INTRODUCTION:

Over the past few years, there has been growing interest in the synthesis of organic compounds under microwave irradiation because of increasing environmental consciousness.¹ Epilepsy is the commonest serious neurological condition, affecting 0.5-1 % of the population, of which 30% have epilepsy that is not adequately treated with our present antiepileptic drugs (AED). Research to find more effective and safer antiepileptic drugs is therefore, imperative and challenging in medicinal chemistry. Phthalazine derivatives were reported to possess anticonvulsant,^2-6 antimicrobial,^7,8 and vasorelaxant, ^9,10 anticancer, ^11-13 hypolipidemic,¹⁴ activities. These facts roted us to synthesize some novel phthalazine derivatives having antiepileptic activity. In this study, we report the synthesis and the pharmacological evaluation of N²-substituted Dihydrophthalazine-1,4-dione Derivatives. The compounds were characterized by IR and 1H-NMR spectral studies. The compounds were investigated for anticonvulsant activity and the rotarod assay was performed in mice to evaluate the neurotoxicity of the compounds at dose of 50mg/kg.

MATERIAL AND METHODS:

Microwave irradiations were carried out on CATA’s Scientific Microwave Synthesis System-700 Watt, 2450 MHz domestic microwave oven. The progress of reaction was monitor using thin layer chromatography. Melting points were determined by open capillary method using Veego (VMP-D) melting point apparatus. The FTIR spectra were recorded on JASCO FTIR 4100 spectrophotometer by using KBr powder method. The ¹H spectra were recorded on Varian-NMR-mercury 300 MHz spectrophotometer using TMS as internal standard using DMSO as solvent and their chemical shift are reported in δ ppm.

General procedure for synthesis of substituted aromatic acid hydrazides (1a-1o)¹.

Substituted aromatic acid (0.01moles) and hydrazine hydrate (0.012 moles) were taken in a 150 ml conical flask. The reaction mixture was irradiated at power level 700 watt for specific period of time. The reaction mixture was kept for overnight in refrigerator. The product obtained were recystalised from methanol.

General procedure for synthesis of N²-(substituted)-2, 3-dihydrophthalazine-1,4-dione (2a-2o)

A mixture of substituted aromatic acid hydrazide (1a-1o) (0.01mole) and phthalic anhydride (0.01mole) was thoroughly mixed in a beaker and DMF (5 drops) was added to it. The reaction mixture was irradiated with microwaves at power level H i.e. 700 Watt for specific time duration using microwave oven. The reaction mixture was allowed to cool to the room temperature and then to it 10-20 ml cold ethanol was added. The precipitated solids were filtered and recrystallized from methanol. Similarly, other compounds of the series were synthesized.

Scheme: synthesis of compounds (2a-2o)

All the synthesized compounds (2a-2o) were characterized by IR and 1H NMR spectral studies. The spectral data of the synthesized compounds are as follows.

2-(phenylcarbonyl)-2,3-dihydrophthalazine-1,4-dione (2a) IR (KBr): 3313.11cm^-1 (NH-), 3162.69 cm^-1 (Ar-CH), 1781.9 cm¹,1708.62cm^-1,1627.63cm^-1 (C=O amide), 1455cm^-1 (C-N), 1600.63 (Ar-C=C)

2-[(2-chlorophenyl)carbonyl]-2,3-dihydrophthalazine-1,4-dione (2b) IR (KBr): 3324.68cm^-1 (-NH-), 3197.4 cm^-1 (Ar-CH), 1689.34cm^-1, 1646.91cm^-1, 1542.77cm^-1 (C=O amide), 1106.94cm^-1 (C-N), 1612.43 (Ar-C=C), 748.24 cm^-1 (C-Cl)

2-[(2-hydroxy-3-methylphenyl)carbonyl]-2,3-dihydrophthalazine-1,4-dione (2c) IR (KBr): 3644.80 cm^-1 (-OH) 3390.25 cm^-1 (-NH-), 3193.54 cm^-1 (Ar-CH), 3112.55 cm^-1(-CH₃), 1685.48cm^-1, 1627.63 cm^-1, 1542.77 cm^-1 (C=O amide), 1172.51cm^-1 (C-N), 1600.83 (Ar-C=C); ¹H NMR (DMSO): 7.976-8.605 ( m 7H Ar-H), 11.446 (s 1H N-H), 2.478 (s 3H CH₃), 3.365 (s 1H Ar-OH).

2-[(4-chlorophenyl)carbonyl]-2,3-dihydrophthalazine-1,4-dione(2d) IR (KBr): 3354.84 cm^-1 (-NH-), 3097.43 cm^-1 (Ar-CH), 1633.24cm^-1, 1675.93 cm^-1, 1612.23 cm^-1 (C=O amide), 1123.92 cm^-1 (C-N), 1623.73 (Ar-C=C), 786.87 cm^-1 (C-Cl)

2-[(4-aminophenyl)carbonyl]-2,3-dihydrophthalazine-1,4-dione (2e) IR (KBr): 3525.24, 1492.63 cm^-1(NH₂), 3201.26 cm^-1 (-NH-), 3116.4 cm^-1 (Ar-CH), 1697.65 cm^-1, 1600.63 cm^-1, 1538.92 cm^-1, (C=O amide), 1288.22 cm^-1 (C-N), 1632.43 (Ar-C=C).

2-[(4-nitrophenyl)carbonyl]-2,3-dihydrophthalazine-1,4-dione (2f) IR (KBr): 3123.23 cm^-1 (-NH-), 3023.28 cm^-1 (Ar-CH), 1712.34 cm^-1, 1654.66 cm^-1, 1534.43 cm^-1 (C=O amide), 1198.43 cm^-1 (C-N), 1456.32 cm^-1, 1398.43 cm^-1 (NO₂), 1632.43 (Ar-C=C).

2-[(3,5-dinitrophenyl)carbonyl]-2,3-dihydrophthalazine-1,4-dione (2g) IR (KBr): 3158.83cm^-1 (-NH-), 3046.98cm^-1 (Ar-CH), 1731.76 cm^-1, 1685.48 cm^-1, 1596.77 cm^-1 (C=O amide), 1214.93 cm^-1 (C-N), 1484.92 cm^-1, 1342.21 cm^-1 (NO₂) 1632.43 (Ar-C=C).

2-[(3,4-dimethoxyphenyl)carbonyl]-2,3-dihydrophthalazine-1,4-dione (2h) IR (KBr): 3292.23 cm^-1 (-NH-), 3097.43 cm^-1 (Ar-CH), 1685.93 cm^-1, 1663.34 cm^-1, 1643.25 cm^-1 (C=O amide), 1121.22 cm^-1 (C-N), 1654.12 (Ar-C=C). ¹H NMR (DMSO): 7.874-8.403 (m 7H Ar-H), 11.456 (s 1H N-H), 3.349 (s 3H OCH₃), 2.500(s 3H OCH₃).

2-[(2-bromophenyl)carbonyl]-2,3-dihydrophthalazine-1,4-dione (2i) IR (KBr): 3334.34 cm^-1 (-NH-), 3098.56 cm^-1 (Ar-CH), 1655.76 cm^-1, 1678.43 cm^-1, 1623.45 cm^-1 (C=O amide), 1145.22 cm^-1 (C-N), 1653.63 (Ar-C=C), 664.23 cm^-1 (C-Br).

2-[(2-hydroxyphenyl)carbonyl]-2,3-dihydrophthalazine-1,4-dione (2j) IR (KBr): 3598.52 (-OH), 3293.82 cm^-1 (-NH-), 3198.54 cm^-1 (Ar-CH), 1681.62 cm^-1, 1596.77 cm^-1, 1546.63 cm^-1 (C=O amide), 1222.51 cm^-1 (C-N), 1610.63 cm^-1 (Ar-C=C).

2-[(5-chloro-2-hydroxyphenyl)carbonyl]-2,3-dihydrophthalazine-1,4-dione (2k)

IR (KBr): 3565.72 (-OH), 3310.36 cm^-1 (-NH-), 3086.34 cm^-1 (Ar-CH), 1698.90 cm^-1, 1696.98 cm^-1, 1643.23 cm^-1 (C=O amide), 1183.64 cm^-1 (C-N), 1634.81 cm^-1 (Ar-C=C), 788.98 cm^-1(C-Cl)

2-[(1,4-dioxo-3,4-dihydrophthalazin-2(1H)-yl)carbonyl]phenyl acetate (2l)

IR (KBr): 3285.37 cm^-1 (-NH-), 3017.46 cm^-1 (Ar-CH), 1735.66 cm^-1, 1688.23 cm^-1, 1668.35 cm^-1, 1623.76 cm^-1 (C=O amide), 1133.54 cm^-1 (C-N), 1043.34(C-O) 1613.34 (Ar-C=C).

2-{[2-(phenylamino)phenyl]carbonyl}-2,3-dihydrophthalazine-1,4-dione (2m)

IR (KBr): 3325.24 cm^-1 , 3271.45 cm^-1 (-NH-), 3106.54 cm^-1 (Ar-CH), 1689.65 cm^-1, 1645.23 cm^-1, 1624.98 cm^-1, (C=O amide), 1212.72 cm^-1 (C-N), 1612.09 (Ar-C=C).

2-{[2-(phenylcarbonyl)phenyl]carbonyl}-2,3-dihydrophthalazine-1,4-dione( 2n)

IR (KBr): 3385.67 cm^-1 (-NH-), 2987.56 cm^-1 (Ar-CH), 1745.56 cm^-1, 1698.78 cm^-1, 1678.65 cm^-1, 1612.56 cm^-1 (C=O amide), 1198.79 cm^-1 (C-N), 1598.09 (Ar-C=C).

2-(pyridin-3-ylcarbonyl)-2,3-dihydrophthalazine-1,4-dione (2o) IR (KBr): 3334.89 cm^-1 (-NH-), 2998.56 cm^-1 (Ar-CH), 1687.56 cm^-1, 1656.23 cm¹, 1643.67 cm^-1, (C=O amide), 1201.93 cm^-1 (C-N), 1589.37 (Ar-C=C).

The physical data of synthesized compounds is presented in table 1

Table 1: Physical data of synthesized compounds (2a-2o)

Compound No.	R	Mol. Formula	Mol. Wt	Melting Point (°C)	% yield	Reaction time (min)
2a	-C₆H₅	C₁₅H₁₀N₂O₃	266.25	168-169	73.54	6.00
2b	2-Cl-C₆H₄-	C₁₅H₁₀N₂O₃Cl	300.69	270-271	88.82	4.00
2c	2OH,3Me-C₆H₃-	C₁₆H₁₂N₂O₄	296.27	<275	70	1.30
2d	4-ClC₆H₄-	C₁₅H₁₀N₂O₃Cl	300.69	225-226	58.71	1.00
2e	4-NH₂-C₆H₃-	C₁₅H₁₁N₃O₃	281.26	<275	69.14	2.00
2f	4-NO₂-C₆H₄-	C₁₅H₉N₃O₅	311.24	210-211	66.62	2.30
2g	3,5-NO₂-C₆H₃-	C₁₅H₈N₄O₇	356.24	250-251	80.39	1.00
2h	3,4-OCH₃-C₆H₃-	C₁₇H₁₄N₂O₅	346.30	181-182	85.84	2.00
2i	2-Br-C₆H₄-	C₁₅H₉BrN₃O₃	345.14	251-252	80.00	2.00
2j	2-OH-C₆H₄-	C₁₅H₁₀N₃O₄	282.25	<275	77.87	2.30
2k	2-OH,5-Cl-C₆H₃-	C₁₅H₉ClN₂O₄	316.69	<275	78.54	2.30
2l	2-OCOCH₃-C₆H₄-	C₁₇H₁₂N₂O₅	324.28	<275	84.81	1.30
2m	2-C₆H₄-NH-C₆H₄-	C₂₁H₁₅N₃O₃	357.36	195-196	70.21	2.00
2n	2-C₆H₄-CO-C₆H₄-	C₂₂H₁₄N₃O₄	370.35	184-186	85.40	2.00
2o	-pyridine	C₁₄H₉N₃O₃	267.39	168-169	79.92	2.00

Table 2: Duration of hind limb extensor of N²-(Substituted)-2,3-dihydrophthalazine-1,4- dione (2a-2o)

Group	Treatment	Dose (mg/kg)	Duration of hind limb extensor in seconds (mean±S.E.M)
Group	Treatment	Dose (mg/kg)	60 minutes	120 minutes
I	Control	-	64.66±2.88	70±2.887
II	Standard	5 mg	10.33±2.603	25±1.155
III	2a	50mg	21±2.517**	41±1.732**
IV	2b	50mg	22±2.175**	34.33±7.764**
V	2c	50mg	28.33±1.856**	45±1.732**
VI	2d	50mg	31.33±1.528**	51.66±2.603**
VII	2e	50mg	27.33±2.186**	42.66±2.906**
VIII	2f	50mg	29±3.906**	45.33±1.764**
IX	2g	50mg	53±2.175^ns	67±1.528^ns
X	2h	50mg	24.33±2.33**	45.66±2.028**
XI	2i	50mg	17.45±2.906**	31.33±1.453**
XII	2j	50mg	24±3.00**	45±1.528**
XIII	2k	50mg	29±2.082**	44±2.309**
XVI	2l	50mg	26.33±1.856**	42±2.082**
XV	2m	50mg	28±1.55**	52±1.732**
XVI	2n	50mg	25.33±3.52**	54.66±3.00**
XVII	2o	50mg	15±1.732**	45.33±2.028**

Data were analyzed by one-way ANOVA followed by Dunnett’s test.

Values are expressed as mean ± S.E.M.

** P<0.01 when compared to control, ns-non significant.

Table 3: Neurotoxicity study at 60 and 120 minutes time interval of N²-(Substituted)-2,3- dihydrophthalazine-1,4- dione (2a-2o)

Group	Treatment	Dose (mg/kg)	Fall of latency in sec
Group	Treatment	Dose (mg/kg)	60 minutes	120 minutes
I	Control	-	-	-
II	Standard	5 mg	15	24
III	2a	50mg	26	-
IV	2b	50mg	-	-
V	2c	50mg	-	-
VI	2d	50mg	-	-
VII	2e	50mg	21	-
VIII	2f	50mg	36	-
IX	2g	50mg	43	-
X	2h	50mg	-	-
XI	2i	50mg	-	-
XII	2j	50mg	-	-
XIII	2k	50mg	-	-
XVI	2l	50mg	-	-
XV	2m	50mg	18	48
XVI	2n	50mg	-	-
XVII	2o	50mg	26	-

The figure in the table indicate fall of latency in seconds.

The dash (-) indicates an absence of neurotoxicity at 60 and 120 minutes time interva

CONCLUSION:

In summary, all synthesized compounds displayed anticonvulsant activity at 50 mg/kg and their structure-activity relationship was discussed in the present study. In particular, we found that 2-[(2- chlorophenyl)-carbonyl]-2,3-dihydro- phthalazine-1,4-dione (2b) and 2-[(2-bromophenyl)-carbonyl]-2,3-dihydro- phthalazine-1,4-dione (2i) possessed the most potential anticonvulsant activity all the tested time intervals, Which was comparable to reference agent diazepam in anticonvulsant activity having no neurologic deficit.

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Received on 09.09.2011 Modified on 18.09.2011

Asian J. Research Chem. 4(11): Nov., 2011; Page 1722-1725