“ANALYTICAL METHOD DEVELOPMENT AND VALIDATION FOR SOME ANTIHYPERTENSIVE DRUGS”
Synopsis for registration of M.Pharm Dissertation
Submitted to
RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES, BENGALURU
KARNATAKA
In partial fulfillment
Of the requirement for the Degree of
Master in Pharmacy Pharmaceutical Analysis
Under the Guidance of
Mr. T. SREENIVASA RAO
Asst. Professor
DEPARTMENT OF PHARMACEUTICAL ANALYSIS
BY
BINA NAMA
I M.PHARM
DEPARTMENT OF PHARMACEUTICAL ANALYSIS
KARNATAKA COLLEGE OF PHARMACY, BENGALURU-64
2012-2013
RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES
KARNATAKA, BENGALURU.
ANNEXURE-II
PROFORMA FOR REGISTRATION OF SUBJECTS FOR DISSERTATION
1. / Name of the candidate and address / BINA NAMAKARNATAKA COLLEGE OF PHARMACY,
#33/2, THIRUMANAHALLI,
HEGDE NAGAR MAIN ROAD,
JAKKUR POST, YELAHANKA HOBLI,
BENGALURU-560064,
KARNATAKA.
PERMANENT ADDRESS
D/O :NEPAL CHANDRA NAMA
NORTH SRIRAMPUR
SIDDHINAGAR(P.O)
BELONIA,SOUTH TRIPURA
TRIPURA 799157
2. / Name of the Institution / KARNATAKA COLLEGE OF PHARMACY,
33/2,THIRUMENAHALLI,
HEGDENAGAR,
BENGALURU-560064,
KARNATAKA, INDIA.
3. /
Course of study and subject
/ MASTER OF PHARMACY(Pharmaceutical Analysis)
4. /
Date of the admission
/ 24-01-20135. /
Title of the topic
“ ANALYTICAL METHOD DEVELOPMENT AND VALIDATION FOR SOME ANTIHYPERTENSIVE DRUGS”6
7
8 /
BRIEF RESUME OF THE INTENDED WORK:
6.1 Need for the study:Hypertension (HTN) is a cardiac chronic medical condition in which the systemic arterial blood pressure is elevated.
WHO-ISH guidelines (2003) have defined it to be 140mm Hg systolic and 90mm Hg diastolic, though risk appears to increase even above 120/80 mm Hg1. [2]
The entire world observing World Hypertension Day every year on the 17th of May. Hypertension is also known as the ‘Silent Killer’. Hypertension had accounted for around 972 million people globally in 2000 alone out of which 693 million belonged to economically developing nations which include India. Experts predict this number to swell further, so much so that by the year 2025, hypertension will affect around 1.56 billion people world over.
The effects of hypertension include stroke, ischemic heart disease, dementia, heart failure and sometimes it leads to death also. This is the reason to increase the development of new anti-hypertensive agents.
The antihypertensive agents are a class of drugs that are used to treat hypertension (elevated blood pressure). The reduction of the blood pressure by 5 mmHg can decrease the risk of stroke by 34%, of ischemic heart disease by 21%, and reduce the likelihood of dementia, heart failure, and mortality from cardiovascular disease. There are many classes of anti hypertensive, which lower blood pressure by different means; among the most important and most widely used are the thiazide diuretics, the ACE inhibitors, the calcium channel blockers, the beta blockers, and the angiotensin II receptor antagonists or ARBs.
Need for new and improved antihypertensive agents:[4]
Antihypertensive agents lower the elevated blood pressure, although the mechanisms of action vary greatly. A Joint National Committee on Prevention, Detection, Evaluation, and Treatment of High Blood Pressure report said that recent clinical trials show that antihypertensive treatment can reduce incidence of stroke by 35-40%, heart attack by 20-25%, and onset of new heart failure by 50%. There are a large number of drugs used to control hypertension. But as the need for antihypertensive agents is rising due to various factors especially high levels of stress. There is a constant need to develop new drugs in this class which can overcome the side effects like dizziness, head ache, anxiety, stomach disorders, decreased heartbeat, and constipation etc. of drugs that were earlier used.
Generally chromatographic methods (HPLC, HPTLC, and GC) are used because it fulfills requirement of various guidelines. The interest of work emphasizes on HPLC because, in the modern pharmaceutical industry HPLC (high performance liquid chromatography) is the major and integral analytical tool applied in all stages of drug discovery, development, and production.
HPLC Method development and validation are performed in order to meet the ICH requirements to prepare for regulatory submissions (e.g., NDA). “The object of validation of an analytical procedure is to demonstrate that it is suitable for its intended purpose.” The advantages of HPLC are simple, accurate, precise, sensitive and specific.
The need of the proposed project is to develop and validate new anti-hypertensive drug and to ensure regulatory compliance as per ICH Guidelines.
Classification of Antihypertensive drugs:[2]
A. Diuretics:
Hydrochlorothiazide, Chlorthiazide, Furosemide
B. ACE Inhibitors:
Ramipril, Captopril, Lisinopril, Enalapril
C. Angiotensin blockers:
Losartan, Valsartan, Telmisartan, Azilsartan
D. Calcium channel blockers:
Amlodipine, Verapramil, Diltiazem,Cilnidipine
E. α - adrenergic blockers :
Prozosin, Terazosin, Phenoxy benzamine
F. α,β adrenergic blockers:
Labetolol, Carvidilol
G. β adrenergic blockers:
Atenolol, Propanolol, Metaprolol
H. Central sympholytics:
Clonidine, Methyl dopa
I. Vasodilator:
Hydralazine, Minoxidil
DRUG PROFILE
1. Furosemide:[3]
IUPAC name: 4-chloro-2-(furan-2-ylmethylamino) - 5-sulfamoylbenzoic acid.
Molecular formula: C12H11ClN2O5S
Mechanism of action:
In the kidneys, salt (composed of sodium and chloride), water, and other small molecules normally are filtered out of the blood and into the tubules of the kidney. The filtered fluid ultimately becomes urine. Most of the sodium, chloride and water that is filtered out of the blood is reabsorbed into the blood before the filtered fluid becomes urine and is eliminated from the body. Furosemide works by blocking the absorption of sodium, chloride, and water from the filtered fluid in the kidney tubules, causing a profound increase in the output of urine (diuresis).
2.Captopril
IUPACname: (2S)-1-[(2S)-2-methyl-3-sulfanylpropanoyl] pyrrolidine-2-carboxylic acid.
Molecular formula: C9H15NO3S
Mechanism of action:
Acts on Kidneys to increase excretion of Na and H2O – decrease in blood volume – decreased BP
3.Losartan
IUPAC name: (2-butyl-4-chloro-1-{[2'-(1H-tetrazol-5-yl)biphenyl-4-yl]methyl}-1H-imidazol-5-yl) methanol.
Molecular formula: C22H23ClN6OMechanism of action:
Losartan competitively inhibits the binding of angiotensin II to AT1 in many tissues including vascular smooth muscle and the adrenal glands. Losartan is metabolized to its active metabolite, E-3174, which is 10 to 40 times more potent than Losartan and acts as a non-competitive AT1 antagonist. Inhibition of angiotensin II binding to AT1 inhibits its AT1-mediated vasoconstrictive and aldosterone-secreting effects and results in decreased vascular resistance and blood pressure. Losartan is 1,000 times more selective for AT1 than AT2. Inhibition of aldosterone secretion may increase sodium and water excretion while decreasing potassium excretion. Losartan is effective for reducing blood pressure and may be used to treat essential hypertension, left ventricular hypertrophy and diabetic nephropathy.
4. Amlodipine :
IUPAC NAME: (RS)-3-ethyl 5-methyl 2-[(2-aminoethoxy)methyl]-4-(2-chlorophenyl)-6-methyl-1,4-dihydropyridine-3,5-dicarboxylate
Molecular formula: C20H25ClN2O5
Mechanism of action: Amlodipine decreases arterial smooth muscle contractility and subsequent vasoconstriction by inhibiting the influx of calcium ions through L-type calcium channels. Calcium ions entering the cell through these channels bind to calmodulin. Calcium-bound calmodulin then binds to and activates myosin light chain kinase (MLCK). Activated MLCK catalyzes the phosphorylation of the regulatory light chain subunit of myosin, a key step in muscle contraction. Signal amplification is achieved by calcium-induced calcium release from the sarcoplasmic reticulum through ryanodine receptors. Inhibition of the initial influx of calcium decreases the contractile activity of arterial smooth muscle cells and results in vasodilation. The vasodilatory effects of amlodipine result in an overall decrease in blood pressure. Amlodipine is a long-acting CCB that may be used to treat mild to moderate essential hypertension and exertion-related angina (chronic stable angina). Another possible mechanism is that Amlodipine inhibits vascular smooth muscle carbonic anhydrase I activity causing cellular pH increases which may be involved in regulating intracellular calcium influx through calcium channels.
5.Atenolol:
IUPAC name: (RS)-2-{4-[2-Hydroxy-3-(propan-2-ylamino)propoxy]phenyl}acetamide
Molecular formula: C14H22N2O3
Mechanism of action: Bind to beta adrenergic receptors and blocks the activity
6.Labetolol:
IUPAC name: (RS)-2-hydroxy-5-{1-hydroxy-2-[(1-methyl-3-phenylpropyl)amino]ethyl}benzamide
Molecular formula: C19H24N2O3
Mechanism of action: Labetolol HCl combines both selective, competitive, alpha-1-adrenergic blocking and nonselective, competitive, beta-adrenergic blocking activity in a single substance. In man, the ratios of alpha- to beta- blockade have been estimated to be approximately 1:3 and 1:7 following oral and intravenous (IV) administration, respectively. The principal physiologic action of Labetolol is to competitively block adrenergic stimulation of β-receptors within the myocardium (β1-receptors) and within bronchial and vascular smooth muscle (β2-receptors), and α1-receptors within vascular smooth muscle. This causes a decrease in systemic arterial blood pressure and systemic vascular resistance without a substantial reduction in resting heart rate, cardiac output, or stroke volume, apparently because of its combined α- and β-adrenergic blocking activity.
7.Prazocin:
IUPAC name: 2-[4-(2-furoyl)piperazin-1-yl]-6,7-dimethoxyquinazolin-4-amin
Molecular formula: C19H21N5O4
Mechanism of action: Prazosin acts by inhibiting the postsynaptic alpha (1)-adrenoceptors on vascular smooth muscle. This inhibits the vasoconstrictor effect of circulating and locally released catecholamine (epinephrine and nor epinephrine), resulting in peripheral vasodilation.
6.2 REVIEW OF LITERATURE:
1. Blessen Philip et al have developed the RP-HPLC method for the simultaneous estimation of Atenolol & Amlodipine Besylate in pharmaceutical dosage form by using Agilent technologies 1200 series HPLC, Inertsil C18 column (5µm x 250mm x 4.6mm); mobile phase buffer: acetonitrile: methanol (4:3.5:2.5), UV detection at 225 nm and linearity range is 50-150 µg/ml for Atenolol and 5-15 µg/ml for Amlodipine besylate.[5]
2. Micheli wrasse sangai et al have developed UV-spectroscopic method for the determination of Aliskiren in tablets, linearity is 40-100 µg/ml.[6]
3. Zaveri Maitreyi et al have developed simultaneous estimation of Atenolol & Hydrochlorothiazide by RP-HPLC using Zorbax SB-CN (250 x 4.6) mm, 5µm column, in isocratic mobile phase water:buffer:methanol (50:35:15), UV detection is 286 nm and linearity is 50-150 µg/ml for Atenolol ang 12-6-75.6 µg/ml for Hydrochlorothiazide respectively.[6]
4. Jignesh Prajapati et al have developed simultaneous estimation of Amlodipine Besylate & Perindopril Erbumine by RP-HPLC using Eclipse C8 column (150mm x 4.6mm); 5µm; mobile phase is buffer:acetonitraile (65:35); pH adjust to 2.6 by orthophosphoric acid, UV detection is 210 nm and linearity is 10-75 µg/ml for Amlodipine Besylate and 8-60 µg/ml for Perindopril.[7]
5. M. I. Maguregui et al have developed simultaneous determination of β- blocker Atenolol & several complementary Antihypertensive agents in pharmaceutical formulation and urine by capillary zone electrophoresis using fused silica capillary (78 cm-75 µm), 70 µm length, borate buffer is used and 25 kV is applied.[8]
6. P. S. Jain et al have developed stability indicating method for Olmesartan medoxomile in pharmaceutical dosage from and in pure drug by RP-HPLC using LC, GC, BDS C18 column (250 mm x 4.5 mm, 5 µm). Mobile phase A (7 ml triethylamine in 1000 ml water) and B acetonitrile in water. UV detection is 257 nm and linearity range is 50-100 µg/ml.[9]
7. Y. Nirupa Rani et al have developed the estimation of Carvedilol in bulk and dosage form by new analytical method such as UV method and also UV along with chromatography. The absorption maxima was detected at 286 nm and linearity range is 50-150 µg/ml.[10]
8. Belal. F et al have developed the determination of atenolol by stability indicating HPLC method using C8 column (250mm x 4.6mm; i.d: 5µm); mobile phase acetonitrile:methanol:0.02M phosphate buffer; UV detection at 226 nm and linearity range 0.05-10 µg/ml.[11]
9. R. Veereswara Rao et al have developed simultaneous estimation of metaprolol succinate and hydrochlorothiazide by RP-HPLC using YMC C18 column; mobile phase acetonitrile and buffer ratio is 32:68; UV detection is 225 nm and linearity range for metaprolol is 1-300 µg/ml and for hydrochlorothiazide is 0.126-37.687 µg/ml.[12]
10. Singh Brijesh et al have developed simultaneous estimation of metaprolol succinate and hydrochlorothiazide by RP-HPLC method using C18 column mobile phase is 50 mM disodium hydrogen phosphate:methanol:acetonitile ratio is 525:225:250; compound is detected at 222 nm and linearity is 2-32 µg/ml.[13]
11. Thamake Sl et al have developed simultaneous estimation of Atorvastatin & Ramipril by first order derivative spectroscopy scanning 10 µg/ml of each drug at 200-350 nm range. At 294 Atorvastatin show max absorbance at Zero point of Ramipril and Ramipril shown max absorbance at 229 nm at zero crossing point of Atorvastatin and linearity for Atorvastatin is 2-20 µg/ml and Ramipril 5-40 µg/ml.[14]
12. Delhiraj et al have developed simultaneous estimation of antihypertensive drugs by Chromatographically method using Qualisil BDS C18 (250 x 4.6mm;I.d. 5µm), mobile phase consist of 1.0ml triethylamine with orthophosphoric acid and acetonitrile (60:40); UV detection is 231 nm and linearity range to HPTLC is 400-1600 ng/ml and 200-800 ng/ml respectively. For second method column used is silica gel 60F254, mobile phase is chloroform:methanol (85:15) and detected at 231 nm and linearity range is 2-25 µg/ml for Amlodipine and 2-30 µg/ml for Olmesartan.[15]
13. Bheru S. Malviya et al have developed simultaneous determination of Irbesartan , Hydrochlorothiazide, Amlodipine besylate and Atorvastatin by UPLC using ACQUITY UPLC BEH C18 (2.1 x150mm, id 1.7µm), mobile phase is potassium hydrogen phosphate buffer:acetonitrile (70:30), UV detection is at 254 nm and linearity is Amlodipine and Hydrochlorothiazide is 4-24 µg/ml, Atorvastatin 5-30 µg/ml & Irbesartan 10-60 µg/ml.[16]
14. Napa delhiraj et al have developed simultaneous estimation of antihypertensive drug by stability indicating gradient RP-HPLC using INERTSIL ODS C18-3V (150 x 4.6mm, 5µm) using mobile phase 1ml TEA in 1L water and pH is adjusted to 2.5 with orthophosphoric acid and acetonitrile, UV detection at 225 nm and linearity is 25-150 µg/ml and 15-90 µg/ml for Olmesartan and Hydrochlorothiazide respectively.[17]
15. Lakshmana Rao et al have developed simultaneous estimation of Atorvastatin and Amlodipine by Spectrophotmetric method by measuring absorptivity at 242 nm & 364 nm ; and linearity is 1-20 µg/ml and 1-50 µg/ml for Atorvastatin and Amlodipine respectively.[18]
16. Jothieswari D et al have developed simultaneous determination of Valsartan, Hydrochlorothiazide and Amlodipine besylate by RP-HPLC using RP C18 column using mobile phase acetonitrile:methanol:phosphate buffer (20:50:30), UV detection is 239 nm and linearity is 0.5-5 µg/ml for Amlodipine, 4-40 µg/ml for Valsartan & 1-10 µg/ml for Hydrochlorothiazide.[19]
17. Boyka G et al have developed simultaneous determination of metaprolol and aspirin by RP-HPLC using Phenomenex – Luna C18 (250 x 4.6mm;id 5µm) mobile phase is orthophosphoric acid and methanol (20:80); UV detection is 230 nm &linearity range is 20-100 µg/ml for aspirin & 10-50 µg/ml for metaprolol.[20]
18. N. Delhiraj et al have developed simultaneous estimation of antihypertensive compound by RP-HPLC using Quilisil BDS C18 column (250 x 4.6mm; id 5µm), mobile phase consist of TEA in 1L water adjusted pH is 2.5 with orthophosphoric acid and acetonitrile (60:40) and UV detection is 281 nm; in 2nd method silica gel 60F254 is used and UV detected at 281 nm using mobile phase methanol:formic acid:chloroform (15:5:85) and linearity rang for telmisartan, amlodipine and hydrochlorothiazide is 10-50 µg/ml, 2-25 µg/ml, 2.5-17.5 µg/ml.[21]