INTRODUCTION:

An overactive Renin-angiotensin system is the main cause of renal hypertension and is being focused in the present study. When blood volume is low, juxtaglomerular cells in the kidneys secrete renin. Renin stimulates the production of angiotensin I, which is then converted to angiontensin II. Angiotensin II causes blood vessels to constrict, resulting in increased blood pressure. Angiotensin II also stimulates the secretion of the hormone aldosterone from the adrenal cortex. Aldosterone causes the tubules of the kidneys to increase the reabsorption of sodium and water into the blood. This increases the volume of fluid in the body, which also increases blood pressure. ^1,2

Angiotensin II receptor antagonists, also known as angiotensin receptor blockers (ARBs), AT₁-receptor antagonists or sartans, are a group of pharmaceuticals which modulate the renin-angiotensin-aldosterone system.

These substances are AT₁-receptor antagonists – that is, they block the activation of angiotensin II AT₁ receptors. Blockade of AT₁ receptors directly causes vasodilation, reduces secretion of vasopressin, reduces production and secretion of aldosterone, amongst other actions – the combined effect of which is reduction of blood pressure.^3,4 The available agents of this category are Losartan, Candesartan. Irbesartan, Eprosartan, Telmisartan, Olmesartan and Valsartan which have been proved very efficient in controlling hypertension. But, it has been noticed that they possess a number of serious side effects like orthostatic hypotension, dyspepsia, decreased haemoglobin level, insomnia, renal impairement and hyperkalemia. Unfortunately, the frequency of side effects makes it difficult for some patients to continue their long term use. While mono therapy is preferable, it is often necessary to use a second or even third drug to control blood pressure effectively, further increasing the likelihood that the patient will experience untoward reaction.^{5, 6}

This factor promotes the need to invent the better and safer drugs for long term use.

Irbesartan molecule has pyrazolidinone structure and many thiazolidinone derivatives, are reported for their interesting profile of activities. As these two compounds could prove isosteric with the moiety present in Irbesartan, we thought it is worthwhile to condense these two moieties and form their Mannish bases as the resultant compound may act as angiotensin II antagonists. For this reason Mannish bases were prepared and evaluated for their activity.

General Structure of FP

MATERIAL AND METHOD:

Prerparation of Schiff Bases (1): Sulfanilamide (25 mmol) was dissolved in 40 ml boiling ethanol and aromatic aldehyd (25 mmol) was added to this solution. This mixture was refluxed for 3-4 hrs and was then cooled. The solid obtained was filtered, dried and crystallized from 95% ethanol.⁷

Preparation of Thiazolidinones (A):

0.01 mol Schiff base and 0.02 mol thioglycollic acid were dissolved in 30 ml glacial acetic acid. This mixture was refluxed for 4-5 hrs. The reaction mixture was then poured in an ice cool saturated solution of sodium bicarbonate. It was then kept overnight at refrigeration. The product obtained was washed with cold water to remove alkali and crystallized with appropriate solvent.⁸ (Figure 1)

Figure 1 – Synthesis of Thiazolidinones

Preparation of Chalcones (2):

Equimolar mixture of substituted acetophenone (0.08mol) and substituted benzaldehyde (0.08 mol) was added to a mixture of 4.2g sodium hydroxide in 40ml water and 25ml ethanol. The resulting mixture was stirred for 3-4 hrs in an ice bath. The stirred mix was kept under refrigeration overnight. The product was filtered and was crystallized from 95% ethanol.⁹

Preparation of Pyrazolines (B):

Chalcone (0.01 mol) and hydrazine hydrate (0.02 mol) were taken in 20ml glacial acetic acid and the mixture was refluxed for 10-12 hrs. the reaction mixture was poured in 300ml ice cold water and was kept aside for 12 hrs. the product obtained was filtered and crystallized from 95% ethanol.¹⁰ (Figure 2)

Figure 2 – Synthesis of Pyrazolines

Preparation of Mannich Bases (C):

An equimolar mixture of thiazolidinone (0.005 mol) and pyrazoline (0.005 mol) in an appropriate solvent was refluxed for 4-5-hrs. The reaction mixture was poured in 200-300 ml ice cold water and was kept aside for 12 hrs. The product obtained was filtered and crystallized from appropriate solvent.¹¹ (Figure 3)

Mannich Bases:

4-{2-(4-nitrophenyl)-5-[(3-phenyl-5-(4-nitrophenyl))-1,2-pyrazolidin-1-yl)methyl]-4-oxo-thiazolidin-3-yl}benzene sulphonamide; FP1

Mol formula: C₃₁H₂₆O₇N₆S_2; Mol Wt: 658; m.p. 146˚C; Yield 71%; IR (KBr), cm^-1: 3430 (-NH), 1570 (C=N), 1710 (C=O), 1014.56 (C-N), 1508.53 (-N-O), 1448 (S=O), 1363.67 (-NO₂), 690.52 (-C-S); ¹H NMR (DMSO) δppm: 5.683 (-NH₂), 2.337 (-CH₂), 2.507 (-CH₂of pyrazoline), 3.234 (-CH of Thiazolidinone adj to N), 3.167 (-CH of Thiazolidinone adj. to C=O), 3.934-4.097 (-CH of Pyrazoline), 7.327-8.194 (aromatic H); Rf value: 0.72.

4-{2-(4-chlorophenyl)-5-[(3-phenyl-5-(4-bromophenyl)-1,2-pyrazolidin-1-yl) methyl]-4-oxo-thiazolidin-3-yl}benzene sulphonamide; FP2

Mol formula: C₃₁H₂₆O₃N₄S₂ClBr; Mol Wt: 679.5; m.p. 110˚C; Yield 56%; IR (KBr), cm^-1: 3242.34 (-NH), 1590.76 (C=N), 1645.28 (C=O), 1163.08 (C-N), 1496.76 (S=O), 833.25 (-C-S), 750.31 (C-Cl), 617.22 (C-Br); Rf value: 0.45.

4-{2-(4-chlorophenyl)-5-[(3-(4-hydroxyphenyl)-5-(4-methylphenyl)-1,2-pyrazolidin-1-yl) methyl]-4-oxo-thiazolidin-3-yl}benzene sulphonamide; FP3

Mol formula: C₃₃H₃₂O₄N₄S₂; Mol Wt: 612; m.p. 180˚C; Yield 49%; IR (KBr), cm^-1: 3145.90 (OH), 3126.54 (NH), 2814.14 (C-H), 1575.84 (C=N), 1803.44 (C=O), 1022.27 (C-N), 1442.75 (S=O), 1336 (C-S); Rf value: 0.30.

4-{2-[4-(N,N-dimethylaminophenyl)]-5-[(3-(4-chlorophenyl)-5-(4-methoxyphenyl)-1,2-pyrazolidin-1-yl) methyl]-4-oxo-thiazolidin-3-yl}benzene sulphonamide, FP4

Mol formula: C₃₄H₃₄O₄N₅S₂Cl; Mol Wt: 675.5; m.p. 118˚C; Yield 58%; IR (KBr), cm^-1: 3331.07 (-NH), 3255.84 (-CH), 1658.78 (C=O), 1595.13 (C=N), 1444.48 (S=O), 1251.80 (C-O), 1155.36 (C-N), 1024.20 (C-Cl); Rf value: 0.59.

4-{2-(4-bromophenyl)-5-[(3,5-(4,4’-dimethoxyphenyl)-1,2-pyrazolidin-1-yl) methyl]-4-oxo-thiazolidin-3-yl}benzene sulphonamide, FP5

Mol formula: C₃₃H₃₁O₅N₄S₂; Mol Wt: 627; m.p. 110˚C; Yield 64%; IR (KBr), cm^-1: 3257.77 (-OH), 3022.45 (-NH), 2937.59 (C-H), 1907.60 (C=O), 1444.68 (S=O), 1089.78 (C-O), 1024.20 (C-N), 827.46 (C-Br); Rf value: 0.75.

PHARMACOLOGICAL SCREENING¹²:

Method: Induction of Experimental Hypertension- Albino rats weighing 200-250g were anaesthetized with anaesthetic ether. The fur on the back was shaved and the skin was disinfected. In the left lumbar area of flank incision was made parallel to a long axis of rat. The renal pedicel was exposed with the kidney retracted to the abdomen. The renal artery was located and a U-shaped silver clip was slipped around it near the aorta using special forceps, the size of the clip was adjusted so that the internal gap ranges from 0.25-0.38 mm. the right kidney is removed after tying of the renal pedicel. The skin incisions were closed and appropriate treatment was given to prevent infection.

Blood pressure was measured 4-5 weeks after clipping and rats with values higher than 150 mm Hg were selected for the experiments. Blood pressure readings were taken on each of 3 days prior to the drug treatment. Rats were divided into 6 animals per dose and each animal was used as its own control. One of the groups will serve as standard and given 27mg/kg of Irbesartan. Initial reading (prior) and 1 hr post-drug blood pressure readings were taken.

Measurement of Systolic Blood Pressure- The systolic blood pressure is measured by tail cuff method by Harvard non invasive BP apparatus. The acquisition of the data was done by bio-pack data acquisition system and visualized on the computer screen.

Tail cuff method is a common and convenient means to measure systolic blood pressure in rats. The tail cuff in inflated and then deflated. Pulsations disappear when cuff is inflated. When cuff starts deflating pulsations start appearing when pressure in the cuff equals systolic pressure. The cuff is attached to Harvard non-invasive BP monitor and BP is recorded. The results were analyzed by one-way ANOVA followed by Dunnet’s test (p-value ≤ 0.05 was taken as significant).

Table 1:

Sr. No	Comp. Code	R₁	R₂	R₃	Mol. Formula	% Yield	M.P. (˚C)	R_f
1.	FP1	4- NO₂	4- NO₂	H	C₃₁H₂₆O₇N₆S₂	71	146	0.72
2.	FP2	4- Cl	4- Br	H	C₃₁H₂₆O₃N₄S₂ClBr	56	110	0.45
3.	FP3	4-CH₃	4-CH₃	4-OH	C₃₃H₃₂O₄N₄S₂	49	180	0.30
4.	FP4	4- N(CH₃)₂	4- OCH₃	4-Cl	C₃₄H₃₄O₄N₅S₂Cl	58	118	0.59
5.	FP5	4- Br	4- OCH₃	4- OCH₃	C₃₃H₃₁O₅N₄S₂	64	110	0.75

Table 2: Anti-hypertensive activity of synthesized compounds

Group No.	Drug	Mean blood Pressure in mm of Hg Mean ± SE		P value
Group No.	Drug	Before Treatment	After Treatment	P value
1.	Control	178.8 ± 1.986	178.2 ± 1.325	-----
2.	Irbesartan	166.4 ± 2.456	94.52 ± 3.482	<0.05
3.	FP-1	164.3 ± 3.485	140. 4 ± 2.761	<0.05
4.	FP-2	168.9 ± 2.445	132.6 ± 2.687	<0.05
5.	FP-3	176.6 ±2.121	144.0 ± 4.886	<0.05
6.	FP-4	165.3 ± 3.845	136.4 ± 2.102	<0.05
7.	FP-5	170.1 ± 2.098	134.2 ± 2.354	<0.05

RESULT AND DISCUSSION:

Mannich condensation is very useful method which acts as a bridge to combine two different moieties. The Thiazolidinones were made to combine with pyrazolines, the yields were optimum. Toxicity studies were performed along with 1K-1C Goldblatt method and the effect of the newly synthesized compounds was comparable to standard drug Irbesartan, which gave the results shown in table 2. The compound synthesized appeared to be quite promising to the extent and that they may be less toxic.

SUMMARY AND CONCLUSION:

We have synthesized various thiazolidinone and pyrazoline derivatives and combined them to form their Schiff bases and then carried out test for their angiotensin II blocking action. The Mannich base compounds of thiazolidinones and pyrazolines proved to be useful for anti-hypertensive activity. Further design may prove an alternative and very useful and fruitful in the discovery of new angiotensin II blocking activity in comparison to losartan, Irbesartan, etc.

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Received on 20.05.2013 Modified on 16.06.2013

Asian J. Research Chem. 6(7): July 2013; Page 641-644