Synthesis and antimicrobial evaluation of 1-[(2-substituted phenyl)carbamoyl]naphthalen-2-yl carbamates

Article abstract of DOI:10.3390/molecules21091189

Series of thirteen 1-[(2-chlorophenyl)carbamoyl]naphthalen-2-yl carbamates and thirteen 1-[(2-nitrophenyl)carbamoyl]naphthalen-2-yl carbamates with alkyl/cycloalkyl/arylalkyl chains were prepared and characterized. Primary in vitro screening of the synthesized compounds was performed against Staphylococcus aureus, two methicillin-resistant S. aureus strains, Mycobacterium marinum, and M. kansasii. 1-[(2-Chlorophenyl)carbamoyl]naphthalen-2-yl ethylcarbamate and 1-[(2-nitrophenyl) carbamoyl]naphthalen-2-yl ethylcarbamate showed antistaphylococcal (MICs = 42 μM against MRSA) and antimycobacterial (MICs = 21 μM) activity against the tested strains comparable with or higher than that of the standards ampicillin and isoniazid. In the case of bulkier carbamate tails (R > propyl/isopropyl), the activity was similar (MICs ca. 70 μM). Screening of the cytotoxicity of both of the most effective compounds was performed using THP-1 cells, and no significant lethal effect was observed (LD50 >30 μM). The structure-activity relationships are discussed.

Full text of DOI:10.3390/molecules21091189

molecules
Article
Synthesis and Antimicrobial Evaluation of
1-[(2-Substituted phenyl)carbamoyl]naphthalen-2-yl
Carbamates †
Tomas Gonec 1,*, Sarka Pospisilova 2, Lucie Holanova 3, Josef Stranik 1, Aneta Cernikova 1,
Valeria Pudelkova 2, Jiri Kos 1, Michal Oravec 4, Peter Kollar 3, Alois Cizek 2 and Josef Jampilek 5,
*
1
Department of Chemical Drugs, Faculty of Pharmacy, University of Veterinary and Pharmaceutical Sciences,
Palackeho 1, Brno 61242, Czech Republic; F15334@vfu.cz (J.S.); cernikova.aneta@gmail.com (A.C.);
jurd@email.cz (J.K.)
2
3
Department of Infectious Diseases and Microbiology, Faculty of Veterinary Medicine,
University of Veterinary and Pharmaceutical Sciences, Palackeho 1, Brno 61242, Czech Republic;
sharka.pospisilova@gmail.com (S.P.); valeria.pudelkova@gmail.com (V.P.); cizeka@vfu.cz (A.C.)
Department of Human Pharmacology and Toxicology, Faculty of Pharmacy,
University of Veterinary and Pharmaceutical Sciences, Palackeho 1, Brno 61242, Czech Republic;
lucieholanova9@gmail.com (L.H.); kollarp@vfu.cz (P.K.)
Global Change Research Institute CAS, Belidla 986/4a, Brno 60300, Czech Republic; oravec.m@czechglobe.cz
Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Comenius University, Odbojarov 10,
Bratislava 83232, Slovakia
4
5
*
†
Correspondence: t.gonec@seznam.cz (T.G.); josef.jampilek@gmail.com (J.J.); Tel.: +420-541-562-938 (T.G.)
Preliminary Results Were Presented at the 8th Central European Conference “Chemistry towards Biology”
(CTB-2016), Brno, Czech Republic, 28 August–1 September 2016 (Paper P-18).
Academic Editor: Derek J. McPhee
Received: 24 August 2016; Accepted: 5 September 2016; Published: 7 September 2016
Abstract: Series of thirteen 1-[(2-chlorophenyl)carbamoyl]naphthalen-2-yl carbamates and thirteen
1-[(2-nitrophenyl)carbamoyl]naphthalen-2-yl carbamates with alkyl/cycloalkyl/arylalkyl chains were
prepared and characterized. Primary in vitro screening of the synthesized compounds was performed
against Staphylococcus aureus, two methicillin-resistant S. aureus strains, Mycobacterium marinum, and
M. kansasii. 1-[(2-Chlorophenyl)carbamoyl]naphthalen-2-yl ethylcarbamate and 1-[(2-nitrophenyl)
carbamoyl]naphthalen-2-yl ethylcarbamate showed antistaphylococcal (MICs = 42
MRSA) and antimycobacterial (MICs = 21 M) activity against the tested strains comparable with
or higher than that of the standards ampicillin and isoniazid. In the case of bulkier carbamate tails
R > propyl/isopropyl), the activity was similar (MICs ca. 70 M). Screening of the cytotoxicity of
µM against
µ
(
µ
both of the most effective compounds was performed using THP-1 cells, and no significant lethal
effect was observed (LD50 >30 µM). The structure-activity relationships are discussed.
Keywords: carbamates; hydroxynaphthalene-carboxamides; in vitro antibacterial activity; in vitro
antimycobacterial activity; in vitro cytotoxicity assay; structure-activity relationships
1. Introduction
Infectious diseases represent an increasing worldwide threat. The number of untreatable diseases
decreased after the 1950s due to the introduction of new antimicrobial agents. However, since the
1980s, morbidity has risen again. The increase in the number of new infections is caused by general
immunosuppression, a significant increase in the number of diabetic or HIV-positive patients, and the
development of resistance to commonly used drugs. The resistance of common pathogens to first-choice
drugs increased by up to 100% during the last few decades. Moreover, the resistance of some strains to
second- or third-choice drugs can be found. Development of cross-resistant and multidrug-resistant
Molecules 2016, 21, 1189; doi:10.3390/molecules21091189
www.mdpi.com/journal/molecules

Molecules 2016, 21, 1189
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strains is a serious global problem. Selection of resistant pathogens is especially caused by irrational and
unavailing application of anti-invasive agents in human, veterinary medicine, and in agriculture [ ].
Resistance may complicate the treatment of infections regardless of how mild these infections were
at the early stage [ ]. This increasing resistance highlights the urgency of designing new effective
1–4
5,6
anti-invasive drugs and developing strategies focused on overcoming drug resistance [5–9].
The discovery of salicylanilides dates back to early phenol applications; currently, salicylanilides
are well-known organic compounds exhibiting a broad spectrum of biological activities such as
anthelmintic, antibacterial, antimycobacterial, and antiviral, among others. Salicylanilides have been
described to affect a wide range of targets, although the appropriate mechanism of action responsible
for overall biological activities of these compounds has not been proposed so far. Thus, salicylanilides
seem to be promising candidates for antibacterial agents, which could be a solution to the resistance
challenges [10–17].
In addition, the presence of an amide (–CONH–) or carbamate (–OCONH–) group with
a hydrophobic residue in its close vicinity is characteristic not only of a number of clinically used
drugs [18], but also applied pesticides [19]. These moieties are important functional groups that are
able, due to their electron properties, to interact and bind with a number of enzymes/receptors and,
by means of these target sites, affect the biological response. The properties of the amide and the
carbamate moieties can easily be modified by various substitutions [20,21]. Therefore, the reason for
the widespread occurrence of amides and carbamates among new biologically active compounds is
obvious [22–36].
1-[(2-Substituted phenyl)carbamoyl]naphthalen-2-yl carbamates, described in the present work,
can be considered as cyclic analogues of salicylanilides that have expressed promising results as potential
antibacterial and antimycobacterial agents ([14
carbamates–N-(2-chlorophenyl)-2-hydroxynaphthalene-1-carboxamide (
hydroxynaphthalene-1-carboxamide ( ) showed antimycobacterial or antibacterial activity with
–17], and refs. therein). Pattern compounds of these
1
) and N-(2-nitrophenyl)-2-
2
insignificant cytotoxicity on human cells in the previous screening [30]; therefore, the aim of this
contribution was to describe the preparation of various alkyl, cycloalkyl, and arylalkyl carbamates and
the investigation of their antimicrobial activity.
2. Results and Discussion
2.1. Chemistry
All the studied compounds were prepared according to Scheme 1. In the first step,
N-(2-chlorophenyl)-2-hydroxynaphthalene-1-carboxamide (
naphthalene-1-carboxamide ( ) were synthesized by the microwave-assisted method [30]. In the
second step, a modified method using triethylamine for activation of the phenolic group was used [24].
The addition of activated compounds and to appropriate alkyl, cycloalkyl, and arylalkyl carbamates
1) and N-(2-nitrophenyl)-2-hydroxy-
2
1
2
yielded a series of thirteen 1-[(2-chlorophenyl)carbamoyl]naphthalen-2-yl carbamates 3–15 and thirteen
1-[(2-nitrophenyl)carbamoyl]naphthalen-2-yl carbamates 16–28.
Scheme 1. Synthesis of 1-[(2-chlorophenyl)carbamoyl]naphthalen-2-yl carbamates
1-[(2-nitrophenyl)carbamoyl]naphthalen-2-yl carbamates 16 28. Reagents and conditions: (a) PCl3,
chlorobenzene, MW; (b) TEA, acetonitrile, ambient temperature.
3–15 and
–

Molecules 2016, 21, 1189
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In a number of studies examining the biological activity of potential drugs, the relationship
between lipophilicity or other descriptors and their potency have been investigated. In the current
investigation, the calculated lipophilicity (log P) of the compounds as well as the electronic parameters
and molar volume of R2 substituents (see Table 1) were used to determine if these factors play
a role in their biological activity. All the predicted molecular descriptors were calculated using
the ACD/Percepta ver. 2012 program, see Table 1. The lipophilicity, expressed as log P values,
of the chlorine substituted compounds
1, 3–15 was higher (ranged from 3.94 to 5.75) than that of
the nitro substituted derivatives 16 28 (ranged from 3.58 to 5.55). Lipophilicity increases with
2,
–
lengthening of the alkyl tail. Isopropyl showed a lower lipophilicity value than propyl. Significantly
lower lipophilicity values were calculated for the cycloalkyl derivatives compared with their N-alkyl
isomers. For individual R2 substituents, alkyl/cycloalkyl/arylalkyl tails of the discussed compounds,
and also electronic properties expressed as electronic constants
σ
* were predicted; they ranged from
parameters) of the 2-Cl moiety
(compound 1) and the 2-NO2 moiety (compound 2) were 0.22 and 0.77, respectively. Molar volume
−
0.25 to 0.08. The electronic parameters (expressed as Hammett’s
σ
MV [cm3], a parameter representing the bulk of R2 substituents (i.e., tail length/branching) of each
compound, was also calculated for the hydrophobic tail.
Table 1. Structures of the discussed anilides
1, 2 and carbamates 3–28; calculated values of log P,
electronic constants
σ
*, and molar volume (MV [cm3]) of R2 substituents; in vitro antibacterial activity
(MIC) of the compounds in comparison with the ampicillin (APC) standard; in vitro antimycobacterial
activity (MIC) of the compounds in comparison with the isoniazid (INH) standard and in vitro
cytotoxicity assay (LD50) of the chosen compounds.
MIC [µM]
log P a
a
a
MVR2 [cm3]
R1
R2
Compd.
σ*R2
LD50 [µM]
SA
MRSA SA 630
MRSA 3202
MM MK
1
3
4
5
6
7
8
9
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
−
Et
Pr
5.03
3.94
4.41
4.71
5.47
6.03
6.67
7.19
4.20
4.60
5.03
5.46
5.19
5.75
4.45
3.58
3.96
4.32
5.15
5.71
6.81
7.22
3.80
4.21
4.60
5.05
4.67
5.55
−
–
–
215
21.6
83.5
645
623
602
583
565
83.5
313
303
146
288
271
26
5.27
10.1
628
607
587
569
552
40.6
153
74
860
43.3
83.5
645
623
602
583
565
167
313
605
586
575
541
104
42.1
81.3
628
607
587
569
552
162
610
591
572
562
529
45.8
–
860
43.3
83.5
645
623
602
583
565
167
626
303
586
575
541
52
42.1
81.3
628
607
587
569
552
162
610
591
572
562
529
45.8
–
107
21.6
41.7
80.6
77.8
75.3
72.9
70.6
41.7
78.2
75.6
73.2
71.9
67.6
104
21.0
40.6
78.5
75.9
73.4
71.1
69.0
40.6
76.2
73.8
71.5
70.2
66.1
–
107
21.6
36.5
80.6
77.8
75.3
72.9
70.6
41.7
78.2
75.6
73.2
71.9
67.6
51.9
21.0
40.6
78.5
75.9
73.4
71.1
69.0
40.6
76.2
73.8
71.5
70.2
66.1
–
>30
>30
>30
–
–
–
–
–
>30
–
–
–
–
–
>30
>30
>30
–
–
–
–
–
>30
–
–
–
–
–
–
47.29
63.80
80.31
96.81
113.32
129.83
146.33
64.18
84.10
100.87
117.56
108.00
141.01
–
47.29
63.80
80.31
96.81
113.32
129.83
146.33
64.18
84.10
100.87
117.56
108.00
141.01
–
−0.11
−0.12
−0.25
−0.23
−0.25
−0.23
−0.23
−0.19
−0.20
−0.15
−0.14
0.08
Bu
Pen
Hex
Hep
Oct
10
11
12
13
14
15
2
16
17
18
19
20
21
22
23
24
25
26
27
28
APC
INH
i-Pr
c-Pent
c-Hex
c-Hep
PhEt
PhBu
−
Cl
−0.21
NO2
NO2
NO2
NO2
NO2
NO2
NO2
NO2
NO2
NO2
NO2
NO2
NO2
NO2
–
–
Et
Pr
Bu
−0.11
−0.12
−0.25
−0.23
−0.25
−0.23
−0.23
−0.19
−0.20
−0.15
−0.14
0.08
Pen
Hex
Hep
Oct
i-Pr
c-Pent
c-Hex
c-Hep
PhEt
PhBu
72
281
132
5.72
–
−0.21
–
–
−
–
–
467
29.2
–
a
calculated using ACD/Percepta ver. 2012 (Advanced Chemistry Development, Toronto, ON, Canada);
SA = S. aureus ATCC 29213, MRSA = clinical isolates of methicillin-resistant S. aureus SA 630 and 3202 (National
Institute of Public Health, Prague, Czech Republic); MM = M. marinum CAMP 5644, MK = M. kansasii DSM 44162.
2.2. In Vitro Antibacterial Susceptibility Testing
The in vitro antibacterial activity of the discussed compounds was evaluated against two clinical
isolates of methicillin-resistant Staphylococcus aureus (MRSA) and S. aureus ATCC 29213 as a reference

Molecules 2016, 21, 1189
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and quality control strain. All the compounds showed only moderate or negligible activity, except
(R1 = Cl) and 16 (R1 = NO2). The activity of both ethylcarbamates
ethylcarbamates and 16 was
comparable with that of the standard ampicillin; furthermore, the effect of compound 16 against
S. aureus was ca. 4-fold higher in comparison with that of the chloro derivative . In addition,
nitrated propylcarbamate 17 showed ca. 8-fold higher activity than the corresponding chlorinated
compound , and nitro-anilide had ca. 8-fold higher effectivity against S. aureus than chloro-anilide 1.
3
3
3
4
2
The observation that anilides substituted by the nitro moiety on the anilide ring showed higher
potency only against S. aureus and not against MRSA strains in comparison with chlorine substituted
derivatives was also described by Pauk et al. [37]. It can be supposed that nitro derivatives may
interact by hydrogen bonding with specific biological structures in S. aureus (that are not present in
MRSA strains). These interactions are limited only for spatially small molecules, such as anilide and
ethyl-, propyl-, and isopropylcarbamate, while carbamates with bulkier (long or branched) tails are
not capable of these interactions. This fact was also confirmed by Zadrazilova et al., where short
ethylcarbamates demonstrated higher bacteriostatic activity against various Staphylococcus strains
than longer (C9–C12) alkylcarbamates [16]. Due to the moderate activity of the rest of the compounds,
no thorough structure-activity relationships could be established. Nevertheless, in general, it can be
stated that the activity is influenced by similar factors/parameters as those discussed below.
2.3. In Vitro Antimycobacterial Evaluation
The evaluation of the in vitro antimycobacterial activity of the compounds was performed
against Mycobacterium marinum CAMP 5644 (MM) and M. kansasii DSM 44162 (MK), see Table 1.
To lower risks and make manipulation in the laboratory easier, surrogate model pathogens for
M. tuberculosis can be used in laboratory studies. Mycobacterium marinum is very closely related
to M. tuberculosis and is the cause of tuberculosis-like infections in poikilothermic organisms, especially
frogs and fish. M. marinum is a good model for study especially because of the lower risk for
laboratory workers, genetic relatedness, and pathology similar to human tuberculosis [38]. However,
because of M. tuberculosis, the pathogenic role of nontuberculous mycobacteria (NTM) in humans was
underestimated for a long time [39]. M. kansasii, the most virulent of the NTM, causes nontuberculous
mycobacterial lung infections that are very common nowadays and can be indistinguishable from
tuberculosis [40]. Therefore, additionally M. kansasii was chosen as a model species for screening of
prospective antimycobacterial drugs to control mycobacterial diseases. The activity of the compounds
was expressed as the minimum inhibitory concentration (MIC) that is defined for mycobacteria as
a 90% or greater (IC90) reduction of growth in comparison with the control [41].
It can be stated that the potency of almost all discussed compounds is the same against both
mycobacterial strains (see Table 1). Similarly, as mentioned above, ethylcarbamates
and 16 (R1 = NO2) showed the highest antimycobacterial activity (MICs = 21
M). In addition,
propylcarbamates M).
(R1 = Cl) and 17 (R2 = NO2) expressed substantial activity (MICs ca. 40
3
(R1 = Cl)
µ
4
µ
The dependences of the antimycobacterial activity expressed as log (1/MIC) on the lipophilicity
(log P) of the compounds as well as on the bulkiness/molar volume (MV [cm3]) of individual
alkyl/cycloalkyl/arylalkyl chains are illustrated in Figure 1A,B. Practically the same structure-activity
relationships can be found for both series; therefore they are illustrated only for M. kansasii. The activity
rapidly decreases with lipophilicity and with substituent bulkiness, increasing up to log P ca. 4.4
or MV ca. 80 cm3 (R2 = butyl or cyclopentyl), respectively, and then remains practically constant
(or increases insignificantly) with increasing lipophilicity/bulkiness (see Figure 1A,B). For example,
correlation factor r = 0.9919, n = 10 (compounds
on the bulkiness of the R2 substituent can be calculated. It seems that the activity is also secondarily
influenced by the electronic properties of R2 substituents expressed as electronic constants
*, because
it can be stated that the more the * values approximate 0.11 (i.e., the less electron donor properties
are), the higher the activity is (see Figure 2).
3–5, 10, 11, 16–18, 23, 24) of the dependence of activity
σ
σ
−

Molecules 2016, 21, 1189
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B
4.8
4.7
4.6
4.5
4.4
4.3
4.2
4.1
4.0
3.9
A
4.8
4.7
4.6
4.5
4.4
4.3
4.2
4.1
4.0
3.9
40 50 60 70 80 90 100 110 120 130 140 150 160
3.0
3.5
4.0
4.5
5.0
5.5
6.0
6.5
7.0
7.5
MV [cm3]
log P
comp.3-9 comp.10-15 comp.16-22 comp.23-28
comp.1 comp.3-9 comp.10-15 comp.2 comp.16-22 comp.23-28
Figure 1. Dependence of the in vitro antimycobacterial activity against M. kansasii DSM 44162 log
(1/MIC [M]) of the tested compounds on lipophilicity, expressed as log P (
A) and bulkiness/molar
volume (MV [cm3]) of individual R2 substituents (B).
Bu (5/18) cPent (11/24) cHex (12/25) cHep (13/26) Pr (4/17) Et (3/16)
σ* = -0.25 -0.20 -0.15 -0.14 -0.12 -0.11
Figure 2. Increase of the general antimicrobial activity owing to the R2 substituent in dependence on
the electronic properties are expressed as electronic constants σ*.
In the previous studies [24,42], the most potent carbamates were substituted by a longer tail, which
is connected with their surface activity. In this case, no dependence of antibacterial/antimycobacterial
effects on surface activity was observed. As ethylcarbamates showed the highest activity in the
discussed screening and propyl- and isopropyl-carbamates showed medium activity, it seems
that steric requirements (spatially small molecules) play a significant role for a good biological
effect. Similar facts were recently described, for example, by Kratky et al., where carbamates with
spatially bulky tails expressed significantly lower activity than short-tail carbamates [43]. On the
other hand, the blocking of the phenolic moiety by the carbamate group significantly increased
the antimicrobial effect of these compounds. The mode of action (targets) of these compounds is
questionable; however, due to the structural similarity with salicylanilides, interactions with vital
enzymes, energy metabolism, as well as disturbing of the membrane architecture of prokaryotic cells
may be supposed [10,11,13–17,32,33,36,44].
2.4. In Vitro Cytotoxicity Assay
The preliminary in vitro screening of the cytotoxicity of the compounds was performed using
the human monocytic leukemia THP-1 cell line. The cytotoxicity was evaluated as the LD50 value
(LD50—lethal dose to 50% of the cell population), see Table 1. A compound is considered cytotoxic
when it demonstrates a toxic effect on cells at concentrations up to 10
tested concentration that was used for the toxicity assay was 3-fold this value. Treatment with 30
compounds did not lead to lethal effects on the THP-1 cells. Based on these observations, it can
be concluded that the most potent compounds and 16 can be considered as promising agents for
µM [45], and the highest
µM
3
subsequent design of novel antibacterial and antimycobacterial agents.
3. Experimental Section
3.1. General Information
All reagents were purchased from Aldrich (Sigma-Aldrich, St. Louis, MO, USA), and Alfa
(Alfa-Aesar, Ward Hill, MA, USA). TLC experiments were performed on alumina-backed silica gel 60
F254 plates (Merck, Darmstadt, Germany). The plates were illuminated under UV light (254 nm) and
evaluated in iodine vapour. The melting points were determined on a Kofler hot-plate apparatus HMK
(Franz Kustner Nacht KG, Dresden, Germany) and are uncorrected. Infrared (IR) spectra were recorded

Molecules 2016, 21, 1189
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on a Smart MIRacle
™
ATR ZnSe for Nicolet
™
Impact 410 FT-IR spectrometer (Thermo Scientific,
−1
West Palm Beach, FL, USA). The spectra were obtained by the accumulation of 256 scans with a 2 cm
resolution in the region of 4000–650 cm−1. All 1H- and 13C-NMR spectra were recorded on a JEOL
ECZR 400 MHz NMR spectrometer (400 MHz for 1H and 100 MHz for 13C, JEOL, Tokyo, Japan) in
1
DMSO-d6. H and 13C chemical shifts (
δ) are reported in ppm. High-resolution mass spectra were
measured using a high-performance liquid chromatograph Dionex UltiMate® 3000 (Thermo Scientific)
coupled with a LTQ Orbitrap XL Hybrid Ion Trap-Orbitrap Fourier Transform Mass Spectrometer
™
(Thermo Scientific) with injection into HESI II in the positive mode. The lipophilicity (log P) of the
final compounds, the surface tension, and the molar volume of R substituents were predicted using
ACD/Percepta ver. 2012 (Advanced Chemistry Development, Inc., Toronto, ON, Canada).
3.2. Synthesis
General Procedure for Synthesis of Carbamates 3–28
The synthetic pathway and characterization of N-(2-chlorophenyl)-2-hydroxynaphthalene-
1-carboxamide (
1) and N-(2-nitrophenyl)-2-hydroxynaphthalene-1-carboxamide (2) were described
recently by Gonec et al. [30]. Anilides
1
and (1.0 mmol) and triethylamine (1.1 mmol) were dissolved
2
in dry acetonitrile (10 mL). The solution of the appropriate alkyl isocyanate (1.2 mmol) in acetonitrile
(5 mL) was added in four portions within 2 h, and the reacting mixture was stirred for 24 h at ambient
temperature. The solvent was evaporated under reduced pressure, and the solid residue was washed
with methanol and ethyl acetate to give pure product. All the studied compounds are presented
in Table 1.
1-[(2-Chlorophenyl)carbamoyl]naphthalen-2-yl ethylcarbamate (
3264, 3214, 1718, 1675, 1540, 1523, 1510, 1481, 1435, 1300, 1256, 1229, 1006, 826, 741, 723, 688, 667;
1H-NMR (DMSO-d6)
: 10.14 (s, 1H), 7.99–8.06 (m, 3H), 7.88 (t, J = 5.5 Hz, 1H), 7.82 (dd, J = 8.2 Hz,
3
). Yield 33%; Mp 137–139 ◦C; IR (cm−1):
δ
1.3 Hz, 1H), 7.64 (ddd, J = 8.2 Hz, 6.9 Hz, 1.0 Hz, 1H), 7.55–7.59 (m, 2H), 7.43 (td, J = 7.7 Hz, 1.6 Hz,
1H), 7.42 (d, J = 9.0 Hz, 1H), 7.30 (td, J = 7.8 Hz, 1.3 Hz, 1H), 3.08–3.15 (m, 2H), 1.08 (t, J = 7.3 Hz, 3H);
13C-NMR (DMSO-d6),
δ: 164.33, 153.97, 142.20, 134.65, 130.54, 130.31, 129.77, 129.60, 128.40, 128.04,
127.58, 127.30, 127.43, 127.28, 126.60, 125.78, 124.67, 122.62, 35.42, 14.83; HR-MS: for C20H16O3N2Cl
[M + H]+ calculated 367.08440 m/z, found 367.08545 m/z.
1-[(2-Chlorophenyl)carbamoyl]naphthalen-2-yl propylcarbamate (4
). Yield 26%; Mp 132–134 ◦C; IR (cm−1):
3259, 3224, 1716, 1659, 1546, 1526, 1510, 1484, 1441, 1298, 1259, 1232, 1060, 991, 809, 741, 694; 1H-NMR
(DMSO-d6) δ: 10.10 (s, 1H), 7.99–8.05 (m, 3H), 7.89 (t, J = 5.7 Hz, 1H), 7.83 (dd, J = 8.2 Hz, 1.3 Hz, 1H),
7.64 (ddd, J = 8.2 Hz, 6.9 Hz, 1.0 Hz, 1H), 7.55–7.59 (m, 2H), 7.42 (td, J = 7.7 Hz, 1.6 Hz, 1H), 7.41 (d,
J = 9.0 Hz, 1H), 7.30 (td, J = 7.8 Hz, 1.3 Hz, 1H), 3.04 (q, J = 6.4 Hz, 2H), 1.47 (sx, J = 7.2 Hz, 2H), 0.86 (t,
J = 7.6 Hz, 3H); 13C-NMR (DMSO-d6),
δ: 164.33, 154.20, 145.23, 134.67, 130.56, 130.31, 129.81, 129.59,
128.33, 128.04, 127.48, 127.42, 127.30, 127.28, 126.60, 125.78, 124.67, 122.62, 42.35, 22.45, 11.17; HR-MS:
for C21H18O3N2Cl [M + H]+ calculated 381.10005 m/z, found 381.10123 m/z.
1-[(2-Chlorophenyl)carbamoyl]naphthalen-2-yl butylcarbamate (
5
). Yield 55%; Mp 156–159 ◦C; IR (cm−1):
: 10.08
3258, 3228, 1724, 1665, 1553, 1526, 1506, 1479, 1439, 1232, 1007, 818, 751; 1H-NMR (DMSO-d6)
δ
(s, 1H), 7.99–8.05 (m, 3H), 7.88 (t, J = 5.5 Hz, 1H), 7.85 (dd, J = 8.2 Hz, 1.3 Hz, 1H), 7.64 (ddd, J = 8.2 Hz,
6.9 Hz, 1.0 Hz, 1H), 7.55–7.59 (m, 2H), 7.42 (td, J = 7.7 Hz, 1.6 Hz, 1H), 7.41 (d, J = 9.0 Hz, 1H), 7.30
(td, J = 7.8 Hz, 1.3 Hz, 1H), 3.08 (q, J = 6.4 Hz, 2H), 1.44 (qi, J = 7.2 Hz, 2H), 1.28 (sx, J = 7.3 Hz, 2H),
0.85 (t, J = 7.3 Hz, 3H); 13C-NMR (DMSO-d6),
δ: 164.31, 154.19, 145.23, 134.67, 130.57, 130.31, 129.84,
129.60, 128.20, 128.05, 127.41, 127.32, 127.31, 127.22, 126.59, 125.79, 124.65, 122.60, 40.26, 31.32, 19.37,
13.63; HR-MS: for C22H20O3N2Cl [M + H]+ calculated 395.11570 m/z, found 395.11639 m/z.
1-[(2-Chlorophenyl)carbamoyl]naphthalen-2-yl pentylcarbamate (6). Yield 36%; Mp 137–138 ◦C; IR (cm−1):
1
3227, 3215, 1724, 1669, 1550, 1521, 1508, 1480, 1434, 1233, 1037, 997, 910, 817, 746, 686; H-NMR
(DMSO-d6) δ: 10.08 (s, 1H), 7.99–8.05 (m, 3H), 7.88 (t, J = 5.5 Hz, 1H), 7.85 (dd, J = 8.2 Hz, 1.3 Hz, 1H),

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7.64 (ddd, J = 8.2 Hz, 6.9 Hz, 1.0 Hz, 1H), 7.55–7.59 (m, 2H), 7.39–7.44 (m, 2H), 7.30 (td, J = 7.8 Hz,
1.3 Hz, 1H), 3.07 (q, J = 6.7 Hz, 2H), 1.45 (qi, J = 7.0 Hz, 2H), 1.24–1.28 (m, 4H), 0.84 (t, J = 7.0 Hz, 3H);
13C-NMR (DMSO-d6),
δ: 164.39, 154.24, 145.26, 134.68, 130.62, 130.36, 129.90, 129.65, 128.27, 128.10,
127.46, 127.40, 127.37, 127.31, 126.64, 125.87, 124.70, 122.65, 40.59, 28.91, 28.42, 21.87, 13.92; HR-MS: for
C23H22O3N2Cl [M + H]+ calculated 409.13135 m/z, found 409.13303 m/z.
1-[(2-Chlorophenyl)carbamoyl]naphthalen-2-yl hexylcarbamate (
2954, 2924, 1725, 1673, 1587, 1525, 1516, 1463, 1440, 1301, 1241, 1210, 1037, 995, 813, 756, 747, 684;
1H-NMR (DMSO-d6)
: 10.08 (s, 1H), 7.99–8.05 (m, 3H), 7.88 (t, J = 5.5 Hz, 1H), 7.85 (dd, J = 8.2 Hz,
7
). Yield 50%; Mp 103–106 ◦C; IR (cm−1):
δ
1.3 Hz, 1H), 7.64 (ddd, J = 8.2 Hz, 6.9 Hz, 1.0 Hz, 1H), 7.55–7.59 (m, 2H), 7.39–7.44 (m, 2H), 7.30 (td,
J = 7.8 Hz, 1.3 Hz, 1H), 3.07 (q, J = 6.4 Hz, 2H), 1.45 (qi, J = 7.0 Hz, 2H), 1.20–1.31 (m, 6H), 0.85 (t,
J = 6.9 Hz, 3H); 13C-NMR (DMSO-d6),
δ: 164.39, 154.26, 145.26, 134.70, 130.63, 130.37, 129.93, 129.66,
128.25, 128.13, 127.46, 127.39, 127.36, 127.30, 126.66, 125.88, 124.71, 122.65, 40.64, 31.03, 29.21, 25.92,
22.07, 13.96; HR-MS: for C24H24O3N2Cl [M + H]+ calculated 423.14700 m/z, found 423.14767 m/z.
1-[(2-Chlorophenyl)carbamoyl]naphthalen-2-yl heptylcarbamate (
3279, 3227, 2956, 2933, 2923, 1720, 1662, 1538, 1520, 1479, 1463, 1438, 1296, 1266, 1253, 1234, 1192, 1060,
986, 911, 816, 750; 1H-NMR (DMSO-d6)
: 10.08 (s, 1H), 7.99–8.05 (m, 3H), 7.88 (t, J = 5.5 Hz, 1H), 7.86
8
). Yield 66%; Mp 89–90 ◦C; IR (cm−1):
δ
(dd, J = 8.2 Hz, 1.3 Hz, 1H), 7.64 (ddd, J = 8.2 Hz, 6.9 Hz, 1.0 Hz, 1H), 7.55–7.59 (m, 2H), 7.39–7.44
(m, 2H), 7.30 (td, J = 7.8 Hz, 1.3 Hz, 1H), 3.07 (q, J = 6.4 Hz, 2H), 1.45 (qi, J = 7.0 Hz, 2H), 1.20–1.29
(m, 8H), 0.85 (t, J = 6.9 Hz, 3H); 13C-NMR (DMSO-d6),
δ: 164.42, 154.29, 145.28, 134.71, 130.65, 130.39,
129.96, 129.68, 128.27, 128.14, 127.48, 127.41, 127.39, 127.31, 126.67, 125.90, 124.73, 122.67, 40.65, 31.26,
29.26, 28.48, 26.22, 22.11, 14.02; HR-MS: for C25H26O3N2Cl [M + H]+ calculated 437.16265 m/z, found
437.16431 m/z.
1-[(2-Chlorophenyl)carbamoyl]naphthalen-2-yl octylcarbamate (
3297, 3286, 2924, 2851, 1724, 1673, 1584, 1548, 1540, 1509, 1467, 1441, 1434, 1299, 1246, 1215, 1160, 993,
827, 756, 747, 734, 677; 1H-NMR (DMSO-d6)
: 10.08 (s, 1H), 7.99–8.05 (m, 3H), 7.88 (t, J = 5.5 Hz, 1H),
9
). Yield 52%; Mp 115–116 ◦C; IR (cm−1):
δ
7.85 (dd, J = 8.2 Hz, 1.3 Hz, 1H), 7.64 (ddd, J = 8.2 Hz, 6.9 Hz, 1.0 Hz, 1H), 7.54–7.59 (m, 2H), 7.39–7.43
(m, 2H), 7.30 (td, J = 7.8 Hz, 1.3 Hz, 1H), 3.07 (q, J = 6.4 Hz, 2H), 1.45 (qi, J = 7.0 Hz, 2H),1.21–1.30 (m,
10H), 0.86 (t, J = 6.9 Hz, 3H); 13C-NMR (DMSO-d6),
δ: 164.33, 154.19, 145.23, 134.68, 130.57, 130.33,
129.87, 129.60, 128.22, 128.07, 127.40, 127.35, 127.33, 127.22, 126.61, 125.81, 124.67, 122.60, 40.59, 31.25,
29.20, 28.73, 28.64, 26.21, 22.10, 13.96; HR-MS: for C26H28O3N2Cl [M + H]+ calculated 451.17830 m/z,
found 451.18018 m/z.
◦
1-[(2-Chlorophenyl)carbamoyl]naphthalen-2-yl isoproylcarbamate (10). Yield 39%; Mp 167–169 C; IR (cm−1):
3249, 3216, 1971, 1716, 1668, 1581, 1544, 1530, 1510, 1481, 1462, 1439, 1324, 1300, 1256, 1234, 1066, 1022,
962, 910, 820, 742, 697; 1H-NMR (DMSO-d6)
δ: 10.09 (s, 1H), 7.99–8.06 (m, 3H), 7.86 (t, J = 5.5 Hz, 1H),
7.84 (dd, J = 8.2 Hz, 1.3 Hz, 1H), 7.64 (ddd, J = 8.2 Hz, 6.9 Hz, 1.0 Hz, 1H), 7.56–7.59 (m, 2H), 7.43 (td,
J = 7.7 Hz, 1.6 Hz, 1H), 7.41 (d, J = 9.0 Hz, 1H), 7.30 (td, J = 7.8 Hz, 1.3 Hz, 1H), 3.69 (sx, J = 6.6 Hz, 1H),
1.12 (d, J = 6.6 Hz, 6H); 13C-NMR (DMSO-d6),
δ: 164.42, 153.41, 145.28, 134.70, 130.60, 130.37, 129.90,
129.65, 128.34, 128.10, 127.49, 127.48, 127.36, 127.35, 126.67, 125.85, 124.70, 122.71, 42.91, 22.40; HR-MS:
for C21H18O3N2Cl [M + H]+ calculated 381.10005 m/z, found 381.10123 m/z.
◦
1-[(2-Chlorophenyl)carbamoyl]naphthalen-2-yl cyclopentylcarbamate (11). Yield 60%; Mp 135–137 C;
IR (cm−1): 3224, 2951, 1713, 1659, 1544, 1525, 1509, 1485, 1467, 1440, 1290, 1256, 1233, 1151, 1060,
1052, 1013, 911, 815, 784, 740, 688, 680; 1H-NMR (DMSO-d6)
δ: 10.09 (s, 1H), 7.99–8.05 (m, 3H), 7.93 (d,
J = 6.9 Hz, 1H), 7.83 (dd, J = 8.2 Hz, 1.3 Hz, 1H), 7.64 (ddd, J = 8.2 Hz, 6.9 Hz, 1.0 Hz, 1H), 7.55–7.59
(m, 2H), 7.39–7.43 (m, 2H), 7.30 (td, J = 7.8 Hz, 1.3 Hz, 1H), 3.87 (sx, J = 5.9 Hz, 1H), 1.76–1.85 (m, 2H),
1.60–1.70 (m, 2H), 1.46–1.55 (m, 4H); 13C-NMR (DMSO-d6),
δ: 164.39, 153.66, 145.26, 134.69, 130.58,
130.34, 129.84, 129.62, 128.44, 128.07, 127.56, 127.43, 127.33, 127.33, 126.69, 125.81, 124.88, 122.74, 52.49,
32.14, 23.91; HR-MS: for C23H20O3N2Cl [M + H]+ calculated 409.13135 m/z, found 409.13258 m/z.

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1-[(2-Chlorophenyl)carbamoyl]naphthalen-2-yl cyclohexylcarbamate (12). Yield 34%; Mp 129–132 ◦C;
IR (cm−1): 3245, 3218, 2931, 2854, 1717, 1664, 1544, 1525, 1510, 1479, 1463, 1436, 1317, 1295, 1272,
1254, 1229, 1150, 1061, 1027, 1016, 911, 816, 778, 747, 682; 1H-NMR (DMSO-d6)
δ: 10.08 (s, 1H), 7.99–8.05
(m, 3H), 7.86 (d, J = 7.7 Hz, 1H), 7.84 (dd, J = 8.2 Hz, 1.3 Hz, 1H), 7.64 (ddd, J = 8.2 Hz, 6.9 Hz,
1.0 Hz, 1H), 7.55–7.59 (m, 2H), 7.39–7.45 (m, 2H), 7.30 (td, J = 7.8 Hz, 1.3 Hz, 1H), 3.36–3.44 (m, 1H),
1.77–1.87 (m, 2H), 1.66–1.72 (m, 2H), 1.53–1.58 (m, 1H), 1.21–1.29 (m, 4H), 1.07–1.15 (m, 1H); 13C-NMR
(DMSO-d6), δ: 164.38, 153.39, 145.27, 134.69, 130.56, 130.35, 129.85, 129.61, 128.38, 128.07, 127.50, 127.43,
127.32, 127.31, 126.67, 125.80, 124.68, 122.68, 49.99, 32.50, 25.12, 24.60; HR-MS: for C24H22O3N2Cl
[M + H]+ calculated 423.14700 m/z, found 423.14818 m/z.
◦
1-[(2-Chlorophenyl)carbamoyl]naphthalen-2-yl cycloheptylcarbamate (13). Yield 60%; Mp 166–168 C;
IR (cm−1): 3295, 3204, 2919, 2855, 1732, 1708, 1656, 1582, 1544, 1521, 1506, 1475, 1441, 1429, 1342,
1291, 1244, 1217, 1205, 1160, 1039, 1008, 991, 824, 772, 746, 712, 688; 1H-NMR (DMSO-d6)
δ: 10.04 (s,
1H), 7.99–8.05 (m, 3H), 7.90 (d, J = 8.2 Hz, 1H), 7.85 (dd, J = 8.2 Hz, 1.3 Hz, 1H), 7.64 (ddd, J = 8.2 Hz,
6.9 Hz, 1.0 Hz, 1H), 7.55–7.59 (m, 2H), 7.39–7.45 (m, 2H), 7.30 (td, J = 7.8 Hz, 1.3 Hz, 1H), 3.52–3.60 (m,
1H), 1.80–1.86 (m, 2H), 1.43–1.65 (m, 8H), 1.32–1.42 (m, 2H); 13C-NMR (DMSO-d6),
δ: 164.36, 153.29,
145.29, 134.68, 130.57, 130.34, 129.87, 129.60, 128.24, 128.06, 127.43, 127.34, 127.32, 127.25, 126.66, 125.80,
124.67, 122.70, 52.17, 34.35, 27.82, 23.58; HR-MS: for C25H24O3N2Cl [M + H]+ calculated 437.16265 m/z,
found 437.16342 m/z.
1-[(2-Chlorophenyl)carbamoyl]naphthalen-2-yl (2-phenylethyl)carbamate (14). Yield 38%; Mp 115–118 ◦C;
IR (cm−1): 3316, 3221, 1731, 1661, 1583, 1538, 1511, 1467, 1443, 1307, 1248, 1229, 1221, 1200, 1162, 1059,
1031, 984, 817, 751, 739, 698; 1H-NMR (DMSO-d6)
δ: 10.13 (s, 1H), 7.99–8.06 (m, 4H), 7.84 (dd, J = 8.1 Hz,
1.4 Hz, 1H), 7.64 (ddd, J = 8.2 Hz, 6.9 Hz, 1.0 Hz, 1H), 7.55–7.59 (m, 2H), 7.43 (td, J = 7.7 Hz, 1.6 Hz,
1H), 7.37 (d, J = 9.1 Hz, 1H), 7.26–7.32 (m, 4H), 7.21–7.23 (m, 2H), 3.28–3.34 (m, 2H), 2.78 (t, J = 6.4 Hz,
2H); 13C-NMR (DMSO-d6),
δ: 164.33, 154.13, 145.16, 139.13, 134.67, 130.57, 130.32, 129.81, 129.63, 128.70,
128.35, 128.28, 128.07, 127.50, 127.47, 127.33, 127.27, 126.55, 126.14, 125.82, 124.69, 122.55, 42.21, 35.23;
HR-MS: for C26H20O3N2Cl [M + H]+ calculated 445.13135, found 445.13208 m/z.
1-[(2-Chlorophenyl)carbamoyl]naphthalen-2-yl (4-phenylbutyl)carbamate (15). Yield 58%; Mp 132–134 ◦C;
IR (cm−1): 3220, 3024, 2938, 1719,1652, 1582, 1520, 1505, 1478, 1462, 1431, 1290, 1256, 1232, 1060, 985,
822, 760, 745, 699; 1H-NMR (DMSO-d6)
δ: 10.10 (s, 1H), 7.99–8.05 (m, 3H), 7.90 (t, J = 5.5 Hz, 1H), 7.83
(dd, J = 8.2 Hz, 1.3 Hz, 1H), 7.63 (ddd, J = 8.2 Hz, 6.9 Hz, 1.0 Hz, 1H), 7.54–7.59 (m, 2H), 7.36–7.43
(m, 3H), 7.24–7.31 (m, 3H), 7.15–7.19 (m, 2H), 3.10 (q, J = 6.0 Hz, 2H), 2.55 (t, J = 7.3 Hz, 2H), 1.59
(qi, J = 7.3 Hz, 2H), 1.48 (qi, J = 7.3 Hz, 2H); 13C-NMR (DMSO-d6),
δ: 164.34, 154.19, 145.22, 142.09,
134.68, 130.57, 130.32, 129.83, 129.62, 128.31, 128.23, 128.22, 128.07, 127.44, 127.42, 127.32, 127.26, 126.61,
125.81, 125.64, 124.68, 122.62, 40.39, 34.80, 28.88, 28.19; HR-MS: for C28H24O3N2Cl [M + H]+ calculated
473.16265, found 473.16342 m/z.
1-[(2-Nitrophenyl)carbamoyl]naphthalen-2-yl ethylcarbamate (16). Yield 40%; Mp 157–159 ◦C; IR (cm−1):
3320, 3216, 2963, 2917, 2843, 1743, 1705, 1656, 1645, 1608, 1580, 1532, 1504, 1462, 1432, 1357, 1341,
1294, 1270, 1250, 1216, 1199, 1162, 1144, 1079, 1034, 993, 822, 791, 779, 732, 667; 1H-NMR (DMSO-d6)
δ
: 10.94 (s, 1H), 8.07 (d, J = 9.1 Hz, 1H), 8.01–8.05 (m, 3H), 7.71–7.80 (m, 3H), 7.65 (ddd, J = 8.2 Hz,
6.9 Hz, 1.4 Hz, 1H), 7.59 (ddd, J = 8.2 Hz, 6.9 Hz, 1.4 Hz, 1H), 7.41–7.48 (m, 2H), 3.04–3.11 (m, 2H), 1.03
(t, J = 7.1 Hz, 3H); 13C-NMR (DMSO-d6),
: 164.12, 153.71, 145.56, 142.98, 133.89, 130.62, 130.51, 130.33,
δ
130.26, 128.10, 127.42, 125.91, 125.82, 125.70, 124.96, 124.90, 124.43, 122.62, 35.39, 14.75; HR-MS: for
C20H16O5N3 [M + H]+ calculated 378.10845, found 378.10934 m/z.
1-[(2-Nitrophenyl)carbamoyl]naphthalen-2-yl propylcarbamate (17). Yield 75%; Mp 153–155 ◦C; IR (cm−1):
3298, 3223, 2960, 2930, 2873, 1727, 1717, 1662, 1589, 1537, 1520, 1505, 1488, 1463, 1440, 1353, 1285, 1257,
1224, 1146, 1106, 1051, 996, 981, 914, 865, 818, 779, 731, 666; 1H-NMR (DMSO-d6)
δ: 10.93 (s, 1H), 8.07
(d, J = 9.1 Hz, 1H), 8.01–8.05 (m, 3H), 7.72–7.82 (m, 3H), 7.65 (ddd, J = 8.2 Hz, 6.9 Hz, 1.4 Hz, 1H), 7.59
(ddd, J = 8.2 Hz, 6.9 Hz, 1.4 Hz, 1H), 7.41–7.48 (m, 2H), 3.00 (q, J = 6.5 Hz, 2H), 1.42 (sx, J = 7.1 Hz, 2H),

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0.80 (t, J = 7.3 Hz, 3H); 13C-NMR (DMSO-d6),
δ
: 164.14, 153.94, 145.61, 142.89, 133.91, 130.69, 130.53,
130.33, 130.33, 128.11, 127.43, 125.93, 125.79, 125.72, 125.64, 124.91, 124.46, 122.62, 42.29, 22.37, 11.10;
HR-MS: for C21H18O5N3 [M + H]+ calculated 392.12410, found 392.12521 m/z.
1-[(2-Nitrophenyl)carbamoyl]naphthalen-2-yl butylcarbamate (18). Yield 82%; Mp 171–174 ◦C; IR (cm−1):
3293, 3222, 2957, 2873, 1724, 1661, 1589, 1549, 1532, 1520, 1505, 1471, 1463, 1439, 1350, 1279, 1257,
1
1227, 1145, 1109, 1006, 914, 865, 818, 779, 769, 731, 680, 667; H-NMR (DMSO-d6)
δ
: 10.92 (s, 1H),
8.07 (d, J = 9.1 Hz, 1H), 8.01–8.05 (m, 3H), 7.73–7.80 (m, 3H), 7.65 (ddd, J = 8.2 Hz, 6.9 Hz, 1.4 Hz,
1H), 7.58 (ddd, J = 8.2 Hz, 6.9 Hz, 1.4 Hz, 1H), 7.45 (td, J = 8.2 Hz, 1.4 Hz, 1H), 7.38 (d, J = 9.1 Hz,
1H), 3.03 (q, J = 6.2 Hz, 2H), 1.38 (qi, J = 7.0 Hz, 2H), 1.23 (sx, J = 7.0 Hz, 2H), 0.81 (t, J = 7.1 Hz, 3H);
13C-NMR (DMSO-d6),
δ: 164.13, 153.91, 145.58, 142.77, 133.91, 130.74, 130.52, 130.32, 130.31, 128.10,
127.41, 125.91, 125.74, 125.73, 125.56, 124.91, 124.44, 122.59, 40.18, 31.23, 19.27, 13.58; HR-MS: for
C22H20O5N3 [M + H]+ calculated 406.13975, found 406.14081 m/z.
1-[(2-Nitrophenyl)carbamoyl]naphthalen-2-yl pentylcarbamate (19). Yield 70%; Mp 141–143 ◦C; IR (cm−1):
3298, 3226, 2935, 2874, 1723, 1667, 1588, 1548, 1532, 1519, 1505, 1485, 1475, 1436, 1347, 1322, 1284, 1253,
1226, 1142, 1108, 1037, 1025, 996, 913, 866, 819, 778, 767, 730, 680, 660; 1H-NMR (DMSO-d6) δ: 10.93 (s,
1H), 8.07 (d, J = 9.1 Hz, 1H), 8.01–8.05 (m, 3H), 7.75–7.81 (m, 3H), 7.65 (ddd, J = 8.2 Hz, 6.9 Hz, 1.4 Hz,
1H), 7.58 (ddd, J = 8.2 Hz, 6.9 Hz, 1.4 Hz, 1H), 7.45 (td, J = 8.2 Hz, 1.4 Hz, 1H), 7.41 (d, J = 9.1 Hz, 1H),
3.02 (q, J = 6.4 Hz, 2H), 1.40 (qi, J = 7.0 Hz, 2H), 1.09–1.33 (m, 4H), 0.82 (t, J = 6.2 Hz, 3H); 13C-NMR
(DMSO-d6), δ: 164.12, 153.89, 145.58, 142.80, 133.88, 130.72, 130.51, 130.31, 130.30, 128.10, 127.42, 125.91,
125.74, 125.72, 125.58, 124.91, 124.44, 122.61, 40.47, 28.77, 28.29, 21.76, 13.83; HR-MS: for C23H22O5N3
[M + H]+ calculated 420.15540, found 420.15643 m/z.
1-[(2-Nitrophenyl)carbamoyl]naphthalen-2-yl hexylcarbamate (20). Yield 58%; Mp 122–123 ◦C; IR (cm−1):
3332, 3272, 2929, 2868, 1709, 1665, 1609, 1589, 1541, 1514, 1472, 1443, 1352, 1295, 1250, 1215, 1205, 1158,
1148, 1113, 1046, 993, 978, 911, 863, 825, 779, 761, 738, 697, 672; 1H-NMR (DMSO-d6)
δ: 10.93 (s, 1H),
8.07 (d, J = 9.1 Hz, 1H), 8.01–8.05 (m, 3H), 7.75–7.81 (m, 3H), 7.65 (ddd, J = 8.2 Hz, 6.9 Hz, 1.4 Hz, 1H),
7.58 (ddd, J = 8.2 Hz, 6.9 Hz, 1.4 Hz, 1H), 7.45 (td, J = 8.2 Hz, 1.4 Hz, 1H), 7.41 (d, J = 9.1 Hz, 1H), 3.02 (q,
J = 6.0 Hz, 2H), 1.38 (qi, J = 6.4 Hz, 2H), 1.18–1.25 (m, 6H), 0.83 (t, J = 6.6 Hz, 3H); 13C-NMR (DMSO-d6),
δ
: 164.65, 154.44, 146.08, 143.25, 134.42, 131.24, 131.04, 130.83, 130.82, 128.61, 127.94, 126.44, 126.23,
126.23, 126.06, 125.42, 124.94, 123.10, 41.03, 31.42, 29.56, 26.28, 22.49, 14.38; HR-MS: for C24H24O5N3
[M + H]+ calculated 434.17105, found 434.17236 m/z.
1-[(2-Nitrophenyl)carbamoyl]naphthalen-2-yl heptylcarbamate (21). Yield 46%; Mp 101–102 ◦C; IR (cm−1):
3333, 3265, 2957, 2925, 2852, 1709, 1665, 1652, 1609, 1588, 1541, 1511, 1472, 1464, 1441, 1350, 1294, 1250,
1215, 1205, 1148, 1045, 979, 911, 824, 779, 755, 737, 696, 667; 1H-NMR (DMSO-d6)
δ: 10.93 (s, 1H), 8.07
(d, J = 9.1 Hz, 1H), 8.01–8.05 (m, 3H), 7.75–7.81 (m, 3H), 7.65 (ddd, J = 8.2 Hz, 6.9 Hz, 1.4 Hz, 1H),
7.58 (ddd, J = 8.2 Hz, 6.9 Hz, 1.4 Hz, 1H), 7.45 (td, J = 8.2 Hz, 1.4 Hz, 1H), 7.41 (d, J = 9.1 Hz, 1H),
3.02 (q, J = 6.3 Hz, 2H), 1.39 (qi, J = 6.4 Hz, 2H), 1.17–1.28 (m, 8H), 0.85 (t, J = 6.6 Hz, 3H); 13C-NMR
(DMSO-d6), δ: 164.12, 153.89, 145.58, 142.77, 133.88, 130.74, 130.53, 130.31, 130.31, 128.10, 127.42, 125.91,
125.73, 125.72, 125.58, 124.90, 124.44, 122.61, 40.50, 31.16, 29.11, 28.36, 26.07, 22.04, 13.93; HR-MS: for
C25H26O5N3 [M + H]+ calculated 448.18670, found 448.18848 m/z.
◦
1-[(2-Nitrophenyl)carbamoyl]naphthalen-2-yl octylcarbamate (22). Yield 37%; Mp 92–94 C; IR (cm−1):
3315, 3230, 2924, 2850, 1704, 1653, 1589, 1528, 1508, 1485, 1463, 1435, 1360, 1293, 1271, 1251, 1219, 915,
834, 780, 759, 736, 701, 667; 1H-NMR (DMSO-d6)
δ: 10.93 (s, 1H), 8.07 (d, J = 9.1 Hz, 1H), 8.01–8.05
(m, 3H), 7.75–7.81 (m, 3H), 7.65 (ddd, J = 8.2 Hz, 6.9 Hz, 1.4 Hz, 1H), 7.59 (ddd, J = 8.2 Hz, 6.9 Hz,
1.4 Hz, 1H), 7.46 (td, J = 8.2 Hz, 1.4 Hz, 1H), 7.41 (d, J = 9.1 Hz, 1H), 3.03 (q, J = 5.5 Hz, 2H), 1.39 (qi,
J = 6.0 Hz, 2H), 1.18–1.29 (m, 10H), 0.86 (t, J = 6.4 Hz, 3H); 13C-NMR (DMSO-d6),
δ: 164.12, 153.89,
145.58, 142.79, 133.86, 130.72, 130.51, 130.32, 130.31, 128.10, 127.42, 125.91, 125.75, 125.72, 125.58, 124.90,
124.44, 122.59, 40.50, 31.22, 29.09, 28.67, 28.59, 26.12, 22.07, 13.95; HR-MS: for C26H28O5N3 [M + H]+
calculated 462.20235, found 462.20410 m/z.

Molecules 2016, 21, 1189
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◦
1-[(2-Nitrophenyl)carbamoyl]naphthalen-2-yl isoproylcarbamate (23). Yield 69%; Mp 172–176 C; IR (cm−1):
3355, 3320, 2974, 1738, 1665, 1586, 1532, 1505, 1489, 1440, 1362, 1245, 1213, 1173, 1149, 1056, 1018,
953, 924, 826, 778, 763, 732; 1H-NMR (DMSO-d6)
δ: 10.93 (s, 1H), 8.07 (d, J = 9.1 Hz, 1H), 8.01–8.05
(m, 3H), 7.75–7.80 (m, 3H), 7.66 (ddd, J = 8.2 Hz, 6.9 Hz, 1.4 Hz, 1H), 7.59 (ddd, J = 8.2 Hz, 6.9 Hz,
1.4 Hz, 1H), 7.46 (td, J = 8.2 Hz, 1.4 Hz, 1H), 7.42 (d, J = 9.1 Hz, 1H), 3.65 (sx, J = 6.4 Hz, 2H), 1.07 (d,
J = 6.4 Hz, 6H); 13C-NMR (DMSO-d6),
δ: 164.18, 153.10, 145.61, 142.94, 133.92, 130.69, 130.54, 130.37,
130.33, 128.14, 127.46, 125.96, 125.85, 125.80, 125.72, 124.94, 124.46, 122.71, 42.86, 22.32; HR-MS: for
C21H18O5N3 [M + H]+ calculated 394.13975, found 394.14058 m/z.
1-[(2-Nitrophenyl)carbamoyl]naphthalen-2-yl cyclopentylcarbamate (24). Yield 28%; Mp 138–142 ◦C;
IR (cm−1): 3295, 3219, 2960, 1719, 1661, 1588, 1519, 1487, 1438, 1351, 1283, 1255, 1222, 1144, 1078,
1
998, 822, 779, 732, 667; H-NMR (DMSO-d6)
δ: 10.92 (s, 1H), 8.07 (d, J = 9.1 Hz, 1H), 8.01–8.05 (m,
3H), 7.84 (d, J = 6.9 Hz, 1H), 7.72–7.79 (m, 2H), 7.65 (ddd, J = 8.2 Hz, 6.9 Hz, 1.4 Hz, 1H), 7.59 (ddd,
J = 8.2 Hz, 6.9 Hz, 1.4 Hz, 1H), 7.46 (td, J = 8.2 Hz, 1.4 Hz, 1H), 7.42 (d, J = 9.1 Hz, 1H), 3.83 (sx,
J = 5.5 Hz, 1H), 1.72–1.80 (m, 2H), 1.56–1.63 (m, 2H), 1.42–1.49 (m, 4H); 13C-NMR (DMSO-d6),
δ: 164.18,
153.41, 145.64, 142.94, 133.92, 130.69, 130.55, 130.36, 130.34, 128.14, 127.46, 125.97, 125.86, 125.80, 125.72,
124.95, 124.45, 122.73, 52.47, 32.06, 23.29; HR-MS: for C23H20O5N3 [M + H]+ calculated 420.15540,
found 420.15620 m/z.
◦
1-[(2-Nitrophenyl)carbamoyl]naphthalen-2-yl cyclohexylcarbamate (25). Yield 54%; Mp 140–143 C; IR
(cm−1): 3328, 3297, 2937, 2911, 2858, 1705, 1662, 1589, 1536, 1514, 1505, 1472, 1440, 1354, 1312, 1292,
1209, 1146, 1049, 1014, 823, 779, 761, 733, 697; 1H-NMR (DMSO-d6)
1H), 8.01–8.05 (m, 3H), 7.72–7.79 (m, 3H), 7.65 (ddd, J = 8.2 Hz, 6.9 Hz, 1.4 Hz, 1H), 7.58 (ddd, J = 8.2 Hz
δ
: 10.91 (s, 1H), 8.07 (d, J = 9.1 Hz
,
,
6.9 Hz, 1.4 Hz, 1H), 7.45 (td, J = 8.2 Hz, 1.4 Hz, 1H), 7.41 (d, J = 9.1 Hz, 1H), 3.26–3.33 (m, 1H), 1.71–1.76
(m, 2H), 1.63–1.68 (m, 2H), 1.51–1.56 (m, 1H), 1.02–1.27 (m, 5H); 13C-NMR (DMSO-d6),
δ
: 164.17, 153.14,
145.65, 142.90, 133.91, 130.70, 130.53, 130.36, 130.35, 128.13, 127.46, 125.96, 125.82, 125.79, 125.69, 124.93,
124.45, 122.67, 49.96, 32.39, 25.10, 24.56; HR-MS: for C24H22O5N3 [M + H]+ calculated 434.17105, found
434.17208 m/z.
1-[(2-Nitrophenyl)carbamoyl]naphthalen-2-yl cycloheptylcarbamate (26). Yield 58%; Mp 142–145 ◦C;
IR (cm−1): 3324, 2912, 2855, 1703, 1662, 1588, 1528, 1505, 1472, 1440, 1353, 1291, 1245, 1218, 1203,
1141, 1041, 997, 909, 823, 779, 758, 737, 697; 1H-NMR (DMSO-d6)
δ: 10.91 (s, 1H), 8.06 (d, J = 9.1 Hz,
1H), 8.01–8.05 (m, 3H), 7.81 (d, J = 8.2 Hz, 1H), 7.72–7.78 (m, 2H), 7.65 (ddd, J = 8.2 Hz, 6.9 Hz, 1.4 Hz,
1H), 7.58 (ddd, J = 8.2 Hz, 6.9 Hz, 1.4 Hz, 1H), 7.45 (td, J = 8.2 Hz, 1.4 Hz, 1H), 7.41 (d, J = 9.1 Hz,
1H), 3.46–3.54 (m, 1H), 1.73–1.80 (m, 2H), 1.39–1.60 (m, 8H), 1.29–1.38 (m, 2H); 13C-NMR (DMSO-d6),
δ
: 164.18, 153.01, 145.67, 142.82, 133.92, 130.73, 130.54, 130.35, 130.35, 128.13, 127.45, 125.95, 125.80,
125.80, 125.63, 124.93, 124.45, 122.70, 52.13, 34.26, 27.80, 23.54; HR-MS: for C25H24O5N3 [M + H]+
calculated 448.18670, found 448.18784 m/z.
1-[(2-Nitrophenyl)carbamoyl]naphthalen-2-yl (2-phenylethyl)carbamate (27). Yield 32%; Mp 127–130 ◦C;
IR (cm−1): 3336, 1742, 1708, 1683, 1608, 1578, 1535, 1510, 1495, 1434, 1398, 1334, 1270, 1211, 1144, 1130,
967, 824, 758, 743, 698; 1H-NMR (DMSO-d6)
δ: 10.96 (s, 1H), 8.07 (d, J = 9.1 Hz, 1H), 8.01–8.05 (m, 3H),
7.91 (t, J = 5.7 Hz, 1H), 7.71–7.79 (m, 2H), 7.65 (ddd, J = 8.2 Hz, 6.9 Hz, 1.4 Hz, 1H), 7.59 (ddd, J = 8.2 Hz,
6.9 Hz, 1.4 Hz, 1H), 7.46 (td, J = 8.2 Hz, 1.4 Hz, 1H), 7.38 (d, J = 9.1 Hz, 1H), 7.17–7.26 (m, 5H), 3.26 (q,
J = 6.4 Hz, 2H), 2.73 (t, J = 7.3 Hz, 2H); 13C-NMR (DMSO-d6),
δ: 164.10, 153.87, 145.54, 143.01, 139.13,
133.94, 130.63, 130.54, 130.35, 130.31, 128.88, 128.72, 128.31, 128.14, 127.46, 126.10, 125.98, 125.82, 123.69,
124.94, 124.48, 122.58, 42.16, 35.11; HR-MS: for C26H20O5N3 [M + H]+ calculated 456.15540, found
456.15712 m/z.
1-[(2-Nitrophenyl)carbamoyl]naphthalen-2-yl (4-phenylbutyl)carbamate (28). Yield 47%; Mp 109–110 ◦C;
IR (cm−1): 3351, 3233, 2934, 1714, 1677, 1593, 1519, 1505, 1482, 1458, 1435, 1352, 1287, 1244, 1222, 1051,
1007, 816, 779, 752, 731, 967; 1H-NMR (DMSO-d6)
δ
: 10.94 (s, 1H), 8.07 (d, J = 8.7 Hz, 1H), 8.00–8.05
(m, 3H), 7.82 (t, J = 5.7 Hz, 1H), 7.70–7.72 (m, 2H), 7.65 (ddd, J = 8.2 Hz, 6.9 Hz, 1.4 Hz, 1H), 7.58 (ddd,

Molecules 2016, 21, 1189
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J = 8.2 Hz, 6.9 Hz, 1.4 Hz, 1H), 7.43–7.46 (m, 1H), 7.41 (d, J = 9.1 Hz, 1H), 7.24–7.28 (m, 2H), 7.15–7.18
(m, 3H), 3.06 (q, J = 5.9 Hz, 2H), 2.52 (t, J = 7.3 Hz, 2H), 1.54 (qi, J = 7.3 Hz, 2H), 1.43 (qi, J = 7.3 Hz, 2H);
13C-NMR (DMSO-d6),
δ: 164.13, 153.93, 145.59, 142.96, 142.09, 133.88, 130.65, 130.54, 130.35, 130.32,
128.30, 128.22, 128.13, 127.45, 125.95, 125.80, 125.72, 125.64, 125.64, 124.93, 124.47, 122.63, 40.32, 34.77,
28.80, 28.11; HR-MS: for C28H24O5N3 [M + H]+ calculated 484.18670, found 484.18801 m/z.
3.3. In Vitro Antibacterial Susceptibility Testing
The synthesized compounds were evaluated for in vitro antibacterial activity against
representatives of multidrug-resistant bacteria and clinical isolates of methicillin-resistant
Staphylococcus aureus (MRSA) SA 630 and SA 3202, that were obtained from the National Institute of
Public Health (Prague, Czech Republic). Staphylococcus aureus ATCC 29213 was used as a reference
and quality control strain. Ampicillin (Sigma-Aldrich) was used as the standard. Prior to testing,
each strain was passaged onto nutrient agar (Oxoid, Hampshire, UK) with 5% of bovine blood,
and bacterial inocula were prepared by suspending a small portion of bacterial colony in sterile
phosphate buffered saline (pH 7.2–7.3). The cell density was adjusted to 0.5 McFarland units using
a densitometer (Densi-La-Meter, LIAP, Riga, Latvia). The final inoculum was made to a 1:20 dilution of
the suspension with the Mueller-Hinton broth (MH broth). The compounds were dissolved in DMSO
(Sigma), and the final concentration of DMSO in the MH broth (Oxoid) did not exceed 2.5% of the total
solution composition. The final concentrations of the evaluated compounds ranged from 256
0.008 g/mL. The broth dilution micro-method, modified according to NCCLS (National Committee
for Clinical Laboratory Standards) guidelines [46 47] in MH broth, was used to determine the minimum
µg/mL to
µ
,
inhibitory concentration (MIC). Drug-free controls, sterility controls, and controls consisting of MH
broth and DMSO alone were included. The determination of results was performed visually after 24 h
of static incubation in the darkness at 37 ◦C in an aerobic atmosphere. The MICs were defined as the
lowest concentration of the compound at which no visible bacterial growth was observed. The results
are summarized in Table 1.
3.4. In Vitro Antimycobacterial Evaluation
The evaluation of the in vitro antimycobacterial activity of the compounds was performed against
Mycobacterium marinum CAMP 5644 and M. kansasii DSM 44162. The broth dilution micro-method
in Middlebrook 7H9 medium (Difco, Lawrence, KS, USA) supplemented with ADC Enrichment
(Becton, Dickinson & Comp., Franklin Lakes, NJ, USA) was used to determine the minimum
inhibitory concentration (MIC), as previously described [48]. The compounds were dissolved in
DMSO (Sigma-Aldrich), and the final concentration of DMSO did not exceed 2.5% of the total solution
composition. The final concentrations of the evaluated compounds, ranging from 256
µg/mL to
0.125 g/mL, were obtained by twofold serial dilution of the stock solution in a microtiter plate with
µ
sterile medium. Bacterial inocula were prepared by transferring colonies from the culture to sterile
water. The cell density was adjusted to 0.5 McFarland units using a densitometer (Densi-La-Meter,
LIAP, Riga, Latvia). The final inoculum was made by 1:1000 dilution of the suspension with sterile
water. Drug-free controls, sterility controls, and controls consisting of the medium and DMSO alone
were included. The determination of results was performed visually after 7 days of static incubation in
the darkness at 37◦◦C in an aerobic atmosphere for M. kansasii and after 21 days of static incubation in
the darkness at 28 C in an aerobic atmosphere for M. marinum. The minimum inhibitory concentration
(MIC) was defined as the lowest concentration of the compound at which no visible bacterial growth
was observed. The MIC value is routinely and widely used in bacterial assays and is a standard
detection limit according to the Clinical and Laboratory Standards Institute (CLSI) [49]. Isoniazid
(Sigma-Aldrich) was used as the reference antibacterial drug. The results are summarized in Table 1.

Molecules 2016, 21, 1189
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3.5. In Vitro Cytotoxicity Assay
Human monocytic leukemia THP-1 cells were obtained from the European Collection of Cell
Cultures (ECACC, Salisbury, UK; Methods of characterization: DNA Fingerprinting (Multilocus
probes) and isoenzyme analysis). These cells were routinely cultured in RPMI 1640 (Lonza, Verviers,
Belgium) medium supplemented with 10% fetal bovine serum (FBS, Sigma-Aldrich), 2% L-glutamine,
◦
1% penicillin, and streptomycin (Lonza, Verviers, Belgium) at 37 C with 5% CO2. Cells were passaged
at approximately 1-week intervals. Cells were routinely tested for the absence of mycoplasma (Hoechst
33258 staining method). The tested compounds were dissolved in DMSO (Sigma-Aldrich) and
added in five increasing concentrations to the cell suspension in the culture medium. The maximum
concentration of DMSO in the assays never exceeded 0.1%. Subsequently, the cells were incubated
for 24 h at 37 ◦C with 5% CO2 at various compound concentrations ranging from 0.37 to 30
µmol/L
in RPMI 1640 medium. Cell toxicity was determined using a Cytotoxicity Detection KitPLUS Lactate
dehydrogenase (LDH) assay kit (Roche Diagnostics, Mannheim, Germany), and used according to the
manufacturer’s instructions, as described previously [14
,30,32,33]. For LDH assays, cells were seeded
into 96-well plates (5 L culture medium) in triplicate in serum-free RPMI
×
104 cells well−1 in 100
·
µ
1640 medium, and measurements at 492 nm wavelength (Synergy 2 Multi-Mode Microplate Reader,
BioTek, Winooski, VT, USA) were taken 24 h after the treatment with tested compounds. The median
lethal dose values, LD50, were deduced through the production of a dose-response curve. All data were
evaluated using GraphPad Prism 5.00 software (GraphPad Software, San Diego, CA, USA). The results
are summarized in Table 1.
4. Conclusions
Series of thirteen 1-[(2-chlorophenyl)carbamoyl]naphthalen-2-yl carbamates and thirteen
1-[(2-nitrophenyl)carbamoyl]naphthalen-2-yl carbamates were prepared and subsequently characterized.
All compounds were tested for their in vitro antimicrobial activity against S. aureus, two methicillin-resistant
S. aureus strains, M. marinum, and M. kansasii. 1-[(2-Chlorophenyl)-carbamoyl]naphthalen-2-yl
ethylcarbamate (
3
) and 1-[(2-nitrophenyl)carbamoyl]naphthalen-2-yl ethylcarbamate (16) showed
M against all methicillin-resistant S. aureus strains. In addition, both compounds expressed
M against M. marinum and M. kansasii. Propyl- ( 17) and isopropyl- (10 23) carbamates
MICs = 42
MICs = 21
µ
µ
4
,
,
demonstrated medium antimicrobial activity, while bulkier-substituted carbamates showed only
moderate or no activity, which was similar in both the chlorinated and nitrated series. Screening of the
cytotoxicity of the most potent and discussed compounds, performed using the THP-1 cells, proved
no significant lethal effect (LD50 >30 µM). The biological activity of the compounds is dependent
on the alkyl substitution of carbamate nitrogen. Spatially small N-alkyl substituents are important
for the activity. Antimicrobial activity rapidly decreases with chain prolongation or with a bulkier
N-cycloalkyl, N-arylalkyl substituent.
Acknowledgments: This study was supported by IGA VFU Brno 311/2016/FaF, 328/2016/FaF, 302/2015/FaF,
320/2015/FaF and by the Slovak Research and Development Agency (Grant No. APVV-0516-12). The
HPLC/HRMS system forms a part of the National Infrastructure CzeCOS (LM2015061); Michal Oravec was
supported by the National Sustainability Program (NPU I; Grant No. LO1415 POLYMAT).
Author Contributions: Tomas Gonec, Josef Stranik, Jiri Kos, Josef Jampilek—design, synthesis of the
compounds, SAR, writing of the paper. Michal Oravec, Aneta Cernikova—analysis/characterization of the
compounds. Sarka Pospisilova, Valeria Pudelkova, Alois Cizek—antimicrobial evaluation. Lucie Holanova,
Peter Kollar—cytotoxicity assay.
Conflicts of Interest: The authors declare no conflict of interest.
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2016 by the authors; licensee MDPI, Basel, Switzerland. This article is an open access
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