Synthesis of Some New 3-(4-Phthalimidophenyl)-5-(Substituted Phenyl) 4- Isoxazoline Derivatives with Analgesic and Anti-Inflammatory Activities.

 

Deepak Lande*, Meenakshi Deodhar, Ashok Bhosle and Ganesh Bhawal

Dep. of Pharmaceutical Chemistry, SGRS College of Pharmacy, Saswad, Pune, Maharashtra 412301, India

*Corresponding Author E-mail: deepak_lande@rediffmail.com

ABSTRACT:

The derivatives of 3-(4-phthalimidophenyl)-5-(substituted-phenyl)-4-isoxazoline were synthesized, which were screened for their analgesic and anti-inflammatory activity in test animals. The analgesic activity of all compounds were found to be higher than their anti-inflammatory activity, Compounds 3b, 3e, 3h and 3l were found to be more active among the synthesized compounds.

 

 

 

INTRODUCTION:

Thalidomide(1) was introduced as a sedative/hypnotic drug in 1956 but was subsequently withdrawn from the market due to its teratogenicity¹. Later Thalidomide was found to cause a rapid and noticeable improvement of painful neuritis in patients suffering from lepromatous leprosy.² This activity was attributed to a selective blockade of tumor necrosis factor-α (TNF-α) production^3,4, thus suggesting use of thalidomide  as anti-inflammatory under restricted conditions.

 

(1)

 

Isoxazoline derivatives have also been found to possess analgesic, anti-inflammatory activity besides antiviral⁵, antiepileptic⁶ and antibacterial activity.^7-8

In present work, we report synthesis of phthalimide derivatives incorporating isoxazoline ring that is substituted with variously substituted phenyl rings.

 

RESULTS AND DISCUSSION:

The synthetic pathway leading to the title compounds were synthesized as per Scheme1. 2-(4-Acetylphenyl) isoindoline-1,3-dione (1), was synthesized using p-amino acetophenone and phthalic anhydride using glacial acetic acid as solvent. (1) was reacted with substituted benzaldehydes to obtain N-[4-substituted cinnamoyl- phenyl)]phthalimide (2a-2l) which was subsequently reacted with hydroxyl amine hydrochloride in presence of sodium hydroxide to produce 3-(4-phthalimidophenyl)-5-(substituted phenyl) 4- isoxazoline derivatives (3a-3l)

 

The structures of the compounds have been elucidated by IR and 1H-NMR. Reaction time, crystallization solvents, melting points, % yields and spectral data of the compounds are given in the Experimental Part.

 

The IR spectrum of 2-(4-acetylphenyl) isoindoline-1,3-dione (1) showed aromatic stretch at 3076.56 cm^-1 and sharp carbonyl stretching vibration for phthalimide and the acetyl group at around 1714.77 cm^-1 and at 1670.06 cm^-1 respectively. The stretching vibrations for C-N group of phthalimide are seen at around 1365.95 cm^-1.

 

The IR spectrum of (2a-2l) in KBr showed aromatic stretch between 2989-3078 cm^-1 and carbonyl stretching vibrations at around 1670-1765 cm^-1, Sharp aliphatic C=C stretching vibrations for the chlacone compounds were seen at around at 1630-1663cm^-1 and The stretching vibrations for C-N of phthalimide are seen at around 1343-1453 cm^-1.

 

The IR spectrum of (3a-3l) in KBr showed aromatic stretch at 2928-3099 cm^-1 and carbonyl stretching vibrations at around 1694-1733 cm^-1  Disappearance of carbonyl peak of chalcone and appearance of C=N streaching confirms formation of isoxazoline ring and C-N group of  phthalimide ring showed stretching vibrations at around 1313-1599 cm^-1.

 

In the ¹H NMR (δ ppm, CDCl₃) spectrum of (1), methyl group being adjacent to the carbonyl group gives the singlet slightly downfield that is at δ 2.63 ppm. The aromatic protons showed peaks at δ 7.58, 8.05, 7.79 and 7.93 ppm as doublet, doublet, doublet and triplet respectively. The ¹H NMR (δ ppm, CDCl₃) spectrum of (2a) bearing the olefinic carbons, revealed characteristic peaks as two doublets at δ 6.45  and δ 6.85 ppm, aromatic protons revealed characteristic peaks as multiplet at 7.25 - 7.853 ppm. The ¹H NMR (δ ppm, CDCl₃) spectrum of (3a) having the aromatic protons showed peaks at δ 7.15-7.45 ppm as multiplet. Disappearance of doublets of olefinic protons of chalcones (6.45 and 6.85) and appearance of triplet at δ 6.13 ppm and  doublet of isoxazoline 4-CH₂  at δ 3.704 ppm confirmed the formation of isoxazoline ring.

 

The results of the Pharmacological screening of the synthesized compounds indicate that all the synthesized cmpounds exhibit significant anti-inflammatory and analgesic activity when compaired with standard. Four compounds of these serise namely 3-(4-phthalimidophenyl)-5-(4-chlorophenyl)isoxazoline (3b), 3-(4-phthalimidophenyl)-5-(4-nitrophenyl)isoxazoline (3e), 3-(4-phthalimidophenyl)-5-(4-bromo- phenyl)isoxazoline (3h), and 3-(4-phthalimidophenyl)-5-(2-hydroxyl phenyl)isoxazoline (3l) were found to significant decrease the inflammation whereas all the compounds showed good activity in the acetic acid. This  indicates that electron withdrowing groups –NO₂, -Br, -Cl placed at 4^th position of the phenyl group are favorable for anti-inflammatory and analgesic activity. These groups when shifed to 2^nd position resulted in a decrease in activity. Where as –OH group is tolarated at this position.

 

Experimental Section:

All the chemicals used in the synthesis were of laboratory grade. The melting points were determined in open capillary on Veego (model:-VMP-D) electronic apparatus and are uncorrected. The IR spectra of synthesized compounds were recorded on Shimadzu 8400-S FT-IR Spectrophotometer using potassium bromide. The ¹H NMR were recorded in CDCl₃ using NMR Varian-Mercury 300MHz spectrometer and chemical shifts are given in parts per millions, downfield from tetramethylsilane (TMS) as an internal standard. Splitting patterns were designated as follows: s: singlet, d: doublet, t: triplet, q: quartet, and m: multiplet. All chemical shift values were recorded as δ (ppm). The purity of the compounds was determined by thin layer chromatography (Merck, silicagel, HF254−366, Type 60, 0.25 mm).

 

2-(4-Acetylphenyl) isoindoline-1,3-dione (1):

To the solution of 4-amino acetophenone 0.5 g (0.01mol) in glacial acetic acid (5 ml) kept in 250 ml Round Bottomed Flask phthalic anhydride 0.5 g (0.01 mol) was added. The reaction mixture was refluxed for 30 min. The solid separated out on cooling was collected by vacuum filtration, dried and recrystalised from ethanol.

Yield 86%, mp  245-248⁰ C, R_f = 0.72 (chloroform-ethyl acetate = 9:1), IR (KBr), 1714.77 cm−1 (C=O), 3076.56 (Ar stretch), 1466.56 (CN). ¹H-NMR (CDCl₃) δ: 2.63 (3H; s; CH₃), 7.58-7.93 (8H; m; Arom-H.).

 

General procedure for N-[4-substituted cinnamoyl phenyl)] phthalimide. (2a-2l):

Compound (1) 2.2 g (0.01 mol) dissolved in ethanol (25 ml) was stirred with 2 ml (10%) sodium hydroxide solution in a conical flask, under cold conditions for 10 minutes. Substituted benzaldehyde 0.01 mol was added and the mixture was stirred for 3-4 hours. The resulting mixture was poured onto ice. The separated solid was filtered, washed with water dried and recrystalised from ethanol. The physical characteristics of all the intermediates are summarized in Table 1.

 

Table 1: Physical characterization data of all the intermediates compounds

Comp.	R1	m.pa (oC)	Yield (%)	Rf*
2a 2b 2c 2d 2e 2f 2g 2h 2i 2j 2k 2l	H 4-Cl 3-Cl 2-Cl 4-NO2 3-NO2 2-NO2 4-Br 3,4,5(OCH3)3 4-N(CH3)2 4-OCH3 2-OH	134-136 142-144 149-150 160-162 152-154 165-167 170-172 148-152 180-182 187-190 168-170 167-170	86.79 84.90 89.43 87.54 81.13 83.39 84.15 86.03 86.79 88.67 85.66 89.81	0.76 0.80 0.72 0.50 0.41 0.79 0.73 0.86 0.63 0.73 0.84 0.71

a.   The value is uncorrected

*    Solvent system - benzene: methanol = 9:1

 

N- [4-(Cinnamoyl phenyl)] phthalimide. (2a):

Yield 86.79 % mp 134-136⁰C, R_f = 0.76 (benzene: methanol = 9:1), Reaction time is 4 h. IR (KBr), cm⁻¹ 3053.42 (Ar stretch); 1670, 1716 (C=O), 1630.11 (C=C stretch); 1343.78 (C-N stretch). ¹H-NMR (CDCl₃) δ: 6.521-7.233 (m, 13H, Ar-H aromatic protons), 6.85 (d, 1H,  HC=CH ), 6.45 (d, 1H, Ar-CH=CH ).

N- [(4-Chlorocinnamoyl) phenyl] phthalimide (2b)

IR (KBr), cm⁻¹ 3064.59, 3020.89 (Ar stretch), 1670.41, 1765.77 (C=O stretch), 1647.26(C=C stretch), 1453.23 (C-N stretch).

N- [(3-Chlorocinnamoyl) phenyl] phthalimide (2c)

IR (KBr), cm⁻¹ 3078.49, 2989.26 (Ar stretch), 1670.41, 1763.34 (C=O stretch), 1647.26 (C=C stretch), 1445.65 (C-N stretch).

N- [(2-Chlorocinnamoyl) phenyl] phthalimide (2d)

IR (KBr), cm⁻¹ 3073.49, 2989.26, (Ar stretch), 1670.41,1763.34 (C=O stretch), 1647.26 (C=C stretch), 1445.65 (C-N- stretch).

N- [(4-Nitrocinnamoyl) phenyl] phthalimide (2e)

IR (KBr), cm⁻¹ 3078.49, 3064.59 (Ar stretch), 1670.41, 1733.84 (C=O stretch), 1635.78 (C=C stretch), 1445.65 (C-N stretch), 1545.12 (NO₂ stretch)

N- [(3-Nitrocinnamoyl) phenyl] phthalimide (2f)

IR (KBr), cm⁻¹ 3078.49, 2989.26 (Ar stretch), 1670.41, 1763.34 (C=O stretch), 1647.26 (C=C stretch), 1445.65 (C-N stretch), 1552.22 (NO₂ stretch)

N- [(2-Nitrocinnamoyl) phenyl] phthalimide (2g)

IR (KBr), cm⁻¹ 3064.59, 3020.89 (Ar stretch), 1670.41, 1765.77 (C=O stretch), 1647.26(C=C stretch), 1453.23 (C-N stretch); 1543.12 (NO₂ stretch)

N- [(4-Bromocinnamoyl) phenyl] phthalimide (2h)

IR (KBr), cm⁻¹ 3064.59, 3020.89 (Ar stretch), 1670.41, 1765.77 (C=O stretch), 1647.26(C=C stretch), 1453.23 (C-N stretch).

N- [(3,4,5 Trimethoxycinnamoyl) phenyl] phthalimide (2i)

IR (KBr), cm⁻¹ 3064.59, 3020.89 (Ar stretch), 1670.41, 1765.77 (C=O stretch), 1647.26(C=C stretch), 1453.23 (C-N stretch).

N- [(4-Dimethylaminocinnamoyl) phenyl] phthalimide (2j)

IR (KBr), cm⁻¹ 3064.59, 3020.89 (Ar stretch), 1670.41, 1765.77 (C=O stretch), 1647.26(C=C stretch), 1453.23 (C-N stretch).

N- [(4-Methoxycinnamoyl) phenyl] phthalimide (2k)

IR (KBr), cm⁻¹ 3095.67, 3061.35 (Ar stretch), 1698.49, 1723.42 (C=O stretch), 1663.91 (C=C stretch), 1370.25 (C-N stretch).

N-[(4-Hydroxycinnamoyl) phenyl] phthalimide (2l)

IR (KBr), cm⁻¹ 3092.69, 3066.35 (Ar stretch), 1689.49, 1730.32 (C=O stretch), 1663.91 (C=C stretch), 1370.25 (C-N stretch).

 

General procedure for synthesis of 3-(4-Phthalimidophenyl)-5-(substituted phenyl) 4- isoxazoline Derivatives. (3a-3l):

A mixture of N-[4-substituted cinnamoyl phenyl)] phthalimide. (2a-2l) (0.005 mol), Hydroxyl amine hydrochloride (1.2 g, 0.02 mol) and sodium hydroxide (0.4 g, 0.01 mol) in ethanol (25  ml) was taken in a RBF. Reaction mixture was heated under reflux for 10-12 hours. The resulting mixture was concentrated, cooled and poured into ice water. The solid separated was collected and washed several times with water, dried and recrystallised from ethanol. The physical characteristics of all the intermediates are summarized in Table 2.

 

Table 2: Physical characterization data of synthesized compounds

Comp.	R1	m.pa (oC)	Yield (%)	Rf*
3a 3b 3c 3d 3e 3f 3g 3h 3i 3j 3k 3l	H 4-Cl 3-Cl 2-Cl 4-NO2 3-NO2 2-NO2 4-Br 3,4,5(OCH3)3 4-N(CH3)2 4-OCH3 2-OH	143-145 172-175 138-140 147-149 187-190 140-144 189-191 155-158 168-170 190-192 191-193 173-175	67.35 67.35 63.73 66.83 69.84 65.82 68.34 83.33 87.33 79.79 80.43 78.21	0.77 0.86 0.87 0.73 0.70 0.89 0.60 0.84 0.77 0.54 0.81 0.59

a.   The value is uncorrected

*    Solvent system – chloroform-ethyl acetate -9:1

3-(4-Phthalimidophenyl)-5(phenyl)-4-isoxazoline. (3a)

Recrystallized from ethanol to yield 65.90 % mp: 143-145^oC, Rf = 0.77 (chloroform-ethyl acetate = 9:1), IR (KBr), cm⁻¹ 3030.27, 3056.35 (Ar stretch); 1694.25 (C=O), 1585.02 (C=N stretch); 1313.57 (C-N stretch). ¹H-NMR (CDCl₃) δ: 7.15-7.45 (m, 13H, Ar-H aromatic protons),  6.13 (t, 1H, isoxazoline 5-H), 3.704 (d, 2H, isoxazoline 4-CH₂).

 

3-(4-Phthalimidophenyl)-5-(4-chlorophenyl)-4-isoxazoline. (3b)

Recrystallized from ethanol to yield 67.35% mp: 172-175^oC, R_{f =} 0.86 (chloroform-ethyl acetate = 9:1),  IR (KBr), cm⁻¹ 3044.70, 2980.34 (Ar stretch); 1697.67 (C=O), 1597.14 (C=N stretch); 1369.81 (C-N stretch). ¹H-NMR (CDCl₃) δ: 7.15-7.45 (m, 12H, Ar-H aromatic protons),  6.13 (t, 1H, isoxazoline 5-H), 3.704 (d, 2H, isoxazoline 4-CH₂).

 

3-(4-Phthalimidophenyl)-5-(3-chlorophenyl)-4-isoxazole. (3c)

Recrystallized from ethanol to yield 63.73 % mp: 138-140^oC, R_{f =} 0.87 (chloroform-ethyl acetate = 9:1), IR (KBr), cm⁻¹ 3057.27 (Ar stretch); 1707.25 (C=O), 1597.11 (C=N stretch); 1375.59 (C-N stretch). ¹H-NMR (CDCl₃) δ: 7.15-7.45 (m, 12 H, Ar-H aromatic protons),  6.13 (t, 1H, isoxazoline 5-H), 3.704 (d, 2H, isoxazoline 4-CH₂).

 

3-(4-Phthalimidophenyl)-5-(2-chlorophenyl)-4-isoxazoline. (3d)

Recrystallized from ethanol to yield 66.83 % mp: 147-149^oC, R_{f =} 0.73 (chloroform-ethyl acetate = 9:1), IR (KBr), cm⁻¹ 3099.20, 3088.71 (Ar stretch); 1702.81 (C=O), 1599.04 (C=N stretch); 1456.30 (C-N stretch). ¹H-NMR (CDCl₃) δ: 7.15-7.45 (m, 12H, Ar-H aromatic protons), 6.13 (t, 1H, isoxazoline 5-H), 3.704 (d, 2H, isoxazoline 4-CH₂).

 

3-(4-Phthalimidophenyl)-5-(4-nitrophenyl)-4-isoxazoline. (3e)

Recrystallized from ethanol to yield 69.84 % mp: 187-190^oC, R_{f =} 0.70 (chloroform-ethyl acetate = 9:1), IR (KBr), cm⁻¹ 3056.35, 3059.71 (Ar stretch); 1717.54 (C=O), 1572.20 (C=N stretch); 1456.30 (C-N stretch), 1572.20 (NO₂ stretch). ¹H-NMR (CDCl₃) δ: 7.12-7.53 (m, 12H, Ar-H aromatic protons), 6.12 (t, 1H, isoxazoline 5-H), 3.715 (d, 2H, isoxazoline 4-CH₂).

 

3-(4-Phthalimidophenyl)-5-(3-nitrophenyl)-4-isoxazoline. (3f)

Recrystallized from ethanol to yield 65.82 % mp: 140-144^oC, R_{f =} 0.89 (chloroform-ethyl acetate = 9:1), IR (KBr), cm⁻¹ 3055.35, 2966.56 (Ar stretch); 1713.57 (C=O), 1597.11 (C=N stretch); 1473.66 (C-N stretch), 1562.50 (NO₂ stretch). ¹H-NMR (CDCl₃) δ: 7.12-7.53 (m, 12H, Ar-H aromatic protons),  6.12 (t, 1H, isoxazoline 5-H), 3.715 (d, 2H, isoxazoline 4-CH₂).

 

 

3-(4-Phthalimidophenyl)-5-(2-nitrophenyl)-4-isoxazoline. (3g)

Recrystallized from ethanol to yield 68.34 % mp: 189-191^oC, R_{f =} 0.60 (chloroform-ethyl acetate = 9:1), IR (KBr), cm⁻¹ 3053.42 (Ar stretch); 1737.76 (C=O), 1597.11 (C=N stretch); 1473.66 (C-N stretch), 1549.25 (NO₂ stretch). ¹H-NMR (CDCl₃) δ: 7.12-7.53 (m, 12H, Ar-H aromatic protons),  6.12 (t, 1H, isoxazoline 5-H), 3.715 (d, 2H, isoxazoline 4-CH₂).

 

3-(4-Phthalimidophenyl)-5-(4-bromophenyl)-4-isoxazoline. (3h)

Recrystallized from ethanol to yield 83.33 % mp: 191-193^oC, R_{f =} 0.84 (chloroform-ethyl acetate = 9:1), IR (KBr), cm⁻¹ 3055.35, 2966.56 (Ar stretch); 1713.57 (C=O), 1597.11 (C=N stretch); 1473.66 (C-N stretch). ¹H-NMR (CDCl₃) δ: 7.20-7.43 (m, 12 H, Ar-H aromatic protons),  6.16 (t, 1H, isoxazoline 5-H), 3.70 (d, 2H, isoxazoline 4-CH₂).

 

3-(4-Phthalimidophenyl)-5-(3,4,5-trimethoxy phenyl)-4-isoxazoline. (3i)

Recrystallized from ethanol to yield 87.33 % mp: 168-170^oC, R_{f =} 0.77 (chloroform-ethyl acetate = 9:1), IR (KBr), cm⁻¹ 3055.36, 3099.71 (Ar stretch); 1733.76 (C=O), 1597.11 (C=N stretch); 1473.66 (C-N stretch). ¹H-NMR (CDCl₃) δ: 7.18-7.48 (m, 12 H, Ar-H aromatic protons),  6.18 (t, 1H, isoxazoline 5-H), 3.27 (d, 2H, isoxazoline 4-CH₂), 3.90 (s, 9H, (OCH₃) ₃).

 

3-(4-Phthalimidophenyl)-5-(4-dimethylamino phenyl)-4-isoxazoline (3j)

Recrystallized from ethanol to yield 79.79 % mp: 191-193^oC, R_{f =} 0.54 (chloroform-ethyl acetate = 9:1), IR (KBr), cm⁻¹ 3099.20, 3088.71 (Ar stretch); 1702.81 (C=O), 1599.04 (C=N stretch); 1456.30 (C-N stretch). ¹H-NMR (CDCl₃) δ: 7.13-7.42 (m, 12 H, Ar-H aromatic protons),  6.08 (t, 1H, isoxazoline 5-H), 3.26 (d, 2H, isoxazoline 4-CH₂), 2.90 (s, 6H, -N(CH₃)₂)_.

 

3-(4-Phthalimidophenyl)-5-(4-methoxy phenyl)-4-isoxazoline (3k)

Recrystallized from ethanol to yield 80.43 % mp: 191-193^oC, Rf = 0.81 (chloroform-ethyl acetate = 9:1), IR (KBr), cm⁻¹ 3044.70, 2980.34 (Ar stretch); 1697.67 (C=O), 1597.14 (C=N stretch); 1369.81 (C-N stretch). ¹H-NMR (CDCl₃) δ: 7.11-7.44 (m, 12 H, Ar-H aromatic protons),  6.05 (t, 1H, isoxazoline 5-H), 3.21 (d, 2H, isoxazoline 4-CH₂), 3.98 (s, 3H, OCH₃).

 

3-(4-Phthalimidophenyl)-5-(2- hydroxyl phenyl)-4-isoxazoline (3l)

Recrystallized from ethanol to yield 78.21 % mp: 191-193^oC, R_{f =} 0.59 (chloroform-ethyl acetate = 9:1), IR (KBr), cm⁻¹ 3059.47, 2980.96 (Ar stretch); 1722.67 (C=O), 1597.14 (C=N stretch); 1369.81 (C-N stretch). ¹H-NMR (CDCl₃) δ: 7.15-7.45 (m, 12 H, Ar-H aromatic protons),  6.05 (t, 1H, isoxazoline 5-H), 3.35 (d, 2H, isoxazoline 4-CH₂), 5.12 (s, 1H, OH).

Anti-inflammatory activity:^9,10

The Suspension of test compounds was prepared in sterile 0.9% NaCl solution. Anti-inflammatory activity was evaluated by carrageenin induced rat paw edema method of Winter et al. Albino rats of either sex weighing between 150-250 g were randomly distributed in control and experimental group of six animals. At 0 hr the test compounds (2 mg/kg) and indomethacin (2 mg/kg) doses were administered orally. Control received the same quantity of sterile 0.9% NaCl solution as vehicle. 1hr after compounds and standards were administered orally. 0.1 ml of 1 % (w/v) suspension of Carrageenan in distilled water was injected into the planter tissue of right paw of rat by using 27 gauge needles. The paw was marked with ink at the level of the tibia-tarsal junction and the initial volume of paw was measured by plethysmometer within 30 sec. of injection. The relative increase in paw volume was found by remeasuring the paw volume after 3 hr of carrageenan injection.

The % inhibition of edema was calculated by following formula and the results are shown in Table-3

 

% inhibition of edema =   1-   Vt₃-Vt₀       X 100

              3 hr                              Vc₃-Vc_o

Where Vt = mean relative change in paw volume in test group.

Vc= mean relative change in paw volume in control group.

 

Table 3: Carrageenan induced inflammation in rat model

Compound	Dosage inflammation	% Decrease in (mean)
Control 3a 3b 3c 3d 3e 3f 3g 3h 3i 3j 3k 3l Indomethacin	Vehicle 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 2 mg/kg	- 29** 58** 19* 17* 62** 19** 21.85* 56.28** 27.90* 27.32* 21.85* 53.5** 64.48**

N=6, ** - P < 0.01 compared with vehicle treated group,

Data expressed as Mean ± SEM­.

Data was analysed by one-way ANOVA followed by Dunnett’s test

 

 

Table 4: Acetic acid induced writhing response in mices

Compound	Dosage	Number of writhings in 20 minutes   (mean ± s.e.m)	% Analgesic activity
Vehicle 3a 3b 3c 3d 3e 3f 3g 3h 3i 3j 3k 3l Diclofenac sodium	- 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg	66.33 ± 1.202 22 ± 1.155 9.0 ± 0.5774 26.33 ± 0.8819 35 ± 1.732 12 ± 1.155 26.66 ± 0.8819 32 ± 1.732 9.66 ± 0.333 25.66 ± 1.202 35.66 ± 3.33 29.66 ± 1.202 10 ± 1.155 4.0 ± 0.577	0 66.66** 86.36** 60.10** 46.96** 81.81** 59.60** 51.51** 85.36** 61.12** 45.96** 55.06** 84.84** 93.93**

N=6, * - P < 0.05, ** - P < 0.01 compared with vehicle treated group, Data expressed as Mean ± SEM­.

Data was analysed by one-way ANOVA followed by Dunnett’s test.

 

Acetic acid induced writhing in mice¹¹: -

Analgesic activity was determined in vivo by calculating total number of writhings, following intraperitonial (I.P) administration of 0.6 % (0.1 ml/10 g) acetic acid in mice. Albino mice of either sex (25-30 g) were used. Synthesized compounds (3a-3l) were administered intraperitonialy (20 mg/kg) as a suspension in sterile 0.9% NaCl solution as vehicle.

 

Diclofenac sodium (20 mg/kg) was used as the standard drug under same conditions. Acetic acid solution was administered intraperitonialy 30 min after administration of the compounds. 10 min after intraperitonial injection of the acetic acid solution, the number of writhings per animal was recorded for 20 min. Control animals received an equal volume of vehicle.

 

Analgesic activity was expressed as percentage of inhibition of number of writhings, when compared with the vehicle control group.

Percentage analgesic activity of compounds was calculated using following formula and the results are shown in Table-4

 

                       No. of writhings  _  No. of writhings for

% Analgesic =     for control            test compound           X 100

    activity                  No. of writings for control

Formalin induced licking and biting in mice¹²

The analgesic activity was determined by calculating total licking and biting time (sec) in mice, following administration of 0.1% (0.1 ml/10 g) formalin into the sub planter area of right hind paw of mice. Albino mice of either sex (25-30 g) were used. Synthesized compounds (3a-3l) were administered intraperitonialy (20 mg/kg) as a suspension in sterile 0.9 % Nacl solution as vehicle. Diclofenac sodium (20 mg/kg) was used as the standard drug under same conditions. 30 min after administration of the compounds, the duration of paw licking, an index of noniception, was measured at 0-5 min (first phase, which indicates central analgesic activity) and 15-25 min (second phase, which indicates peripheral analgesic activity) after formalin administration. Control animals received an equal volume of vehicle.

Analgesic activity was expressed as percentage of inhibition of duration of paw licking, when compared with the vehicle control group and Results are shown in Table-5

                         Duration of paw   -         Duration of paw

%Analgesic = licking for control      licking test compound 100          

     activity             Duration of paw licking for control]

 

 

Table 5: Formalin induced licking and biting response in mice.

Compound	Dosage	Licking / biting (time in secs) Mean ± S.E.M		% Analgesic activity
		0-5 min	5-25 min	0-5 min	5-25 min
Control 3a 3b 3c 3d 3e 3f 3g 3h 3i 3j 3k 3l Diclofenac sodium	20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg 20 mg/kg	18 ± 0.5774 15 ± 1.732 12.66 ± 1.202 19 ± 1.155 18.33 ± 0.8819 12 ± 0.5774 19 ± 1.732 21.66 ± 2.728 14± 1.00 20 ± 0.5774 25 ± .0155 16 ± 0.5774 11 ± 1.155 10 ± 0.5774	26 ± 3.1215 14 ± 0.5774 10± 2.082 17 ± 1.155 13.33 ± 0.8819 12 ± 1.155 15 ± 2.082 17 ± 3.606 9 ± 1.155 20 ± 2.082 17.33 ± 1.764 16 ± 1.00 8.0 ± 1.528 7.0 ± 1.528	0 16.66 27.77 - - 33.33 - - 22.22 - - 11.11 38.88 44.44	0 46.15 61.53 34.61 30.76 53.84 42.30 34.61 65.68 23.07 33.34 38.46 69.23 73.07

N=6, * - P < 0.05, ** - P < 0.01 compared with vehicle treated group, Data expressed as Mean ± SEM­.

Data was analysed by one-way ANOVA followed by Dunnett’s test.

 

SCHEME

 

R= H, 2-NO₂, 3-NO₂, 4-NO₂, 2-Cl, 3-Cl, 4-Cl, 4-N(CH₃) ₂, 4-CH₃, 4-OCH₃, 4-Br, 3,4,5(OCH₃)₃.

 

 

 

REFERENCES:

1.       McBride W G, Lancet, 1961, 2, pp1358

2.       Sheskin. J, Clin. Pharmacol. Ther, 1965, 6, pp 303.

3.       Sampaio E. P, Sarno E N, Galily R Cohn, Kaplan Z A, G J, Exp. Med. 1991, pp 173, 699.

4.       Peuckmann V, Fisch M, Bruera E, Drugs 2000, pp 60, 273.

5.       Krishnamurthy A, Krishnamohan Rao K S R, Subbarao N V, J Indian Chem Soc, 49, 1972, pp1025.

6.       Gardner T S, Wenis E, Le J, J org Chem, 26, 1961, 1514.

7.       Nitti V, Tanzi P L, Arch Tisiol Mal App, 12, 1957, 41, Chem Abstr, 51, 1957, 13069.

8.       Teubner H, Quittschorr G, Stopsack H, Schmidt M, Chemnitins K H, Tauchnitz C, Ezoldand W H, Ger (east) DD , 156, 428, Chem Abstr, 98, 1983, 89068.

9.       Vogel H.G, In Drug Discovery and Evaluation Pharmacological assays, 2 ed, Springer 2002, pp 345-346.

10.     Winter C A, Risley E A, Nuss C W, Proceedings of Society Experimental Biology Journal, 1962, 3(NY), 544-547.

11.     Turner R A, In Screening methods in pharmacology, Academic Press, New York, 1965, Vol 1, pp 125-127.

12.     Leal L K, A M Ferreira, A A G Bezerra, G. A.; Matos, F. J. A. J Ethnopharmacology , 2000, 70, 151-159.

 

 

Received on 18.11.2010        Modified on 28.11.2010

Accepted on 05.12.2010        © AJRC All right reserved