Design, synthesis and antibacterial evaluation of novel AHL analogues

Article abstract of DOI:10.1016/j.bmcl.2013.05.035

Two series of novel AHL analogues were designed, synthesized and evaluated for antibacterial activity under cell membrane conditions in vitro. Analogues 4a-c and 4g-m presented potent activity against Gram-positive bacteria. Especially the analogue 4l exe

Full text of DOI:10.1016/j.bmcl.2013.05.035

Accepted Manuscript
Design, synthesis and antibacterial evaluation of novel AHL analogues
Jing-Li Ren, En Zhang, Xian-Wei Ye, Meng-Meng Wang, Bin Yu, Wen-Hua
Wang, Ya-Zhuo Guo, Hong-Min Liu
PII:
S0960-894X(13)00630-6
DOI:
http://dx.doi.org/10.1016/j.bmcl.2013.05.035
BMCL 20496
Reference:
To appear in:
Bioorganic & Medicinal Chemistry Letters
Received Date:
Revised Date:
Accepted Date:
11 April 2013
8 May 2013
10 May 2013
Please cite this article as: Ren, J-L., Zhang, E., Ye, X-W., Wang, M-M., Yu, B., Wang, W-H., Guo, Y-Z., Liu, H-
M., Design, synthesis and antibacterial evaluation of novel AHL analogues, Bioorganic & Medicinal Chemistry
Letters (2013), doi: http://dx.doi.org/10.1016/j.bmcl.2013.05.035
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Design, synthesis and antibacterial evaluation of novel AHL analogues
Jing-Li Ren, En Zhang, Xian-Wei Ye, Meng-Meng Wang, Bin Yu, Wen-Hua Wang, Ya-Zhuo Guo,
Hong-Min Liu.
School of Pharmaceutical Sciences and New Drug Research & Development Center, Zhengzhou
University, Zhengzhou 450001, P. R. China
Abstract: Two series of novel AHL analogues were designed, synthesized and evaluated for
antibacterial activity under cell membrane conditions in vitro. Analogues 4a-c and 4g-m presented
potent activity against Gram-positive bacteria. Especially the analogue 4l exerted the most potent
inhibition against Bacillus subtilis with MIC50 value of 1.443 μg/ml. To our surprise, analogues
6a-c and 6g showed weak inhibition against Gram-negative bacteria with MIC50 values ranging
from 17.589 to 67.840 μg/ml. This was the first report about synthesis and antibacterial evaluation
in vitro of AHL analogues containing dithioester linkage.
Key words: antibacterial; quorum sensing; dithiocarbamates; AHLs.
Bacterial resistance to the existing drugs is still a global problem, which develops at
increasingly fast speed in hospitals and community settings during these years. Thus research of
more effective antibacterial agents is of considerable significance to public health especially in
developing countries.
Bacterial quorum sensing (QS) refers to a chemical communication system, in which bacteria
use diffusible, small molecular signals namely autoinducers to monitor their local population
densities. QS regulates diverse phenotypes, such as bioluminescence, biofilm formation, virulence
expression and antibiotic production, in various bacteria survived in plants and mammals.1,2
Different kinds of compounds are employed by bacteria as QS signals.3 To Gram-negative bacteria,
N-acylhomoserine lactones (AHLs) (Figure 1,Ⅰand Ⅱ) are the most studied class of autoinducers.
Numerous researches on synthetic analogues (Figure 1, Ⅲ-Ⅴ) and their signaling action
mechanisms
have
been
carried
out
during
these
years.4,5,6,7 The
AHL,
3-oxo-dodecanoyl-L-homoserine lactone (OdDHL, figure 1,Ⅰ) produced by the prevalent
pathogen Pseudomonas aeruginosa, has been shown to induce apoptosis of macrophages and
human breast cancer cell lines but not common ones and even to enhance sensitivity of H630
colon cancer cells to 5-Fluorouracil.8,9,10 Except for the action to eukaryotic cells, OdDHL and
some other synthetic 3-oxo derivatives have been reported to inhibit the growth of Gram-positive
bacteria (Bacillus cereus, Bacillus subtilis and Staphylococcus aureus) selectively in high
concentration range.3 So far, strategies to intercept QS have been considered as potential
anti-infective therapies.
 Corresponding author. Tel.: +86 371 67781739; fax: +86 371 67781739. E-mail address: liuhm@zzu.edu.cn
(H.-M. Liu).

Figure 1. Some natural occurring AHLs Ⅰ-Ⅱ and synthetic quorum sensing molecules Ⅲ-Ⅳand the natural
dithiocarbamate Brassinin Ⅵ.
Dithiocarbamates have received considerable attention for their excellent biological activities
during these years.11,12,13 The natural product brassinin (Figure 1, Ⅵ) was reported benign
antifungal activity and as a moderate inhibitor of a novel cancer immunosuppression target
(Indoleamine 2,3-dioxygenase, IDO).11 Lots of trials have been done in order to obtain more novel
dithocarbamates with good biological activities. Dithiocarbamates were efficiently prepared by
multicomponient one-pot reaction, in which the primary or secondary amines reacted with carbon
disulfide and halides in the presence of weak bases.14,15
The intrinsic liposolubility of natural AHLs enables QS signals to cross cell membranes in
unregulated diffusion way. In considering this key element, we designed to prepare two series of
AHL analogues containing dithioester linkage by introduction of N-phenyl acetamide a benign
antimicrobial pharmacophore16,17 and hydrophobic alkyl chains as side chains respectively. Herein,
in this letter we first reported the synthesis and antibacterial evaluation of AHL analogues
containing dithoester lingkage.
The synthetic route towards target AHL Analogues (4a-m and 6a-g) was shown in Scheme 1.
The starting material L-homoserine lactone hydrochloride 3 and the key intermediates 2a-m were
prepared according to the literature reported methods.16,18 The target analogues were easily
obtained in high yields with the mature reaction conditions developed by our group14,15
.
Specifically, 2a-m reacted with compound 3 and carbon disulfide to afford 4a-m. Compound 3
reacted with commercially available halids 5a-g to synthesize 6a-g. Finally, all the structures of
4a-m and 6a-g were fully characterized by 1H NMR, 13C NMR and HRMS.
Scheme 1. Synthesis of AHL analogues 4a-m and 6a-g. Reagents and conditions: (a) ClCH2COCl, Anhyd.K2CO3,
.
.
Acetone, rt. (b) 2a-m, CS2, Na3PO4 12H2O, Acetone, rt. (c) 5a-g, CS2, Na3PO4 12H2O, Acetone, rt.
Analogues 4a-m and 6a-g were screened against Gram-positive bacteria (Staphylococcus

aureus, Bacillus subtilis) and Gram-negative bacteria (Pseudomonas aeruginosa, Escherichia coli)
in vitro by microdilution broth method with levofloxacin as the positive control. MIC50 values of
the tested compounds were listed in Table 1. It was observed that 4a-m displayed benign
inhibitory action against Gram-positive bacteria and relatively weak inhibition against
Gram-negative bacteria, while 6a-g showed weak inhibition against most of the tested bacteria.
Specifically, compounds 4a-c and 4g-m could potently inhibit the growth of S. aureus with the
MIC50 values ranging from 3.498 to 9.515 μg/ml. Analogues 4h (2-Cl ), 4j (4-OCH3), 4l (3-CF3)
and 4m (3-NO2) showed potent inhibition against B. subtilis with the MIC50 of 9.626, 5.600, 1.443
and 6.289 μg/ml, respectively. It is worth mentioning that compound 4l presented the most
excellent inhibition against B. subtilis with the MIC50 of 1.443 μg/ml. To our surprise, the
analogues 6a-c showed weak inhibitory effect against Gram-negative bacteria with the MIC50
range of 17.589-44.631 μg/ml.
Table 1
Antibacterial activity (MIC50, μg/ml) of compouds 4a-m, 6a-g.
Gram+ bacteria
Gram- bacteria
compound
S. aureus
B. subtilis
>128
>128
30.233
>128
>128
>128
47.907
9.629
>128
5.600
>128
1.443
6.289
>128
>128
>128
>128
>128
>128
>128
<0.25
P. aeruginosa
E. coli
40.488
>128
4a
6.534
3.498
3.691
>128
>128
>128
7.722
5.621
3.489
4.898
9.515
5.807
8.674
>128
>128
>128
>128
>128
50.569
>128
<0.25
>128
>128
4b
4c
54.525
>128
24.859
>128
4d
4e
37.192
>128
>128
4f
>128
4g
48.467
>128
>128
4h
>128
4i
63.765
>128
30.589
20.995
54.211
>128
4j
4k
>128
4l
>128
4m
59.935
31.934
34.557
44.631
>128
12.808
17.589
25.224
>128
6a
6b
6c
6d
>128
6e
>128
>128
6f
>128
>128
6g
>128
67.840
<0.25
Levofloxacina
<0.25
a the control drug for antibacterial assays.
The synthesized compounds showed varying degrees of antibacterial activity against
Gram-positive and Gram-negative bacteria. The analogues with hydrophobic alkyl chains
stimulated the growth of Gram-positive bacteria except for 6f, while against Gram-negative
bacteria they exerted weak inhibitory activity and the activity reduced with the length of

hydrophobic alkyl chains increased. The introduction of substituted N-phenyl acetamides as side
chains contributed to antibacterial activities immensely when compared with hydrophobic alkyl
chains. Analogues with different substituents on phenyl ring exemplified varying degrees of
inhibition against Gram-positive bacteria. Analogues 4l and 4m with strong electron-withdrawing
groups -CF3 and -NO2 as meta-substitutions showed benign antibacterial activities, while no
activity was exerted by 4e with the electron-donating group -CH3 at meta-position. Analogues 4b
(4-Cl), 4c (4-Br), 4i (4-CH3), 4j (2, 4-dichloride), 4h (2-Cl) and 4l (3-CF3) presented potent
inhibition against S. aureus. The antibacterial inhibition of 4b (4-Cl) and 4i (4-CH3) with the
substituents at para-position were better than 4h (2-Cl), 4f (3-Cl) and 4e (3-CH3) respectively. No
activity improvement was exhibited by 4j (2, 4-dichloride) compared to 4b (4-Cl). Compound 4l
(3-CF3) presented the best activity against B. subtilis, and the activity disappeared when the aryl
hydrogen atom was replaced by the electron-donating groups. Compound 4j (2, 4-dichloride) had
better antibacterial activity than compound 4h (2-Cl).
In conclusion, we synthesized two series of novel AHL analogues and evaluated their
antibacterial activity in vitro against Gram-positive and Gram-negative bacteria. The nature of the
side chains and the substitutions on the phenyl ring influenced the antibacterial activities
remarkably. Analogues 6a-g with hydrophobic alkyl chains showed weak inhibition against most
of the tested bacteria. The analogues 4a-m obtained via introduction of different substituted
N-phenyl amides as side chains presented benign antibacterial activity to the tested bacterial
strains. Amongst them, compound 4l presented excellent inbititory effect against B. subtilis
(MIC50 = 1.443 μg/ml), which is under further investigation as lead structure. Further
modifications and SAR study are undergoing and will be reported elsewhere.
Acknowledgments
This work was supported by the National Natural Sciences Foundations of China (No.
8112937, U1204206), China Postdoctoral Science Foundation (No. 20100480857, 201104402)
and New Teachers’ Fund for Doctor Stations, Ministry of Education (No. 20114101120013).
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