CHAPTER 9

TELMESARTAN

242

9.1. DRUG PROFILE

Telmisartan is a member of a family of drugs called angiotensin receptor

blockers (ARBs), which includes losartan (Cozaar), valsartan (Diovan), irbesartan

(Avapro), and candesartan (Atacand). Telmisartan was approved by the FDA in

November of 2000. Telmisartan is used alone or in combination with other

medications to treat high blood pressure.

Telmisartan is an angiotensin II receptor blocker that shows high affinity for

the angiotensin II receptor type 1 (AT1), with a binding affinity 3000 times greater for

AT1 than AT2. It has the longest half-life of any ARB (24 hours) [1-2]

and the largest

volume of distribution]. In addition to blocking the RAs, Telmisartan acts as a

selective modulator of peroxisome proliferator-activated receptor gamma (PPAR-γ), a

central regulator of insulin and glucose metabolism. It is believed that Telmisartan’s

dual mode of action may provide protective benefits against the vascular and renal

damage caused by diabetes and cardiovascular disease (CVD).[2]

It is used to treat

high blood pressure (hypertension [1]

). High blood pressure reduction helps prevent

strokes, heart attacks, and kidney problems. This drug works by blocking the hormone

angiotensin thereby relaxing blood vessels, causing them to widen. This drug may

also be used to treat congestive heart failure and to help protect the kidneys from

damage due to diabetes.

Figure: 9.A: Structure of Telmisartan

243

Systematic (IUPAC) name : 2-(4-{[4-methyl-6-(1-methyl-1H-1,3-benzodiazol-

2-yl)- 2-propyl-1H-1,3-benzodiazol-1 yl] methyl}

phenyl) benzoic acid

Formula : C33H30N4O2

Mol. Mass : 514.617 g/mol

Routes : Oral

Excretion : Faecal 97%

This drug can cause serious (possibly fatal) harm to an unborn baby if used

during pregnancy. Therefore, it is important to prevent pregnancy while taking this

medication. Other Side effects are similar to other angiotensin II receptor antagonists

and include tachycardia and bradycardia (fast or slow heartbeat), hypotension (low

blood pressure), edema (swelling of arms, legs, lips, tongue, or throat, the latter

leading to breathing problems), and allergic reactions.[3]

The ONTARGET Trial

Programme (The Ongoing Telmisartan Alone and in combination with Ramipril

Global Endpoint Trial) was one of the largest ARB clinical study programmes ever

undertaken.[4]

25,620 patients from 733 centers in 41 countries were randomized for

5.5 years of treatment of either Telmisartan the ACE inhibitor ramipril or a

combination of the two.

List of brand names of Telmisartan:

S.No. BRAND NAME

FORMULATIO

N

COMBINATION AVAILABLE

STRENGTH

Mg

MANUFACTURER

1 ARBITEL-AV Tab Atorvastatin 50 MICRO

2 CRESAR 80 Tab 80 CIPLA

3 CRESAR AM Tab Amlodipine 50 CIPLA

4 CRESLIP Tab Atorvastatin 50 CIPLA

5 INDITEL Tab 20 ZYDUS ALIDAC

6 INDITEL AM Tab Amlodipine 50 ZYDUS ALIDAC

244

7 LIPISAR Tab Atorvastatin 50 INTAS

8 LIPISAR 40 Tab Atorvastatin 50 INTAS

9 STAMLO-T Tab Amlodipine 50 DR. REDDY'S

LABS

10 TALY-40 Tab 40 GENETIC PHARMA

11 TALY-H Tab Hydrochlorothiazide 60 GENETIC PHARMA

12 TARGIT H Tab Hydrochlorothiazide 60 PFIZER

13 TARGIT Tab 20 PFIZER

14 TARGIT Tab 80 PFIZER

15 TELAST Tab 20 INTAS

16 TELDAY AV Tab Atorvastatin 50 TORRENT

17 TELEACT AM Tab Amlodipine 50 RANBAXY

18 TELIPRIL Tab Ramipril 50 SUN PHARMA

19 TELIPRIL Tab Ramipril 50 SUN PHARMA

20 TELISTA 20 Tab 20 LUPIN

21 TELISTA-H Tab Hydrochlorothiazide 100 LUPIN

22 TELMA Tab 20 ZOLTAN

(GLENMARK)

23 TELMIKIND Tab 80 MANKIND

24 TELMIKIND-

AM

Tab Amlodipine 50 MANKIND

25 TELMIKIND-H Tab Hydrochlorothiazide 100 MANKIND

26 TELMINORM Tab 20 IPCA

27 TELMISAT 40 H Tab Hydrochlorothiazide 60 BIOCON

28 TELMISAT 80 Tab Hydrochlorothiazide 100 BIOCON

29 TELMITOP HC Tab Hydrochlorothiazide 60 ELLIFE (ELDER)

30 TELMITOP Tab 20 ELLIFE (ELDER)

31 TELMITOP Tab 40 ELLIFE (ELDER)

32 TELPRES Tab 20 NICHOLAS

33 TELSAR Tab 40 UNISEARCH

245

Table 9.1

34 TELSARTAN Tab 20 DR. REDDY'S

LABS

35 TELSARTAN-R Tab Ramipril 50 DR. REDDY'S

LABS

36 TELSITE Tab 20 AVENTIS PHARMA

37 TETAN Tab 80 ALEMBIC

38 TELVAS H Tab Hydrochlorothiazide 100 ARISTO

39 TEMAX Tab 20 WOCKHARDT

40 TEMAX Tab 40 WOCKHARDT

41 TETAN Tab 80 ALEMBIC

42 TETAN AV Tab Atorvastatin 50 ALEMBIC

43 TSARTAN Tab 40 OCTANE BIOTECH

44 TSARTAN-H Tab Hydrochlorothiazide 60 OCTANE BIOTECH

45 VASIZITEL-H Tab Hydrochlorothiazide 60 SYNTONIC LIFE

SCIENCES

46 ZITELMI Tab 40 FDC

47 ZITELMI-H tab Hydrochlorothiazide 60 FDC

48 CRESAR TAB 20 CIPLA

246

9.2. LITREATURE SURVEY

Several analytical methods have been reported for the determination of

Telmisartan in pure drug, pharmaceutical dosage forms and in biological samples

using spcetrophotometry liquid chromatography, electro kinetic chromatography high

performance thin layer chromatography either in single or in combined forms.

Sujana K et al [5]

has developed and validated a simple, selective, precise and

stability indicating RP high Performance Liquid Chromatographic (HPCL) method of

an analysis of Telmisartan in pure and Pharmaceutical dosage form. The

chromatographic conditions comprised of a reversed phase C8 column (4.6 x 150mm,

3.5m, Make: XTerra), with a mobile phase Composed of Buffer and Methanol

(40:60v/v, Adjusted the pH to 3.0 with ortho Phosphoric acid).

Flow rate was 0.5 mL / min. Detection was carried out at 230 nm. The retention time

of Telmisartan was 2.6 min. The linear regression analysis data for the calibratin plots

showed good linear relationship in the concentration range 20-100. The values of

correclation coefficient, slope and intercept were, 0.9998, 2.326 and -6.708,

repectively. The method was successfully validated in accordance to

ICH guidelines acceptance criteria for specificity, linearity, precision, recovery, rugge

dnessand robustness. The drug undergoes degradation under acidic, basic, Peroxide a

nd thermal degradation conditions. All the peaks of degraded product were resol

ved from the active pharmaceutical ingredient with significantly different retent

ion time. As the method could effectively separate the drug from its degradation

product, it can be employed as a stability-indicating one.

R. Vijayamirtharaj et al [6]

has developed a RP-HPLC method for the

determination of Telmisartan and Atorvastatin calcium in bulk and in formulation

using UV detector. Selected mobile phase was a combination of Acetonitrile: Buffer

(0.01M Potassium dihydrogen phosphate) 65:35 PH 4.00 (adjusted with

Orthophosphoric acid) and the wavelength selected was 250nm. The flow rate was

kept at 2.0 ml/min, and the injection volume was 10µl. The separation was performed

at ambient temperature. Retention time of Telmisartan and Atorvastatin calcium was

found to be 3.72 and 6.14 minutes respectively. Linearity of the method was found to

be 319-480 µg/ml for Telmisartan and 86-130 µg/ml for Atorvastatin calcium. The

correlation co-efficient of Telmisartan was found to be 0.9998 and the correlation co-

efficient of Atorvastatin calcium was found to be 0.9999. Accuracy of the method

247

was determined through recovery studies by adding known quantities of standard drug

to the pre analyzed test solution and was found to be 98.92-100.02 for Telmisartan

and 99.93-100.96 for Atorvastatin calcium respectively The system suitability

parameters such as theoretical plates and tailing factor were found to be 6347, 1.652

and 9720, 1.394 respectively for Telmisartan and Atorvastatin calcium. This method

was validated according to ICH guidelines.

V.A. Patel et al [7]

has developed and validated thin layer liquid

chromatography (TLC) method for the simultaneous estimation of Telmisartan and

ramipril in a combined dosage form. Procedure does not require prior separation of

components from the sample. Telmisartan and Ramipril were determined by High

Performance Thin Layer chromatography method (HPTLC) in tablet dosage form.

The method was carried out in TLC Precoated silica gel on aluminum plate 60 F 254,

(10 cm ×10 cm, prewashed by methanol and activated at 60° C for 5 min prior to

chromatography). The solvent system was Acetone: Benzene: Ethyl acetate: Glacial

acetic acid in the proportion of 5:3:2:0.03, (v/v/v/v) with R Value for Telmisartan and

ramipril was 0.673 and 0.353 respectively. The linearity regression analysis for

calibration showed 0.996 and 0.998 for Telmisartan and ramipril with respect to peak

area and height in the concentration range of 100- 1800 ng/spot and 300-1800 ng/spot

respectively. The method developed can be used for routine analysis of drugs content

in tablet dosage form.

V. P. Kurade et al [8]

described a rapid high performance liquid

chromatographic method has for the estimation of ramipril and Telmisartan

simultaneously in combined dosage form. A Genesis C18 column having dimensions

of 4.6×250 mm and particle size of 5μm in isocratic mode, with mobile phase

containing a mixture of 0.01 M potassium dihydrogen phosphate buffer (adjusted to

pH 3.4 using orthophosphoric acid): methanol: acetonitrile (15:15:70 v/v/v) was used.

The mobile phase was pumped at a flow rate of 1.0 ml/min and the eluents were

monitored at 210 nm. The selected chromatographic conditions were found to

effectively separate ramipril (Rt: 3.68 min) and Telmisartan (Rt: 4.98 min) having a

resolution of 3.84. The method was validated in terms of linearity, accuracy,

precision, and specificity, limit of detection and limit of quantitation. Linearity for

ramipril and Telmisartan were found in the range of 3.5-6.5 μg/ml and 28.0-52.0

μg/ml, respectively. The percentage recoveries for ramipril and Telmisartan ranged

248

from 99.09-101.64% and 99.45-100.99%, respectively. The limit of detection and the

limit of quantitation for ramipril were found to be 0.5μg/ml and 1.5 μg/ml

respectively and for Telmisartan were found to be 1.5 μg/ml and 3.0 μg/ml,

respectively. The method was found to be robust and can be successfully used to

determine the drug content of marketed formulations.

SB Wankhede et al [9]

has described a validated reverse phase high

performance liquid chromatographic method for simultaneous estimation of

Telmisartan and hydrochlorothiazide in tablet formulation. Chromatography was

performed on a ODS Hypersil C18 (25 cm×4.6 mm I.D) column from thermo in

isocratic mode with mobile phase containing acetonitrile: 0.05 M KH2PO4 pH 3.0

(60:40). The flow rate was 1.0 ml/min and the eluent was monitored at 271 nm. The

selected chromatographic conditions were found to effectively separate Telmisartan

(RT- 5.19 min) and hydrochlorothiazide (RT- 2.97 min). Linearity for Telmisartan

and hydrochlorothiazide were found in the range of 4.1-20.48 µg/ml and 1.28-6.4

µg/ml, respectively. The proposed method was found to be accurate, precise,

reproducible and specific and can be used for simultaneous analysis of these drugs in

tablet formulation.

A. Gupta et al [10]

has developed and validated a simple, rapid, precise,

sensitive and reproducible reverse phase high performance liquid chromatographic

(RP-HPLC) method for determination of Telmisartan (TELM) in tablet dosage form.

Chromatographic separation was achieved on a 250 × 4.6 mm, 5μ, Waters symmetry

column in gradient mode, with mobile phase consisting of a mixture of solution (10

mM potassium dihydrogen phosphate, pH 3.5 ± 0.01): acetonitrile (64:40) was used.

The quantitation performed at flow rate of 1.0 mL/min at 230 nm and run time was 12

min. The analytical method was validated as per ICH guideline for linearity, accuracy,

precision, specificity, limit of detection, limit of quantification, robustness and

stability and method can be extended to the analysis of TELM in tablet formulations.

The relative standard deviation values for precision was less than 2%, and % recovery

was greater than 98% for TELM. The drug undergoes oxidative degradation, thermal

degradation and in alkali medium.

249

Pengfei Li et al [11]

has developed a rapid, selective and sensitive method for

the determination of the angiotensin II receptor antagonist, Telmisartan, in human

plasma. Telmisartan and the internal standard, diphenhydramine, were extracted from

plasma using diethyl ether–dichloromethane (60:40, v/v), and separated on a Zorbax

extend C18 column using methanol–10 mM ammonium acetate (85:15, v/v) adjusted

to pH 4.5 after mixing with formic acid as mobile phase. Detection was carried out by

multiple reactions monitoring on a Q-trap™ LC–MS/MS system with an ESI

interface. The assay was linear over the range 0.5–600.0 ng/ml with a limit of

quantitation of 0.5 ng/ml and a limit of detection of 0.05 ng/ml. Intra- and inter-day

precision were <6.7% and <8.1%, respectively, and the accuracy was in the range

88.9–111.0%. The assay was applied to a pharmacokinetic study of Telmisartan given

as a single oral dose (80 mg) to healthy volunteers.

Zhang H et al [12]

has been developed a rapid HPLC method using a

monolithic column with fluorescence detection has been for determination of

Telmisartan in human plasma. Sample preparation was done by protein precipitation

with acetonitrile and naproxen was used as IS. The compounds were detected by

fluorescence detection, using an excitation wavelength of 300 nm and emission

wavelength of 385 nm. Calibration curves of Telmisartan were linear in the range of

1-200 ng/mL. The assay was high throughput, sensitive and precise, and it was

successfully applied to a bioequivalence study of two formulations of Telmisartan.

S. V. Londhe et al [13]

developed a sensitive and reproducible HPLC method

for quantitative analysis of Telmisartan. The drug was separated from its degradation

products on a C 18 column at ambient temperature with methanol-water 80:20 (v / v ),

pH 4.0 (adjusted by addition of orthophosphoric acid), as mobile phase at a flow rate

of 1.0 mL min −1

. Under these conditions the retention time of Telmisartan was 4.85

± 0.05 min. Quantification on the basis of peak area was achieved by UV detection at

225 nm; calibration plots were linear in the concentration range 10–60 μg mL −1

.

When the method was applied to a pharmaceutical formulation there was no

chromatographic interference from tablet excipients. The method was validated for

precision, robustness, recovery, and limits of detection and quantification. The drug

was subjected to acidic and alkaline hydrolysis, and oxidising, dry heat, wet heat, and

250

photodegrading conditions. Because the method could effectively separate the drug

from its degradation products, it can be regarded as stability indicating.

L. R. Bhat et al [14]

has been developed a simple, selective, and precise

reverse phase high performance liquid chromatographic method for the simultaneous

determination of Telmisartan and hydrochlorothiazide from pharmaceutical

formulation. The mobile phase consisted of methanol and acetonitrile (70: 30 v/v) at a

flow rate of 1 mL/min and the wavelength of detection was 270 nm. Rabeprazole was

used as an internal standard. The retention times of Telmisartan, hydrochlorothiazide

and rabeprazole were 1.79 ± 0.01, 2.80 ± 0.01, and 3.19 ± 0.01 minutes, respectively.

The developed method was validated according to ICH guidelines. The proposed

method can be used for determination of these drugs in combined dosage forms.

251

9.3. EXPERIMENTAL

9.3.1. Instrumentation

Peak HPLC containing LC 20AT pump and variable wavelength

programmable UV-Visible detector and Rheodyne injector was employed for

investigation. The chromatographic analysis was performed on a Chromosil C18

column (250 mm × 4.6 mm, 5µm). Degassing of the mobile phase was done using a

Loba ultrasonic bath sonicator. A Denwar Analytical balance was used for weighing

the materials.

9.3.2. Chemicals and Solvents

The reference sample of Telmisartan (API) was obtained from Cipla, Mumbai.

The Formulation CRESAR (Telmisartan) was procured from the local market.

Methanol, Acetonitrile used was of HPLC grade and purchased from Merck

Specialities Private Limited, Mumbai, India. And orthophosphoric acid used was AR

grade purchased from local market.

9.3.3. The mobile phase

A mixture of Methanol: 0.1% orthophosphoric acid :Acetonitrile in the ratio of

40: 50:10 v/v/v was prepared and used as mobile phase.

9.3.4. The buffer solution

About 1.0 mL of orthophosphoric acid was diluted to 1000 mL with water and

filtered through 0.45μ nylon filter.

9.3.5. Standard solution of the drug

For analysis 100 ppm standard solution was prepared, required concentrations

were obtained from 100 ppm solution by appropriate dilution.

9.3.6. Sample (tablet) solution

The formulation tablets of Telmisartan (CRESAR - 20 mg) were crushed to

give finely powdered material. From the Powder prepared a 12 ppm solution in

mobile phase and then filtered through Ultipor N66 Nylon 6, 6 membrane sample filter

paper.

252

9.4. METHOD DEVELOPMENT

For developing the method (as described in Chapter 1 and 2), a systematic

study of the effect of various factors was undertaken by varying one parameter at a

time and keeping all other conditions constant. Method development consists of

selecting the appropriate wave length and choice of stationary and mobile phases. The

following studies were conducted for this purpose.

9.4.1. Detection wavelength

The spectrum of 10ppm solution of the Telmisartan in methanol was recorded

separately on UV spectrophotometer. The peak of maximum absorbance wavelength

was observed. The spectra of Telmisartan were showed maximum absorbance at

256nm.

9.4.2. Choice of stationary phase

Preliminary development trials have performed with octadecyl columns with

different types, configurations and from different manufacturers. Finally the expected

separation and peak shapes were obtained on Chromosil C18 (250 mm x 4.6 mm,

5μm) column.

9.4.3. Selection of the mobile phase

In order to get sharp peak, low tailing factor and base line separation of the

separation of the components, a number of experiments were carried out by varying

the composition of various solvents and flow rate. To have an ideal separation of the

drug under isocratic conditions, mixtures of solvents like methanol, water and

Acetonitrile with or without different buffers indifferent combinations were tested as

mobile phases on a Chromosil C18 column. A mixture of Methanol:0.1%

orthophosphoric acid :Acetonitrile in the ratio of 80:05:15 v/v/v was proved to be the

most suitable of all the combinations since the chromatographic peak obtained was

better defined and resolved and almost free from tailing.

253

9.4.4. Flow rate

Flow rates of the mobile phase were changed from 0.5 – 1.5 mL/min for

optimum separation. A minimum flow rate as well as minimum run time gives the

maximum saving on the usage of solvents. It was found from the experiments that 1.5

mL/min flow rate was ideal for the successful elution of the analyte.

9.4.5. Optimized chromatographic conditions

Chromatographic conditions as optimized above were shown in Table.9.2.

These optimized conditions were followed for the determination of Telmisartan in

bulk samples and in its Formulations. The chromatogram of standard (4ppm) shown

in Figure 9.B.

Mobile phase MEOH: 0.1 % OPA: ACN: 80:05:15 v/v/v

Pump mode Isocratic

Mobile phase PH

5.8

Diluent

Mobile phase

Column

chromosil C18 column (250 mm x 4.6 mm, 5μ)

Column Temp

Ambient

Wavelength

256 nm

Injection Volume

20 μl

Flow rate 1.5 mL/min

Run time 6 min

Retention Time 2.7 min

Table 9.2: Optimized chromatographic conditions for estimation of Telmisartan

254

Figure 9.B: Chromatogram of standard solution

255

9.5. VALIDATION OF THE PROPOSED METHOD

The proposed method was validated as per ICH guidelines (as described in

Chapter 1 and 2). The parameters studied for validation were specificity, linearity,

precision, accuracy, robustness, system suitability, limit of detection, limit of

quantification, and solution stability.

9.5.1. Specificity

The specificity of method was performed by comparing the chromatograms of

blank, standard and sample (Prepared from Formulation). It was found that there is no

interference due to excipients in the tablet formulation and also found good

correlation between the retention times of standard and sample. The specificity results

are shown in Table 9.3.

NAME OF THE SOLUTION Retention Time in Min

Blank NO PEAKS

Telmisartan 2.790

Table 9.3: Specificity study

9.5.2 Linearity

Linearity was performed by preparing mixed standard solutions of Telmisartan

at different concentration levels including working concentration mentioned in

experimental condition i.e. 12 ppm. Twenty micro liters of each concentration was

injected in duplicate into the HPLC system. The response was read at 256 nm and the

corresponding chromatograms were recorded. From these chromatograms, the mean

peak areas were calculated and linearity plots of concentration over the mean peak

areas were constructed individually. The regressions of the plots were computed by

least square regression method. Linearity results were presented in Table 9.4.

256

Table 9.4: Linearity results

Figure 9.C: On X axis concentration of sample, On Y axis peak area response

Level Concentration of Telmisartan in ppm Mean peak

area

Level -1 2 85426

Level -2 4 160991.9

Level -3 6 247225.6

Level -4 8 353178.8

Level -5 10 437519.9

Level -6 12 534746.3

Range: 2-12ppm

Slope

Intercept

Correlation coefficient

45459.12

-150323.5

0.999

-100000

0

100000

200000

300000

400000

500000

600000

0 5 10 15

Series1

Linear (Series1)

257

9.5.3. Precision

Precision is the degree of repeatability of an analytical method under normal

Operational conditions. Precision of the method was performed as intraday precision,

Inter day precision.

9.5.3.1. Intraday precision

To study the intraday precision, six replicate standard solutions (12ppm) of

Telmisartan were injected. The percent relative standard deviation (% RSD) was

calculated and it was found to be 0.039, which are well within the acceptable criteria

of not more than 2.0. Results of system precision studies are shown in Table 9.5.

SAMPLE

CONC(PPM) INJECTION

No.

PEAKS

AREA

R.S.D

(Acceptance

criteria ≤

2.0%)

Telmisartan

12

1 588925.5

0.039

2 588289.1

3 588470.8

4 588815.8

5 588548.2

6 588557.4

Table 9.5: System Precision (Intra Day)

9.5.3.2. Inter Day precision

To study the interday precision, six replicate standard solution of Telmisartan

was injected on third day of sample preparation. The percent relative standard

deviation (% RSD) was calculated and it was found to be 0.373, which are well within

the acceptable criteria of not more than 2.0. Results of system precision studies are

shown in Table 9.6.

258

SAMPLE

CONC(PPM) INJECTION

No.

PEAKS

AREA

R.S.D

(Acceptance

criteria ≤ 2.0%)

Telmisartan

12

1 586338.7

0.123

2 585740.8

3 582157.1

4 582307.0

5 581967.1

6 586231.3

Table 9.6: System Precision (Inter Day)

9.5.4. Accuracy

The accuracy of the method was determined by standard addition method. A

known amount of standard drug was added to the fixed amount of pre-analyzed tablet

solution. Percent recovery was calculated by comparing the area before and after the

addition of the standard drug. The standard addition method was performed at 50%,

100% and 150% level of 12ppm. The solutions were analyzed in triplicate at each

level as per the proposed method. The percent recovery and % RSD was calculated

and results are presented in Table 9.7. Satisfactory recoveries ranging from 99.0 to

102.0 were obtained by the proposed method. This indicates that the proposed method

was accurate.

259

Level Amount of

Telmisartan

spiked (ppm)

Amount of

Telmisartan

recovered(ppm)

% Recovery

%RSD

50 %

6

6

6

5.98

5.95

5.95

99.66

99.16

99.16

0.289

100%

8

8

8

7.98

7.94

7.93

99.75

99.75

99.12

0.370

150%

10

10

10

9.98

9.91

9.92

99.8

99.1

99.2

0.380

Mean % of

recovery 99.41

Mean

RSD =

0.346

Table 9.7: Percentage Recovery and % RSD

9.5.5. Robustness

The robustness study was performed by slight modification in flow rate of

Mobile phase, pH of the buffer and composition of the mobile phase. Telmisartan at 6

ppm concentration was analyzed under these changed experimental conditions. It was

observed that there were no marked changes in chromatograms, which demonstrated

that the developed method was robust in nature. The results of robustness study are

shown in Table 9.8.

260

Condition Mean area % assay % difference

Unaltered 590802.8 100.0 0.0

Flow rate at 1.4 mL/min

Flow rate at 1.6mL/min

585143.0

586252.8

99.04

99.22

0.96

0.72

Mobile phase:

MEOH: ACN: 0.1 % OPA

75% 20% 05%

85% 10 % 05%

589314.9

596219.5

99.74

100.9

0.26

0.10

pH of mobile phase at 5.6 592326.2 100.25 0.25

pH of mobile phase at 6.0 593572.1 100.4 0.4

Table 9.8: Robustness

9.5.6. System suitability

System suitability was studied under each validation parameters by injecting

six replicates of the standard solution 2 ppm). The results obtained were within

acceptable limits (Tailing factor ≤2 and Theoretical plate’s ≥2000) and are

represented in Table 9.9. Thus, the system meets suitable criteria.

Parameter Tailing factor Theoretical plates

Specificity study 1.71 6638

Linearity study 1.20 7694.97

Precision study 1.71 6705.46

Table 9.9: System Suitability

261

9.5.7. Limit of detection and Limit of quantification

Limit of detection (LOD) is defined as the lowest concentration of analyte that

gives a detectable response. Limit of quantification (LOQ) is defined as the lowest

Concentration that can be quantified reliably with a specified level of accuracy and

Precision. For this sample was dissolved by using Mobile Phase and injected until

peak was disappeared. After 15ng/ml dilution, Peak was not clearly observed. So it

confirms that 15ng is limit of Detection and 50ng dilution is Limit of Quantification.

For this study six replicates of the analyte at lowest concentration were Measured and

quantified. The LOD and LOQ of Telmisartan are given in Table 9.10.

Table 9.10: LOQ and LOD

Formulation:

The proposed method has been applied to the assay of commercial

tablets containing Telmisartan. Sample was analyzed for five times after

extracting the drug as mentioned in assay sample preparation of the

experimental section. After analysis test result assay of Telmisartan in Tablet

is18.57% and is very close to the labeled amount.

Parameter Measured volume

Limit of Quantification 50ng

Limit of Detection 15ng

262

9.6. DISCUSSION ON THE RESULT

To optimize the RP-HPLC parameters, several mobile phase

compositions were tried. A satisfactory separation and good peak symmetry was

found in a mixture of Acetonitril: Methanol: Acetonitrile: 0.1%OPA in the ratio of

80:15:05 v/v and 1.5 mL/min flow rate proved to be better than the other mixtures in

terms of resolution and peak shape. The optimum wavelength for detection was set at

256nm at which much better detector responses for drug was obtained. As it was

shown in Figure 9.B the retention times were 2.7 min for Telmisartan. The number of

theoretical plates was found to be 6638.20, which indicates efficient performance of

the column. A system suitability test was applied to representative chromatograms for

various parameters. The results obtained were within acceptable limits and are

represented in Table 9.9. Thus, the system meets suitable criteria.

The calibration curve was obtained for a series of concentration in the

range of 2-12µg/ml and it was found to be linear. Seven points graphs was

constructed covering a concentration range 2-12µg/ml. The standard deviation of the

slope and intercept were low. The data of regression analysis of the calibration curves

are shown in Table. Calibration curve found to be linear with r2=0.999, Intercept (-

150323.5) and Slope (45459.12) respectively. The results obtained were within

acceptable limits where capacity factor >2.0, tailing factor ≤2.0 and theoretical plates

>2000 13. In all cases, the relative standard deviation (R.S.D) for the analytic peak

area for two consecutive injections was < 2.0%. The data of regression analysis of the

calibration curve was shown in Table 9.4.

Precision was evaluated by carrying out six independent sample preparation

of a single lot of formulation. Low values of standard deviation denoted very good

repeatability of the measurement. Thus it was showing that the equipment used for the

study was correct and hence the developed analytical method is highly repetitive.

RSD of intraday precision was found to 0.039. For the interday precision a study

carried out on consecutive days indicated a RSD of 0.373. This indicates good method

precision. Results are shown in Table 9.5, 9.6.

Standard addition method at 50%, 100% and 150% to the proposed

HPLC method is carried out to find the Accuracy of the Telmisartan. The results

showed good recoveries ranging from 99.00 to 101.45%. The mean recovery data

263

obtained for each level as well as for all levels combined (Table 9.7) were within

2.0% of the label claim for the active substance with an R.S.D. < 2.0%, which

satisfied the acceptance criteria set for the study.

The proposed method has been applied to the assay of commercial

tablets containing Telmisartan. Sample was analyzed for five times after extracting

the drug as mentioned in assay sample preparation of the experimental section. The

results (18.57%) presented good agreement with the labeled content. Low values of

standard deviation denoted very good repeatability of the measurement.

The statistical evaluation of the proposed method was revealed its

good linearity, reproducibility and its validation for different parameters and let us to

the conclusion that it could be used for the rapid and reliable determination of

Telmisartan in tablet formulation. All these factors lead to the conclusion that the

proposed method is accurate, precise, simple, sensitive and rapid and can be applied

successfully for the estimation of Telmisartan in bulk and in pharmaceutical

formulations without interference and with good sensitivity.

264

9.7. BIBILOGRAPHY

1. Pritor prescribing information.

2. Benson, S. C.; Pershadsingh, H.; Ho, C.; Chittiboyina, A.; Desai, P.; Pravenec,

M.; Qi, N.; Wang, J. et al; "Identification of Telmisartan as a Unique

Angiotensin II Receptor Antagonist with Selective PPAR -Modulating

Activity". Hypertension 2004, 43 (5): 993.

3. Drugs.com: Micardis.

4. Ontarget, I.; Yusuf, S.; Teo, K.; Pogue, J.; Dyal, L.; Copland, I.; Schumacher,

H; Dagenais, G. et al; "Telmisartan, Ramipril, or Both in Patients at High Risk

for Vascular Events"; New England Journal of Medicine; 2008, 358 (15):

1547.

5. ―Stability indicating rp­hplc method for the determination of Telmisartan in

pure and pharmaceutical formulation‖; International Journal of Pharmacy and

Pharmaceutical Sciences; 2011; 3(2).

6. ―Development and validation of RP-HPLC method for the simultaneous

estimation of Telmisartan and atorvastatin calcium in tablet dosage forms‖;

Pharmacie Globale (IJCP); 2010, 4 (03).

7. V.A. Patel et al; International Journal on Pharmaceutical and Biological

Research; 2010; 1(1): 18-24.

8. ―RP-HPLC Estimation of Ramipril and Telmisartan in Tablets‖; Indian J

Pharm Sci; 2009; 71(2): 148–151.

9. ―RP-HPLC method for simultaneous estimation of Telmisartan and

hydrochlorothiazide in tablet dosage form‖; Indian Journal of Pharmaceutical

science; 2007; 69(2): 298-300.

10. ―Determination of Telmisartan and forced degradation behavior by RP-HPLC

in tablet dosage form‖; Journal of Pharmacy Research; 2011; 4(4): 1270-1273.

11. ―Determination of Telmisartan in human plasma by liquid chromatography–

tandem mass spectrometry‖; Journal of Chromatography B; 2005; 828(1-2),

15, 126-129.

12. ―Rapid determination of Telmisartan in human plasma by HPLC using a

monolithic column with fluorescence detection and its application to a

bioequivalence study‖; 2009; 877(29): 3729-33.

265

13. ―Stability-indicating RP-HPLC method for analysis of Telmisartan in the bulk

drug and in formulations‖; Acta Chromatographica,; 2010; 22(4): 539-548.

14. ―Validated RP-HPLC method for simultaneous determination of Telmisartan

and hydrochlorothiazide in pharmaceutical formulation‖; J. Liq. Chromatogr.

Relat. Technol; 2007; 30(17-20): 3059-3067.

15. ―Q2A: Text on Validation of Analytical Procedures‖; International Conference

on Harmonization; Federal Register; 1995; 60(40): 11260–11262.

16. ―Q2B: Validation of Analytical Procedures: Methodology; Availability‖;

International Conference on Harmonization; Federal Register; 1997; 62(96):

27463–27467.

17. "Analytical Procedures and Methods Validation: Chemistry, Manufacturing

and Controls Documentation; Availability"; FDA; Federal Register (Notices);

2000; 65(169): 52776 – 52777.

18. www.fda.gov/cder/guidance/cmc3.pdf.

19. USP 25–NF 20; ―Validation of Compendial Methods Section (1225) (United

States Pharmacopeal Convention, Rockville, Maryland, USA, 2002)‖; 2256.

20. G.A. Shabir; ―Validation of HPLC Chromatography Methods for

Pharmaceutical Analysis‖; Understanding the Differences and Similarities

between Validation Requirements of FDA, the US Pharmacopeia and the ICH;

J. Chromatogr. A; 2003; 987(1-2): 57-66.

21. C.E. Wood; "Medicare Program; Changes to the Hospital Outpatient

Prospective"; Med. J. Aust; 1996; 165: 510–514.

22. A. Prentice; "Medical Management of Menorrhagia": Br. Med. J; 1999; 319,

1343–1345.

23. D.T. Baired and A.F. Glasier; "Hormonal Contraception"; New Engl. J. Med;

1993; 328: 1543–1549.

24. P.E. Belchetz; "Hormonal Treatment of Postmenopausal Women"; New Engl.

J. Med; 1994; 330: 1062–1071.

25. International Conference on Harmonization (ICH) of Technical Requirements

for the Registration of Pharmaceuticals for Human Use; ―Validation of

analytical procedures: definitions and terminology‖; Geneva (1996).

266

26. U.S. FDA; Title 21 of the U.S. Code of Federal Regulations:

21 CFR 211—Current good manufacturing practice for finished

pharmaceuticals.

27. U.S. FDA - Guidance for Industry (draft) Analytical Procedures and Methods

Validation: Chemistry, Manufacturing, and Controls and Documentation,

2000.

28. ISO/IEC 17025, General requirements for the competence of testing and

calibration laboratories, 2005.

29. International Conference on Harmonization (ICH) of Technical Requirements

for the Registration of Pharmaceuticals for Human Use, Validation of

analytical procedures: Methodology, adopted in 1996, Geneva.

30. U.S. EPA, Guidance for methods development and methods validation for the

Resource Conservation and Recovery Act (RCRA) Program, Washington,

D.C. (1995).http://www.epa.gov/sw-846/pdfs/methdev.pdf.

31. General Chapter 1225, Validation of compendial methods, United States

Pharmacopeia 30, National Formulary 25, Rockville, Md., USA, The United

States Pharmacopeial Convention, Inc., (2007).

32. U.S. FDA - Guidance for Industry, Bioanalytical Method Validatio.n

33. G. C. Hokanson, A life cycle approach to the validation of analytical methods

during pharmaceutical product development, Part I: The initial validation

process, Pharm. Tech., Sept. 1994, pp. 118–130.