Article
Figure 2. (A) Eradication of the preformed S. aureus ATCC 29213 biofilms by compound 7d. (B) Images of S. aureus ATCC 29213 treated with
d after staining by crystal violet.
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When the number of carbon atoms increased, octyl and decyl
compounds 4e and 4f were essentially inactive toward the
tested bacteria, which might result from large lipophilicity. The
data strongly indicated that the lengths of the alkyl chains
significantly affected antibacterial activity and moderate lengths
might be helpful for improving antibacterial activities. Inspired
by the improved biological potentiality by incorporating
ATCC 29213 growth with a low concentration of 0.006 mM.
Furthermore, this molecule possessed 114- and 4-fold
antibacterial potency than berberine and norfloxacin in
inhibiting MRSA growth, respectively. Additionally, compound
7d gave a low MIC value (0.013 mM) toward S. aureus and E.
coil, which was more than 2-fold higher than norfloxacin.
Moreover, it showed relatively good inhibitory effect on P.
aeruginosa with a MIC value of 0.051 mM, which was 27 times
more potent than berberine. However, the 3,4-dichlorobenzyl
derivative 7e only exhibited good inhibitory effect (MIC =
0.025 mM) against S. aureus and S. aureus ATCC 29213, which
suggested that the number and location of halogen atoms on
benzyl group would result in different effects on bioactivities.
Encouraged by the enhanced antimicrobial activity through
the incorporation of amino derivatives, compounds 8 and 9
were synthesized to study the impact of amide groups on
biological activities. Unfortunately, all of them abolished the
activity completely or partially, indicating that the introduction
of amides at the N-1 position of benzimidazole nucleus was not
favorable for antibacterial potency. Stimulated by the lipophilic
ester group, tert-butyl ester derivative 10 was synthesized and
tested the antibacterial activity; however, it was not active
toward the tested strains.
2
5
cyclanes, compounds 5a and 5b were synthesized and tested
for their antibacterial potencies, where cyclopropyl derivative
5a was active toward the tested bacteria except for MRSA and
K. pneumoniae. It was highly sensitive to S. aureus, A.
baumannii, E. coli, and S. aureus ATCC 29213 at low
concentrations (0.015−0.030 mM). Moreover, this compound
exerted comparable activity (MIC = 0.004 mM) to norfloxacin
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(
MIC = 0.006 mM) for P. aeruginosa. When cyclopropane was
expanded to cyclohexane, the antibacterial activity of
compound 5b significantly reduced.
Compounds 5c and 5d bearing allyl and propargyl groups
were prepared to investigate the effects of unsaturated bonds
on the antibacterial effect, respectively. Molecules 5c and 5d
both showed a 3-fold decrease in anti-S. aureus with a low MIC
value (0.031 mM) in comparison to berberine (0.081 mM).
To explore the effect of hydrophilic groups on the antibacterial
activity of berberine derivatives, compounds 6a−c were
prepared. Both of them were inactive toward the tested
strains, suggesting that the increase of water solubility was
deleterious to the antibacterial activities.
Biofilm Disruption Evaluation. The biofilm formation of
S. aureus made conventional antibiotics hard to adequately
33−35
attack and destroy infectious biofilm populations.
Thus,
new antibacterial agents inhibiting biofilm formation were
imperative to confront Staphylococcus infection. The ability of
compound 7d to disrupt the preformed biofilm of S. aureus
ATCC 29213 was evaluated. As shown in Figure 2, the 2,4-
dichlorobenzyl derivative 7d could eradicate more than 40%
biofilm of S. aureus ATCC 29213 at concentrations of 4 × and
8 × MIC. Especially, when its concentration was increased to
16 × and 32 × MIC, the disruption of S. aureus ATCC 29213
biofilm exceeded 90%. These results demonstrated that
compound 7d could effectively disrupt bacterial biofilm to
exert antibacterial activity.
Metabolic Activity Evaluation. The alamar blue assay
was conducted to determine the viability of S. aureus ATCC
29213 within the matured biofilm. As shown in Figure S1, the
cell viability of S. aureus ATCC 29213 within biofilm reduced
about 47% in the presence of compound 7d at the
concentration of 8 × MIC, which indicated that active
molecule 7d possessed certain ability to disrupt matured
biofilm and then inhibit the viability of S. aureus ATCC 29213
cells.
The structure and activity relationship (SAR) investigations
were further moved onto the effect of halogen-containing
benzyl groups, which were helpful for increasing lipid solubility
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and membrane penetrability,
and compounds 7a−e were
synthesized. Fluorobenzyl derivative 7a gave relatively low
MIC values (0.014−0.055 mM) against MRSA, S. aureus, S.
aureus ATCC 29213, P. aeruginosa, and A. baumannii, which
were comparable to or even higher than norfloxacin. When the
number of fluorine atom was increased, the antibacterial
potency of compound 7b dropped obviously except that the
inhibitory concentration toward S. aureus ATCC 29213 was
decreased by 8-fold up to 0.007 mM compared to the
fluorobenzyl molecule 7a (MIC = 0.055 mM). It seemed that
the displacement of the fluorine atom (7a) by a chlorine atom
(7c) reduced the activity of the molecule, but surprisingly
enhanced its effect against S aureus (8 times in comparison to
norfloxacin) and A. baumannii (the same as for norfloxacin).
The introduction of more chlorine atoms resulted in
compound 7d, which could effectively suppress S. aureus
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J. Agric. Food Chem. 2021, 69, 7831−7840