Anticonvulsant and toxicity evaluation of some 7-alkoxy-4,5-dihydro-[1,2,4]triazolo[4,3-a]quinoline-1(2H)-ones

Article abstract of DOI:10.1016/j.bmc.2006.06.044

To further investigate anticonvulsant activity of quinoline derivatives, a series of 7-alkoxy-4,5-dihydro-[1,2,4]triazolo[4,3-a]quinoline-1(2H)-one derivatives was synthesized starting from 7-hydroxyl-3,4-dihydro-2(1H)-quinoline. In initial (phase I) scre

Full text of DOI:10.1016/j.bmc.2006.06.044

Bioorganic & Medicinal Chemistry 14 (2006) 6868–6873
Anticonvulsant and toxicity evaluation of some 7-alkoxy-4,5-
dihydro-[1,2,4]triazolo[4,3-a]quinoline-1(2H)-ones
Hong-Guang Jin,d Xian-Yu Sun,b Kyu-Yun Chai,c Hu-Ri Piaob and Zhe-Shan Quana,b,
*
aKey Laboratory of Organism Functional Factors of the Changbai Mountain, Ministry of Education,
Yanbian University, Yanji, Jilin 133002, PR China
bCollege of Pharmacy, Yanbian University, Yanji, Jilin 133000, PR China
cDepartment of Chemistry, Wonkwang University, Iksan 570-749, Republic of Korea
dDepartment of Pharmacy, Jilin Medical College, Jilin 132130, PR China
Received 18 May 2006; revised 17 June 2006; accepted 19 June 2006
Available online 14 July 2006
Abstract—To further investigate anticonvulsant activity of quinoline derivatives, a series of 7-alkoxy-4,5-dihydro-[1,2,4]triazolo[4,3-
a]quinoline-1(2H)-one derivatives was synthesized starting from 7-hydroxyl-3,4-dihydro-2(1H)-quinoline. In initial (phase I) screen-
ing and quantitative (phase II) evaluation, compound 7-benzyloxyl-4,5-dihydro-[1,2,4]thiazolo[4,3-a]quinoline-1(2H)-one (3f) was
among the most active but also has the lowest toxicity. In the anti-MES potency test, it showed median effective dose (ED50) of
12.3 mg/kg, median toxicity dose (TD50) of 547.5 mg/kg, and the protective index (PI) of 44.5, which is much greater than PI of
the prototype drugs phenytoin, phenobarbital, carbamazepin, and valproate. Compound 3f was chosen for further evaluation.
In phase III pharmacological test, the compound had median hypnotic dose (HD50) and median lethal dose (LD50) of 1204 mg/
kg and >3000 mg/kg, respectively, thus demonstrating much greater margin of safety compared to prototype drugs. The compound
3f also showed significant oral activity against MES-induced seizures and low oral neurotoxicity in mice in phase IV pharmacolog-
ical test. Possible structure–activity relationship was discussed.
Ó 2006 Elsevier Ltd. All rights reserved.
1. Introduction
anticonvulsant activities, among which 6-benzyloxy-
3,4-dihydro-2(1H)-quinoline (compound I) showed the
strongest activity with an ED50 value of 29.6 mg/kg in
the maximal electroshock test (MES) and a TD50 value
of greater than 300 mg/kg.6 Introduction of triazole ring
to the first and second position of this compound I
caused a remarkable increase in the anticonvulsant
activity, as seen in 7-benzyloxyl-4,5-dihydro-[1,2,4]triaz-
olo[4,3-a]quinoline (compound II), which showed ED50
values of 17.3 and 24 mg/kg in the MES and the
sc-PTZ tests, respectively.7 Another derivative in the
group of 7-alkoxy-4,5-dihydro-[1,2,4]triazolo[4,3-a]quin-
oline, 7-(4-fluoro-benzyloxyl)-4,5-dihydro-[1,2,4]triazolo-
[4,3-a]quinoline, showed ED50 values of 11.1 or 6.7 mg/
kg, protective index (PI = TD50/ED50) values of 4.6 or
8.1 in the MES or the sc-PTZ tests, respectively, and thus
demonstrated comparable anticonvulsant potency to that
of phenobarbital in the corresponding tests.8
Epilepsy, a ubiquitous disease characterized by recur-
rent seizures, inflicts more than 60 million people world-
wide according to epidemiological studies.1 For epilepsy
treatment, nearly 95% of clinically available drugs were
approved before 1985 and they could provide satisfacto-
ry seizure control for 60–70% of patients. These drugs,
however, also cause notable adverse side effects such
as drowsiness, ataxia, gastrointestinal disturbance, hep-
atotoxicity, and megaloblastic anemia,2–4 and even life-
threatening conditions.5 Research to find more effective
and safer antiepileptic drugs is, therefore, imperative
and challenging in medicinal chemistry.
In our previous work, a series of derivatives of 6-alkoxy-
3,4-dihydro-2(1H)-quinoline was first found to have
Aimed at exploring effective compounds with lower
neurotoxicity, compound III was designed and synthe-
sized which substituted by triazolone the triazole in com-
pound II. The hypothesis was that triazolone compound
Keywords: 4,5-dihydro-[1,2,4]thiazolo[4,3-a]quinoline-1(2H)-one; Syn-
thesis; Anticonvulsant activity; MES; sc-PTZ.
*
Corresponding author. Tel.: +86 433 2660606; fax: +86 433
2660568; e-mail: zsquan@ybu.edu.cn
0968-0896/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2006.06.044

H.-G. Jin et al. / Bioorg. Med. Chem. 14 (2006) 6868–6873
HO
6869
may have higher affinity to the receptor due to the
carbonyl group, and thus may increase the anticon-
vulsant activity. There were some similar design
reports.9–11 For instance, when the triazole in the pre-
cursor of compound IV (8,9-dimethoxy-6-phenyl-11H-
[1,2,4]triazolo[4,5-c][2,3]benzodiazepine) was substituted
by triazolone, the anticonvulsant activity was remark-
ably increased. So a series of 7-alkoxy-4,5-dihydro-
RO
RX, K CO
2
3
P S
5 2
N
O
N
O (C H ) N
2 5 3
H
H
1a-1l
O
R
RO
[1,2,4]triazolo[4,3-a]quinoline-1(2H)-one
derivatives
H NNHCOOCH
was prepared. The new compounds were evaluated as
anticonvulsant agents in experimental epilepsy models,
that is, maximal electroshock test (MES) and pentylene-
tetrazol (sc-PTZ)-induced seizure in mice. The rotarod
assay was performed in mice to evaluate the neurotox-
icity of the compounds. From these tests compound 3f
was selected to undergo further (phases III and IV)
pharmacological tests to evaluate its primary toxicology
profile (median hypnotic dose (HD50), median lethal
dose (LD50)), oral activity against MES-induced
seizures, and oral neurotoxicity in mice.
2
3
N
N
N
S
NH
H
O
2a-2l
3a-3l
R:
3a = n-C4H9
3b = n-C5H11
3c = n-C6H13
3d= n-C7H15
3e = n-C8H17
3i = CH2C6H4(p-F)
3f = CH2C6H5
3j = CH2C6H4(p-CH3)
3k = CH2C6H4(p-Cl)
3l = CH2C6H4(p-OCH3)
3g = CH2C6H4(o-F)
3h = CH2C6H4(m-F)
Scheme 1. The synthesis route of compounds 3a–l.
2.2. Pharmacological evaluations
Pharmacological tests of the 7-alkoxy-4,5-dihydro-
[1,2,4]triazolo[4,3-a]quinoline-1(2H)-ones (3a–l) were
conducted at the Epilepsy Branch of the National
Institute of Neurological Disorders and Stroke
(NINDS) following the protocol adopted by the Antiep-
ileptic Drug Development (ADD) program.14,15
The results of preliminary (phase I) screening of 3a–l are
summarized in Table 1. All synthesized compounds
exhibited strong anticonvulsant activity, among which
seven compounds, including all of five alkyl-substituted
derivatives 3a–e, benzyl-substituted derivative 3f, and
Table 1. Phase I anticonvulsant and toxicity data in mice (ip)a
Compound
MESb
scPTZc
Rotarod
toxicity
0.5 h
4 h
0.5 h
4 h
0.5 h
4 h
e
2. Results and discussion
2.1. Synthesis
3a
3b
3c
3d
3e
3f
30d
30
30
30
30
30
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
100
—
30
—
—
—
—
—
—
100
100
100
—
Target compounds were prepared along the reaction
sequence in Scheme 1. The compound 1a–l was
synthesized using the method described in a former
paper of our group.6 6-Hydroxy-3,4-dihydro-2(1H)-
quinoline and appropriate alkyl halide reacted in the
solution of sodium hydroxide methanol and produced
the compound 1a–l. Compound 1a–l then reacted with
phosphorus pentasulfide in acetonitrile in the presence
of triethylamine under protection of nitrogen,7 and the
resulting compound (2a–1) reacted further with methyl
hydrazine carboxylate in cyclohexanol to produce the
target compounds 7-alkoxy-4,5-dihydro-[1,2,4]triazolo-
[4,3-a]quinoline-1(2H)-ones (3a–l) according to the Refs.
12 and 13.
—
3g
3h
3i
100
100
100
—
100
30
3j
100
100
100
300
—
3k
3l
100
a All of tested compounds were dissolved in Polyethylene glycol-400.
b The maximal electroshock test was carried out 30 min after admin-
istration of the test compounds.
c Subcutaneous pentylenetetrazol (85 mg/kg) was administered 30 min
after administration of the test compounds.
d Doses are denoted in milligrams per kilogram.
e —, no activity at 300 mg/kg.

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H.-G. Jin et al. / Bioorg. Med. Chem. 14 (2006) 6868–6873
(p-F) benzyl-substituted derivative 3i, possessed anticon-
vulsant activity against MES-induced seizure at the dose
of 30 mg/kg, and the remaining five benzyl-substituted
compounds, 3g–h and 3j–l, were active at the dose of
100 mg/kg. However, none of the 12 compounds exhib-
ited any potency to the convulsion induced by sc-PTZ at
the dose of 300 mg/kg, for which we do not yet formu-
lated a plausible activity–structure relation-based expla-
nation. Rotarod toxicity test result indicated that five
compounds (3a, 3e, 3f, 3i, and 3k) showed no toxicity
at the dose of 300 mg/kg, meaning that they did not
cause ataxia or other neurotoxic reactions and thus pos-
sessed high safety profile.
when the alkyl chain had more than seven carbon num-
bers as seen with compound 3e. Compound 3d, with
ED50 of 9.8 mg/kg, was better than phenobarbital and
close to carbamazepin and phenytoin in anti-MES activ-
ity. With PI value of 20.8, compound 3d could be consid-
ered superior to all the other drugs compared in terms of
combined efficacy and safety profile. In addition, com-
pound 3c, with ED50 of 11.8 mg/kg and lower neurotox-
icity and thus a high PI value of 44.7, could also be
considered a potentially useful and safe therapeutic.
Among the 7-benzyloxy derivatives, the anticonvulsant
potency of compounds containing substituted benzyloxy
was lower than that of the compound with non-substi-
tuted benzyloxy (i.e., 3f). The potency order of three
F-substituted derivatives was p-F > o-F > m-F. The
p-F-substituted derivative 3i exhibited the strongest
anticonvulsant activity, and has a higher PI value of
9.3 compared to the other two (3g and 3h), thus its supe-
riority in terms of safety as well. In another three substi-
tuted-benzyl derivatives 3j–l, p-OCH3 derivative 3l
exhibited the weakest anticonvulsant activity with anti-
MES ED50 of 50.9 mg/kg. Nevertheless, it possessed
the lowest neurotoxicity with PI value of 12.9 and thus
higher safety profile.
In the phase II pharmacology test, 12 compounds were
quantitatively evaluated for their anticonvulsant activity
(indicated by ED50) and neurotoxicity (indicated by
TD50). Among these compounds, four compounds, 3c,
3d, 3e, and 3f, were more active than compound II in
the MES test, and all four had lower neurotoxicity than
compound II. In fact, all 12 compounds except one (3b)
had markedly lower neurotoxicity than compound II.
But all 12 compounds had no activity in the sc-PTZ test
at the dose of 300 mg/kg. For the higher anticonvulsant
activity of some compounds, we believe it may be due to
the carbonyl group in the triazolone ring which en-
hanced affinity of the triazole ring to the receptor. For
the general lower neurotoxicity, it is possible that com-
pound hydrophobicity decreased when triazole was
substituted by triazolone, which led to decreased drug
concentration in brain and thus lower neurotoxicity.
The compound 3f possessed strong anti-MES activity,
with ED50 of 12.3 mg/kg, which was close to current epi-
lepsy drugs phenytoin and carbamazepin. Its neurotox-
icity was one of the lowest in all the synthesized
compounds and was markedly lower than all current
drugs compared (Table 2). It had a protective index as
high as 44.5, which was many folds higher than current
drugs whose PI values were in the range of 1.6–8.1.
Compared with compound 3d, compound 3f possessed
slightly lower anticonvulsant but nearly threefold lower
neurotoxicity and thus a much higher PI value (44.5 vs.
20.8). Therefore, the compound 3f was chosen to be
Length of the alkyl chain appeared to have a direct
impact on anticonvulsant activity of the 7-alkyloxyl
derivatives. From compound 3a to 3d, as alkyl chain
length increased, ED50 gradually increased with the
compound 3d (with the n-heptyl-substituted group)
being the most active. The trend reversed, however,
Table 2. Phase II quantitative anticonvulsant data in mice (test drug administered ip)
PIb
a
a
c
Compound
MES, ED50
sc-PTZ, ED50
Tox, TD50
MES
sc-PTZ
II
3a
17.3
24.0
61.4
132.0 (112.5–154.9)
52.8 (44.9–61.9)
3.5
6.7
2.6
—
19.7 (17.2–22.6)
18.2 (15.2–21.8)
11.8 (10.2–13.7)
9.8 (8.5–11.4)
>300
>300
>300
>300
>300
>300
>300
>300
>300
>300
>300
>300
>300
>100
3b
3c
2.9
—
—
527.9 (440.0–633.4)
204.6 (170.5–245.4)
203.6 (170.9–242.5)
547.5 (459.5–652.2)
204.6 (178.9–223.8)
306.0 (261.3–358.4)
229.1 (195.3–268.8)
184.1 (155.7–217.7)
183.3 (160.4–209.5)
659.9 (550.0–786.4)
65.5 (52.5–72.9)
44.7
20.8
14.3
44.5
4.2
3d
3e
—
—
14.2 (12.3–16.4)
12.3 (10.8–13.9)
49.1 (43.2–55.8)
65.4 (57.8–73.8)
24.5 (21.2–28.4)
33.9 (29.2–39.6)
35.3 (29.4–42.4)
50.9 (46.4–55.8)
9.5 (8.1–10.4)
3f
3g
—
—
3h
3i
4.7
9.3
—
—
3j
3k
5.4
5.2
—
—
3l
12.9
6.9
—
Phenytoin
Carbamazepin
Phenobarbital
Valproate
<0.22
<0.22
5.2
2.9
8.8 (5.5–14.1)
21.8 (21.8–25.5)
272 (247–338)
71.6 (45.9–135)
69 (62.8–72.9)
426 (369–450)
8.1
3.2
13.2 (5.8–15.9)
149 (123–177)
1.6
a The dose is in milligrams per kilogram.
b PI = TD50/ED50
c Minimal neurotoxicity was determined by the rotarod test 30 min after the tested compounds were administered.
.

H.-G. Jin et al. / Bioorg. Med. Chem. 14 (2006) 6868–6873
6871
Table 3. Phase III quantitative toxicity profile of 3f and prototype
anticonvulsant drugs
ED50/ip ED50 ratios for 3f were >5.48 and 5.54 calculat-
ed by rotarod and MES test, respectively. These ratios
suggested that the compound 3f was adequately ab-
sorbed in mice after oral administration. No neurotox-
icity was found after oral administration at the very
high dose of 3000 mg/kg, so protective index in po
administration was 44.0, which was very close to that
in ip administration and was better than any of the
prototype drugs.
a,b
a,c
Compound
HD50
LD50
HD50/ED50
97.8
3f
1204 (1085–1373) >2000
178 (153–196)
172 (134–198)
Phenytoin
Carbamazepin
230 (216–259) 18.8
629 (556–708) 19.5
a Median hypnoitic dose (HD50) in milligrams per kilogram; deter-
mined by loss of righting reflex.
b 95% confidence intervals in parentheses.
c Median lethal dose (LD50) in milligrams per kilogram; mortality was
determined 24 h after ip injection.
3. Conclusions
further evaluated in phase III and phase IV pharmaco-
logical tests.
A
new anticonvulsant compound, 7-benzyloxyl-4,
5-dihydro-[1,2,4]triazolo[4,3-a]quinoline-1(2H)-one, was
synthesized and its pharmacological properties were
evaluated. The compound has stronger anticonvulsant
activity compared with reference compound II which
is likely due to the addition of a carbonyl group.
The compound not only has higher anticonvulsant
activity, but also has marked lower neurotoxicity. So
The toxicity profile of 3f (Table 3) was determined in
phase III testing by administering the drug intraperito-
neally (ip) to mice at different doses (1TD50, 2TD50,
and 4TD50). The toxicity induced by 3f was character-
ized by decreased motor activity, ataxia, sedation, pto-
sis, muscular relaxation, loss of righting reflex,
decreased respiration, and cyanosis. Animals given
doses of 2TD50 and 4TD50 also experienced hypnosis,
analgesia, and anesthesia.
a
larger protective index was observed for this
compound in mice compared with the prototype
drugs phenytoin, phenobarbital, carbamazepin, and
valproate.
The median hypnotic dose (HD50) for 3f was established
as 1204 mg/kg, which is twice the TD50 (547.5 mg/kg).
Compound 3f exhibited a greater margin of safety
(HD50/ED50 = 97.8) against MES-induced seizures than
any of the prototype drugs whose HD50/ED50 value was
below 20. The 24 h median lethal dose (LD50) could not
be accurately determined, but it was in excess of
2000 mg/kg. The toxicity of compound 3f was obviously
much lower than any of the prototype drugs.
4. Experimental
4.1. Chemistry
Melting points were determined in open capillary tubes
and are uncorrected. IR spectra were recorded (in KBr)
1
on a FT-IR1730, H NMR and 13C NMR spectra were
measured on a BRUKER-300, and all chemical shifts
were given in parts per million relative to tetramethylsi-
lane. Mass spectra were measured on an AP12000 (EIS,
70 eV).
As in phase II, phase IV tests involved the evaluation of
ED50 and TD50 for 3f, except that the candidate drug
was administered orally (po) rather than ip in mice.
With consideration for enhancing drug bioavailability,
the compound 3f was evaluated as hydrochloride salt.
As shown in Table 4, the orally anticonvulsant time to
peak effect (TPE) was 2 h, and this was comparable with
phenytoin and carbamazepin. The data in Table 4 clear-
ly indicated a decrease in anticonvulsant potency and
neurotoxicity of 3f administered po compared to that
in ip administration. Nevertheless, the PI values were
comparable in the two modes of drug delivery. The po
4.1.1. General procedure for preparation of 7-alkoxy-4,
5-dihydro-[1,2,4]triazolo[4,3-a]quinoline-1(2H)-ones. Com-
pounds 2a–l (0.010 mol) reacted with methyl hydrazino-
carboxylate (0.015 mol) in cyclohexanol 60 mL under
nitrogen atmosphere for 30 h, the solvent was removed
under reduced pressure, and the residue dissolved with
dichloromethane and washed with water three times.
The dichloromethane layer was dried with anhydrous
MgSO4, filtered, and concentrated the solvent, and the
residue was chromatographed on silica gel using ether
acetate and hexane (5:3) to afford target compounds
3a–l.
Table 4. Phase IV pharmacological evaluation of 3f and prototype
drugs administered in mice op
a
a
4.1.2. 7-Butyloxyl-4,5-dihydro-[1,2,4]triazolo[4,3-a]quino-
line-1(2H)-one (3a). Yield 58%, mp 181–183 °C. 1H
NMR (CDCl3) d 0.99 (t, 3H, J = 7.1 Hz, CH3), 1.27–
1.78 (m, 4H, CH2), 2.89 (t, 2H, J = 7.7 Hz, 4-CH2),
2.93 (t, 2H, J = 7.8 Hz, 5-CH2), 3.98 (t, 2H,
J = 7.0 Hz, 7-OCH2), 6.80 (d, 1H, J = 2.75 Hz, 6-H),
6.88 (dd, 1H, J = 9.15, 9.15 Hz, 8-H), 8.23 (d, 1H,
J = 8.7 Hz, 9-H), 9.98 (s, 1H, N-H). 13C NMR (CDCl3)
d 14.1 21.9, 22.4, 26.5, 29.3, 68.4, 113.2, 115.0, 118.4,
126.2, 127.8, 144.7, 153.3, 156.9. MS m/z 260 (M+1).
IR (KBr) cmÀ1: 3186, 1699.
Compound
TPE MES, ED50
(h)
TD50
PI
3fb
2
2
2
68.2
(51.3–81.6)
9.04
>3000
>44.0
Phenytoin
Carbamazepin
86.7
9.59
14.1
(7.39–10.6)
15.4
(12.4–17.3)
(80.4–96.1)
217
(131.5–270.1)
a ED50 and TD50 values are in units of milligrams per kilogram and
determined at the indicated time.
b The compound was evaluated as hydrochloride salt.

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H.-G. Jin et al. / Bioorg. Med. Chem. 14 (2006) 6868–6873
4.1.3. 7-Pentyloxyl-4,5-dihydro-[1,2,4]triazolo[4,3-a]quin-
oline-1(2H)-one (3b). Yield 56%, mp 197–199 °C. 1H
NMR (CDCl3) d 0.91 (t, 3H, J = 7.3 Hz, CH2),
1.37–1.76 (m, 6H, CH2), 2.87 (t, 2H, J = 7.7 Hz,
4-CH2), 2.93 (t, 2H, J = 8.7 Hz, 5-CH2), 3.92 (t, 2H,
J = 6.5 Hz, 7-OCH2), 6.76 (d, 1H, J = 2.75 Hz, 6-H),
6.82 (dd, 1H, J = 9.15, 9.15 Hz, 8-H), 8.21 (d, 1H,
J = 8.7 Hz, 9-H), 9.99 (s, 1H, N-H). 13C NMR (CDCl3)
d 14.1 21.9, 22.5, 26.5, 28.3, 29.0, 68.4, 113.2, 115.0,
118.4, 126.2, 127.8, 144.7, 153.3, 156.9. MS m/z 274
(M+1). IR (KBr) cmÀ1: 3189, 1698.
(s,1H). 13C NMR (CDCl3) d 25.8, 30.7, 64.3, 113.6,
114.9, 114.2 (d, JC–F = 20.3 Hz), 115.1, 115.2, 115.3,
116.3, 124.3, 124.7, 124.4, 128.0, 129.8 (d, JC–F
7.5 Hz), 154.5, 160.5 (d, JC–F = 254 Hz). MS m/z 312
=
(M+1). IR (KBr) cmÀ1: 3201, 1688.
4.1.9. 7-(3-Fluoro-benzyloxy)-4,5-dihydro-[1,2,4]triazolo-
[4,3-a]quinoline-1(2H)-one (3h). Yield 46%, mp 144–
148 °C. 1H NMR (CDCl3) d 2.61 (t, 2H, J = 7.3 Hz,
4-CH2), 2.92 (t, 2H, J = 7.8 Hz, 5-CH2), 5.01 (s, 2H,
7-OCH2), 6.72–8.27 (m, 7H, Ar-H), 10.01 (s, 1H). 13C
NMR (CDCl3) d 25.8, 30.7, 69.7, 113.6, 114.2, 114.4,
114.8 (d, JC–F = 20.9 Hz), 115.0, 115.1, 116.3, 122.7,
125.3, 130.3 (d, JC–F = 7.5 Hz), 132.4, 139.7, 154.5,
163.1 (d, JC–F = 254 Hz). MS m/z 312 (M+1). IR
(KBr) cmÀ1: 3201, 1689.
4.1.4. 7-Hexyloxyl-4,5-dihydro-[1,2,4]triazolo[4,3-a]quin-
oline-1(2H)-one (3c). Yield 62%, mp 148–151 °C. 1H
NMR (CDCl3) d 0.91 (t, 3H, J = 7.3 Hz, CH3),
1.34–1.76 (m, 8H, CH2), 2.89 (t, 2H, J = 7.7 Hz,
4-CH2), 2.92 (t, 2H, J = 8.6 Hz, 5-CH2), 3.93 (t, 2H,
J = 6.5 Hz, 7-OCH2), 6.77 (d, 1H, J = 2.75 Hz, 6-H),
6.83 (dd, 1H, J = 9.15, 9.15 Hz, 8-H), 8.22 (d, 1H,
J = 8.7 Hz, 9-H), 9.95 (s, 1H, N-H). 13C NMR (CDCl3)
d14.1, 21.9, 22.7, 25.8, 26.5, 29.3, 31.7, 68.4, 113.2,
115.0, 118.4, 126.2, 127.8, 144.7, 153.3, 156.9. MS m/z
288 (M+1). IR (KBr) cmÀ1: 3191, 1698.
4.1.10. 7-(4-Fluoro-benzyloxy)-4,5-dihydro-[1,2,4]triazolo-
[4,3-a]quinoline-1(2H)-one (3i). Yield 53%, mp
171–174 °C. 1H NMR (CDCl3)
d 2.89 (t, 2H,
J = 7.7 Hz, 4-CH2), 2.95 (t, 2H, J = 7.35 Hz, 5-CH2),
5.02 (s, 2H, 7-OCH2), 6.84–8.26 (m, 7H, Ar-H), 9.83
(s,1H). 13C NMR (CDCl3) d 21.8, 26.5, 69.7, 113.5,
115.4, 115.6 (d, JC–F = 20.9 Hz), 118.4, 126.7, 127.9,
129.3 (d, JC–F = 7.5 Hz), 132.4, 144.5, 153.2, 156.2,
162.2 (d, JC–F = 254 Hz). MS m/z 312 (M+1). IR (KBr)
cmÀ1: 320, 1689.
4.1.5. 7-Heptyloxyl-4,5-dihydro-[1,2,4]triazolo[4,3-a]quino-
line-1(2H)-one (3d). Yield 59%, mp 120–121 °C. 1H NMR
(CDCl3) d 0.88 (t, 3H,J = 7.1 Hz, CH3), 1.29–1.78 (m,
10H,
CH2),
2.89
(t,
2H,
J = 6.8 Hz,
4-CH2), 2.93 (t, 2H, J = 7.3 Hz, 5-CH2), 3.93 (t, 2H,
J = 6.3 Hz, 7-OCH2), 6.77 (d, 1H, J = 2.75 Hz, 6-H),
6.84 (dd, 1H, J = 8.7, 8.7 Hz, 8-H), 8.23 (d, 1H,
J = 8.7 Hz, 9-H), 10.07 (s, 1H, N-H). 13C NMR (CDCl3)
d 14.2, 21.9, 22.7, 26.1, 26.5, 29.1, 29.3, 31.8, 68.4, 113.2,
115.0, 118.4, 126.2, 127.8, 144.7, 153.3, 156.9. MS m/z 302
(M+1). IR (KBr) cmÀ1: 3192, 1695.
4.1.11. 7-(4-Methyl-benzyloxy)-4,5-dihydro-[1,2,4]triazolo-
[4,3-a]quinoline-1(2H)-one (3j). Yield 43%, mp
159–162 °C. H NMR (CDCl3) d 2.38 (s, 3H, Ar-CH3),
2.62 (t, 2H, J = 7.5 Hz, 5-CH2), 2.94 (t, 2H, J = 7.8 Hz,
5-CH2), 5.04 (s, 2H, 7-OCH2), 6.70–8.28 (m, 7H, Ar-
H), 9.32 (s, 1H); 13C NMR (CDCl3) d 21.3, 21.8, 26.5,
70.4, 113.6, 115.5, 118.5, 127.7, 129.6, 128.7, 133.9,
137.8, 144.6, 153.3, 154.9, 156.6. MS m/z 308 (M+1).
IR (KBr) cmÀ1: 3202, 1695.
1
4.1.6. 7-Octyloxyl-4,5-dihydro-[1,2,4]triazolo[4,3-a]quin-
oline-1(2H)-one (3e). Yield 65%, mp 114–116 °C. 1H
NMR (CDCl3) d 0.87 (t, 3H, J = 7.1 Hz, CH3),
1.25 ꢀ 1.79 (m, 12H, CH2), 2.89 (t, 2H, J = 7.3 Hz,
4-CH2), 2.93 (t, 2H, J = 6.65 Hz, 5-CH2), 3.94 (t, 2H,
J = 6.6 Hz, 7-OCH2), 6.77 (d, 1H, J = 2.75 Hz, 6-H),
6.84 (dd, 1H, J = 8.7, 8.7 Hz, 8-H), 8.23 (d, 1H,
J = 8.7 Hz, 9-H), 10.16 (s, 1H, N-H). 13C NMR d
(CDCl3) 14.2, 21.9, 22.7, 26.1, 26.5, 29.4, 29.3, 29.4,
31.9, 68.4, 113.2, 115.0, 118.4, 126.2, 127.8, 144.7,
4.1.12. 7-(4-Chloro-benzyloxy)-4,5-dihydro-[1,2,4]triazolo-
[4,3-a]quinoline-1(2H)-one (3k). Yield 52%, mp
189–192 °C. 1H NMR (CDCl3)
d 2.62 (t, 2H,
J = 7.3 Hz, 4-CH2), 2.93 (t, 2H, J = 7.8 Hz, 5-CH2),
5.03 (s, 2H, 7-OCH2), 6.70–8.29 (m, 7H, Ar-H), 9.49 (s,
1H). 13C NMR (CDCl3) d 21.8, 26.5, 69.7, 113.6, 115.0,
115.5, 118.5, 126.8, 128.0, 128.9, 131.3, 144.6, 153.1,
156.2, 171.3. MS m/z 329 (M+1). IR (KBr) cmÀ1: 3200,
1691.
153.3, 156.9. MS m/z 316 (M+1). IR (KBr) cmÀ1
:
3185, 1698.
4.1.13. 7-(4-Methoxy-benzyloxy)-4,5-dihydro-[1,2,4]triaz-
olo[4,3-a]quinoline-1(2H)-one (3l). Yield 51%, mp 177–
179 °C. H NMR (CDCl3) d 2.62 (t, 2H, J = 7.2 Hz, 4-
CH2), 2.95 (t, 2H, J = 7.8 Hz, 5-CH2), 3.84 (s, 3H,
Ar-OCH3), 5.01 (s, 2H, 7-OCH2), 6.67–8.28 (m, 7H,
Ar-H), 9.56 (s, 1H). 13C NMR (CDCl3) d 21.8, 26.5,
30.7, 70.4, 113.6, 115.0, 118.5, 127.5, 127.6, 128.7,
128.8, 136.7, 144.6, 153.3, 154.9, 156.5. MS m/z 324
(M+1). IR (KBr) cmÀ1: 3203, 1689.
4.1.7. 7-Benzyloxy-4,5-dihydro-[1,2,4]triazolo[4,3-a]quin-
oline-1(2H)-one (3f). Yield 52%, mp 187–189 °C. 1H
NMR (CDCl3) d 2.89 (t, 2H, J = 6.9 Hz, 4-CH2), 2.94
(t, 2H, J = 7.8 Hz, 5-CH2), 5.06 (s, 2H, 7-OCH2),
6.86–8.26 (m, 8H, Ar-H), 10.03 (s, 1H). 13C NMR
(CDCl3) d 21.8, 26.5, 70.4, 113.6, 115.5, 118.5, 127.6,
128.1, 128.6, 128.7, 136.8, 144.6, 153.3, 154.9, 156.5.
MS m/z 294 (M+1). IR (KBr) cmÀ1: 3202, 1695.
1
4.1.8. 7-(2-Fluoro-benzyloxy)-4,5-dihydro-[1,2,4]triazolo-
[4,3-a]quinoline-1(2H)-one (3g). Yield 48%, mp
4.2. Pharmacology
173–176 °C. 1H NMR (CDCl3)
J = 7.8 Hz, 4-CH2), 2.93 (t, 2H, J = 8.25 Hz, 5-CH2),
5.07 (s, 2H, 7-OCH2), 6.74–8.26 (m, 7H, Ar-H), 10.16
d
2.88 (t, 2H,
The MES test, sc-PTZ test, and rotarod test were carried
out by the Antiepileptic Drug Development Program
(ADD), Epilepsy Branch, National Institutes of Health,

H.-G. Jin et al. / Bioorg. Med. Chem. 14 (2006) 6868–6873
6873
Bethesda, MD, USA.11,12 All compounds were tested
for anticonvulsant activity with C57B/6 mice in the
18–25 g weight range purchased from the Laboratory
of Animal Research, College of Pharmacy, Yanbian
University. The tested compounds were dissolved in
polyethylene glycol-400.
the test compound. The median hypnotic dose (HD50),
assessed by loss of righting reflex, and the 24 h median
lethal dose (LD50) were determined (Table 3) using the
procedure described previously for evaluation of the
ED50 and TD50 values.
Phase IV testing (Table 4) involved the same procedures
for determining ED50 and TD50 as used in phase II
screening, except that the test drug was administered
po to mice.
In phase I screening (Table 1) each compound was
administered at three dose levels (30, 100, and 300 mg/
kg ip, to a total of six mice, using two for each dose)
with anticonvulsant activity and neurotoxicity assessed
at 30 min and 4 h intervals after administration. Anti-
convulsant efficacy was measured in the MES test and
the sc-PTZ test. In the MES test, seizures were elicited
with a 60 Hz alternating current of 50 mA intensity in
mice. The current was applied via corneal electrodes
for 0.2 s. Abolition of the hind-leg tonic-extensor com-
ponent of the seizure indicated protection against the
spread of MES-induced seizures. The sc-PTZ test in-
volved subcutaneous injection of a convulsant dose
(CD97) of pentylenetetrazol (85 mg/kg in mice). Eleva-
tion of the pentylenetetrazol-induced seizure threshold
was indicated by the absence of clonic spasms for at
least 5 s duration over a 30 min period following admin-
istration of the test compound. Anticonvulsant drug-
induced neurologic deficit was detected in mice using
the rotarod ataxia test.
Acknowledgment
This work was supported by the National Science Foun-
dation of China (No. 30460151).
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