Design, Synthesis and Biological Evaluation for Anti-Inflammatory Activities of Oxadiazole Substituted of Phenothiazine Derivatives

The inflammatory response represents a generalized response to infection or tissue damage and is designed to remove cellular debris, to localize invading 16 organisms and arrest the spread of infection. The inflammatory response is characterized by the following symptoms: Reddening of the localized area, swelling, pain and elevated temperature. Reddening results from capillary dialation that allows more blood to flow to the damaged tissue. Elevated temperature results from capillary dialation which permits increased blood flow through these vessels, with associated high metabolic activities of neutrophils and macrophages. The dialationof blood vessels is accompanied by increased capillary permeability causing swelling as fluid accumulates inthe spaces surrounding tissue and cells.  Oxadiazoles are an important class of bioactive and industrially important organic compounds. Steroidsand non-steroidal anti-inflammatory drugs are globally used for reducing inflammation in the body.[1]

Phenothiazine is a nitrogen and sulfur containing heterocyclic ring which is having the dopamine antagonist activity.It is a basic anti-psychotic drug, but it have many activity like antidepressant[2], antimicrobial,[3], [8] , antioxidant [4], [5] , antifungal [6] , [7] , antibacterial [12], anti-tuberculosis[9] [12], antipsychotic[11],  anti-inflammatory[8],[13], [14], [15], etc.

They have been utilized as a source of useful chemicals since the dawn of civilization. Plants are the basis of many traditional medicine systems throughout the world for thousands of years and continue to provide with new remedies. Plant based medicines, dispensed in the form of crude drugs now serve as  the basis of novel drug discovery.

The use of plants in modern medicine started with the isolation of active compounds, beginning with the isolation of morphine from opium in the early nineteenth century. This was closely followed by the isolation of early drugs such as cocaine, codeine and quinine of which some are still in use. Isolation and characterization of pharmacologically active compounds from medicinal plants continue to this day. The chance of finding new compounds from the plant kingdom is enormous. Till date, only 1 % of the total tropical plants have been studied for their pharmaceutical potential. Drug discovery from plants has evolved to include numerous interdisciplinary fields and various methods of analyses. These involve random selection of plants followed by chemical screening or biological assays, follow up of biological activity reports and follow up of ethno medical use of plants etc

Experimental data

MATERIALS AND METHODS:

CHEMISTRY: The chemicals which are used in this study were supplied by E. Merck and LOBA Co. All the reactions were monitored by TLC using silica gel G. The melting point determinations were done by using in open glass capillary using Kjeldahl flask containing liquid paraffin. IR spectra were recorded on the (JASCO) FTIR-Spectrophotometer using KBr pellets. 1HNMR spectra were recorded on BRUKER AVANCE II 400 NMR spectrometer in DMSO using tetra methyl silane (TMS) as internal reference.

SYNTHESIS OF PHENOTHIAZINE DERIVATIVES:

Procedure for Scheme:

Step 1) General procedure for the preparation of  4-(Cyclohexylideneamino)  benzoic acid derivative.   

Equimolar amount of substituted p-amino benzoic acid added to cyclohexanone and the reaction mixture was heated under refluxed at about 80 C temperature ,for 2 h. the mixture was cooled by addition of a water/ice mixture. the solid was filtered in excellent.

Step 2) General procedure for  the preparation of ethyl 4-(cyclohexylideneamino) benzoate.

Equimolar amount 4-(cyclohexylideneamino )benzoic acid added to ethyl alcohol and the reaction mixture was heated 2 h. poured into ice/water mixture. The precipitate was filter and wash with cold water.

Step 3) General  procedure  for the   preparation of 4-(cyclohexylideneamino) benzohydrazide.

Equimolar   amount  ethyl 4-(cyclohexylideneamino) benzoate added to hydrazine hydrate . and  the reaction mixture was heated 2 h. poured into ice/water. The precipitate was filter and wash   with cold water.

Step 4) General   procedure  for the preparation of 10 H-Phenothiazine-2-carbohydrazide.

Equimolar  amount  4-(cyclohexylideneamino)  benzohydrazide  added  sulfer  .and  the  reaction  mixture  was  heated 2 h. poured  into ice/water  .the precipitate  was filter and was with cold water.

Step 5) General   procedure for  the prepration  (E) –N (argiomethylene)-10H phenothiazine -2-carbohydrazide.

Equimolar  amount  10 H –phenothiazine -2-carbohydrazide  added to various  aldehyde  and  the reaction  mixture was heated  2 h . poured into ice/water .the precipitate was filter and wash with  cold water.

Step 6) General  procedure for the prepration  (z)-N-(argiomethylene)-10-(2 chloroacetyl)

Equimolar  amount  (E) –N-(argiomethylene) -10H-phenothiazine -2-carbohydrazide added to  2-chloroacetyl  chloride  and the reaction  mixture was heated 2 h. poured  into ice/water .the precitate was filter  and was filter  and wash with cold water.

SCHEME:

For Compounds: A-1 To A-12

Analytical & Physicochemical Data of the Synthesized Compound

• IR DATA

A1: IR (cm-1) : 3134.80(-OH str.), 3023.15(-ArCH str.), 1725.20 (-C=O str.), 1580.70(-C=N str.), 1335.42 (-C-O str.), 1404.10(-C-N str.), 678.12(-C-S str.)

1HNMR: 7.41 1H of CHphenothiazine, 6.7-7.3= 14 H m of ArCH , 9.71= 1H s of-CHO of aldehyde,   4.73= 1H s of CH of thiazine               

A2: IR (cm-1) : 822.50(-C-Cl str.), 3083.15(-Ar-CH str.), 1725.11(-C=O str.), 1518.32(- C=N str.), 1256.36(-C-N str.), 3255.23(-N-H str.), 1360.32(- C-O str.), 676(-C-S str.)

1HNMR: 6.7-7.3= 14 H m of ArCH , 8.0= 1H s of-CHO of aldehyde, 4.73= 1H s of CH of thiazine 6.42 = 1H s of CH phenothiazine, 12.74= 1H of  OHcarboxylic acid

A3: IR (cm-1) : 822.50(-C-Cl str.), 3023.15(-Ar-CH str.), 1725.11(-C=O str.), 1518.32(-C=N str.) 1256.36(-C-N str.), 3255.23(-N-H str.), 1360.32(-C-O str.), 676(-C-S str ).

1HNMR: 6.7-7.3= 14 H m of ArCH , 9.71= 1H s of-CHO of aldehyde, 4.73= 1H s of CH of thiazine  6.42 = 1H s of CH phenothiazine, 12.74= 1H of OH carboxylic acid.

A4: IR (cm-1) : 3356(-N-H str.), 3030 (- Ar-CH str.), 1760(-C=C str.), 1725(-C=O str.), 1540(-C=N str.), 1454 -C-N str., 1043 (-C-O str.), 676(-C-S str.)

1HNMR: 6.42 = 1H s of CH phenothiazine, 12.74= 1H of OH carboxylic acid, 4.73= 1H s of CH of diazine.

A5: IR (cm-1) : 3023.10(-Ar-CH str.), 1731.15 (-C=O str.), 1532.11(-C=N str.), 1236.60 (-C-N str.), 3223.50-N-Hstr., 1321.21–C-Ostr., 676(-C-S str.)

1HNMR: 1.45=1H s of CH methine, 7.00=1H s of CH phenothiazine, 2.56 of 1H s CH2 methylene,7.40 of 1H s CH1-benzylidinimine.

A6: IR (cm-1) : 3023.15(-Ar-CH str.), 1716.11(-C=O str.), 712.12(-Br str.), 1518.32(-C=str.), 1256.36(-C-Nstr.), 3255.23(-N-H str.), 1360.32(- C-O str.), 676(-C-S str.).

1HNMR: 6.42 = 1H s of CH phenothiazine, 12.74= 1H of OH carboxylic acid, 4.73= 1H s of CH of thiazine,7.40= of 1H s CH 1-benzene.

A7: IR (cm-1) : 3356.80(-N-H str.), 3030.10(-Ar-CH str.), 1760.12(-C=C str.), 1360(-N-O str.), 1725.15(-C=O str.), 1540(-C=N str.), 1454 (-C-N str.), 1043(-C-O str.), 676(-C-S str.). 

1HNMR:7.41 of 1H s CH  phenothiazine, 7.24 of 1H s CH 1-benzene, 12.74= 1H of OH carboxylic acid, 4.73= 1H s of CH of thiazine.

A8: IR (cm-1) : 822.50(-C-Cl str.), 3023.15(-Ar-CH str.), 1725.11(-C=O str.), 1518.32(-C=N str.), 1256.36(-C-N str.), 3255.23(-N-H str.), 1360.32(-C-O str.), 676(-C-S str.).

1HNMR: 6.7-7.3= 14 H m of ArCH , 9.71= 1H s of-CHO of aldehyde, 4.73= 1H s of CH of thiazine, 6.42 = 1H s of CH phenothiazine, 12.74= 1H of  OH carboxylic acid

A9: IR (cm-1) : 3112.80(-OH str.), 3023.15(-ArCH str.), 1725.20(-C=O str.), 1580.70(-C=N str.), 1325.42 (-C-O str.), 1404.10 -C-N str., 676.12(-C-S str.).

1HNMR: 6.7-7.3= 14 H m of ArCH, 9.71= 1H s of-CHO of aldehyde, 4.73= 1H s of CH of thiazine,  6.42 = 1H s of CH phenothiazine, 12.74= 1H of  OH carboxylic acid.

A10: IR (cm-1) : 3112.80(-OH str.), 3023.15(-ArCH str.), 1725.20(-C=O str.), 1580.70(-C=N str.), 1325.42 (-C-O str.), 1404.10 -C-N str., 676.12(-C-S str.).

1HNMR: 6.7-7.3= 14 H m of ArCH, 9.71= 1H s of-CHO of aldehyde, 4.73= 1H s of CH of thiazine,  6.42 = 1H s of CH phenothiazine, 12.74= 1H of OH carboxylic acid.

Anti-inflammatory activity-

• Carrageenan-Induced Paw Edema method

Carrageenan - induced rat paw edema method was employed for evaluating the anti inflammatory activity of the synthesized compounds (A1- A8). Wister Albino rats of either sex weighing approx 200- 300 gm, were housed in clean polypropylene cages and kept under room temperature (25±2OC), and relative humidity 40-50% in a 12 h light-dark cycle. Food was withdrawn 12 h before and during experimental hours. In this study, the animals were divided into 4 groups containing 5 animal  per groups .first group (Negative  control) received normal saline solution (10ml/ kg . Acute inflammation was produced by sub plantar injection of 0.1ml of 1%  w/v solution  of Carrageenan  in the right hind paw of the rats. After oral administration of the test compounds, the paw volume was measured at 30 min ,60 min, 90 min and 120min intervals. Ibuprofen 100mg/kg of was used as standard drug. The percentage inhibition of denaturation was calculated by using following formula.

% of Inhibition = 100 X [1- Vt / Vc]

Where,

Vt = Mean absorbance of test sample.

Vc = Mean absorbance of control