Design and Synthesis of 5-Substituted Benzo[d][1,3]dioxole Derivatives as Potent Anticonvulsant Agents

Article abstract of DOI:10.1002/ardp.201600274

A series of 5-substituted benzo[d][1,3]dioxole derivatives was designed, synthesized, and tested for anticonvulsant activity using the maximal electroshock (MES) and subcutaneous pentylenetetrazole (scPTZ) screens. Neurotoxicity was determined by rotarod test. In the preliminary screening, six compounds, 3a, 3c, 3d, and 4d–f, showed promising anticonvulsant activities in the MES model, and compounds 4c and 4d exhibited full protection against seizures at doses of 300 mg/kg in the scPTZ model. Among the synthesized compounds, 3c as the most active compound showed high protection against the MES-induced seizures with an ED₅₀ value of 9.8 mg/kg and a TD₅₀ value of 229.4 mg/kg after intraperitoneal injection into mice, thus providing compound 3c with a high protective index (TD₅₀/ED₅₀) of 23.4 comparable to those of reference antiepileptic drugs.

Full text of DOI:10.1002/ardp.201600274

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Full Paper
Design and Synthesis of 5-Substituted Benzo[d][1,3]dioxole
Derivatives as Potent Anticonvulsant Agents
Shiyang Dong1,2ꢀ, Tiantian Wang1,3ꢀ, Chundi Hu2, Xiaodong Chen1, Yi Jin1,3, and Zengtao Wang1
1
College of Pharmacy, JiangXi University of Traditional Chinese Medicine, Nanchang, China
College of Pharmacy, Hubei University of Science and Technology, Xianning, China
The National Pharmaceutical Engineering Center for Solid Preparation in Chinese Herbal Medicine,
Nanchang, China
2
3
A series of 5-substituted benzo[d][1,3]dioxole derivatives was designed, synthesized, and tested for
anticonvulsant activity using the maximal electroshock (MES) and subcutaneous pentylenetetrazole
(scPTZ) screens. Neurotoxicity was determined by rotarod test. In the preliminary screening, six
compounds, 3a, 3c, 3d, and 4d–f, showed promising anticonvulsant activities in the MES model, and
compounds 4c and 4d exhibited full protection against seizures at doses of 300 mg/kg in the scPTZ
model. Among the synthesized compounds, 3c as the most active compound showed high protection
against the MES-induced seizures with an ED50 value of 9.8 mg/kg and a TD50 value of 229.4 mg/kg
after intraperitoneal injection into mice, thus providing compound 3c with a high protective index
(TD50/ED50) of 23.4 comparable to those of reference antiepileptic drugs.
Keywords: Anticonvulsant activity / Benzo[d][1,3]dioxole derivatives / Design / Synthesis
Received: September 17, 2016; Revised: December 1, 2016; Accepted: December 7, 2016
DOI 10.1002/ardp.201600274
Additional supporting information may be found in the online version of this article at the publisher’s web-site.
:
anticonvulsants for the development of more effective
and safer AEDs [8–10]. An earlier report indicated that
Introduction
stiripentol (STP) was effective against maximal electro-
Epilepsy is
a
chronic neurological disorder affecting
shock (MES)-induced convulsions in mice with ED50 value
of 240 mg/kg [11], and a recent study suggested that STP
as a lactate dehydrogenase (LDH) inhibitor suppressed
seizures in vivo in a mouse model of epilepsy by inhibition
of the metabolic pathway via LDH. Moreover, by further
0.5–1% of the population worldwide [1]. In spite of many
efforts have been made in epilepsy research, the currently
available antiepileptic drugs (AEDs) still present two major
limitations: Firstly, a significant group of patients (up to
30%) are resistant to the available AEDs which fail to
adequately control seizures [2, 3]. Secondly, many AEDs
exhibit an unfavorable side effect profile, such as
headache, nausea, anorexia, ataxia, hepatotoxicity,
drowsiness, gastrointestinal disturbance, gingival hyper-
plasia, and hirsutism [4–7]. These facts drive the ongoing
search of novel AEDs, there is a substantial need to design
modifying STP,
(isosafrole) was identified, which shared
a
previously unknown LDH inhibitor
common
a
skeleton of benzo[d][1,3]dioxole and also potently sup-
pressed seizures in vivo (Fig. 1) [12].
These results hinted that the skeleton of benzo[d][1,3]-
dioxole should play an important role in keeping
anticonvulsant activity. Prompted by these observations,
we herein synthesized a series of benzo[d][1,3]dioxole
derivatives, and evaluated their anticonvulsant activities
with a view to explore their potency as anticonvulsant
agents.
Correspondence: Dr. Zengtao Wang, Department of Medicinal
Chemistry, College of Pharmacy, JiangXi University of Traditional
Chinese Medicine, Nanchang 330004, China.
E-mail: zengtaowang@126.com
ꢀThese two authors contributed equally to this article.
Fax: þ86-791-87118911
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4a–f carrying different condensation reaction. 3a–c were
synthesized by Knoevenagel condensation reaction, that is
treatment of appropriate intermediate (2) with five-
membered heterocyclic ring containing an internal amide
(such as imidazolidine-2,4-dione, thiazolidine-2,4-dione, and
2-thioxothiazolidin-4-one) at refluxing temperature in pres-
ence of sodium acetate (AcONa) and acetic acid (AcOH).
Similarly, the condensation of aldehyde (2) with oxindole
was accomplished with subsequent addition of piperidine at
refluxing temperature in presence of ethanol (EtOH), which
easily formed methyl 3-(benzo[d][1,3]dioxol-5-ylmethylene)-
indolin-2-one 3d. Finally, imines 4a–f were obtained by the
condensation of aldehyde (2) with different anilines,
respectively. The detailed synthesis, physical data, spectral
data are listed in the Experimental section.
Figure 1. Reported and designed benzo[d][1,3]dioxole
derivatives as anticonvulsants.
Pharmacology
Results and discussion
The anticonvulsant activity and neurotoxicity of all synthesized
compounds 3a–d and 4a–f were evaluated according to the
standard protocols within the Antiepileptic Drug Development
(ADD) Program at the National Institute of Neurological
Disorders and Stroke, National Institutes of Health, Rockville,
USA [14, 15]. The phase I evaluation included the maximal
electroshock seizure (MES), the subcutaneous pentylenetetra-
zole (scPTZ), and neurotoxicity [16]. In our study, the test
compounds were administered via ip injection at the doses of 30,
Chemistry
The synthesis of compounds 3a–d and 4a–f was accomplished
as shown in Scheme 1. Initially, 3,4-dihydroxy benzaldehyde
(1) reacted with dichloromethane (CH2Cl2) in presence of
dimethylformamide (DMF) according to the published
method that yielded benzo[1,3]dioxole-5-carbaldehyde
(2) [13]. Later on, the intermediate (2) was used as active
scaffolds for the synthesis of target compounds 3a–d and
Scheme 1. Synthesis route of the target
compounds.
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Arch. Pharm. Chem. Life Sci. 2017, 349, e1600274
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Table 1. Phase I anticonvulsant screening and minimal motor impairment of the synthesized compounds (3a–d and
4a–f).
MES screena)
scPTZ screenb)
NT screenc)
Compounds
0.5 h
4 h
0.5 h
4 h
0.5 h
4 h
3a
3b
3c
3d
4a
4b
4c
4d
30
300
30
30
–
30
300
30
30
–
–
300
–
–
–
100
300
300
300
300
300
100
100
300
300
100
100
–
100
300
300
300
300
300
300
300
300
–
–
–
–
300
300
300
300
–
–
100
–
300
–
–
300
300
–
100
100
300
30
100
–
300
300
100
30
30
–
4e
4f
–
–
Phenytoind)
Phenobarbitald)
Ethosuximided)
–
–
100
300
–
30
100
300
300
The dash (ꢁ) indicates the absence of activity at the maximum dose administered (300 mg/kg).
a)
Maximal electroshock test.
Subcutaneous pentylenetetrazole test.
Neurotoxocity screening (rotarod test).
Data from refs. [17–19].
b)
c)
d)
100, and 300 mg/kg in the MES screening whereas 100 and
300 mg/kg in the scPTZ screening and neurotoxicity. An
observation was carried out at two different time intervals,
namely 0.5 and 4 h. The screening results of anticonvulsant
activity and neurotoxocity studies were summarized in Table 1.
Most of the compounds (except 4a and 4c) showed
significant protection in MES screen which reflects the
potential of these compounds against generalized tonic–
clonic seizure. Compounds 3a, 3c, and 3d showed protection
from seizure at the lowest dose of 30 mg/kg at 0.5 and 4 h,
which indicated the promising nature of the compound
having quick onset and long duration of action at lower
doses. The slight difference between the structures of
compounds 3b and 3c that the oxygen atom of 3b is
replaced by sulfur isostere, resulting in compound 3c that is
more potent as anticonvulsant agents. This substitution
interestingly renders about 100-fold increase in activity from
3b to 3c. Obtained imines derivatives 4b, 4d, 4e, and 4f
showed moderate activity at doses of 100 mg/kg at 0.5 or 4 h
post administration.
In the scPTZ screen, compounds 3b and 4a–d inhibited
seizures, which identifies substances elevating seizure thresh-
old. Especially, imines derivatives 4c and 4d exhibited full
protection against seizures at doses of 300 mg/kg at 0.5 and
4 h post administration.
In the neurotoxicity screening, majority of compounds were
devoid of minimal motor impairment at the doses of 30 and
100 mg/kg after 0.5 and 4 h. While 4c and 4d elucidated
neurotoxicity at 100 mg/kg at 0.5 h followed by an increase in
non-toxic potential after delayed absorption at 4 h by
preventing minimal motor impairment at 300 mg/kg, and
compound 3a equalized the effect of the standard drug
phenytoin.
Since the results of preliminary screening indicated the
compounds that contain hydantoin or thiazolidinones ring
displayed remarkable activity compared with imines deriva-
tives in MES model, therefore, the compounds 3a–d were
further subjected to phase II screening for quantification of
their anticonvulsant activity (indicated by ED50) and neuro-
toxicity (indicated by TD50) in mice, and the results are
presented in Table 2.
Compound 3a with an ED50 value of 28.8 mg/kg in the MES
model exhibited neurotoxicity at a dose of 100 mg/kg. It is
worth mentioning that compound 3b has little difference in
the structure compared with 3c, which showed especially low
activity (ED50 > 200). 3c and 3d had ED50 values of 9.8 and
11.0 mg/kg in the MES model and TD50 values of 229.4 and
247.5 mg/kg resulting in high protective index (PI) values of
TD50/ED50 ¼ 23.4 and 22.5, respectively. The PI values of the 3c
(23.4) and 3d (22.5) were higher than the reference drugs as
compared to 6.9 for phenytoin and 3.2 for phenobarbital. The
PI value is considered to be an index representing the margin
of safety and tolerability between anticonvulsant doses and
doses of anticonvulsant drugs exerting acute adverse effects.
The order of anticonvulsant activities in the MES test as
judged from ED50 values is 3c > 3d > 3a > 3b. Actually, earlier
structure–activity relationship (SAR) studies with stiripen-
tol [21] have uncovered remarkable anticonvulsant activity for
skeleton of benzo[d]-[1,3]dioxole. Aboul-Enein et al. [21]
attached semicabazone moiety to the backbone of STP
(strategy a) and cyclization of the semicarbazone (strategy
b) to afford the semicarbazone 5 and racemic pyrazoline 6,
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Table 2. Phase II quantitative anticonvulsant evaluation in MES model in mice.
TPE (h)a)
ED50
TD50
PId)
b)
c)
Compounds
3a
3b
3c
3d
1
1
1
3
2
1
28.8 (14.4–82.6)e)
>200
9.8 (3.0–18.1)
11.0 (4.7–20.7)
9.5 (8.1–10.4)
21.8 (21.8–25.5)
ndf)
ndf)
229.4 (197.8–264.8)
247.5 (216.1–283.5)
65.5 (52.5–72.9)
69 (62.8–72.9)
23.4
22.5
6.9
Phenytoing)
Phenobarbitalg)
3.2
a) Time to peak effect.
b) ED50: median effective dose affording anticonvulsant protection in 50% of animals, the dose is measured in mg/kg.
c) TD50: median toxic dose eliciting minimal neurological toxicity in 50% of animals, the dose is measured in mg/kg.
d) PI: protective index (TD50/ED50).
e) 95% confidence intervals given in parentheses.
f) Not determined.
g) Data from ref. [14, 20].
Conclusion
respectively (Fig. 2). Interestingly, compound 5 showed a
good ED50 value of 87 mg/kg in the MES model. In contrast to
5, a good activity against scPTZ-induced seizures was observed
in compound 6 (ED50 ¼ 110 mg/kg).
In the present study, we have synthesized a series of
5-substituted benzo[d][1,3]dioxole derivatives as anticonvul-
sant agents. All the synthesized compounds were screened for
their anticonvulsant activity by MES and scPTZ induced
seizures test. Six compounds 3a, 3c, 3d, and 4d–f showed
promising anticonvulsant activities in MES model, and imines
derivatives 4c and 4d exhibited full protection against seizures
at doses of 300 mg/kg in scPTZ model. The most active
compound 3c showed the MES-induced seizures with
ED50 value of 9.8 mg/kg and TD50 value of 229.4 mg/kg, which
In this study, we conducted the strategy (c) that a five-
membered ring could be used to modify neopentyl alcohol
based on the cyclization of the amide linker, which gave
products 3a–d. The SAR of 3a–d, 5, and 6 are presented in
Fig. 2. Collectively, the SAR analyses indicated that the
modification of neopentyl alcohol in the strategy (c) was
important, and enhanced the anticonvulsant activities of
STP analogs in the MES model, such as compounds
3a (ED50 ¼ 28.8 mg/kg), 3c (ED50 ¼ 9.8 mg/kg), and 3d
(ED50 ¼ 11.0 mg/kg), which were more active than 5. 3c
was the most potent of all tested and promising compounds
in this work, which exhibited quick absorption with short
time to peak effect (1 h).
provided compound 3c with
a high protective index
(TD50/ED50) of 23.4 comparable to reference antiepileptic
drugs. These obtained results showed that certain compounds
could be useful as a template for future design, modification,
and investigation to produce more active analogs.
Figure 2. The SAR of 3a–d and reported STP
analogs.
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11.19 (s, br, 1H, NH). 13C-NMR (DMSO-d6, 600 MHz,): d 101.88
(CH2), 109.15 (Ar–C7), 109.20 (Ar–C4), 109.23 (CH), 125.29
Experimental
–
Chemistry
(Ar–C6), 126.77 (C CH), 127.51(Ar–C5), 148.02 (Ar–C–O),
–
–
–
–
General
148.28 (Ar–C–O), 156.09 (C O), 166.05 (C O). ESI-HRMS calcd
–
Melting points were determined in open capillary tubes and
are uncorrected. Infrared spectra were measured on a
PerkinElmer infrared (FTIR) spectroscopy instrument over a
wave number range of 4000–450 cmꢁ1. 1H-NMR spectra were
measured on a Bruker Avance 600 MHz NMR spectrometer,
with all chemical shifts given in ppm relative to tetramethyl-
silane. Chemical shift values are in hertz. Splitting patterns are
indicated as s, singlet; d, doublet; t, triplet; m, multiplet; dd,
doublet of doublets for 1H-NMR data. High resolution mass
spectra were recorded on an AB SCIEX Triple TOFTM 5600
equipped with an electrospray ionization source. Dichloro-
methane/methanol (15:1) (3a–d) and ethyl acetate/nhexane
(1:5) (2 and 4a–f) were used as developing solvent for TLC. The
major chemicals were purchased from Xiya Reagent Co., Ltd
(China).
for C11H8N2O4 ([MþH]þ): 233.0480; found: 233.0556.
(Z)-5-Benzo[d][1,3]dioxole-5-ylmethylene-thiazolidine-
2,4-dione (3b)
Rf ¼ 0.61 (CH2Cl2/CH3OH ¼ 15:1); yield: 85%; brown solid; mp:
256–258°C; FT-IR: 3400–2900 (br, NH), 3145, 3008, 2919, 1741,
1694, 1587, 1500, 1443, 1340, 1263, 1154, 1097, 1036, 923,
804 cmꢁ1 1H-NMR (DMSO-d6, 600 MHz): d 6.13 (s, 2H, CH2),
;
7.08 (d, 1H, J ¼ 7.8 Hz, Ar–H7), 7.11 (d, 1H, J ¼ 1.2 Hz, Ar–H4),
7.14 (dd, 1H, J ¼ 1.2, 7.8 Hz, Ar–H6), 7.70 (s, 1H, CH), 12.53 (s, br,
1H, NH). 13C-NMR (DMSO-d6, 600 MHz): d 102.48 (CH2), 109.59
–
–
(Ar–C ), 109.66 (Ar–C ), 121.38 (C CH), 126.34 (Ar–C ), 127.63
7
4
6
(Ar–C5), 132.38 (CH), 148.61 (Ar–C–O), 149.73 (Ar–C–O), 167.82
(C O), 168.30 (C O). ESI-HRMS calcd for C11H7NO4S ([MþH]þ):
–
–
–
–
250.0096; found: 250.0168.
The InChI codes of the investigated compounds together
with some biological activity data are provided as Supporting
Information.
(Z)-5-Benzo[d][1,3]dioxole-5-ylmethylene-2-thioxo-
thiazolidine-4-dione (3c)
Rf ¼ 0.67 (CH2Cl2/CH3OH ¼ 15:1); yield:87%; brown solid; mp:
301–302°C; FT-IR: 3400–2800 (br, NH), 3145, 2985, 1689, 1579,
1496, 1428 1368, 1307, 1262, 1216, 1190, 1097, 1032, 918,
Synthesis of benzo[d][1,3]dioxole-5-carbaldehyde (2)
A solution of 3,4-dihydroxybenzaldehyde (2.77 g, 20 mmol) in
DMF (50 mL) was added dropwise to a suspension of CH2Cl2
(1.9 mL, 30 mmol) and K2CO3 (5.5 g, 40 mmol) in DMF (50 mL).
The mixture was heated under reflux for overnight, then
cooled and filtered. The filtrate was washed by water and
extracted with diethyl ether (3 ꢂ 100 mL). The organic layers
werecombinedandwashedby10%NaOH,thendried(Na2SO4)
and evaporated to give product 2 as white solid. Rf ¼ 0.45; mp:
36–38°C; yield: 83%; FT-IR: 3081, 3000, 2918, 2852, 2793, 2752,
802 cmꢁ1 1H-NMR (DMSO-d6, 600 MHz): d 6.15 (s, 2H, CH2),
;
7.11 (d, 1H, J ¼ 8.4 Hz, Ar–H7), 7.14 (d, 1H, J ¼ 1.8 Hz, Ar–H4),
7.18 (dd, 1H, J ¼ 1.8, 8.4 Hz, Ar–H6), 7.59 (s, 1H, CH), 13.78 (s, br,
1H, NH). 13C-NMR (DMSO-d6, 600 MHz): d 102.63 (CH2), 109.79
–
(Ar–C ), 109.99 (Ar–C ), 123.38 (C CH), 127.19 (Ar–C ), 127.65
–
7
4
6
(Ar–C5), 132.41 (CH), 148.80 (Ar–C–O), 150.17 (Ar–C–O), 169.88
(C O), 195.88 (C S). ESI-HRMS calcd for C11H7NO3S2 ([MþH]þ):
–
–
–
–
265.9867; found: 265.9946.
2722, 1673, 1600, 1490, 1443, 1250, 1032, 928, 860, 812 cmꢁ1
;
1H-NMR (CDCl3, 600 MHz): d 6.05 (s, 2H, CH2), 6.90 (d, 1H,
J ¼ 8.4 Hz, Ar–H7), 7.29 (d, 1H, J ¼ 1.2 Hz, Ar–H4), 7.38 (dd, 1H,
J ¼ 1.2, 8.4 Hz, Ar–H6), 9.77 (s, 1H, CHO). ESI-HRMS calcd for
C8H6O3 ([MþH]þ): 151.0317; found: 151.0383.
(E)-3-Benzo[d][1,3]dioxol-5-ylmethylene-indolin-2-one
(3d)
A mixture of benzo[d][1,3]dioxole-5-carbaldehyde 2 (0.48 g,
3.2 mmol), piperidine, and 1,3-dihydroindol-2-one (0.425 g,
3.2 mmol) was dissolved in EtOH (20 mL), then heated at 90°C
for 4 h. The reaction mixture was cooled and the solvent
removed under vacuum. The resulting residue was further
washed with hexane, and recrystallized by ethanol to give the
product 3d. Rf ¼ 0.6; yield: 64%; brown solid; mp: 218–220°C;
FT-IR: 3400–2700 (br, n(N-H)), 3132, 3073, 3021, 2900, 1704,
1610, 1496, 1448, 1345, 1264, 1219, 1102, 1031, 925, 870, 812,
General procedure for the synthesis of 3a–c
A mixture of benzo[d][1,3]dioxole-5-carbaldehyde 2 (0.314 g,
2.1 mmol), acetic acid glacial (15 mL), and AcONa (0.413 g,
5.0 mmol) was heated with stirring until complete dissolution
occured, then hydantoin or thiazolidinones (2.3 mol) were
added and refluxed for 24 h. The reaction mixture was poured
into cold water and filtered, the resulted solids were
recrystallized with ethanol.
738 cmꢁ1 1H-NMR (DMSO-d6, 600 MHz): d 6.13 (s, 2H, CH2),
.
6.88 (d, 1H, J ¼ 7.8, Ar–H7), 6.88 (t, 1H, J ¼ 7.8, Ar0–H5), 7.07 (d,
1H, J ¼ 7.8, Ar0–H7), 7.22 (t, 1H, J ¼ 7.8, Ar0–H6), 7.29 (s, 1H,
Ar–H4), 7.29 (d, 2H, J ¼ 7.8 Hz, Ar–H6), 7.54 (s, 1H, CH), 7.64 (d,
1H, J ¼ 7.8, Ar0–H4), 10.57 (s, br, 1H, NH). 13C-NMR (DMSO-d6,
600 MHz): d 102.15 (CH2), 109.13 (Ar–C7), 109.78 (Ar00–C7),
(Z)-5-Benzo[d][1,3]dioxole-5-ylmethylene-imidazolidine-
2,4-dione (3a)
Rf ¼ 0.43; yield: 84%; reddish brown solid; mp: 258–260°C; FT-
–
IR: 3400–2900 (br, NH), 3045, 2901, 1708, 1659, 1503, 1448,
110.56 (Ar–C ), 121.46 (C CH), 121.55 (Ar–C ), 122.69 (Ar –C ),
–
4
6
4
1347, 1285, 1261, 1232, 1107, 1035, 1106, 927, 871 cmꢁ1
;
1H-
125.06 (Ar0–C5), 126.57 (Ar0–C–C), 128.67 (Ar0–C6), 130.30
NMR (DMSO-d6, 600 MHz): d 6.07 (s, 2H, CH2), 6.37 (s, 1H, CH),
6.96 (d, 1H, J ¼ 8.4 Hz, Ar–H7), 7.13 (dd, 1H, J ¼ 1.8, 8.4 Hz,
Ar–H6), 7.28 (d, 1H, J ¼ 1.8 Hz, Ar–H4), 10.46 (s, br, 1H, NH),
(Ar–C5), 136.46 (CH), 143.23 (Ar0–C–N), 148.08 (Ar–C–O),
–
149.13 (Ar–C–O), 169.31 (C O). ESI-HRMS calcd for
–
C
16H11NO3 ([MþH]þ): 266.0739; found: 266.0814.
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General procedure for the synthesis of 4a–f
927, 874, 771, 706 cmꢁ1
.
1H-NMR (CDCl3, 600 MHz): d 6.08 (s,
A mixture of benzo[d][1,3]dioxole-5-carbaldehyde 2 (0.61 g,
4.0 mmol), few drops of glacial acetic acid, aniline or
substituted anilines (4.0 mmol) was dissolved in dry ethanol
(15 mL), then the mixture was refluxed for 4–5 h. After the
reaction was completed, the reaction mixture was cooled and
poured into ice water, dried, and recrystallized with absolute
ethanol to give compounds 4a–f.
2H, CH2), 6.91 (d, 1H, J ¼ 7.8 Hz, Ar1–H7), 6.95–7.00 (m, 2H,
Ar2–H3, and Ar2–H5), 7.04–7.09 (m, 1H, Ar2–H4), 7.31 (dd, 1H,
J ¼ 1.2, 7.8 Hz, Ar1–H6) 7.60 (d, 1H, J ¼ 1.2 Hz, Ar1–H4), 8.50 (s,
1H, CH). ESI-HRMS calcd for C14H9F2NO2 ([MþH]þ): 262.0601;
found: 262.0672.
(E)-N-(Benzo[d][1,3]dioxol-5-ylmethylene)-2-
fluoroaniline (4f)
(E)-N-(Benzo[d][1,3]dioxol-5-ylmethylene)aniline (4a)
Rf ¼ 0.50; yield: 56.4%; pale yellow solid; mp: 64–66°C; FT-IR:
3062, 2785, 1623, 1601, 1495, 1448, 1261, 1210, 1104, 1038, 934,
812, 767,697cmꢁ1. 1H-NMR(CDCl3, 600 MHz):d6.07(s, 2H,CH2),
6.91 (d, 1H, J ¼ 7.8 Hz, Ar1–H7), 7.20–7.25 (m, 3H, Ar2–H), 7.30
(dd,1H,J ¼ 1.2 Hz,7.8 Hz,Ar1–H6),7.39–7.42(m,2H,Ar2–H),7.56
(d, 1H, J ¼ 1.2 Hz, Ar1–H4), 8.36 (s, 1H, CH). ESI-HRMS calcd for
Rf ¼ 0.62; yield: 74.3%; pale yellow solid; mp: 72–74°C; FT-IR:
3020, 2907, 2788, 1623, 1585, 1496, 1450, 1262, 1228, 1103,
1030, 927, 822, 750 cmꢁ1. 1H-NMR (CDCl3, 600 MHz): d 6.07 (s,
2H, CH2), 6.91 (d, 1H, J ¼ 7.8 Hz, Ar1–H7), 7.13–7.18 (m, 4H,
Ar2H), 7.31 (dd, 1H, J ¼ 1.2, 7.8 Hz, Ar1–H6), 7.59 (d, 1H,
J ¼ 1.2 Hz, Ar1–H4), 8.42 (s, 1H, CH). ESI-HRMS calcd for
C
14H10FNO2 ([MþH]þ): 244.0696; found: 244.0772.
C
14H11NO2 ([MþH]þ): 226.0790; found: 226.0861.
Pharmacology
Maximal electroshock test (MES)
(E)-N-(Benzo[d][1,3]dioxol-5-ylmethylene)-3-
chloroaniline (4b)
The maximal electroshock seizure test was carried out
according to the standard protocol [14]. Male mice (Kunming,
China), weighing 20–25 g, were used as experimental animals.
Mice were housed under temperature-controlled conditions
(25–30°C) and a 12-h light/dark cycle. They were allowed to
acclimatize with free access to food and water for a 24-h
period before testing except during the experiment. Aboli-
tion of hind limb tonic extension spasm was recorded as
anticonvulsant activity. The test compounds were dissolved in
an aqueous solution of 50% polyethylene glycol. In prelimi-
nary screening, each compound was administered as an ip
injection at three dose levels (30, 100, 300 mg/kg body mass)
and the anticonvulsant activity was assessed after 0.5 and 4 h
intervals of administration.
Rf ¼ 0.64; yield: 62.0%; white solid; mp: 60–61°C; FT-IR: 3060,
3011, 2887, 1629, 1601, 1501, 1440, 1254, 1210, 1089, 1032,
929, 862, 821, 775, 683 cmꢁ1. 1H-NMR (CDCl3, 600 MHz): d 6.07
(s, 2H, CH2), 6.91 (d, 1H, J ¼ 7.8 Hz, Ar1–H7), 7.08–7.10 (m, 1H,
Ar2–H), 7.19–7.22 (m, 2H, Ar2–H), 7.29 (dd, 2H, J ¼ 1.2 Hz,
7.8 Hz, Ar1–H6), 7.32 (t, 1H, J ¼ 7.8 Hz, Ar2–H5), 7.53 (d, 1H,
J ¼ 1.2 Hz, Ar1–H4), 8.32 (s, 1H, CH). ESI-HRMS calcd for
C
14H10ClNO2 ([MþH]þ): 260.0400; found: 260.0472.
(E)-N-(Benzo[d][1,3]dioxol-5-ylmethylene)-3-
nitroaniline (4c)
Rf ¼ 0.45; yield: 79.9%; pale yellow solid; mp: 116–118°C; FT-IR:
3076, 2783, 1628, 1596, 1498, 1447, 1523, 1344, 1261, 1211, 1081,
1036,932,816,800,689 cmꢁ1.1H-NMR(CDCl3,600 MHz):d6.10(s,
2H, CH2), 6.94 (d, 2H, J ¼ 7.8 Hz, Ar1–H7), 7.33 (d, 2H, J ¼ 1.2,
7.8 Hz, Ar1–H6), 7.52–7.54 (m, 1H, Ar2–H6), 7.56 (d, 1H, J ¼ 1.2 Hz,
Ar1–H4), 7.57 (t, 1H, J ¼ 7.8 Hz, Ar2–H5), 8.04 (t, 1H, J ¼ 1.8 Hz,
Ar2–H2), 8.09–8.10 (m, 1H, Ar2–H4), 8.39 (s, 1H, CH). ESI-HRMS
calcd for C14H10N2O4 ([MþH]þ): 270.0641; found: 271.0713.
Subcutaneous pentylenetetrazole seizure test (scPTZ)
This test involved treating the mice with metrazol (penty-
lenetetrazole, 85 mg/kg in mice). This produced clonic
seizures lasting for a period of at least 5 s in 97% of the
animals tested. At the anticipated time of testing, the
convulsant was subcutaneously administered. The test
compound was intraperitoneally administered in mice and
the animals were observed over a 30-min period. Mice were
tested 0.5 and 4 h after doses of 100 and 300 mg/kg of the
test compound were administered. The absence of clonic
spasms over the period of observation indicated the
compound’s ability to abolish the effect of pentylenetetrazol
on the seizure threshold.
(E)-N-(Benzo[d][1,3]dioxol-5-ylmethylene)-4-(tert-butyl)-
aniline (4d)
Rf ¼ 0.70; yield: 91.8%; pale yellow solid; mp: 92–93°C; FT-IR:
3084, 2962, 2863, 2776, 1624, 1589, 1501, 1442, 1252, 1214,
1100, 1038, 932, 875, 830, 812 cmꢁ1
.
1H-NMR (CDCl3,
600 MHz): d 1.37 (s, 9H, 3CH3), 6.06 (s, 2H, CH2), 6.90 (d, 1H,
J ¼ 7.8 Hz, Ar1–H7), 7.17 (dt, 2H, J ¼ 2.4, 8.4 Hz, Ar2–H2, and
Ar2–H6), 7.29 (dd, 1H, J ¼ 1.2, 7.8 Hz, Ar1–H6), 7.43 (dt, 2H,
J ¼ 2.4, 8.4 Hz, Ar2–H3, and Ar2–H5), 7.56 (d, 1H, J ¼ 1.2 Hz,
Ar1–H4), 8.38 (s, 1H, CH). ESI-HRMS calcd for C18H19NO2
([MþH]þ): 282.1416; found: 282.1486.
Neurotoxicity screening
The rotarod test was used to evaluate neurotoxicity. The mice
were trained to stay on an accelerating rotarod that rotated
at six revolutions per minute. The rod diameter was 3.2 cm.
Trained animals were given ip injection of the test compounds
in doses of 100 and 300 mg/kg. Neurotoxicity was indicated by
the inability of the animal to maintain equilibrium on the rod
for at least 1 min in each of the three trials.
(E)-N-(Benzo[d][1,3]dioxol-5-ylmethylene)
-2,6-difluoroaniline (4e)
Rf ¼ 0.64; yield: 57.9%; white solid; mp: 89–90°C; FT-IR: 3076,
2913, 2797, 1631, 1600, 1500, 1451, 1262, 1236, 1212, 1040,
ß 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
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RCHH HARM
A P
Arch. Pharm. Chem. Life Sci. 2017, 349, e1600274
Benzo[d][1,3]dioxole Derivatives as Anticonvulsant Agents
Archiv der Pharmazie
Quantification studies
[7] Z. Lin, P. K. Kadaba, Med. Res. Rev. 1997, 17, 537–572.
[8] M. Bialer, S. I. Johannessen, R. H. Levy, E. Perucca,
T. Tomson, H. S. White, Epilepsy Res. 2010, 92, 89–124.
[9] M. Bialer, B. Yagen, Neurotherapeutics: J. Am. Soc. Exp.
NeuroTherapeutics 2007, 4, 130–137.
[10] H. Nau, W. Loscher, Fundam. Appl. Toxicol. 1986, 6,
669–676.
[11] M. Poisson, F. Huguet, A. Savattier, F. Bakri-Logeais,
G. Narcisse, Arzneim. Forsch. 1984, 34, 199–204.
[12] N. Sada, S. Lee, T. Katsu, T. Otsuki, T. Inoue, Science
(New York, N.Y.) 2015, 347, 1362–1367.
The ED50 was calculated using the Bliss method. The ED50
values were presented as the mean with 95% confidence
intervals. Neurotoxicity was expressed as the median toxic
dose (TD50 in mg/kg) eliciting minimal neurological toxicity in
50% of animals. The quantitative determinations of ED50 and
TD50 values were performed at the previously estimated time
of peak effect after ip injection into mice. Groups of six
animals received various doses of compounds 3a–d until at
least three points were established in the range of 10–90%
seizure protection. The results are shown in Table 2.
[13] D.-D. Li, F. Fang, J.-R. Li, Q.-R. Du, J. Sun, H.-B. Gong,
H.-L. Zhu, Bioorg. Med. Chem. Lett. 2012, 22, 5870–5875.
[14] R. L. Krall, J. K. Penry, B. G. White, H. J. Kupferberg,
E. A. Swinyard, Epilepsia 1978, 19, 409–428.
[15] R. J. Porter, J. J. Cereghino, G. D. Gladding, B. J. Hessie,
H. J. Kupferberg, B. Scoville, B. G. White, Cleve. Clin. Q.
1984, 51, 293–305.
This work was supported by the Science and Technology
Research Project of Jiangxi Provincial Education Department
(No. GJJ150845), Traditional Chinese Medicine Research
Project of Jiangxi Provincial Health and Family Planning
Commission (No.2016A035) and PhD Start-up Fund of Jiangxi
University of Traditional Chinese Medicine (No. 2014BS016).
HYPERLINK “http://dict.cn/original%20text“ .
[16] N. W. Dunham, T. S. Miya, J. Am. Pharm. Assoc. 1957, 46,
208–209.
The authors have declared no conflict of interest.
[17] V. Jatav, P. Mishra, S. Kashaw, J. P. Stables, Eur. J. Med.
Chem. 2008, 43, 1945–1954.
[18] H. Rajak, R. Deshmukh, N. Aggarwal, S. Kashaw,
M. D. Kharya, P. Mishra, Arch. Pharm. 2009, 342,
453–461.
References
[1] B. Malawska, Curr. Top Med. Chem. 2005, 5, 69–85.
[2] J. A. French, Epilepsia 2007, 1 (48 Suppl), 3–7.
[3] P. Kwan, M. J. Brodie, N. Engl. J. Med. 2000, 342, 314–
319.
[4] E. Perucca, J. French, M. Bialer, Lancet Neurol. 2007, 6,
793–804.
[19] J. R. Dimmock, S. N. Pandeya, J. W. Quail,
U. Pugazhenthi, T. M. Allen, G. Y. Kao, J. Balzarini,
E. DeClercq, Eur. J. Med. Chem. 1995, 30, 303–314.
[20] P. Yogeeswari, D. Sriram, R. Thirumurugan, J. V.
Raghavendran, K. Sudhan, R. K. Pavana, J. Stables,
J. Med. Chem. 2005, 48, 6202–6211.
[5] M. L. Wagner, Am. J. Hosp. Pharm. 1994, 51, 1657–1666.
[6] G. Zaccara, D. Franciotta, E. Perucca, Epilepsia 2007, 48,
1223–1244.
[21] M. N. Aboul-Enein, A. A. El-Azzouny, M. I. Attia,
Y. A. Maklad, K. M. Amin, M. Abdel-Rehim, M. F.
El-Behairy, Eur. J. Med. Chem. 2012, 47, 360–369.
ß 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
www.archpharm.com
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