Novel 2-(E)-substituted benzylidene-6-(N-substituted aminomethyl) cyclohexanones and cyclohexanols as analgesic and anti-inflammatory agents

Article abstract of DOI:10.1007/s00044-012-0362-x

Twenty-two new 2-(E)-substituted benzylidene-6-(N-substituted aminomethyl)cyclohexanones (6a-6j) and cyclohexanols (7a-7l) were designed and synthesized. Target compounds were obtained through Stork enamine, Mannich, and Grignard reactions taking cyclohexanone as starting material. The structures were confirmed by the application of IR, ¹H NMR, MS, and HR-MS data. The analgesic activities were evaluated by acetic acid-induced writhing test and hot plate method. The anti-inflammatory activities were assayed by xylene-induced ear swelling and carrageenan-induced paw edema in mice model. All tested compounds showed analgesic and anti-inflammatory capacities in oral administration. Some compounds (6a, 6c, 6h, 6i, 7c, 7h, and 7i) displayed the moderate analgesic activity compared with positive control ibuprofen, and some compounds (6a, 6b, 6d, 6h, 7a, and 7d) exhibited more anti-inflammatory activity than ibuprofen. Among them, compound 6a could be a potential nonsteroidal anti-inflammatory agent with significant analgesic activities and remarkable anti-inflammatory activities. Further research is being conducted.

Full text of DOI:10.1007/s00044-012-0362-x

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res (2013) 22:3779–3786
DOI 10.1007/s00044-012-0362-x
ORIGINAL RESEARCH
Novel 2-(E)-substituted benzylidene-6-(N-substituted
aminomethyl)cyclohexanones and cyclohexanols
as analgesic and anti-inflammatory agents
•
Dan Liu Weishe Yu Jingjing Li
•
•
•
Cong Pang Linxiang Zhao
Received: 29 March 2012 / Accepted: 10 November 2012 / Published online: 9 December 2012
Ó Springer Science+Business Media New York 2012
Abstract Twenty-two new 2-(E)-substituted benzyli-
dene-6-(N-substituted aminomethyl)cyclohexanones (6a–6j)
and cyclohexanols (7a–7l) were designed and synthesized.
Target compounds were obtained through Stork enamine,
Mannich, and Grignard reactions taking cyclohexanone as
starting material. The structures were confirmed by the
Introduction
Nonsteroidal anti-inflammatory drugs (NSAIDs) are com-
mon medication for treatment of pain, inflammation, and
fever. However, the significant side effects have been shown
with a long-term usage of NSAIDs including serious gas-
trointestinal lesions, kidney injury, and cardiovascular risk.
Cycloketo compounds with exo-cyclic a,b-unsaturated
double bond have multifunctional pharmacological bioac-
tivities, such as anti-inflammatory, antiviral, and antitumor
(Tham et al., 2011; Li et al., 2011; Wang et al., 2005). In our
previous research, we found that cyclopentanones (cyclo-
hexanones) with 2-alkylaminomethyl, the simplified struc-
ture of bioactivity diterpenes and sesquiterpenes, showed
powerful inhibiting effects on inflammation and pain, as well
as cancer cell growth, and explored the preliminary antitu-
mor mechanisms (Wang et al., 2004, 2005; Zhang et al.,
2006). The stability of cyclohexanones is more powerful
than the cyclopentanones. So, in view of these studies,
introduction of substituted phenyl aminomethyl group into
the ortho position of cyclohexanones and cyclohexanols is
expected to increase the stability and enhance the anti-
inflammatory and analgesic activity. A series of novel 2-(E)-
substituted benzylidene-6-(N-substituted aminomethyl)
cyclohexanones and cyclohexanols were designed and syn-
thesized. Their analgesic and anti-inflammatory effects were
measured in mice model.
1
application of IR, H NMR, MS, and HR-MS data. The
analgesic activities were evaluated by acetic acid-induced
writhing test and hot plate method. The anti-inflammatory
activities were assayed by xylene-induced ear swelling and
carrageenan-induced paw edema in mice model. All tested
compounds showed analgesic and anti-inflammatory
capacities in oral administration. Some compounds (6a, 6c,
6h, 6i, 7c, 7h, and 7i) displayed the moderate analgesic
activity compared with positive control ibuprofen, and
some compounds (6a, 6b, 6d, 6h, 7a, and 7d) exhibited
more anti-inflammatory activity than ibuprofen. Among
them, compound 6a could be a potential nonsteroidal anti-
inflammatory agent with significant analgesic activities and
remarkable anti-inflammatory activities. Further research is
being conducted.
Keywords 2-(E)-Substituted benzylidene-6-(N-
substituted aminomethyl)cyclohexanones ꢀ
2-(E)-Substituted benzylidene-6-(N-substituted
aminomethyl)-1-aryl-cyclohexanols ꢀ Analgesia ꢀ
Anti-inflammation
Results and discussion
D. Liu ꢀ W. Yu ꢀ J. Li ꢀ C. Pang ꢀ L. Zhao (&)
Key Laboratory of Structure-Based Drugs Design & Discovery
of Ministry of Education, Shenyang Pharmaceutical University,
103 Wenhua Road, Shenhe District, Shenyang 110016,
Liaoning, People’s Republic of China
Chemistry
The synthetic route to the 2-(E)-substituted benzylidene-6-
(N-substituted aminomethyl) cyclohexanones 6a–6j and
e-mail: zhaolinxiang@syphu.edu.cn
123

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Med Chem Res (2013) 22:3779–3786
2-(E)-substituted benzylidene-6-(N-substituted amino-
methyl)-1-aryl cyclohexanols 7a–7l are presented in
Scheme 1. The substituents of compounds 6a–6j and 7a–7l
are also listed in Scheme 1. Enamine 3 was produced as a
colorless liquid through Stork enamine synthesis by com-
mercially available cyclohexanone condensed with mor-
pholine (Hunig et al., 1973). 2-Benzylidene cyclohexanone
4a–4c were synthesized by fresh enamine 3 refluxing in
toluene with corresponding benzaldehyde (Stork et al.,
1963). Compounds 4a–4c were reacted with fresh Esc-
henmoser’s salt (N,N-dimethylmethylene-ammonium
chloride), synthesized based on the literature procedure
(Brand et al., 2003), to yield the Mannich base hydro-
chlorides 5a–5c. After amine exchange reaction of com-
pounds 5a–5c with aromatic amine, target compounds
6a–6j were produced. a,b-Unsaturated ketones 6a–6j were
reacted with appropriate Grignard reagent to get the target
compounds 7a–7l as the racemic mixture with the specific
rotator power 0° in 1 % MeOH.
Biologic results and discussion
The synthesized compounds, 6a–6j and 7a–7l, were eval-
uated for analgesic and anti-inflammatory, and the results
are summarized in Tables 1 and 2. The results indicated
that all the tested compounds exhibited different degree of
analgesic and anti-inflammatory capacities under the
experimental conditions.
The analgesic capacities were evaluated by acetic acid-
induced writhing test and hot plate test using Kunming
male mice, ibuprofen as position drug. As shown in
Table 1, all the tested compounds exhibited moderate
Scheme 1 The synthetic
O
N
O
1
O
pathway of target compounds.
Reagents and conditions: a
toluene, p-toluenesulfonic acid,
reflux, 20 h; b substituted
benzaldehyde, toluene, reflux,
36 h; c Eschenmoser’s salt, dry
acetonitrile, r.t., 5 h; d
substituted aromatic amine,
50 % ethanol, 30 °C, 12 h; e
grignard reagent, THF, r.t., 24 h
.
O
H
N
HCl
b
a
c
N
+
R1
R1
O
2
4a-4c
R3
5a-5c
3
O
OH
d
e
R2
R2
R1
R1
7a-7l
6a-6j
HN
R1
R2
R3
R1
H
R2
N
CH3
CH3
7a
H
H
6a
6b
6c
6d
6e
6f
CH3
N
7b
7c
7d
7e
7f
H
H
N
CH3
H
CH3
CH3
N
CH3
H
3-OCH3
3-OCH3
3-OCH3
4-OCH3
4-OCH3
3-OCH3
3-OCH3
4-OCH3
4-OCH3
4-OCH3
N
CH3
CH3
H
H
H
N
CH3
CH3
N
H
CH3
CH3
N
N
CH3
4-CH3
4-CH3
CH3
CH3
N
CH3
H
N
CH3
CH3
N
CH3
CH3
CH3
CH3
7g
H
N
6g
4-CH3
CH3
CH3
N
CH3
7h 4-CH3
7i 4-CH3
N
CH3
6h 4-SO2CH3
6i 4-SO2CH3
6j 4-SO2CH3
N
CH3
N
CH3
CH3
7j
4-CH3
N
CH3
N
CH3
CH3
7k 4-CH3
N
CH3
N
7l
4-CH3
CH3
CH3
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Med Chem Res (2013) 22:3779–3786
3781
Table 1 The analgesic activity
of the target compounds 6a–7l
Code
Acetic acid-induced writhing test
Hot plate test
Writhing number (/150)
Inhibition rate (%)
Pain threshold (s)
Increasing rate (%)
Control
Ibuprofen
6a
47.83 13.84
7.60 3.21**
8.21 1.80**
14.50 6.17**,#
9.07 3.12**
11.14 5.22**
15.21 9.80**,#
11.21 6.38**
11.57 6.29**
9.07 3.74**
7.40 3.07**
13.00 7.47**
17.14 5.76**,##
14.07 6.57**
8.50 3.27**
10.64 6.10**
9.60 4.67**
11.64 4.79**
10.21 5.25**
9.14 2.09**
8.78 2.48**
13.36 5.59**
18.07 5.38**,##
12.36 4.40**
0
16.80 5.08
4.20
158.56
166.19
109.59
146.12
108.45
79.04
84.11
82.82
69.69
81.04
76.70
68.19
76.55
75.81
81.04
84.53
72.82
64.16
70.58
82.23
77.75
79.93
75.66
78.65
80.89
81.63
72.08
62.22
74.17
41.11 7.98**
43.88 3.55**
35.66 6.98**
41.62 8.69**
33.84 3.43#
28.68 6.34**,##
26.97 4.76**,##
27.51 8.38*
6b
6c
6d
6e
6f
71.60
6g
75.50
6h
38.41 4.71**
40.61 8.01**
33.80 5.65**
26.84 7.41**,##
29.57 10.64*,#
42.63 2.73**
33.79 3.89**
34.62 7.63**
35.68 9.07**
34.99 3.49**
39.75 4.12**
35.58 6.25**
32.83 10.46**
31.50 3.64**,#
31.22 4.40**,#
142.70
155.12
109.98
70.08
6i
6j
7a
7b
83.22
7c
162.71
110.54
116.52
121.45
117.46
153.01
125.34
106.84
93.41
7d
7e
7f
7g
7h
Values are expressed as
mean SEM. n = 10
7i
7j
* p \ 0.05, ** p \ 0.01
compared with control;
7k
#
##
p \ 0.05, p \ 0.01
compared with ibuprofen
7l
93.52
analgesic capacity. Cyclohexanones with methylsulfonyl
group at the para-position of benzylidene group (6h–6j)
increased the analgesic activity, in which compound 6i
was the best analgesic one with inhibition of 84.53 %
(ibuprofen 84.11 %) and improvement pain threshold
([150 %). Cyclohexanones with no substituent at the para-
position of benzylidene group (6a–6d) could also increase
the pain threshold ([100 %). Cyclohexanols with methoxy
group at the meta-position of benzene ring (7c–7e, 7h, 7i)
showed better analgesic activity than that of no or para-
position substituent.
tested compounds exhibited preferable analgesic activity,
pain threshold increasing rate was more than 70 %. And,
they did not display the more powerful anti-inflammatory
activity, except compounds 6a, 6b, 7a, and 7d. So, we will
do the further study to focus their analgesic activity.
Experimental
Chemistry
The anti-inflammatory activities were assayed in mice
model of xylene-induced ear swelling using Kunming male
mice and carrageenan-induced paw edema on Wistar rats,
ibuprofen as position drug. As a whole, cyclohexanols
(7a–7l) exhibited less anti-inflammatory activity than
cyclohexanones (6a–6j), whereas compound 7a and 7d had
the most potent activity during both experiments. The
activity of cyclohexanones was altered by the substitution of
the aromatic amines, and the order was methyl group at the
para-position (R2: 4-CH3) \ meta-position (R2: 3-CH3) \
no substituent (R2: H).
1H NMR spectra were recorded on a BRUKER ARX-300
instrument in CDCl3 solution with Me4Si as internal stan-
dard. MS were determined on Waters Quattro micro API
LC–MS mass spectrometer. HR-MS were obtained on
Finnigan MAT-711 mass spectrometer in ESI mode. Infra-
red spectra were recorded on a BRUKER IFS-55 FTIR
spectrometer. Optical rotation was recorded on Pekin-
Elmer 241 instrument. The melting points were determined
on an electrically heated X4 digital visual melting point
apparatus and were uncorrected. Unless specified, other-
wise, all reagents and solvents were used as supplied by the
manufacturer.
In summary, the synthesized compounds had certain
analgesic and anti-inflammatory activity. It looked that all
123

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Med Chem Res (2013) 22:3779–3786
Paw edema test
Table 2 The anti-inflammatory
activity of the target compounds
6a–7l
Code
Ear swelling test
Swelling (mg)
Inhibition rate (%)
Edema (mL)
Inhibition rate (%)
Control
Ibuprofen
6a
21.00 4.74
12.57 2.30**
9.17 4.02**
10.57 3.46**
17.17 3.19
11.57 5.56**
12.71 3.99**
15.20 6.76*
13.57 8.02**
11.14 5.81**
15.33 2.81*
13.29 5.41*
7.00 4.62**,#
16.14 6.74
13.71 7.34*
12.29 2.50**
13.43 6.78*
15.43 4.54*
15.43 3.69*
17.21 7.50
16.20 5.63
17.57 5.38
14.71 4.61*
15.86 7.11*
0
0.47 0.05
0
40.14
56.35
49.66
18.25
44.90
39.45
27.62
35.37
46.94
26.98
36.73
66.67
23.13
34.69
41.50
36.05
26.53
26.53
18.04
22.86
16.33
29.92
24.49
0.22 0.03**
53.19
72.34
76.60
25.53
46.81
44.68
38.30
36.17
51.06
31.91
36.17
59.57
36.17
38.30
57.45
44.68
38.30
21.28
25.53
21.28
17.02
29.79
29.79
0.13 0.03**,##
0.11 0.03**,##
0.35 0.04**,##
0.25 0.05**
0.26 0.01**,##
0.29 0.03**,##
0.30 0.03**,##
0.23 0.08**
0.32 0.06**,##
0.30 0.07**,##
0.19 0.03**,#
0.30 0.08**,##
0.29 0.09**,#
0.20 0.07**
6b
6c
6d
6e
6f
6g
6h
6i
6j
7a
7b
7c
7d
7e
0.26 0.08**
7f
0.29 0.05**,##
0.37 0.08**,##
0.35 0.10**,##
0.37 0.02**,##
0.39 0.09*,##
0.33 0.01**,##
0.33 0.05**,##
7g
7h
Values are expressed as
mean SEM. n = 10
7i
7j
* p \ 0.05, ** p \ 0.01
compared with control;
7k
#
##
p \ 0.05, p \ 0.01
compared with ibuprofen
7l
N-(1-Cyclohexen-1-yl)morpholine (3)
to dryness. The solid was recrystallized from methanol or
ethanol.
A mixture of cyclohexanone 1 (58.8 g, 0.6 mol), mor-
pholine 2 (87.2 g, 1.0 mol), and p-toluenesulfonic acid
(0.5 g, 0.003 mol) in toluene (200 mL) was heated to
boiling 20 h in a 500 mL round-bottomed flask to which a
water separator under the reflux condenser was attached.
Cooled and distilled at atmospheric pressure to remove the
most toluene. The residue was evaporated under reduced
pressure. Compound 3 was obtained as a colorless liquid at
117–119 °C/10 mmHg (118–120 °C/10 mmHg, Hunig
et al., 1973). Yield: 85.5 %.
2-Benzylidene cyclohexanone (4a) Yield: 55 %, pale
yellow powder; m.p.: 52–53 °C (52–54 °C, Falck et al.,
2006).
2-(4-Methylbenzylidene) cyclohexanone (4b) Yield:
64 %, pale yellow powder; m.p.: 66–68 °C (71 °C, Das
et al., 2008).
2-(4-Methylsulfonylbenzylidene)
cyclohexanone
(4c)
Yield: 70 %, pale yellow powder; m.p.: 94–95 °C
(97–98 °C, Ao et al., 2004).
General procedure for the synthesis of 4a–4c
The corresponding benzaldehyde (0.20 mol) was dissolved
in toluene (80 mL) and fresh N-(1-cyclohexen-1-yl)mor-
pholine 3 (50.2 g, 0.30 mol) was added. The solution was
heated to boiling 36 h in a 250 mL round-bottomed flask
attached a water separator under the reflux condenser.
Cooled and added 6 mol/L HCl (40 mL). The organic layer
was washed with 10 % NaHCO3 solution, H2O, and dried
by Na2SO4. It was filtered, and the filtrate was evaporated
General procedure for the synthesis of 5a–5c
Fresh
N,N-dimethylmethylene-ammonium
chloride
(0.02 mol), synthesized based on a literature procedure
(Gaudry et al., 1988), was added to a solution of 2-ben-
zylidene cyclohexanone 4a, 4b, or 4c (0.01 mol) in ace-
tonitrile at 80 °C. The mixture was reacted under reflux
monitored by TLC using a solvent system of chloroform–
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Med Chem Res (2013) 22:3779–3786
3783
2-(E)-Benzylidene-6-(N-methyl-N-(p-tolyl)aminomethyl)cyc-
lohexanone (6c) Yield: 56 %. m.p.: 130–131 °C; MS (ESI,
m/z): 319.6 (M?); IR (KBr) cm-1: 3,434, 2,944, 1,694, 1,622,
methanol (7:1). After 5 h, the mixture was cooled and fil-
tered. The residue was crystallized from chloroform-tetra-
hydrofuran to give 5a–5c as white solid.
1
1,134, 756; H NMR (300 MHz, CDCl3) d: 7.36–7.14 (m,
9H), 6.82 (s, 1H), 2.95–2.86 (m, 2H), 2.64–2.62 (d,
J = 6.0 Hz, 2H), 2.28–2.18 (m, 3H), 2.10–2.08 (m, 3H),
2.05–2.00 (m, 4H), 1.68–1.64 (m, 4H); HR-MS: m/z, calcd,
C22H25NO (M?): 319.1936, found: 319.1948.
2-Dimethylaminomethyl-6-benzylidene-cyclohexanonehydro
chloride (5a) Yield: 61.2 %; m.p.: 152–154 °C; MS (ESI)
m/z: 243.2 (M?); 1H NMR (300 MHz, DMSO-d6) d: 10.31
(brs, 1H), 7.46–7.42 (m, 5H), 7.34 (s, 1H), 3.57–3.55
(q, J = 7.6 Hz, 1H), 3.09–3.06 (d, J = 6.4 Hz, 2H), 2.94
(brs, 1H), 2.78 (s, 6H), 2.71 (s, 1H), 2.31 (m, 1H), 1.86
(m, 1H), 1.69 (m, 2H).
2-(E)-Benzylidene-6-(N-methyl-N-(m-tolyl)aminomethyl)
cyclohexanone (6d) Yield: 51 %. m.p.: 128–130 °C; MS
(ESI, m/z): 319.5 (M?); IR (KBr) cm-1: 3,428, 2,942,
1,692, 1,578, 1,133, 756; 1H NMR (300 MHz, CDCl3)
d: 7.36–7.12 (m, 9H), 6.82 (s, 1H), 2.96 (m, 1H), 2.85 (m,
1H), 2.64–2.62 (d, J = 6.6 Hz, 2H), 2.26–2.21 (m, 3H),
2.10–2.18 (m, 3H), 2.00 (m, 1H), 1.79–1.66 (m, 4H),;
HR-MS: m/z, calcd, C22H25NO (M?): 319.1936, found:
319.1939.
2-Dimethylaminomethyl-6-[(4-methyl)benzylidene]-cyclo-
hexanone hydrochloride (5b) Yield: 59.9 %; m.p.: 163–
166 °C; MS (ESI) m/z: 258.2 (M?H)?.
2-Dimethylaminomethyl-6-[(4-methylsulfonyl)benzylidene]-
cyclohexanone hydrochloride (5c) Yield: 71.2 %; m.p.:
142–144 °C; MS (ESI) m/z: 322.4 (M?H)?.
2-(E)-(4-Methyl)benzylidene-6-(N-methyl-N-phenylaminom-
ethyl)cyclohexanone (6e) Yield: 47 %. m.p.: 103–105 °C;
MS (ESI, m/z): 319.6 (M?); IR (KBr) cm-1: 3,444, 2,940,
1,681, 1,578, 1,149, 808; 1H NMR (300 MHz, CDCl3)
d: 7.29–7.25 (m, 3H), 7.18–7.16 (m, 2H), 7.10–7.06 (m, 4H),
6.79 (s, 1H), 2.94 (m, 1H), 2.87 (m, 1H), 2.63–2.61 (m, 2H),
2.37 (m, 1H), 2.32 (s, 3H), 2.26–2.18 (m, 3H), 1.78–1.65 (m,
4H); HR-MS: m/z, calcd, C22H25NO (M?): 319.1936, found:
319.1939.
General procedure for the synthesis of 6a–6j
To a solution of substituted cyclohexanone hydrochlorides
5a–5c (0.01 mol) in 50 % (v/v) ethanol (30 mL), substi-
tuted aromatic amines (0.02 mol) was added and reacted
for 12 h at 30 °C. The reaction was cooled and filtered. The
residue was purified by crystallization from acetone or by a
silica gel column with CHCl3:MeOH (v/v) = 60:1 to give
6a–6j as yellow solid.
2-(E)-(4-Methyl)benzylidene-6-(N-methyl-N-(p-tolyl)amino-
methyl) cyclohexanone (6f) Yield: 49 %. m.p.: 105–107
°C; MS (ESI, m/z): 333.7 (M?); IR (KBr) cm-1: 3,445,
2,940, 1,681, 1,578, 1,149, 809; 1H NMR (300 MHz, CDCl3)
d: 7.28–7.25 (m, 2H), 7.16–7.13 (m, 2H), 7.10–7.06 (q,
J = 8.1 Hz, 4H), 6.79 (s, 1H), 2.94–2.84 (m, 2H), 2.62–2.61
(d, J = 3.6 Hz, 2H), 2.36 (s, 3H), 2.31 (s, 3H), 2.26–2.22
(m, 2H), 2.13–2.09 (m, 2H), 2.10 (m, 2H), 1.78–1.73
(m, 2H), 1.67–1.63 (m, 2H); HR-MS: m/z, calcd, C23H27NO
(M?): 333.2093, found: 333.2099.
2-(E)-Benzylidene-6-(p-toluidinomethy)cyclohexanone (6a)
Yield: 58 %. m.p.: 83–85 °C; MS (ESI, m/z): 305.3 (M?); IR
1
(KBr) cm-1: 3,412, 2,933, 1,669, 1,608, 1,131, 756; H
NMR (300 MHz, CDCl3) d: 7.43 (s, 1H), 7.38–7.30 (m, 5H),
7.01–7.00 (d, J = 8.2 Hz, 2H), 6.62–6.59 (d, J = 8.3 Hz,
2H), 4.45 (brs, 1H), 3.54–3.48 (q, J = 20.2 Hz, 1H),
3.29–3.23 (q, J = 18.4 Hz, 1H), 3.02–2.97 (d, J = 16.0 Hz,
1H), 2.73–2.68 (m, 2H), 2.24 (s, 3H), 2.12 (m, 1H),
1.95–1.91 (m, 1H), 1.79–1.69 (m, 2H); HR-MS: m/z, calcd,
C21H23NO (M?): 305.1780, found: 305.1778.
2-(E-(4-Methyl)benzylidene-6-(N-methyl-N-(m-tolyl)amino-
methyl) cyclohexanone (6g) Yield: 44 %. m.p.: 101–102
°C; MS (ESI, m/z): 333.5 (M?); IR (KBr) cm-1: 3,423,
2,940, 1,682, 1,578, 1,148, 810; 1H NMR (300 MHz, CDCl3)
d: 7.28–7.25 (m, 2H), 7.19–7.16 (m, 2H), 7.13–7.06 (q,
J = 20.6 Hz, 4H), 6.79 (s, 1H), 2.94–2.86 (m, 2H),
2.63–2.61 (d, J = 3.6 Hz, 2H), 2.36 (s, 3H), 2.32 (s, 3H),
2.24–2.19 (m, 2H), 2.18–2.08 (m, 3H), 1.76–1.73 (m, 1H),
1.65–1.63 (m, 2H); HR-MS: m/z, calcd, C23H27NO (M?):
333.2093, found: 333.2097.
2-(E)-Benzylidene-6-(N-methyl-N-phenylaminomethyl)cyclo
hexanone (6b) Yield: 53 %. m.p.: 129–131 °C; MS (ESI,
m/z): 305.2 (M?); IR (KBr) cm-1: 3,426, 2,943, 1,692,
1,622, 1,134, 756; 1H NMR (300 MHz, CDCl3) d: 7.37–7.34
(m, 5H), 7.26–7.15 (m, 5H), 6.83 (s, 1H), 3.00–2.93 (m, 1H),
2.87–2.82 (m, 1H), 2.66–2.60 (q, J = 17.5 Hz, 2H),
2.29–2.19 (m, 2H), 2.13–2.09 (m, 2H), 2.02–1.96 (m, 2H),
1.78–1.66 (m, 2H); HR-MS: m/z, calcd, C21H23NO (M?):
305.1780, found: 305.1782.
123

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Med Chem Res (2013) 22:3779–3786
2-(E)-(4-Methylsulfonyl)benzylidene-6-(N-methyl-N-pheny-
The crude was purified using a silica gel column with
laminomethyl) cyclohexanone (6h) Yield: 47 %. m.p.:
:
CHCl3–MeOH to give 7a–7j as white solid.
143–145 °C; MS (ESI, m/z): 383.2 (M?); IR (KBr) cm-1
1
3,446, 2,925, 1,685, 1,587, 1,306, 1,145, 769; H NMR
(300 MHz, CDCl3) d: 7.95–7.92 (d, J = 8.3 Hz, 2H),
7.85–7.83 (d, J = 8.3 Hz, 2H), 7.52–7.49 (d, J = 8.2 Hz,
2H), 7.39–7.36 (d, J = 8.3 Hz, 2H), 7.21 (s, 1H), 6.82
(s, 1H), 3.08–3.06 (d, J = 5.3 Hz, 3H), 2.87 (m, 1H),
2.64–2.62 (d, J = 6.3 Hz, 2H), 2.15 (m, 3H), 2.05–2.04 (m,
2H), 1.83–1.68 (m, 4H); HR-MS: m/z, calcd, C22H25NO3S
(M?): 383.1555, found: 383.1562.
2-(E)-Benzylidene-6-(N-methyl-N-(p-tolyl)aminomethyl)-1-
phenylcyclohexanol (7a) Yield: 7.1 %. m.p.: 135–138
°C; [a]D 25 = 08 (c = 1 % MeOH); MS (ESI, m/z): 396
([M-H]-); IR (KBr) cm-1: 3,492, 1,598, 1,448, 1,137,
1
751; H NMR (300 MHz, CDCl3) d: 7.57–7.55 (d, J =
6.7 Hz, 2H), 7.36–7.34 (d, J = 4.3 Hz, 4H), 7.29–7.27 (m,
2H), 7.24–7.23 (m, 4H), 7.10–7.08 (m, 2H), 2.63–2.39 (m,
2H), 2.32–2.22 (m, 2H), 2.18–2.05 (m, 3H), 1.98–1.87 (m,
3H), 1.64–1.47 (m, 5H); HR-ESI-MS: m/z, calcd,
C28H31NO ([M-H]-): 397.2406, found: 397.2417.
2-(E)-(4-Methylsulfonyl)benzylidene-6-(N-methyl-N-(p-tolyl)
aminomethyl) cyclohexanone (6i) Yield: 52 %. m.p.: 136–
139 °C; MS (ESI, m/z): 397.6 (M?); IR (KBr) cm-1: 3,446,
2,925, 1,685, 1,587, 1,306, 1,145, 768; 1H NMR (300 MHz,
CDCl3) d: 7.95–7.92 (d, J = 8.1 Hz, 2H), 7.85–7.83
(d, J = 8.4 Hz, 2H), 7.52–7.49 (d, J = 8.4 Hz, 2H),
7.39–7.36 (d, J = 8.4 Hz, 2H), 6.82 (s, 1H), 3.08–3.06
(d, J = 5.4 Hz, 3H), 2.87 (m, 2H), 2.65–2.63 (d, J = 6.0 Hz,
2H), 2.28–2.15 (m, 5H), 2.05–2.04 (m, 2H), 1.84–1.78
(m, 2H), 1.72–1.68 (t, J = 12.0 Hz, 2H); HR-MS: m/z,
calcd, C23H27NO3S (M?): 397.1712, found: 397.1722.
2-(E)-Benzylidene-6-(N-methyl-N-(m-tolyl)aminomethyl)-1-
phenylcyclohexanol (7b) Yield: 6.3 %. m.p.: 143–145 °C;
MS (ESI, m/z): 396 ([M-H]-); IR (KBr) cm-1: 3,493,
1,597, 1,447, 1,138, 753; 1H NMR (300 MHz, CDCl3)
d: 7.57–7.55 (d, J = 6.8 Hz, 2H), 7.36–7.34 (d, J = 4.4 Hz,
4H), 7.24–7.22 (m, 6H), 7.10–7.08 (d, J = 7.5 Hz, 2H),
2.63–2.44 (m, 2H), 2.39–2.32 (m, 2H), 2.25–2.12 (m, 3H),
1.98–1.86 (m, 3H), 1.62–1.47 (m, 5H); HR-ESI-MS: m/z,
calcd, C28H31NO ([M-H]-): 397.2406, found: 397.2445.
2-(E-(4-Methysulfonyll)benzylidene-6-(N-methyl-N-(mtolyl)
aminomethyl) cyclohexanone (6j) Yield: 50 %. m.p.: 140–
142 °C; MS (ESI, m/z): 397.1 (M?); IR (KBr) cm-1: 3,443,
2,925, 1,686, 1,588, 1,305, 1,146, 769; 1H NMR (300 MHz,
CDCl3) d: 7.95–7.92 (d, J = 8.4 Hz, 2H), 7.85–7.83
(d, J = 8.4 Hz, 2H), 7.52–7.49 (d, J = 8.1 Hz, 2H),
7.39–7.36 (d, J = 8.4 Hz, 2H), 6.82 (s, 1H), 3.08–3.06 (d,
J = 5.1 Hz, 3H), 2.86 (m, 2H), 2.64–2.62 (d, J = 6.3 Hz,
2H), 2.24–2.10 (m, 5H), 2.07–2.04 (m, 2H), 1.83–1.78
(m, 2H), 1.72–1.68 (t, J = 12.3 Hz, 2H); HR-MS: m/z,
calcd, C23H27NO3S (M?): 397.1712, found: 397.1718.
2-(E)-Benzylidene-6-(N-methyl-N-phenylaminomethyl)-1-
(3methoxy)phenylcyclohexanol (7c) Yield: 10.2 %. m.p.:
137–140 °C; MS (ESI, m/z): 413 (M?); IR (KBr) cm-1
:
3,455, 2,831, 1,600, 1,437, 1,140, 759; 1H NMR (300 MHz,
CDCl3) d: 7.36–7.35 (d, J = 8.9 Hz, 2H), 7.28–7.27 (m,
3H), 7.23–7.20 (m, 4H), 7.10–7.02 (m, 5H), 3.75 (s, 3H),
2.58 (m, 1H), 2.49–2.38 (m, 3H), 2.22–2.13 (m, 3H), 2.05
(brs, 2H), 1.63–1.52 (m, 6H); HR-ESI-MS: m/z, calcd,
C28H31NO2 (M?): 413.2355, found: 413.2356.
2-(E)-Benzylidene-6-(N-methyl-N-(p-tolyl)aminomethyl)-1-
(3-methoxy)phenylcyclohexanol (7d) Yield: 11.0 %. m.p.:
143–145 °C; [a]D 25 = 08 (c = 1 % MeOH); MS (ESI,
m/z): 427 (M?); IR (KBr) cm-1: 3,453, 2,831, 1,600,
General procedure for the synthesis of 7a–7l
In a 100-mL three-necked flask with thermometer, a
dropping funnel and reflux condenser with calcium chlo-
ride tube, magnesium turnings (0.02 mol), and iodine (a
grain) was covered with 10 mL purified tetrahydrofuran
(THF) and treated with about 1/20 of a total of substituted
bromobenzene (0.02 mol). After the reaction was started,
the remaining bromobenzene, dissolved in 15 mL THF,
was added drop wise with stirring under THF boiling
gently. The mixture was boiled gently until almost all the
magnesium was dissolved (about 30 min).
1
1,437, 1,139, 757; H NMR (300 MHz, CDCl3) d: 7.34–
7.32 (m, 4H), 7.28 (s, 1H), 7.13–7.11 (m, 4H), 7.09–7.07
(m, 4H), 3.60 (s, 3H), 2.61 (m, 1H), 2.50–2.39 (m, 3H),
2.31–2.22 (m, 3H), 2.05 (brs, 2H), 1.63–1.56 (m, 6H); HR-
ESI-MS: m/z, calcd, C29H33NO2 (M?): 427.2511, found:
427.2509.
2-(E)-Benzylidene-6-(N-methyl-N-(m-tolyl)aminomethyl)-
1-(3-methoxy)phenylcyclohexanol (7e) Yield: 10.8 %.
m.p.: 141–144 °C; MS (ESI, m/z): 426 ([M-H]-); IR
To a solution of substituted cyclohexanone 6a–6j
(0.01 mol) in 30 mL purified THF, fresh Grignard reagent
(0.02 mol) was added dropwise under ice-water bath. After
24 h stirring at room temperature, the mixture was poured
into 37 % (w/v) NH4Cl solution and the solid was produced.
1
(KBr) cm-1: 3,452, 2,833, 1,597, 1,449, 1,138, 755; H
NMR (300 MHz, CDCl3) d: 7.35–7.33 (m, 4H), 7.28–7.26
(m, 1H), 7.23–7.18 (m, 4H), 7.13–7.09 (m, 4H), 3.60 (s,
3H), 2.61 (m, 1H), 2.51–2.39 (m, 3H), 2.29–2.22 (m, 3H),
123

Med Chem Res (2013) 22:3779–3786
3785
1.98 (brs, 2H), 1.65–1.52 (m, 6H); HR-ESI-MS: m/z, calcd,
C29H33NO2 ([M-H]-): 427.2511, found: 427.2523.
2H), 7.25–7.21 (m, 3H), 7.10 (m, 6H), 6.93 (s, 1H), 6.87–6.84
(d, J = 8.7 Hz, 2H), 3.80 (s, 3H), 2.86–2.81 (tt, J = 15.3 Hz,
1H), 2.59–2.81 (m, 2H), 2.34–2.33 (m, 6H), 2.13–2.08 (m,
2H), 1.73–1.72 (m, 2H), 1.52 (m, 2H); HR-ESI-MS: m/z,
calcd, C29H33NO2 (M?): 427.2511, found: 427.2583.
2-(E)-Benzylidene-6-(N-methyl-N-phenyl)aminomethyl-1-
(4-methoxy)phenylcyclohexanonol (7f) Yield: 10.2 %.
m.p.: 146–147 °C; MS (ESI, m/z): 413 (M?); IR (KBr)
1
cm-1: 3,583, 2,825, 1,606, 1,443, 1,149, 753; H NMR
2-(E)-(4-Methyl)benzylidene-6-(N-methy-N-(p-tolyl))ami-
nomethyl-1-(4-methoxy)phenylcyclohexanonol (7k) Yield:
13.5 %. m.p.: 153–155 °C; MS (ESI, m/z): 441 (M?); IR
(300 MHz, CDCl3) d: 7.84–7.82 (d, J = 8.9 Hz, 2H),
7.32–7.30 (m, 6H), 7.22–7.20 (m, 4H), 6.97 (s, 1H),
6.89–6.86 (d, J = 8.9 Hz, 2H), 3.80 (s, 3H), 2.88–2.83
(tt, J = 15.1 Hz, 1H), 2.56–2.52 (m, 3H), 2.03–1.99
(m, 3H), 180–1.73 (m, 3H), 1.61–1.56 (m, 3H); HR-ESI-MS:
m/z, calcd, C28H31NO2 (M?): 413.2355, found: 413.2367.
1
(KBr) cm-1: 3,565, 2,835, 1,609, 1,435, 1,155, 755; H
NMR (300 MHz, CDCl3) d: 7.84–7.82 (d, J = 8.9 Hz,
2H), 7.26–7.24 (m, 2H), 7.19–7.08 (m, 6H), 6.87 (m, 1H),
6.85–6.83 (d, J = 9.0 Hz, 2H), 3.79 (s, 3H), 2.86–2.80
(tt, J = 15.2 Hz, 1H), 2.54 (m, 3H), 2.35 (s, 3H),
2.00–1.94 (m, 5H), 1.68–1.50 (m, 6H); HR-ESI-MS: m/z,
calcd, C30H35NO2 (M?): 441.2668, found: 441.2681.
2-(E)-Benzylidene-6-(N-methyl-N-(p-tolyl))aminomethyl-1-
(4-methoxy)phenylcyclohexanonol (7g) Yield: 11.0 %.
m.p.: 148–150 °C; MS (ESI, m/z): 427 (M?); IR (KBr)
1
cm-1: 3,529, 2,827, 1,605, 1,443, 1,149, 753; H NMR
2-(E)-(4-Methyl)benzylidene-6-(N-methy-N-(m-tolyl))ami-
nomethyl-1-(4-methoxy)phenylcyclohexanonol (7l) Yield:
12.5 %. m.p.: 144–147 °C; [a]D 25 = 08 (c = 1 % MeOH);
MS (ESI, m/z): 441 (M?); IR (KBr) cm-1: 3,565, 2,835,
1,609, 1,434, 1,155, 755; 1H NMR (300 MHz, CDCl3)
d: 7.84–7.81 (d, J = 8.9 Hz, 2H), 7.21–7.18 (m, 2H),
7.18–7.10 (m, 6H), 6.93 (m, 1H), 6.88–6.85 (d, J = 9.0 Hz,
2H), 3.80 (s, 3H), 2.87–2.82 (tt, J = 15.1 Hz, 1H), 2.55 (m,
3H), 2.34 (s, 3H), 2.08–1.99 (m, 5H), 1.82–1.79 (m, 4H), 1.55
(m, 2H); HR-ESI-MS: m/z, calcd, C30H35NO2 (M?): 441.
2668, found: 441.2656.
(300 MHz, CDCl3) d: 7.84–7.81 (d, J = 8.9 Hz, 2H),
7.32–7.30 (m, 5H), 7.21–7.19 (m, 4H), 6.97 (s, 1H), 6.89–
6.86 (d, J = 8.9 Hz, 2H), 3.81 (s, 3H), 2.83 (m, 1H),
2.60–2.52 (m, 3H), 2.17–2.10 (m, 2H), 2.03–1.99 (m, 3H),
1.80–1.73 (m, 3H), 1.61–1.55 (m, 3H); HR-ESI-MS: m/z,
calcd, C29H33NO2 (M?): 427.2511, Found: 427.2532.
2-(E)-(4-Methyl)benzylidene-6-(N-methyl-N-phenyl))ami-
nomethyl-1-(3-methoxy)phenylcyclohexanol (7h) Yield:
10.5 %. m.p.: 144–147 °C; [a]D 25 = 08 (c = 1 % MeOH);
MS (ESI, m/z): 427 (M?); IR (KBr) cm-1: 3,451, 2,860,
1,602, 1,433, 1,145, 753; 1H NMR (300 MHz, CDCl3)
d: 7.86–7.84 (d, J = 8.9 Hz, 2H), 7.32–7.29 (m, 5H),
7.21–7.18 (m, 4H), 6.95 (s, 1H), 6.86–6.84 (d, J = 8.9 Hz,
2H), 3.61 (s, 3H), 2.84–2.67 (m, 4H), 2.28–2.23 (m, 3H),
2.10–1.98 (m, 2H), 1.65–1.56 (m, 6H); HR-ESI-MS: m/z,
calcd, C29H33NO2 (M?): 427.2511, found: 427.2553.
Pharmacology
Male Kunming mice (18–22 g) and Wistar rats
(160 20 g) for the analgesic and anti-inflammatory assay
were supplied by the Experimental Animal Center of
Shenyang Pharmaceutical University. Each group consisted
of ten mice, which were fasted for 12 h before performance.
The vehicle (0.5 % sodium carboxymethyl cellulose, 0.5 %
Na-CMC), positive control (ibuprofen), and test compounds
were administered orally at a 200 mg/kg dose.
2-(E)-(4-Methyl)benzylidene-6-(N-methyl-N-(p-tolyl))ami-
nomethyl-1-(3-methoxy)phenylcyclohexanol (7i) Yield:
11.5 %. m.p.: 146–150 °C; MS (ESI, m/z): 441 (M?); IR
1
(KBr) cm-1: 3,449, 2,862, 1,604, 1,438, 1,141, 755; H
NMR (300 MHz, CDCl3) d: 7.25–7.18 (m, 4H), 7.16–7.11
(m, 6H), 6.92–6.89 (d, J = 7.9 Hz, 2H), 6.75–6.73
(m, 1H), 3.61 (s, 3H), 2.61–2.44 (m, 2H), 2.34 (s, 3H),
2.28–2.25 (m, 1H), 2.21 (s, 3H), 2.06–1.81 (m, 3H),
1.59–1.52 (m, 6H); HR-ESI-MS: m/z, calcd, C30H35NO2
(M?): 441.2688, found: 441.2635.
Analgesic evaluation by acetic acid-induced writhing
test
One hour after the administration of the tested compounds,
the mice were given an intraperitoneal injection of 0.7 %
(v/v)aceticacidsolution(10 mL/kg). Awrithewasdefinedas
stretching of the abdomen with swelling of the trunk and hind
limbs, and rising of the rump. The number of writhes was
counted for 15 min and the percent analgesic activity (PAA)
was calculated by the following formula (Wang et al., 2009),
x ꢁ a
2-(E)-(4-Methyl)benzylidene-6-(N-methyl-N-phenyl)amino-
methyl-1-(4-methoxy)phenylcyclohexanonol (7j) Yield:
13.2 %. m.p.: 149–151 °C; MS (ESI, m/z): 427 (M ?); IR
1
(KBr) cm-1: 3,565, 2,834, 1,609, 1,435, 1,155, 755; H
PAA ¼
ꢂ 100 %
x
NMR (300 MHz, CDCl3) d: 7.83–7.80 (d, J = 09.0 Hz,
123

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Med Chem Res (2013) 22:3779–3786
ꢂ
ꢀ
ꢁꢃ
a ꢁ x
b ꢁ y
where, x is the mean writhing number of mice in the
control group for administration of 0.5 % Na-CMC solu-
tion and a is the mean writhing number of mice adminis-
trated the tested compounds or ibuprofen.
RPE ¼ 1 ꢁ
ꢂ 100 %
where, a is the mean volume at time = 3 h of the test
group, x is the mean volume at time = 0 of the test group,
b is the mean volume at time = 3 h of the control group,
and y is the mean volume at time = 0 of the control group.
Analgesic evaluation by hot plate test
Mice were placed on a hot plate maintained at 55 0.5 °C
and the time in seconds between the placement of mice on
the platform and licking of the hind paw or jumping was
recorded as the pain threshold. Pain threshold measures
were recorded before drug and 1 h after the last drug
administration. Mice exhibiting pain threshold time [30 s
or \5 s were excluded. If the mice did not respond to the
stimulus, the latency was recorded as 60 s.
Acknowledgments This work was supported by a grant from the
National Natural Science Foundation of China (No. 81028015).
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S ꢁ S
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ꢀ
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Carrageenan (1 % in saline) was injected into the plantar
surface of the right hind paw of the rats 1 h after the last
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et al., 2012),
123