxybenzotriazole (37.3 mg, 0.28 mmol), 1-ethyl-3-(3-dimethylamino- for C30H47N3O6S2: 610.2985, found: 610.2981; H NMR (400 MHz, CD3OD)
1
propyl)carbodiimide·HCl (53.0 mg, 0.28 mmol) and cyclopropylamine δ 0.88–0.97 (m, 3 H), 1.30–1.42 (m, 4 H), 1.36 (d, J = 6.8 Hz, 3 H), 1.48–1.77
(0.019 ml, 0.28 mmol) and stirred at room temperature for 4 h. The (m, 7 H), 1.85–1.97 (m, 1 H), 1.92 (s, 3 H), 1.99–2.10 (m, 1 H), 2.14–2.27
mixture was diluted with saturated aq NaHCO3 (10 ml), then extracted (m, 2 H), 2.47 (s, 3 H), 3.08–3.17 (m, 1 H), 3.32–3.45 (m, 2 H), 3.57
with EtOAc, washed with water, dried over MgSO4 and concentrated under (dd, J = 10.1, 3.2 Hz, 1 H), 3.61–3.74 (m, 2 H), 3.77 (br d, J = 3.2 Hz, 1 H), 3.95
The Journal of Antibiotics

Synthesis and structure–activity relationships of novel LCM derivatives
Y Wakiyama et al
10
(dq, J =6.8, 2.5 Hz, 1 H), 4.10 (dd, J = 10.1, 5.6 Hz, 1 H), 4.36 (br d, J = 9.8 Hz,
5 M aq NaOH (0.3 ml) and stirred at room temperature for 1 h. The mixture
1 H), 4.50 (br dd, J = 9.8, 2.5 Hz, 1 H), 5.25 (d, J = 5.6 Hz, 1 H), 7.32–7.37 was diluted with 5 N HCl (0.3 ml) and concentrated under reduced pressure.
The resulting residue (crude compound 22), morpholine (0.32 ml, 3.70 mmol),
1-ethyl-3-(3-dimethylaminopropyl)carbodiimide·HCl (212.8 mg, 1.11 mmol),
1-hydroxybenzotriazole (150 mg, 1.11 mmol) and Et3N (1.03 ml, 7.40 mmol)
were treated according to the similar procedure (60 °C, 2 days) as described for
(m, 2 H), 7.44–7.49 (m, 2 H).
7(S)-7-Deoxy-7-(4-morpholinocarbonylphenylthio)lincomycin (28). Compound
19 (200 mg, 0.37 mmol) and morpholine (0.048 ml, 0.55 mmol) were treated
according to the similar procedure as described for the preparation of 23 to
27
the preparation of 23 to afford 32 (29.9 mg, 7%) as a colorless solid. [α]D
31
afford 28 (142 mg, 63%) as a colorless solid. [α]D +78.7° (c 2.08, MeOH);
+84.5° (c 3.39, MeOH); ESI-MS (m/z) 619 (M+H)+ as C26H42N4O7S3;
TOF-ESI-HR-MS (M+H)+ calcd for C26H42N4O7S3: 619.2294, found:
619.2288; 1H NMR (400 MHz, CD3OD) δ 0.86–0.97 (m, 3 H), 1.26–1.41
(m, 4 H), 1.51 (d, J = 7.0 Hz, 3 H), 1.77–1.89 (m, 1 H), 1.96 (s, 3 H), 1.96–2.11
(m, 2 H), 2.11–2.27 (m, 1 H), 2.37 (s, 3 H), 2.99 (dd, J = 10.4, 5.1 Hz, 1 H),
3.24 (dd, J = 8.4, 6.1 Hz, 1 H), 3.56 (dd, J = 10.2, 3.2 Hz, 1 H), 3.66–3.77 (m, 8
H), 3.77–3.82 (m, 1 H), 4.10 (dd, J = 10.2, 5.6 Hz, 1 H), 4.30 (dq, J = 7.0,
3.2 Hz, 1 H), 4.36 (br dd, J = 9.8, 0.5 Hz, 1 H), 4.57 (dd, J = 9.8, 3.2 Hz, 1 H),
5.26 (d, J = 5.6 Hz, 1 H), 7.93 (s, 1 H).
ESI-MS (m/z) 612 (M+H)+ as C29H45N3O7S2; TOF-ESI-HR-MS (M+H)+ calcd
1
for C29H45N3O7S2: 612.2777, found: 612.2772; H NMR (400 MHz, CD3OD)
δ 0.88–0.97 (m, 3 H), 1.28–1.42 (m, 4 H), 1.35 (d, J = 6.8 Hz, 3 H), 1.81–1.91
(m, 1 H), 1.91 (s, 3 H), 1.97–2.06 (m, 1 H), 2.07–2.14 (m, 1 H), 2.14–2.25
(m, 1 H), 2.43 (s, 3 H), 3.03 (dd, J = 10.6, 4.8 Hz, 1 H), 3.27 (dd, J =8.3,
5.7 Hz, 1 H), 3.33–3.57 (m, 2 H), 3.57 (dd, J = 10.2, 3.3 Hz, 1 H), 3.57–3.85
(m, 6 H), 3.77 (br dd, J = 3.3, 0.6 Hz, 1 H), 3.97 (dq, J = 6.8, 2.7 Hz, 1 H), 4.10
(dd, J = 10.2, 5.6 Hz, 1 H), 4.35 (br dd, J = 9.7, 0.6 Hz, 1 H), 4.49 (dd, J = 9.7,
2.7 Hz, 1 H), 5.26 (d, J = 5.6 Hz, 1 H), 7.36–7.42 (m, 2 H), 7.45–7.52 (m, 2 H).
7(S)-7-Acetylthio-7-deoxy-2,3,4-tris-O-(trimethylsilyl)lincomycin (33). To
a
7(S)-7-Deoxy-7-(4-(4-methylpiperazin-1-yl)carbonylphenylthio)lincomycin
(29). Compound 19 (67.8 mg, 0.12 mmol) and 1-methylpiperazine (0.021 ml,
0.19 mmol) were treated according to the similar procedure as described for the
preparation of 23 to afford 29 (54.0 mg, 69%) as a colorless solid. [α]D31 +77.0°
(c 1.31, MeOH); ESI-MS (m/z) 625 (M+H)+ as C30H48N4O6S2; TOF-ESI-HR-
MS (M+H)+ calcd for C30H48N4O6S2: 625.3094, found: 625.3091; 1H NMR
(400 MHz, CD3OD) δ 0.88–0.98 (m, 3 H), 1.27–1.42 (m, 4 H), 1.36
(d, J =6.8 Hz, 3 H), 1.81–1.95 (m, 1 H), 1.91 (s, 3 H), 1.97–2.07 (m, 1 H),
2.08–2.25 (m, 2 H), 2.33 (s, 3 H), 2.36–2.58 (m, 4 H), 2.43 (s, 3 H), 3.05 (dd,
J = 10.6, 4.8 Hz, 1 H), 3.28 (dd, J = 8.1, 5.5 Hz, 1 H), 3.38–3.61 (m, 2 H), 3.57
(dd, J = 10.1, 3.2 Hz, 1 H), 3.61–3.88 (m, 3 H), 3.97 (dq, J = 6.8, 2.7 Hz, 1 H),
4.10 (dd, J = 10.1, 5.5 Hz, 1 H), 4.35 (br dd, J = 9.7, 0.5 Hz, 1 H), 4.49
(dd, J = 9.7, 2.7 Hz, 1 H), 5.25 (d, J = 5.5 Hz, 1 H), 7.35–7.40 (m, 2 H),
7.45–7.51 (m, 2 H).
solution of compound 2 (200 mg, 0.29 mmol) in DMF (0.65 ml) was added
potassium ethanethioate (163 mg, 1.4 mmol) at 60 °C for 4 h. The mixture was
diluted with EtOAc and washed with 10% aq NaHCO3, dried over MgSO4 and
concentrated under reduced pressure. The resulting residue was purified by
silica gel column chromatography (hexane/EtOAc = 3/1) to obtain the title
compound as a colorless solid (170 mg, 88%). ESI-MS (m/z) 681 (M+H)+ as
C29H60N2O6S2Si3; 1H NMR (400 MHz, CDCl3) δ 0.09–0.20 (m, 27 H),
0.84–0.93 (m, 3 H), 1.20–1.47 (m, 7 H), 1.76–1.87 (m, 1 H), 1.90–2.09
(m, 6 H), 2.31 (s, 3 H), 2.40 (s, 3 H), 2.93–3.02 (m, 1H), 3.12–3.20 (m, 1H),
3.56 (dd, J = 9.5, 2.4 Hz, 1 H), 3.72 (d, J = 2.4 Hz, 1 H), 3.94 (d, J = 10.0 Hz,
1 H), 4.07 (dt, J = 7.1, 2.2 Hz, 1 H), 4.15 (dd, J = 9.5, 5.6 Hz, 1 H), 4.55 (ddd,
J = 10.7, 10.0, 2.2 Hz, 1 H), 5.18 (d, J = 5.6 Hz, 1 H), 7.34 (d, J = 10.7 Hz, 1 H).
7(S)-7-Acetylthio-7-deoxylincomycin (34). To a solution of compound 33
(10.6 g, 16 mmol) in MeOH (50 ml) was added 2 N HCl (39 ml) and stirred
at room temperature for 10 min. The mixture was diluted with 10% aq
NaHCO3 (30 ml) and concentrated under reduced pressure. The resulting
residue was diluted with EtOAc and washed with 10% aq NaCl, dried over
MgSO4 and concentrated under reduced pressure. The resulting residue was
purified by silica gel column chromatography (EtOAc/methanol= 19/1) to
obtain the title compound as a colorless solid (7.05 g, 97%). ESI-MS (m/z) 465
(M+H)+ as C20H36N2O6S2; TOF-ESI-HR-MS (M+H)+ calcd for
C20H36N2O6S2: 465.2093, found: 465.2092; 1H NMR (400 MHz, CDCl3)
δ 0.88–0.95 (m, 3 H), 1.22–1.42 (m, 7 H), 1.82–2.13 (m, 7 H), 2.35–2.44
(m, 7 H), 2.72 (d, J = 10.0 Hz, 1 H), 3.05 (dd, J = 10.5, 4.6 Hz, 1 H), 3.19–3.28
(m, 1 H), 3.46–3.56 (m, 1 H), 3.61 (br s, 1H), 3.94 (d, J = 10.2 Hz, 1 H), 4.11
(dd, J = 10.5, 4.6 Hz, 1 H), 4.17 (dq, J = 7.1, 2.4 Hz, 1 H), 4.25 (ddd, J = 10.2,
9.5, 2.4 Hz, 1 H), 5.07 (d, J = 2.9 Hz, 1 H), 5.31 (d, J = 5.6 Hz, 1 H), 7.79
(d, J = 9.5 Hz, 1 H).
7(S)-7-Deoxy-7-(5-molpholinocarbonylpyridin-2-ylthio)lincomycin (30). Com-
pound 20 (96.9 mg, 0.18 mmol) and morpholine (0.024 ml, 0.28 mmol) were
treated according to the similar procedure as described for the preparation of
29
23 to afford 30 (76.7 mg, 70%) as a colorless solid. [α]D +55.4° (c 2.46,
MeOH); ESI-MS (m/z) 613 (M+H)+ as C28H44N4O7S2; TOF-ESI-HR-MS
(M+H)+ calcd for C28H44N4O7S2: 613.2730, found: 613.2735; 1H NMR
(400 MHz, CD3OD) δ 0.88–0.97 (m, 3 H), 1.27–1.41 (m, 4 H), 1.47
(d, J = 6.9 Hz, 3 H), 1.79 (s, 3 H), 1.82–1.92 (m, 1 H), 2.02 (ddd, J = 13.0,
7.9, 5.1 Hz, 1 H), 2.07–2.14 (m, 1 H), 2.14–2.27 (m, 1 H), 2.39 (s, 3 H), 3.03
(dd, J = 10.5, 5.1 Hz, 1 H), 3.26 (dd, J = 8.4, 5.9 Hz, 1 H), 3.40–3.83 (m, 8 H),
3.55 (dd, J =10.2, 3.2 Hz, 1 H), 3.75–3.80 (m, 1 H), 4.10 (dd, J = 10.2, 5.6 Hz,
1 H), 4.34 (br dd, J = 9.8, 0.4 Hz, 1 H), 4.43 (dq, J = 6.9, 3.1 Hz, 1 H), 4.52
(dd, J = 9.8, 3.1 Hz, 1 H), 5.23 (d, J = 5.6 Hz, 1 H), 7.38 (dd, J = 8.3, 0.9 Hz,
1 H), 7.67 (dd, J =8.3, 2.3 Hz, 1 H), 8.49 (dd, J =2.3, 0.9 Hz, 1 H).
7(S)-7-(4-Amino-5-molpholinocarbonylpyrimidin-2-ylthio)-7-deoxylincomycin
(31). To a solution of compound 12 (117.2 mg, 0.2 mmol) in MeOH (1 ml)
was added 1 M aq NaOH (0.3 ml) and stirred at room temperature for 10 h.
The mixture was diluted with 1 N HCl (0.3 ml) and concentrated under
reduced pressure. The resulting residue (crude compound 21), morpholine
(0.008 ml, 0.092 mmol), N,N’-dicyclohexylcarbodiimide (30.7 mg, 0.15 mmol),
1-hydroxybenzotriazole (20.3 mg, 0.15 mmol) and Et3N (0.012 ml, 0.09 mmol)
were treated according to the similar procedure as described for the preparation
7(S)-7-Deoxy-7-mercaptolincomycin (35). To a solution of compound 34
(7.05 g, 15.2 mmol) in MeOH (50 ml) was added sodium methoxide
(2.46 g, 45.5 mmol) and stirred at room temperature for 20 min. The mixture
was diluted with saturated aq NH4Cl and concentrated under reduced pressure.
The resulting residue was diluted with EtOAc and washed with 10% aq
NaHCO3, dried over MgSO4 and concentrated under reduced pressure.
The resulting residue was purified by silica gel column chromatography
(CHCl3/MeOH/28% aq NH4OH= 95/5/0.1) to obtain the title compound as
31
26
of 23 to afford 31 (7.7 mg, 6%) as a colorless solid. [α]D +22.4° (c 0.11,
a colorless solid (6.06 g, 94%). [α]D +152.6° (c 0.98, MeOH); ESI-MS (m/z)
MeOH); ESI-MS (m/z) 629 (M+H)+ as C27H44N6O7S2; TOF-ESI-HR-MS
(M+H)+ calcd for C27H44N6O7S2: 629.2791, found: 629.2792; 1H NMR
(400 MHz, CD3OD) δ 0.84–0.97 (m, 3 H), 1.24–1.39 (m, 4 H), 1.47
(d, J =6.8 Hz, 3 H), 1.78–1.91 (m, 1 H), 1.86 (s, 3 H), 1.94–2.10 (m, 2 H),
2.12–2.26 (m, 1 H), 2.37 (s, 3 H), 2.98 (dd, J = 10.5, 5.1 Hz, 1 H), 3.22
423 (M+H)+ as C18H34N2O5S2; TOF-ESI-HR-MS (M+H)+ calcd for
C18H34N2O5S2: 423.1987, found: 423.1987; 1H NMR (400 MHz, CD3OD)
δ 0.88–0.96 (m, 3 H), 1.28 (d, J =7.1 Hz, 3 H), 1.28–1.41 (m, 4 H), 1.81–1.93
(m, 1 H), 1.96–2.05 (m, 1 H), 2.06–2.23 (m, 2 H), 2.17 (s, 3 H), 2.43 (s, 3 H),
3.02 (dd, J =10.8, 4.6 Hz, 1 H), 3.25 (dd, J = 8.3, 5.6 Hz, 1 H), 3.54 (dd,
(dd, J =8.4, 6.2 Hz, 1 H), 3.50–3.74 (m, 9 H), 3.75–3.80 (m, 1 H), 4.09 J = 10.2, 3.3 Hz, 1 H), 3.58 (dq, J = 7.1, 2.0 Hz, 1 H), 3.70 (br dd, J = 3.3,
(dd, J = 10.2, 5.6 Hz, 1 H), 4.29–4.38 (m, 2 H), 4.49 (dd, J = 9.8, 3.2 Hz, 1 H),
5.22 (d, J = 5.6 Hz, 1 H), 7.96 (s, 1 H).
0.6 Hz, 1 H), 4.04 (br dd, J = 10.0, 0.6 Hz, 1 H), 4.09 (dd, J = 10.2, 5.7 Hz, 1 H),
4.26 (dd, J = 10.0, 2.0 Hz, 1 H), 5.25 (d, J = 5.7 Hz, 1 H).
7(S)-7-Deoxy-7-(5-molpholinocarbonylthiazol-2-ylthio)lincomycin (32). To
a
7(S)-7-Deoxy-7-(4-(methoxy-N-propylacetamido)phenylthio)lincomycin (36).
solution of compound 13 (430 mg, 0.74 mmol) in EtOH (8 ml) was added To
a
solution of compound 35 (70 mg, 0.17 mmol), 4,5-bis
The Journal of Antibiotics

Synthesis and structure–activity relationships of novel LCM derivatives
Y Wakiyama et al
11
(diphenylphosphino)-9,9-dimethylxanthene (Xantphos) (9.7 mg, 0.017 mmol) (c 2.63, MeOH); ESI-MS (m/z) 598 (M+H)+ as C29H47N3O6S2; TOF-ESI-HR-
MS (M+H)+ calcd for C29H47N3O6S2: 598.2985, found: 598.2983; 1H NMR
(400 MHz, CD3OD) δ 0.88–0.97 (m, 3 H), 1.28 (d, J = 6.9 Hz, 3 H), 1.29–1.41
(m, 4 H), 1.80–1.91 (m, 1 H), 1.95–2.04 (m, 1 H), 1.98 (s, 3 H), 2.05–2.12
(m, 1 H), 2.12–2.23 (m, 1 H), 2.35–2.50 (m, 4 H), 2.40 (s, 3 H), 2.99
(dd, J = 10.6, 4.6 Hz, 1 H), 3.25 (dd, J = 8.1, 5.6 Hz, 1 H), 3.49 (s, 2 H), 3.57
(dd, J = 10.3, 3.2 Hz, 1 H), 3.63–3.71 (m, 4 H), 3.74 (br dd, J =3.2, 0.5 Hz,
1 H), 3.85 (dq, J = 6.9, 2.6 Hz, 1 H), 4.10 (dd, J =10.3, 5.5 Hz, 1 H), 4.32
(br dd, J = 9.8, 0.5 Hz, 1 H), 4.41 (dd, J = 9.8, 2.6 Hz, 1 H), 5.26 (d, J = 5.5 Hz,
1 H), 7.28–7.34 (m, 2 H), 7.36–7.43 (m, 2 H).
and tris(dibenzylideneacetone)dipalladium(0) (Pd2(dba)3) (7.6 mg, 0.0084
mmol) in 1,4-dioxane (1 ml) were added N-(4-bromophenyl)-2-methoxy-N-
propylacetamide (94.7 mg, 0.33 mmol) and N,N-diisopropylethylamine
(0.06 ml, 0.33 mmol) and refluxed for 6 h. The mixture was diluted with
saturated aq NaHCO3 (15 ml), then extracted with EtOAc, washed with water,
dried over MgSO4 and concentrated under reduced pressure. The resulting
residue was purified by preparative TLC (CHCl3/MeOH/28% aq NH4OH= 9
30
/2/0.2) to obtain the title compound as a colorless solid (79.0 mg, 76%). [α]D
+83.2° (c 2.21, MeOH); ESI-MS (m/z) 628 (M+H)+ as C30H49N3O7S2; TOF-
1
ESI-HR-MS (M+H)+ calcd for C30H49N3O7S2: 628.3090, found: 628.3086; H
7(S)-7-Deoxy-7-(2-fluoro-4-morpholinocarbonylphenylthio)lincomycin
(41).
NMR (400 MHz, CD3OD) δ 0.90 (t, J = 7.5 Hz, 3 H), 0.88–0.97 (m, 3 H), 1.32–
1.42 (m, 4 H), 1.36 (d, J = 6.9 Hz, 3 H), 1.52 (sxt, J = 7.4 Hz, 2 H),
1.82–1.97 (m, 1 H), 1.93 (s, 3 H), 2.03 (ddd, J = 12.7, 7.6, 5.0 Hz, 1 H),
2.08–2.26 (m, 2 H), 2.45 (s, 3 H), 3.06 (dd, J = 10.5, 4.8 Hz, 1 H), 3.23–3.30
(m, 4 H), 3.58 (dd, J = 10.2, 3.2 Hz, 1 H), 3.61–3.68 (m, 2 H), 3.72–3.80
(m, 3 H), 3.94 (dq, J = 6.9, 2.6 Hz, 1 H), 4.11 (dd, J = 10.2, 5.6 Hz, 1 H), 4.35
(br d, J =9.7, 0.5 Hz, 1 H), 4.49 (dd, J = 9.7, 2.6 Hz, 1 H), 5.27 (d, J = 5.6 Hz,
1 H), 7.20–7.27 (m, 2 H), 7.46–7.52 (m, 2 H).
Compound 35 (190.7 mg, 0.45 mmol) and (4-bromo-3-fluorophenyl)
(morpholino)methanone (260 mg, 0.90 mmol) were treated according to the
similar procedure as described for the preparation of 36 to afford 41 (232 mg,
82%) as a colorless solid. [α]D31 +82.4° (c 8.43, MeOH); ESI-MS (m/z) 630 (M
+H)+ as C29H44FN3O7S2; TOF-ESI-HR-MS (M+H)+ calcd for C29H44FN3O7S2:
630.2683, found: 630.2673; 1H NMR (400 MHz, CD3OD)
δ 0.87–0.97
(m, H), 1.30 (d, J = 6.9 Hz, H), 1.28–1.92 (m, H), 1.79–1.92
3
3
4
(m, 1 H), 1.98 (s, 3 H), 1.98–2.05 (m, 1 H), 2.06–2.13 (m, 1 H), 2.13–2.26
(m, 1 H), 2.42 (s, 3 H), 3.01 (dd, J = 10.6, 4.7 Hz, 1 H), 3.27 (dd, J =8.3,
5.7 Hz, 1 H), 3.35–3.82 (m, 10 H), 4.01 (dq, J = 6.9, 2.7 Hz, 1 H), 4.10
(dd, J = 10.2, 5.5 Hz, 1 H), 4.32 (br dd, J = 9.7, 0.4 Hz, 1 H), 4.49 (dd, J = 9.7,
2.7 Hz, 1 H), 5.26 (d, J = 5.5 Hz, 1 H), 7.22–7.28 (m, 2 H), 7.55–7.62 (m, 1 H).
7(S)-7-Deoxy-7-(4-((S)-2-methoxymethylpyrrolidinocarbonyl)phenylthio)lincomy-
cin (37). Compound 35 (70 mg, 0.17 mmol) and (S)-(4-bromophenyl)
(2-(methoxymethyl)pyrrolidin-1-yl)methanone (98.7 mg, 0.33 mmol) were
treated according to the similar procedure as described for the preparation of
31
36 to afford 37 (84.0 mg, 79%) as a colorless solid. [α]D +46.2° (c 1.85,
MeOH); ESI-MS (m/z) 640 (M+H)+ as C31H49N3O7S2; TOF-ESI-HR-MS
(M+H)+ calcd for C31H49N3O7S2: 640.3090, found: 640.3092; 1H NMR
(400 MHz, CD3OD) δ 0.88–0.97 (m, 3 H), 1.29–1.40 (m, 4 H), 1.36
(d, J = 6.8 Hz, 3 H), 1.69–1.81 (m, 1 H), 1.81–1.90 (m, 1 H), 1.91 (s, 3 H),
1.90–2.27 (m, 7 H), 2.42 (s, 3 H), 3.02 (dd, J = 10.6, 4.8 Hz, 1 H), 3.04–3.14
(m, 1 H), 3.27 (dd, J = 8.1, 5.7 Hz, 1 H), 3.39 (s, 3 H), 3.46–3.67 (m, 4 H), 3.78
(br dd, J = 3.2, 0.6 Hz, 1 H), 3.97 (dq, J = 6.8, 2.6 Hz, 1 H), 4.10 (dd, J = 10.2,
5.6 Hz, 1 H), 4.36 (br d, J = 9.7 Hz, 1 H), 4.49 (dd, J = 9.7, 2.6 Hz, 1 H),
5.26 (d, J = 5.6 Hz, 1 H), 7.41–7.50 (m, 4 H).
7(S)-7-Deoxy-7-(3-fluoro-4-morpholinocarbonylphenylthio)lincomycin
Compound 35 (187 mg, 0.44 mmol) and (4-bromo-2-fluorophenyl)(morpho-
lino)methanone (255 mg, 0.89 mmol) were treated according to the similar
(42).
procedure as described for the preparation of 36 to afford 42 (227 mg, 81%) as
31
a
colorless solid. [α]D +73.4° (c 5.35, MeOH); ESI-MS (m/z) 630
(M+H)+ as C29H44FN3O7S2; TOF-ESI-HR-MS (M+H)+ calcd for
C29H44FN3O7S2: 630.2683, found: 630.2685; 1H NMR (400 MHz, CD3OD)
δ 0.87–0.96 (m, 3 H), 1.27–1.38 (m, 4 H), 1.39 (d, J = 6.8 Hz, 3 H), 1.79–1.89
(m, 1 H), 1.92 (s, 3 H), 1.96–2.12 (m, 2 H), 2.12–2.26 (m, 1 H), 2.40 (s, 3 H),
2.99 (dd, J = 10.6, 4.8 Hz, 1 H), 3.25 (dd, J =8.4, 5.9 Hz, 1 H), 3.31–3.40 (m, 2
H), 3.57 (dd, J = 10.1, 3.2 Hz, 1 H), 3.59–3.66 (m, 2 H), 3.70–3.81 (m, 5 H),
3.99 (dq, J = 6.8, 2.8 Hz, 1 H), 4.11 (dd, J = 10.1, 5.6 Hz, 1 H), 4.33 (br dd,
J = 9.6, 0.4 Hz, 1 H), 4.52 (dd, J = 9.6, 2.8 Hz, 1 H), 5.27 (d, J = 5.6 Hz, 1 H),
7.24 (dd, J = 10.2, 1.5 Hz, 1 H), 7.27–7.32 (m, 1 H), 7.33–7.39 (m, 1 H).
7(S)-7-Deoxy-7-(4-((S)-2-dimethylaminomethylpyrrolidinocarbonyl)phenylthio)
lincomycin (38). Compound 35 (70 mg, 0.17 mmol) and (S)-(4-bromophe-
nyl)(2-((dimethylamino)methyl)pyrrolidin-1-yl)methanone (103.7 mg, 0.33
mmol) were treated according to the similar procedure as described for the
preparation of 36 to afford 38 (90.0 mg, 82%) as a colorless solid. [α]D31 +33.2°
(c 2.39, MeOH); ESI-MS (m/z) 653 (M+H)+ as C32H52N4O6S2; TOF-ESI-HR-
MS (M+H)+ calcd for C32H52N4O6S2: 653.3407, found: 653.3399; 1H NMR
(400 MHz, CD3OD) δ 0.86-0.97 (m, 3 H), 1.25-1.42 (m, 7 H), 1.75–2.25
(m, 13 H), 2.38 (s, 6 H), 2.41 (s 3 H), 2.76 (br dd, J = 11.7, 3.2 Hz, 1 H), 3.00
(dd, J = 10.6, 4.8 Hz, 1 H), 3.26 (dd, J =8.2, 5.9 Hz, 1 H), 3.35–3.46 (m, 1 H),
3.48–3.68 (m, 2 H), 3.75–3.80 (m, 1 H), 3.92–4.20 (m, 1 H), 4.10 (dd, J = 10.1,
5.6 Hz, 1 H), 4.35 (d, J = 9.7 Hz, 1 H), 4.49 (dd, J = 9.7, 2.6 Hz, 1 H), 5.26
(d, J =5.6 Hz, 1 H), 7.40–7.53 (m, 4 H).
7(S)-7-Deoxy-7-(4-morpholinocarbonyl-3-nitrophenylthio)lincomycin
(43).
Compound 35 (70 mg, 0.17 mmol) and (4-bromo-2-nitrophenyl)
(morpholino)methanone (104 mg, 0.33 mmol) were treated according to the
similar procedure as described for the preparation of 36 to afford 43 (87.0 mg,
31
80%) as a colorless solid. [α]D +60.4° (c 2.62, MeOH); ESI-MS (m/z) 657
(M+H)+ as C29H44N4O9S2; TOF-ESI-HR-MS (M+H)+ calcd for C29H44
N4O9S2: 657.2628, found: 657.2632; 1H NMR (400 MHz, CD3OD)
δ 0.87–0.97 (m, 3 H), 1.28–1.39 (m, 4 H), 1.41 (d, J =6.9 Hz, 3 H),
1.79–1.90 (m, 1 H), 1.93 (s, 3 H), 1.97–2.12 (m, 2 H), 2.13–2.26 (m, 1 H),
2.41 (s, 3 H), 3.00 (dd, J = 10.6, 4.8 Hz, 1 H), 3.20–3.34 (m, 3 H), 3.57
(dd, J =10.2, 3.2 Hz, 1 H), 3.62 (t, J = 4.8 Hz, 2 H), 3.67–3.88 (m, 5 H), 4.04
(dq, J = 6.9, 2.9 Hz, 1 H), 4.10 (dd, J =10.2, 5.6 Hz, 1 H), 4.35 (br dd, J = 9.5,
0.7 Hz, 1 H), 4.55 (dd, J = 9.5, 2.9 Hz, 1 H), 5.26 (d, J = 5.6 Hz, 1 H), 7.45
(d, J = 8.0 Hz, 1 H), 7.81 (dd, J = 8.0, 1.8 Hz, 1 H), 8.16 (d, J = 1.8 Hz, 1 H).
7(S)-7-Deoxy-7-(4-(1,4-oxazepane-4-carbonyl)phenylthio)lincomycin
(39).
Compound 35 (70 mg, 0.17 mmol) and (4-bromophenyl)(1,4-oxazepan-4-yl)
methanone (49.0 mg, 0.17 mmol) were treated according to the similar
procedure as described for the preparation of 36 to afford 39 (79.0 mg, 73%)
as a colorless solid. [α]D31 +74.3° (c 3.26, MeOH); ESI-MS (m/z) 626 (M+H)+
as C30H47N3O7S2; TOF-ESI-HR-MS (M+H)+ calcd for C30H47N3O7S2:
626.2934, found: 626.2934; 1H NMR (400 MHz, CD3OD)
δ 0.88–0.97
7(S)-7-Deoxy-7-(3-methyl-4-morpholinocarbonylphenylthio)lincomycin (44).
Compound 35 (70 mg, 0.17 mmol) and (4-bromo-2-methylphenyl)(morpho-
lino)methanone (94.1 mg, 0.33 mmol) were treated according to the similar
(m, H), 1.28–1.41 (m, H), 1.35 (d, J = 6.8 Hz, H), 1.75–1.91
3
4
3
(m, 2 H), 1.92 (s, 3 H), 1.94–2.06 (m, 2 H), 2.07–2.26 (m, 2 H), 2.43
(s, 3 H), 3.04 (dd, J = 10.6, 4.8 Hz, 1 H), 3.28 (dd, J = 5.6, 2.4 Hz, 1 H),
3.49–3.57 (m, 2 H), 3.58 (dd, J =10.1, 2.3 Hz, 1 H), 3.67 (br t, J = 4.9 Hz, 1 H),
3.73–3.88 (m, 6 H), 3.96 (dq, J = 6.8, 2.5 Hz, 1 H), 4.10 (dd, J =10.1, 5.6 Hz,
1 H), 4.35 (br d, J = 9.7 Hz, 1 H), 4.49 (dd, J = 9.7, 2.5 Hz, 1 H), 5.26
(d, J =5.6 Hz, 1 H), 7.34–7.42 (m, 2 H), 7.43–7.52 (m, 2 H).
procedure as described for the preparation of 36 to afford 44 (81 mg, 78%) as a
29
colorless solid. [α]D
+81.7° (c 2.65, MeOH); ESI-MS (m/z) 626
(M+H)+ as C30H47N3O7S2; TOF-ESI-HR-MS (M+H)+ calcd for
C30H47N3O7S2: 626.2934, found: 626.2925; 1H NMR (400 MHz, CD3OD)
δ 0.89–0.97 (m, 3 H), 1.26–1.42 (m, 4 H), 1.33 (d, J = 6.8 Hz, 3 H), 1.80–1.91
(m, 1 H), 1.95 (s, 3 H), 2.01 (ddd, J = 12.8, 7.9, 4.7 Hz, 1 H), 2.06–2.13
(m, 1 H), 2.13–2.24 (m, 1 H), 2.28 (s, 3 H), 2.42 (s, 3 H), 3.00 (dd, J = 10.6,
4.6 Hz, 1 H), 3.21–3.29 (m, 3 H), 3.53–3.64 (m, 3 H), 3.68–3.82 (m, 5 H), 3.93
7(S)-7-Deoxy-7-(4-morpholinomethylphenylthio)lincomycin (40). Compound
35 (70 mg, 0.17 mmol) and 4-(4-bromobenzyl)morpholine (84.8 mg, 0.33
mmol) were treated according to the similar procedure as described for the
preparation of 36 to afford 40 (74.0 mg, 75%) as a colorless solid. [α]D28 +98.2° (dq, J = 6.8, 2.7 Hz, 1 H), 4.10 (dd, J =10.2, 5.6 Hz, 1 H), 4.32 (br dd, J = 9.7,
The Journal of Antibiotics

Synthesis and structure–activity relationships of novel LCM derivatives
Y Wakiyama et al
12
0.5 Hz, 1 H), 4.46 (dd, J = 9.7, 2.7 Hz, 1 H), 5.26 (d, J = 5.6 Hz, 1 H), 7.16
(d, J = 7.8 Hz, 1 H), 7.27–7.34 (m, 2 H).
4
5
6
Schlünzen, F. et al. Structural basis for the interaction of antibiotics with the peptidyl
transferase centre in eubacteria. Nature 413, 814–821 (2001).
Poehlsgaard, J. & Douthwaite, S. The macrolide binding site on the bacterial ribosome.
Curr. Drug Targets Infect. Disord. 2, 67–78 (2002).
Tu, D., Blaha, G., Moore, P. B. & Steitz, T. A. Structures of MLSBK antibiotics bound to
mutated large ribosomal subunits provide a structural explanation for resistance. Cell
121, 257–270 (2005).
7(S)-7-Deoxy-7-(5-morpholinocarbonylthiophen-2-ylthio)lincomycin
(45).
Compound 35 (90.4 mg, 0.21 mmol) and (5-bromothiophen-2-yl)
(morpholino)methanone (69 mg, 0.21 mmol) were treated according to the
similar procedure as described for the preparation of 36 to afford 45
7
8
Hansen, J. L. et al. The structures of four macrolide antibiotics bound to the large
ribosomal subunit. Mol. Cell 10, 117–128 (2002).
29
(100 mg, 76%) as a colorless solid. [α]D +102.9° (c 2.49, MeOH); ESI-MS
Morimoto, S., Takahashi, Y., Watanabe, Y.
& Ōmura, S. Chemical modification
(m/z) 618 (M+H)+ as C27H43N3O7S3; TOF-ESI-HR-MS (M+H)+ calcd for
C27H43N3O7S3: 618.2341, found: 618.2347; 1H NMR (400 MHz, CD3OD)
δ 0.87–0.96 (m, 3 H), 1.25–1.40 (m, 4 H), 1.33 (d, J = 7.0 Hz, 3 H), 1.78–1.89
(m, 1 H), 1.99 (ddd, J = 12.8, 7.9, 4.7 Hz, 1 H), 2.02–2.09 (m, 1 H), 2.11–2.23
(m, 1 H), 2.17 (s, 3 H), 2.37 (s, 3 H), 2.98 (dd, J =10.6, 4.8 Hz, 1 H), 3.22
(dd, J = 8.4, 5.9 Hz, 1 H), 3.58 (dd, J = 10.3, 3.2 Hz, 1 H), 3.66–3.79 (m, 10 H),
4.11 (dd, J = 10.3, 5.6 Hz, 1 H), 4.34 (br dd, J = 9.6, 0.6 Hz, 1 H), 4.42
(dd, J = 9.6, 3.0 Hz, 1 H), 5.29 (d, J = 5.6 Hz, 1 H), 7.18 (d, J = 3.8 Hz, 1 H),
7.32 (d, J = 3.8 Hz, 1 H).
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9
In vitro antibacterial activity
MIC (μg ml− 1) was determined by the agar dilution method, which was
described in Clinical and Laboratory Standards Institute (M7-A5 in 2000). Test
strains of S. pneumoniae and S. pyogenes were subjected to seed culture using
brain heart infusion agar (BHIA; Becton Dickinson and Company, Tokyo,
Japan) and 5% defibrinated horse blood. Test strains of H. influenzae were
subjected to seed culture using sensitivity disk agar-N ‘Nissui’ (SDA; Nissui,
Tokyo, Japan), 5% defibrinated horse blood, 5 μg ml− 1 Hemin and 15 μg ml− 1
NAD. A 5 μl portion of cell suspension of the test strains having about 106
CFU per ml was inoculated into SDA supplemented with 5% defibrinated
horse blood, 5 μg ml− 1 Hemin and 15 μg ml− 1 NAD, and incubated at 37 °C
for 18–22 h. Then, MIC was measured.
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I
23,
19 Hoeksema, H. Octoses from antibiotics. The Upjohn Company, Kalamazoo, Mich.,
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Detroit, p. 9-C (1965)
Docking simulation of the key compound 28
20 Magerlein, B. J., Birkenmeyer, R. D. & Kagan, F. Chemical modification of lincomycin.
Docking simulation was performed with Insight II (Accelrys, San Diego, CA,
USA) using CHARMm force fields. The crystal structure of azithromycin
bound to the 50S ribosomal subunit from Haloarcula marismortui (PDB entry
1M1K)7 was used for the docking template. In preparation for docking
simulation, the azithromycin and the RNA residues other than around the
ligand binding site were removed from the template. In docking simulation, 28
was manually placed in the ligand binding site refer to crystal structure of
CLDM bound to the 50S ribosomal subunit from Haloarcula marismortui (PDB
entry 1YJN),6 and minimized in the template.
Antimicrob. Agents Chemother. 6, 727–736 (1966).
21 Magerlein, B. J.
& Kagan, F. Lincomycin. IX. 7-Thiol and thioamido analogs of
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25 Bannister, B. Modifications of lincomycin involving the carbohydrate portion.
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360–369 (1974).
I 3,
CONFLICT OF INTEREST
The authors declare no conflict of interest.
26 Bannister, B.
& Mydlow, P. K. The S-alkylation of sulphides by an activated
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27 Bannister, B. The S-alkylation of sulphides by an activated carbohydrate epimine
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ACKNOWLEDGEMENTS
We thank Mr A. Tamura, Dr E. Shitara, Dr T. Okutomi, Dr T. Yoshida and
Dr H. Fushimi for encouragement and valuable discussion. We are grateful to
Professor Emeritus Dr M. Konno for supervision through our in-house drug
discovery program in LCM field. We also thank Ms M. Ishii for direction in
intellectual properties, Ms T. Miyara, Ms R. Hiruta, Ms S. Miki and
Ms K. Kaneda for analytical and synthetic chemistry, and Mr Y. Takayama and
Ms K. Yamada for biological studies.
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The Journal of Antibiotics