DOI: 10.1039/C5CC05159B
ChemComm
aureus and E. coli. To evaluate the propensity of developing
Notes and references
resistance, bacteria grown at the sub-MIC level (MIC/2) were
used for successive MIC assay and the process was repeated for
20 passages. The cationic biocide showed no change in MIC
against both the bacteria even after 20 passages, whereas around
805-fold and 250-fold increase in MIC was observed for
antibiotic norfloxacin against S. aureus and lipopeptide colistin
against E. coli respectively (ESI,† Fig. S8). The above results
thus indicated that bacteria were less prone to develop resistance
We thank Prof. C. N. R. Rao, FRS (JNCASR) for his constant
support and encouragement. Dr. Haldar acknowledges the
50 Department of Science and Technology (DST), Govt. of India,
for Ramanujan fellowship [SR/S2/RJN-43/2009]. Jiaul thanks
JNCASR for Senior Research Fellowship (SRF). SS is grateful to
Sheikh Saqr Laboratory (SSL), JNCASR for postdoctoral
5
fellowship.
55
10 against this type of molecules.
In order to further evaluate the toxicity of the small
aChemical Biology and Medicinal Chemistry Laboratory, New Chemistry
Unit, Jawaharlal Nehru Centre for Advanced Scientific Research, Jakkur,
2208-2556.
60 †Electronic Supplementary Information (ESI) available: Experimental
procedures, synthetic protocols and characterizations of the molecules,
molecules, human embryo kidney cells (HEK 293) were treated
with the most potent molecule 2d for 24 h. The half-maximal
inhibitory concentration (IC50) was found to be 220 g/mL in
15 lactate dehydrogenase (LDH) assay following the manufacturer
protocol (Lactate dehydrogenase activity assay kit, Sigma-
Aldrich, catalog number MAK066). Also, the treated cells,
visualized by LIVE/DEAD staining method, showed green
fluorescence even at 128 g/mL (64 times of MIC) and were
20 similar to the untreated cells whereas cell treated with triton-X
were found to have completely red fluorescence (Fig. 3, ESI,†
Fig. S9). These results thus indicated that these biocides are
indeed non-toxic towards mammalian cells.
figures
of
antibacterial
and
antibiofilm
activity.
See
DOI: 10.1039/b000000x/
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25
Fig. 3. Fluorescence microscopy images of HEK cells after
treatment with small molecule 2d for 24 h and staining with
calcein AM and propidium iodide (PI). (a-c) Nontreated cells
(negative control); (d-f) cells treated with 2d (128 μg/mL); and
30 (g-i) cells treated with 0.1% Triton X (positive control). Scale bar
is 20 μm.
In summary, novel membrane-active amphiphilic small
molecules, developed in a facile and cost-effective way, were
35 highly active towards drug-sensitive and drug-resistance
pathogenic bacteria but were less or non-toxic to human
erythrocytes and human kidney cells. The molecules killed
bacteria mainly by disrupting membrane integrity and hindered
the propensity of developing bacterial resistance. Further, the
40 small molecules dispersed the preformed Gram-positive and
Gram-negative bacterial biofilms and reduced viable bacteria in
biofilms. The structure-activity relationship, demonstrated by
varying the nature of the lipophilic alkyl chain and spacer chain
lengths, emphasized the role of optimum amphiphilicity in
45 developing non-toxic yet potent membrane-active antibacterial
agents.
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