Adamantane-isothiourea hybrid derivatives: Synthesis, characterization, in vitro antimicrobial, and in vivo hypoglycemic activities

Article abstract of DOI:10.3390/molecules22050710

A new series of adamantane-isothiourea hybrid derivatives, namely 4-arylmethyl (Z)-N¹-(adamantan-1-yl)-morpholine-4-carbothioimidates 7a-e and 4-arylmethyl (Z)-N0-(adamantan-1-yl)-4 -phenylpiperazine-1-carbothioimidates 8a-e were prepared via the reaction of N-(adamantan-1- yl)morpholine-4-carbothioamide 5 and N-(adamantan-1-yl)-4-phenylpiperazine-1-carbothioamide 6 with benzyl or substituted benzyl bromides, in acetone, in the presence of anhydrous potassium carbonate. The structures of the synthesized compounds were confirmed by ¹H-NMR, ¹³C-NMR, electrospray ionization mass spectral (ESI-MS) data, and X-ray crystallographic data. The in vitro antimicrobial activity of the new compounds was determined against certain standard strains of pathogenic bacteria and the yeast-like pathogenic fungus Candida albicans. Compounds 7b, 7d and 7e displayed potent broad-spectrum antibacterial activity, while compounds 7a, 7c, 8b, 8d and 8e were active against the tested Gram-positive bacteria. The in vivo oral hypoglycemic activity of the new compounds was carried on streptozotocin (STZ)-induced diabetic rats. Compounds 7a, 8ab, and 8b produced potent dose-independent reduction of serum glucose levels, compared to the potent hypoglycemic drug gliclazide.

Full text of DOI:10.3390/molecules22050710

molecules
Article
Adamantane-Isothiourea Hybrid Derivatives:
Synthesis, Characterization, In Vitro Antimicrobial,
and In Vivo Hypoglycemic Activities
Lamya H. Al-Wahaibi 1, Hanan M. Hassan 2, Amal M. Abo-Kamar 2,3, Hazem A. Ghabbour 4 and
Ali A. El-Emam 4,
*
1
Department of Chemistry, College of Sciences, Princess Nourah Bint Abdulrahman University,
Riyadh 11671, Saudi Arabia; lamya-alwahaibi@hotmail.com
2
Department of Pharmaceutical Sciences, College of Pharmacy, Princess Nourah Bint Abdulrahman University,
Riyadh 11671, Saudi Arabia; hananhafila@hotmail.com (H.M.H.); amabokamer@pnu.edu.sa (A.M.A.-K.)
Department of Microbiology, Faculty of Pharmacy, Tanta University, Tanta, Egypt
Department of Pharmaceutical Chemistry, College of Pharmacy, King Saud University,
Riyadh 11671, Saudi Arabia; ghabbourh@yahoo.com
3
4
*
Correspondence: aelemam@ksu.edu.sa; Tel.: +966-11-467-7350; Fax: +966-11-467-6220
Academic Editor: Diego Muñoz-Torrero
Received: 14 April 2017; Accepted: 27 April 2017; Published: 29 April 2017
Abstract: A new series of adamantane-isothiourea hybrid derivatives, namely04-arylmethyl (Z)-N0
-(adamantan-1-yl)-morpholine-4-carbothioimidates 7a–e and 4-arylmethyl (Z)-N -(adamantan-1-yl)-4
-phenylpiperazine-1-carbothioimidates 8a–e were prepared via the reaction of N-(adamantan-1-
yl)morpholine-4-carbothioamide and N-(adamantan-1-yl)-4-phenylpiperazine-1-carbothioamide
5
6
with benzyl or substituted benzyl bromides, in acetone, in the presence of anhydrous potassium
carbonate. The structures of the synthesized compounds were confirmed by 1H-NMR, 13C-NMR,
electrospray ionization mass spectral (ESI-MS) data, and X-ray crystallographic data. The in vitro
antimicrobial activity of the new compounds was determined against certain standard strains of
pathogenic bacteria and the yeast-like pathogenic fungus Candida albicans. Compounds 7b
7e displayed potent broad-spectrum antibacterial activity, while compounds 7a 7c 8b 8d and 8e
were active against the tested Gram-positive bacteria. The in vivo oral hypoglycemic activity of the
new compounds was carried on streptozotocin (STZ)-induced diabetic rats. Compounds 7a 8ab, and
, 7d and
,
,
,
,
8b produced potent dose-independent reduction of serum glucose levels, compared to the potent
hypoglycemic drug gliclazide.
Keywords: synthesis; adamantane; isothiourea; carbothioimidate; antimicrobial activity;
hypoglycaemic activity
1. Introduction
The adamantane nucleus was recognized early as an essential pharmacophore in various
pharmacologically-active drugs. The incorporation of an adamantyl moiety into various bioactive
molecules results in compounds with relatively high lipophilicity which, in turn, modifies the
bioavailability and modulates their therapeutic efficacies [
drug, was approved for the treatment of Influenza A infection [
drug [ ]. Further studies based on amantadine resulted in the development of the potent antiviral drugs
rimantadine [ ] and tromantadine [ ]. Numerous adamantane-based analogues were proved to possess
significant inhibitory activity against human immunodeficiency viruses (HIV) [ 12]. The synthetic
1,2]. Amantadine, the first adamantane-based
3–5] and as an anti-Parkinsonian
6
7
8
9–
retinoid derivative CD437 was developed as a potent inducer of apoptosis in human head and neck
squamous cell carcinoma [13]. Potent bactericidal and fungicidal activities were reported for several
Molecules 2017, 22, 710; doi:10.3390/molecules22050710
www.mdpi.com/journal/molecules

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adamantane derivatives, including SQ109, which was approved for use against drug-susceptible and
drug-resistant TB strains [14]. SQ109 also showed excellent inhibitory activity against Helicobacter
pylori-related duodenal ulcers and carcinomas, and Candida glabrata [15]. The adamantane-based
drugs, vildagliptin [16], and saxagliptin [17] are currently used as oral hypoglycemic agents for the
treatment of type 2 diabetes acting via inhibition of dipeptidyl peptidase IV (DPP-IV). Moreover, the
adamantane derivatives MK-544 [18], PF-877423 [19], and AZD6925 [20] were recently developed as
11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) inhibitors as drug candidates for the treatment
of non-insulin-dependent diabetes and obesity [21] (Figure 1).
O
O
N
N
NH2
Amantadine
H
NH2
Tromantadine
Rimantadine
H
COOH
N
N
H
SQ109
HO
CD437
N
O
H
N
N
HO
N
HO
N
N
H
H2N
H
CN
O
NC
MK-544
Vildagliptin
Saxagliptin
COOH
H
N
N
H
N
N
O
AZD6925
O
S
PF-877423
Figure 1. Biologically-active adamantane-based derivatives.
On the other hand, several compounds containing an isothiourea moiety were reported to
possess significant antiviral [22], histamine-H3 antagonistic [23,24], calcium channel antagonistic [25],
anticancer [26], antibacterial [27], and nitric oxide synthase inhibitory [28,29] activities.
In view of the diverse pharmacological properties of adamantane and isothiourea derivatives,
and following our previous studies on the chemical and biological properties of adamantane
derivatives [10,30–33], we report herein the synthesis and characterization of novel adamantane
derivatives containing an isothiourea moiety as potential antimicrobial and/or hypoglycemic agents.
2. Results and Discussion
2.1. Chemical Synthesis
The key starting material 1-adamantyl isothiocyanate
previously described methods [33]. The reaction of 1-adamantylamine
trimethylamine, in ethanol, yielded the dithiocarbamate salt , which was reacted with di-tert-butyl
dicarbonate (Boc2O) to yield the intermediate . The intermediate was stirred with catalytic
amount of 4-dimethylaminopyridine (DMAP) to furnish the target product . The reaction of
1-adamantyl isothiocyanate with morpholine and 1-phenylpiperazine, in boiling ethanol, yielded
the corresponding N-(adamantan-1-yl)morpholine-4-carbothioamide and N-(adamantan-1-yl)-4-
phenylpiperazine-1-carbothioamide , respectively [33]. The carbothioamides and were reacted
with benzyl or substituted benzyl bromides, in acetone, in the presence of anhydrous potassium
carbonate to yield the corresponding S-arylmethyl derivatives 7a and 8a , respectively, in good
yields (Scheme 1, Table 1). The structures of the target compounds 7a and 8a were confirmed by
4
was prepared in good yield via our
1
with carbon disulphide and
2
3
3
4
4
5
6
5
6
–
e
–e
–
e
–e

Molecules 2017, 22, 710
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elemental analyses (Table S1), in addition to the 1H-NMR and 13C-NMR, and electrospray ionization
mass spectral (ESI-MS) data, which were in full agreement with their structures. The ESI-MS data
showed the correct positive ions (M + H)+ ions for all compounds. In addition, compounds 7d and 8d
were subjected to single crystal X-ray studies.
S
O
S
CS2 / Et3N
Boc2O
S- HN+Et3
N
S
OC4H9(tert)
NH2
R
N
H
H
1
3
2
DMAP
S
HN
O
ArCH2Br, K2CO3
N
H
N
NCS
S
O
5
4
N
N
O
7a-e
N
R
S
N
H
S
ArCH2Br, K2CO3
R
7a, 8a:
7b, 8b: 4-Cl
7c, 8c: 4-Br
7d, 8d: 4-NO2
7e, 8e: 3,5-(CF3)2
H
N
N
H
N
N
N
N
6
8a-e
Scheme 1. Synthetic approach for the target compounds 7a–e and 8a–e.
Table 1. Crystallization solvents, melting points, yield percentages, molecular formulae, and molecular
weights of compounds 7a–e and 8a–e.
◦
Comp. No.
R
Cryst. Solv.
M.p. ( C)
Yield (%)
Mol. Formula (Mol. Wt.)
7a
7b
7c
7d
7e
8a
8b
8c
H
4-Cl
4-Br
4-NO2
3,5-(CF3)2
H
4-Cl
4-Br
4-NO2
3,5-(CF3)2
EtOH/H2O
EtOH
108–110
92–94
98–100
118–120
106–108
137–139
153–155
140–142
145–147
113–115
91
76
85
95
72
88
90
92
96
75
C22H30N2OS (370.55)
C22H29ClN2OS (405.0)
C22H29BrN2OS (449.45)
C22H29N3O3S (415.55)
C24H28F6N2OS (506.55)
C28H35N3S (445.66)
C28H34ClN3S (480.11)
C28H34BrN3S (524.56)
C28H34N4O2S (490.66)
C30H33F6N3S (581.66)
EtOH
EtOH
EtOH/H2O
EtOH/H2O
EtOH
EtOH
EtOH
EtOH/H2O
8d
8e
2.2. Crystallographic Studies
The single crystal X-ray crystallographic data of compounds 7d and 8d are summarized in
Table 2. Compound 7b crystallizes in the centrosymmetric monoclinic space group P21/c with one
molecule in the asymmetric unit (Z = 4). The ORTEP (Oak Ridge Thermal Ellipsoid Plot) is shown in
Figure 2. The morpholine ring adopts a chair conformation and the mean planes of the nitrophenyl
and morpholine rings make a dihedral angle of 52.55 (5). The conformation about the N1=C11 imine
bond is Z (cis) configuration. The crystal packing is mainly controlled by intermolecular C-H . . .
O
hydrogen bonding (Figure 3).
Table 2. Single-crystal X-ray crystallographic data of compounds 7d and 8d.
Data
Compound 7d
Compound 8d
Formula
C22H29N3O3S
415.55
293
0.71073
Monoclinic
C28H34N4O2S
490.66
293
0.71073
Orthorhombi
Formula weight
Temperature (K)
Wavelength (Å)
Crystal system

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Table 2. Cont.
Data
Compound 7d
Compound 8d
Space group
a, b, c (Å)
V (Å3)
P21/c
P212121
6.9204 (5), 29.775 (3), 10.2725 (10)
6.9426 (9), 9.6472 (12), 39.086 (5)
2116.7 (3)
4
2617.8 (6)
4
Z
Radiation type
Mo Kα
0.18
Mo Kα
0.16
−1
µ (mm
)
No. of reflections
No. of unique reflections/obs.
reflections
11033
25091
3718/2253
4609/1447
No. of parameters
262
0
0.28, −0.21
0.939, 0.989
0.073
318
0
0.44, −0.40
0.924, 0.957
0.526
No. of restraints
∆ρmax, ∆ρmin (e Å
Tmin, Tmax
−3
)
Rint
Crystal size (mm)
0.35 × 0.11 × 0.06
0.052, 0.192, 0.65
1525183
0.85 × 0.21 × 0.05
0.128, 0.296, 1.02
1523432
R[F2 > 2σ(F2)], wR(F2), S
CCDC number
Figure 2. ORTEP diagram of compound 7d drawn at 40% ellipsoids for non-hydrogen atoms.
Figure 3. Molecular packing of compound 7d viewed hydrogen bonds, which are drawn as
dashed lines.

Molecules 2017, 22, 710
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Compound 8b crystallizes in the orthorhombic space group P212121 with one molecule in the
asymmetric unit (Z = 4). The ORTEP is shown Figure 4. The piperazine ring also adopts a chair
conformation with a dihedral angle of 39.25 (4). The conformation about the N1A=C11A imine bond is
Z (cis) configuration. The crystal packing is mainly controlled by intermolecular C-H . . . S hydrogen
bonding (Figure 5).
Figure 4. ORTEP diagram of compound 8d drawn at 40% ellipsoids for non-hydrogen atoms.
Figure 5. Molecular packing of compound 8d viewed hydrogen bonds, which are drawn as
dashed lines.
2.3. In Vitro Antimicrobial Activity
The newly synthesized compounds 7a–e and 8a–e were tested for their in vitro growth
inhibitory activity against the standard bacterial strains of the American type culture collection
ATCC, Staphylococcus aureus ATCC 6571, Bacillus subtilis ATCC 5256, Micrococcus luteus ATCC
27141 (Gram-positive bacteria), Escherichia coli ATCC 8726, Pseudomonas aeruginosa ATCC 27853
(Gram-negative bacteia), and the yeast-like pathogenic fungus Candida albicans MTCC 227.
The preliminary antimicrobial activity testing was carried out using the semi-quantitative agar-disc
diffusion method with Müller-Hinton agar medium [34]. The outcomes of the preliminary
antimicrobial screening of compounds 7a
–e
,
8a–
e
(200
µg/disc), the antibacterial antibiotics gentamicin
sulphate, ampicillin trihydrate and the antifungal drug clotrimazole (100
µg/disc) and the calculated
log P values (Clog P) of the tested compounds (calculated using the CS ChemOffice Ultra version 8.0,
CambridgeSoft, Cambridge, MA, USA), are shown in Table 3.

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The main features of the results of the antimicrobial activity testing revealed that the tested
compounds generally displayed marked antibacterial and marginal antifungal activities against the
tested microorganisms, the antibacterial activity against the Gram-positive bacteria was higher than
the activity against the Gram-negative bacteria, and the compound lipophilicity had no influence
on their activity. In addition, the Gram-positive bacteria S. aureus and B. subtilis are more sensitive
than M. luteus, and the Gram-negative bacteria P. aeruginosa was more resistant than E. coli. Potent
antibacterial activity was shown by compounds 7a
zones 20 mm against one or more of the tested microorganisms. In the morpholine derivatives
7a , compounds 7b 7d and 7e retained good broad spectrum antibacterial activity; while compound
–e, 8d and 8e which produced growth inhibition
≥
–
e
,
7a displayed good activity against the Gram-positive bacteria and medium activity against E. coli
(inhibition zones 15–19 mm). The optimum activity was achieved by 7e which was highly active against
all the tested bacterial strains. In the piperazine derivatives 8a–c, compounds 8b, 8d and 8e showed
high activity against the tested Gram-positive bacteria, while compounds 8a and 8c showed medium
activity against the tested Gram-positive bacteria and weak activity (growth inhibition zones 10–14 mm)
against the tested Gram-negative bacteria. The activity against yeast-like pathogenic fungus C. albicans
of the tested compounds was rather lower than that against the tested bacterial strains. Compound 8d
showed medium inhibitory activity, compounds 7b, 7d, 8a, 8b, 8c and 8e displayed weak activity and
compounds 7a, 7c and 7e were practically inactive (growth inhibition zones < 10 mm).
Table 3. Antimicrobial activity of compounds 7a
antibacterial drugs gentamicin sulphate, ampicillin trihydrate and the antifungal drug clotrimazole
(100 g/8 mm disc) against Staphylococcus aureus ATCC 6571 (SA), Bacillus subtilis ATCC 5256 (BS),
–e and 8a–e (200 µg/8 mm disc), the broad-spectrum
µ
Micrococcus luteus ATCC 27141 (ML), Escherichia coli ATCC 8726 (EC), Pseudomonas aeruginosa ATCC
27853 (PA), and the yeast-like pathogenic fungus Candida albicans MTCC 227 (CA).
Diameter of Growth Inhibition Zone (mm) a
Comp. No.
Clog P
SA
BS
ML
EC
PA
CA
22 (2) b
24 (4) b
22 (4) b
31 (0.5) b
33 (0.25) b
18 (8) b
21 (8) b
17 (32) b
24 (1) b
28 (1) b
27 (1) b
22 (2) b
NT
21 (4) b
28 (0.5) b
18 (16) b
32 (0.25) b
34 (0.25) b
18 (8) b
24 (2) b
19 (8) b
28 (1) b
31 (0.5) b
26 (2) b
23 (1) b
NT
20 (4) b
22 (4) b
14 (64) b
28 (0.5) b
28 (1) b
14 (128) b
16 (32) b
14 (64) b
20 (2) b
22 (2) b
20 (2) b
20 (2) b
NT
18 (16) b
22 (20) b
13 (128) b
22 (1) b
14 (64) b
15 (32)
7a
7b
7c
7d
7e
8a
8b
8c
5.584
6.297
6.447
5.327
7.350
7.130
7.843
7.993
6.873
8.896
-
11 (>128) b
12 (128) b
18 (4) b
-
14 (32) b
-
24 (2) b
20 (4) b
12 (>128) b 10 (>128) b 10 (>128) b
16 (64) b
12 (>128) b
13 (64) b
12 (128) b
16 (16) b
14 (64) b
NT
11 (>128) b 10 (>128) b
18 (2) b
19 (4) b
22 (0.5) b
16 (8) b
NT
14 (4) b
18 (8) b
21 (0.5) b
16 (8) b
NT
8d
8e
Gentamicin sulfate
Ampicillin trihydrate
Clotrimazole
NT
21 (4) b
a
b
(-): inactive (inhibition zone < 10 mm), Figures shown in parentheses represent the MIC values (
NT: not tested.
µg/mL),
From the above results, it could be concluded that the antibacterial activity of the morpholine
derivatives 7a is generally superior to their N-phenylpiperazine analogues. In addition, the presence
of the electron-withdrawing substituents NO2 and CF3 (compounds 7d 7e 8d and 8e) greatly enhanced
the antibacterial activity. Unlike the antibacterial activity, the N-phenylpiperazine analogues 8a
were generally more active than the morpholine analogues 7a against C. albicans. The values of
–
e
,
,
–
e
–
e
the minimal inhibitory concentration (MIC) in Müller-Hinton Broth [35] for the tested compounds
(Table 3) were correlated to the results obtained in the preliminary screening.
2.4. In Vivo Hypoglycemic Activity
The oral hypoglycaemic activity of compounds 7a
–c and 8a–c was determined in streptozotocin
(STZ)-induced diabetic rats. STZ induces its hyperglycemic activity via irreversible damage to the

Molecules 2017, 22, 710
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pancreatic beta cells, resulting in loss of insulin secretion [36
,
37]. The compounds were tried in 10 and
20 mg/kg dose levels. The dose levels (10 and 20 mg/kg) were determined after pilot experiments
which showed that increasing the dose to 30 mg/kg was found to produce toxic central nervous
manifestations in the form of severe symmetric convulsions. The hypoglycemic activity testing
experiment (animal treatments, induction of experimental diabetes, and measurement of serum
glucose level) was carried out following the previously reported protocols [32
oral hypoglycemic activity of compounds 7a 8a (10 and 20 mg/kg), and the potent hypoglycemic
drug gliclazide in STZ-induced diabetic rats (10 mg/kg) are shown in Table 4.
The optimum hypoglycemic activity was attained by compounds 7a 8a, and 8b which produced
,33]. The results of the
–
c,
–c
,
potent dose-independent reduction of serum glucose levels, compared to gliclazide at 10 mg/kg
dose level (Potency ratio 71.08, 75.13 and 79.04%, respectively). Compound 7b showed medium
dose-dependent hypoglycemic activity and compound 8c showed weak activity at 10 mg/kg dose
level without significant increase in the activity at 20 mg/kg dose level. Despite the high activity
of the bis(3,5-trifluormethyl) derivative 7e (Potency ratio 82.32%), increasing the dose produced
toxic central nervous manifestations. Similarly, compound 8e showed toxic manifestations at 10 and
20 mg/kg doses.
The hypoglycemic activity of the tested compounds revealed that the piperazine derivatives 8a
are generally more active than their morpholine analogues 7a . In addition, the aryl substituents
greatly influenced the hypoglycemic activity and toxicity. The optimum activity was attained
by the phenyl derivatives (7a 8a) and to a lesser extent the chlorophenyl derivatives (7b 8b).
The bis(3,5-trifluormethyl) derivatives were generally toxic.
–e
–
e
,
,
Table 4. Oral hypoglycemic activity of compounds 7a–c, 8a–c (10 and 20 mg/kg), and gliclazide (10
mg/kg) in STZ-induced diabetic rats.
Results
C24 (mg/dL) a
Treatment
C0 (mg/dL) a
% Glucose Reduction b
Group 1 c
302.8 ± 11.64
299.2 ± 16.50
304.8 ± 13.26
300.6 ± 11.65
288.9 ± 12.15
294.8 ± 9.08
284.8 ± 19.55
290.2 ± 21.64
278.1 ± 16.24
302.6 ± 22.25
306.2 ± 15.20
290.2 ± 18.22
171.6 ± 12.32 *
212.4 ± 12.16 *
134.6 ± 9.75 *
245.2 ± 19.25 *
201.5 ± 9.60 *
281.2 ± 7.19
286.8 ± 19.02
282.2 ± 27.20
299.8 ± 18.80
198.7 ± 19.10 *
Toxic
200.2 ± 9.88 *
108.4 ± 11.05 *
199.8 ± 10.01 *
144.6 ± 10.01 *
277.6 ± 16.20
269.9 ± 20.12
289.2 ± 25.28
309.0 ± 25.95
178.2 ± 16.04 *
Toxic
4.16%
42.65%
Group 1 d
7a (10 mg/kg)
7a (20 mg/kg)
7b (10 mg/kg)
7b (20 mg/kg)
7c (10 mg/kg)
7c (20 mg/kg)
7d (10 mg/kg)
7d (20 mg/kg)
7e (10 mg/kg)
7e (20 mg/kg)
8a (10 mg/kg)
8a (20 mg/kg)
8b (10 mg/kg)
8b (20 mg/kg)
8c (10 mg/kg)
8c (20 mg/kg)
8d (10 mg/kg)
8d (20 mg/kg)
8e (10 mg/kg)
8e (20 mg/kg)
30.31% (71.08)
55.22% (64.74)
15.13% (35.47)
31.65% (37.13)
1.26% (2.69)
2.75% (1.37)
−1.47%
0.93% (1.08)
35.12 (82.32)
294.6 ± 11.30
290.6 ± 8.60
301.4 ± 9.06
296.0 ± 11.02
320.5 ± 22.05
313.5 ± 18.60
295.0 ± 22.45
304.5 ± 27.50
286.6 ± 13.22
32.04% (75.13)
62.70% (73.50)
33.71% (79.04)
51.15% (59.96)
13.39% (31.38)
13.91% (16.30)
1.97% (4.61)
−1.48
37.82% (88.68)
a
b
Results are expressed as mean
±
S.E.M. (n = 5), The figures shown in parentheses are the relative potency
c
compared with gliclazide, Treated with a single oral dose of 0.5% (w/v) aqueous CMC solution (5 mL/kg),
d
Treated with 10 mg/kg gliclazide in 0.5% (w/v) aqueous CMC, * Significant difference at p < 0.01 compared with
the corresponding control.

Molecules 2017, 22, 710
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3. Materials and Methods
3.1. General
Melting points (◦C, uncorrected) were measured in open glass capillaries using a Branstead 9100
electrothermal melting point apparatus (Thermo Fisher Scientific, Waltham, MA, USA). Nuclear
magnetic resonance (NMR) spectra were obtained on a Bruker Ascend 700 NMR spectrometer
(Fällanden, Switzerland) at 700.17 MHz for 1H and 176.08 MHz for 13C, using CDCl3 as the solvent.
The chemical shifts are expressed in
δ (ppm) downfield from tetramethylsilane (TMS) as internal
standard, coupling constants (J) are expressed in Hz. Electrospray ionization mass spectra (ESI-MS)
were recorded on an Agilent 6410 Triple Quad tandem mass spectrometer (Agilent Technologies, Santa
Clara, CA, USA) at 4.0 kV for positive ions. Elemental analyses (C, H, N, and S) were in agreement
with the proposed structures within
±
0.4% of the theoretical values (Table S1). Monitoring the
reactions and checking the purity of the final products were carried out by thin layer chromatography
(TLC) using silica gel precoated aluminum sheets (60 F254; Merck Schuchardt, Darmstadt, Germany),
and visualization with ultraviolet light (UV) at 365 and 254 nm. The reference drugs gentamicin
sulfate (CAS 1405-41-0), ampicillin trihydrate (CAS 7177-48-2), clotrimazole (CAS 23593-75-1) and
gliclazide (CAS 21187-98-4), and streptozotocin (CAS 18883-66-4) were purchased from Sigma-Aldrich
Chemie GmbH, Taufkirchen, Germany. The Sprauge-Dawley rats were purchased from local animal
house. The commercial glucose oxidase (GO) assay kit (Sigma-Aldrich Co., St. Louis, MO, USA)
were used for measurement of serum glucose levels. The animal experiments for the determination
of the hypoglycemic activity were performed in accordance with the guidelines provided by the
Experimental Animal Laboratory (EAL) and approved by the Animal Care and Use Committee
(ACUC) of the College of Pharmacy, King Saud University (Saudi Arabia). The X-ray crystallographic
data of compound 8c was recently described [38].
3.2. Synthesis of 4-Arylmethyl (Z)-N'-(Adamantan-1-yl)-Morpholine-4-Carbothioimidates 7a–e and
4-Arylmethyl (Z)-N'-(Adamantan-1-yl)-4-Phenylpiperazine-1-Carbothioimidates 8a–e
The appropriate arylmethyl bromide (2 mmol) and anhydrous potassium carbonate (276 mg,
2 mmol) were added to a solution of N-(adamantan-1-yl)morpholine-4-carbothioamide
5 (560 mg,
2 mmol) or N-(adamantan-1-yl)-4-phenylpiperazine-1-carbothioamide (711 mg, 2 mmol), in
6
anhydrous acetone (15 mL), and the mixture was heated under reflux for 4 h. The solvent was
then distilled off in vacuo and the resulting residues were washed with water (20 mL), dried, and
crystallized from ethanol or aqueous ethanol.
Benzyl (Z)-N0-(adamantan-1-yl)-morpholine-4-carbothioimidate 7a
:
1H-NMR:
δ 1.61–1.66 (m, 6H,
adamantane-H), 1.85 (m, 6H, adamantane-H), 2.0 (s, 3H, adamantane-H), 3.20–3.21 (m, 4H,
morpholine-H), 3.65–3.72 (m, 4H, morpholine-H), 3.90 (s, 2H, benzylic CH2), 7.07–7.19 (m, 5H, Ar-H).
13C-NMR:
δ 29.02, 29.80, 35.96, 54.18 (adamantane-C), 37.06 (benzylic CH2), 49.66, 66.42 (morpholine-C),
125.60, 126.44, 127.90, 140.02 (Ar-C), 154.46 (C=N). ESI-MS, m/z: 372.3 (M + H)+.
1
4-Chlorobenzyl (Z)-N0-(adamantan-1-yl)-morpholine-4-carbothioimidate 7b: H-NMR:
δ
1.62–1.65 (m, 6H,
adamantane-H), 1.69–1.72 (m, 6H, adamantane-H), 2.05–2.06 (m, 3H, adamantane-H), 3.99 (s, 2H,
benzylic CH2), 3.20–3.22 (m, 4H, morpholine-H), 3.65–3.72 (m, 4H, morpholine-H), 6.99 (d, 2H, Ar-H,
J = 7.5 Hz), 7.14 (d, 2H, Ar-H, J = 7.5 Hz). 13C-NMR:
δ 29.26, 29.99, 35.98, 54.08 (adamantane-C), 36.98
(benzylic CH2), 48.60, 66.44 (morpholine-C), 127.65, 128.65, 133.0, 138.04 (Ar-C), 154.24 (C=N). ESI-MS,
m/z (Rel. Int.): 405.4 (M + H, 100)+, 407.4 (M + 2 + H, 35)+.
4-Bromobenzyl (Z)-N0-(adamantan-1-yl)-morpholine-4-carbothioimidate 7c
: δ 1.63–1.69 (m,
1H-NMR:
6H, adamantane-H), 1.84 (m, 6H, adamantane-H), 2.01 (s, 3H, adamantane-H), 3.25–3.30 (m, 4H,
morpholine-H), 3.69–3.74 (m, 4H, morpholine-H), 3.91 (s, 2H, benzylic CH2), 7.17 (d, 2H, Ar-H,
J = 7.5 Hz), 7.45 (d, 2H, Ar-H, J = 7.5 Hz). 13C-NMR:
δ 29.59, 29.94, 36.57, 54.69 (adamantane-C), 37.76

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(benzylic CH2), 49.70, 66.85 (morpholine-C), 121.0, 130.51, 137.26, 146.91 (Ar-C), 156.46 (C=N). ESI-MS,
m/z (Rel. Int.): 449.4 (M + 2 + H, 100)+, 451.4 (M + 2 + H, 98)+.
: δ 1.62 (s, 6H,
4-Nitrobenzyl (Z)-N0-(adamantan-1-yl)-morpholine-4-carbothioimidate 7d 1H-NMR:
adamantane-H), 1.77–1.79 (m, 6H, adamantane-H), 1.98–2.0 (m, 3H, adamantane-H), 3.24–3.30 (m,
4H, morpholine-H), 3.69–3.72 (m, 4H, morpholine-H), 4.02 (s, 2H, benzylic CH2), 7.42 (d, 2H, Ar-H,
J = 8.2 Hz), 8.21 (d, 2H, Ar-H, J = 8.2 Hz). 13C-NMR:
δ 29.58, 29.84, 36.44, 54.79 (adamantane-C), 37.53
(benzylic CH2), 49.79, 66.78 (morpholine-C), 123.76, 129.64, 146.07, 146.91 (Ar-C), 156.71 (C=N). ESI-MS,
m/z: 416.2 (M + H)+.
3,5-Bis(trifluoromethyl)benzyl (Z)-N0-(adamantan-1-yl)-morpholine-4-carbothioimidate 7e
: δ
1H-NMR:
1.53–1.56 (m, 12H, adamantane-H), 1.85–1.87 (m, 3H, adamantane-H), 3.10–3.12 (m, 4H, morpholine-H),
3.68–3.70 (m, 4H, morpholine-H), 4.17 (s, 2H, benzylic CH2), 7.95 (s, 1H, Ar-H), 8.0 (s, 2H, Ar-H).
13C-NMR:
δ
29.16, 29.66, 35.25, 54.85 (adamantane-C), 36.41 (benzylic CH2), 49.76, 66.20 (morpholine-C),
120.76, 122.72, 130.39, 142.78 (Ar-C), 124.89 (CF3), 148.37 (C=N). ESI-MS, m/z: 507.2 (M + H)+.
Benzyl (Z)-N0-(adamantan-1-yl)-4-phenylpiperazine-1-carbothioimidate 8a 1H-NMR:
1.69 (s, 6H,
:
δ
adamantane-H), 1.74 (s, 6H, adamantane-H), 2.0 (s, 3H, adamantane-H), 2.88–2.92 (m, 4H,
piperazine-H), 3.02–3.06 (m, 4H, piperazine-H), 3.98 (s, 2H, benzylic CH2), 6.84–7.04 (m, 5H,
Ar-H), 7.12–7.16 (m, 5H, Ar-H). 13C-NMR:
δ 29.10, 29.96, 35.68, 53.98 (adamantane-C), 46.90, 48.18
(piperazine-C), 36.90 (benzylic CH2), 114.28, 119.90, 126.92, 128.24, 129.0, 130.58, 139.94, 149.26 (Ar-C),
152.0 (C=N). ESI-MS, m/z: 446.3 (M + H)+.
0
1
4-Chlorobenzyl (Z)-N -(adamantan-1-yl)-4-phenylpiperazine-1-carbothioimidate 8b: H-NMR: δ 1.71–1.76 (m,
12H, adamantane-H), 2.15–2.17 (m, 3H, adamantane-H), 2.63–2.65 (m, 4H, piperazine-H), 3.22–3.24 (m,
4H, piperazine-H), 3.98 (s, 2H, benzylic CH2), 6.89–6.91 (m, 3H, Ar-H), 6.95–6.96 (m, 2H, Ar-H), 7.29–733
(m, 4H, Ar-H). 13C-NMR:
δ 29.17, 29.71, 35.51, 53.07 (adamantane-C), 48.65, 48.10 (piperazine-C), 36.39
(benzylic CH2), 115.55, 116.13, 119.89, 128.49, 129.16, 129.45, 130.57, 150.31 (Ar-C), 151.27 (C=N).
ESI-MS, m/z (Rel. Int.): 380.2 (M + H, 100)+, 382.2 (M + 2 + H, 37)+.
1
4-Bromobenzyl (Z)-N0-(adamantan-1-yl)-4-phenylpiperazine-1-carbothioimidate 8c: H-NMR:
δ
1.65 (s, 6H,
adamantane-H), 1.86 (s, 6H, adamantane-H), 2.02–2.03 (m, 3H, adamantane-H), 3.26–3.27 (m, 4H,
piperazine-H), 3.43–3.44 (m, 4H, piperazine-H), 3.95 (s, 2H, benzylic CH2), 6.92–7.93 (m, 1H, Ar-H),
7.00–7.01 (m, 2H, Ar-H), 7.18 (d, 2H, Ar-H, J = 7.0 Hz), 7.29–7.33 (m, 2H, Ar-H), 7.44 (d, 2H, Ar-H, J = 7.0
Hz). 13C-NMR:
δ 29.95, 36.58, 42.98, 54.70 (adamantane-C), 48.96, 49.17 (piperazine-C), 37.77 (benzylic
CH2), 116.20, 119.97, 120.94, 129.22, 130.59, 131.54, 137.33, 149.26 (Ar-C), 151.29 (C=N). ESI-MS, m/z
(Rel. Int.): 524.4 (M + H, 98)+, 526.4 (M + 2 + H, 100)+ [38].
4-Nitrobenzyl (Z)-N0-(adamantan-1-yl)-4-phenylpiperazine-1-carbothioimidate 8d 1H-NMR:
: δ 1.53 (s,
6H, adamantane-H), 1.72 (s, 6H, adamantane-H), 1.91 (s, 3H, adamantane-H), 3.16–3.18 (m, 4H,
piperazine-H), 3.30–3.33 (m, 4H, piperazine-H), 3.97 (s, 2H, benzylic CH2), 6.80–6.89 (m, 3H, Ar-H),
7.18–7.29 (m, 2H, Ar-H), 7.36 (d, 2H, Ar-H, J = 8.0 Hz), 8.08 (d, 2H, Ar-H, J = 8.0 Hz). 13C-NMR:
δ 29.65,
29.90, 36.52, 54.86 (adamantane-C), 37.54 (benzylic CH2), 43.03, 49.12 (piperazine-C), 116.21, 120.05,
123.72, 129.21, 129.66, 146.14, 146.94, 148.09 (Ar-C), 151.22 (C=N). ESI-MS, m/z: 491.2 (M + H)+.
1
3,5-Bis(trifluoromethyl)benzyl (Z)-N0-(adamantan-1-yl)-4-phenylpiperazine-1-carbothioimidate 8e: H-NMR:
δ
1.58 (s, 6H, adamantane-H), 1.69 (s, 6H, adamantane-H), 1.95–1.97 (m, 3H, adamantane-H), 3.29–3.31
(m, 4H, piperazine-H), 3.40–3.42 (m, 4H, piperazine-H), 4.04 (s, 2H, benzylic CH2), 6.93–7.02 (m, 3H,
Ar-H), 7.29 (s, 1H, Ar-H), 7.32–7.34 (m, 2H, Ar-H), 7.78 (s, 2H, Ar-H). 13C-NMR:
29.80, 35.51, 36.44,
δ
54.80 (adamantane-C), 37.31 (benzylic CH2), 49.0, 49.13 (piperazine-C), 116.28, 120.12, 129.16, 129.24,
131.44, 131.63, 141.30, 147.50 (Ar-C), 124.04 (CF3), 151.19 (C=N). ESI-MS, m/z: 582.2 (M + H)+.

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3.3. Crystal Growth and Single Crystal X-ray Study
Single crystals suitable for X-ray analysis were obtained by slow evaporation of CHCl3:EtOH
(1:1; 5 mL) solution of compounds 7d and 8d at room temperature. Data were collected on a Bruker
APEX-II D8 Venture area diffractometer, equipped with graphite monochromatic Mo K
α radiation
(λ = 0.71073 Å) at 100 K. Unit cell measurement, data collection, integration, scaling, and absorption
corrections for the crystals were conducted using Bruker Apex II software [39]. Data reduction
was done by Bruker SAINT suite [40]. The crystal structures were solved by the full matrix least
squares method using SHELXL 2014 [41]. Absorption correction was applied using SADABS
program [42]. ORTEP (Oak Ridge Thermal Ellipsoid Plot) was generated using Mercury 3.5.1
Cambridge Crystallographic Data Centre (CCDC) program [43].
4. Conclusions
In this study, new adamantane-linked isothiourea derivatives were synthesized and their in vitro
antimicrobial and in vivo hypoglycemic activities were determined. Compounds 7b
, 7d, and 7e
displayed potent broad-spectrum antibacterial activity, while compounds 7a 7c 8b 8d, and 8e were
,
,
,
active against the tested Gram-positive bacteria. The tested compounds were generally inactive
against the yeast-like pathogenic fungus Candida albicans. The in vivo oral hypoglycemic activity of the
synthesized compounds was determined in streptozotocin (STZ)-induced diabetic rats. Compounds
7a, 8a, and 8b produced potent dose-independent reduction of serum glucose levels compared
to gliclazide at 10 mg/kg dose level (potency ratios of 71.08%, 75.13%, and 79.04%, respectively).
The active compounds are considered to be good candidates as newer antibacterial and hypoglycemic
agents, though, further studies including toxicity testing and molecular docking for the exploration
of the mechanism of their biological activity are required for optimization of the activity which are
being undertaken.
Supplementary Materials: Supplementary materials (the experimental details of the determination of in vitro
antimicrobial activity, in vivo hypoglycemic activity and microanalytical data) are available online.
Acknowledgments: The authors would like to thank the Deanship of Scientific Research, Princess Nourah Bint
Abdulrahman University for funding this study (Research Project No. 37-S-173).
Author Contributions: L.H.A.-W. and A.A.E.-E. synthesized and characterized the target compounds and
prepared the single crystals. H.M.H. performed the in vivo hypoglycemic testing, A.M.A.-K. conceived the
antimicrobial testing. H.A.G. carried out the X-ray analysis. All authors discussed the contents of the manuscript
and approved the submission.
Conflicts of Interest: The authors declare no conflict of interest.
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©
2017 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access
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