validated hplc and hptlc method for simultaneous quantitation of amlodipine besylate and olmesartan...

9
This article was downloaded by: [Purdue University] On: 08 June 2013, At: 02:57 Publisher: Taylor & Francis Informa Ltd Registered in England and Wales Registered Number: 1072954 Registered office: Mortimer House, 37-41 Mortimer Street, London W1T 3JH, UK Analytical Letters Publication details, including instructions for authors and subscription information: http://www.tandfonline.com/loi/lanl20 Validated HPLC and HPTLC Method for Simultaneous Quantitation of Amlodipine Besylate and Olmesartan Medoxomil in Bulk Drug and Formulation Asmita Y. Kamble a , Mahadeo V. Mahadik a , Laxman D. Khatal a & Sunil R. Dhaneshwar a a Department of Pharmaceutical Chemistry, Bharati Vidyapeeth University, Poona College of Pharmacy, Pune, India Published online: 13 Jan 2010. To cite this article: Asmita Y. Kamble , Mahadeo V. Mahadik , Laxman D. Khatal & Sunil R. Dhaneshwar (2010): Validated HPLC and HPTLC Method for Simultaneous Quantitation of Amlodipine Besylate and Olmesartan Medoxomil in Bulk Drug and Formulation, Analytical Letters, 43:2, 251-258 To link to this article: http://dx.doi.org/10.1080/00032710903325906 PLEASE SCROLL DOWN FOR ARTICLE Full terms and conditions of use: http://www.tandfonline.com/page/terms-and-conditions This article may be used for research, teaching, and private study purposes. Any substantial or systematic reproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in any form to anyone is expressly forbidden. The publisher does not give any warranty express or implied or make any representation that the contents will be complete or accurate or up to date. The accuracy of any instructions, formulae, and drug doses should be independently verified with primary sources. The publisher shall not be liable for any loss, actions, claims, proceedings, demand, or costs or damages whatsoever or howsoever caused arising directly or indirectly in connection with or arising out of the use of this material.

Upload: sunil-r

Post on 14-Dec-2016

215 views

Category:

Documents


3 download

TRANSCRIPT

This article was downloaded by: [Purdue University]On: 08 June 2013, At: 02:57Publisher: Taylor & FrancisInforma Ltd Registered in England and Wales Registered Number: 1072954 Registeredoffice: Mortimer House, 37-41 Mortimer Street, London W1T 3JH, UK

Analytical LettersPublication details, including instructions for authors andsubscription information:http://www.tandfonline.com/loi/lanl20

Validated HPLC and HPTLC Method forSimultaneous Quantitation of AmlodipineBesylate and Olmesartan Medoxomil inBulk Drug and FormulationAsmita Y. Kamble a , Mahadeo V. Mahadik a , Laxman D. Khatal a &Sunil R. Dhaneshwar aa Department of Pharmaceutical Chemistry, Bharati VidyapeethUniversity, Poona College of Pharmacy, Pune, IndiaPublished online: 13 Jan 2010.

To cite this article: Asmita Y. Kamble , Mahadeo V. Mahadik , Laxman D. Khatal & Sunil R.Dhaneshwar (2010): Validated HPLC and HPTLC Method for Simultaneous Quantitation of AmlodipineBesylate and Olmesartan Medoxomil in Bulk Drug and Formulation, Analytical Letters, 43:2, 251-258

To link to this article: http://dx.doi.org/10.1080/00032710903325906

PLEASE SCROLL DOWN FOR ARTICLE

Full terms and conditions of use: http://www.tandfonline.com/page/terms-and-conditions

This article may be used for research, teaching, and private study purposes. Anysubstantial or systematic reproduction, redistribution, reselling, loan, sub-licensing,systematic supply, or distribution in any form to anyone is expressly forbidden.

The publisher does not give any warranty express or implied or make any representationthat the contents will be complete or accurate or up to date. The accuracy of anyinstructions, formulae, and drug doses should be independently verified with primarysources. The publisher shall not be liable for any loss, actions, claims, proceedings,demand, or costs or damages whatsoever or howsoever caused arising directly orindirectly in connection with or arising out of the use of this material.

Chromatography

VALIDATED HPLC AND HPTLC METHOD FORSIMULTANEOUS QUANTITATION OF AMLODIPINEBESYLATE AND OLMESARTAN MEDOXOMIL IN BULKDRUG AND FORMULATION

Asmita Y. Kamble, Mahadeo V. Mahadik, Laxman D. Khatal,and Sunil R. DhaneshwarDepartment of Pharmaceutical Chemistry, Bharati Vidyapeeth University,Poona College of Pharmacy, Pune, India

Two methods are described for simultaneous determination of amlodipine besylate and

olmesartan medoxomil in formulation. The first method was based on the HPTLC

separation of two drugs on Merck HPTLC aluminium sheets of silica gel 60 F254 using

n-butanol: acetic acid: water (5:1:0.1, v/v/v) as the mobile phase. The second method

was based on the HPLC separation of the two drugs on the RP-PerfectSil-100 ODS-

3–C18 column from MZ-Analysetechnik GmbH, Germany and acetonitrile/0.03M

ammonium acetate buffer (pH¼ 3) in a ratio of 55:45 as the mobile phase. Both methods

have been applied to formulation without interference of excipients of formulation.

Keywords: Amlodipine besylate; HPLC; HPTLC; Olmesartan medoxomil; Validation

INTRODUCTION

Amlodipine besylate (AML), chemically 3-ethyl-5-methyl (4R, S)-2-[(2-aminoethoxy) methyl]-4-(2-chlorophenyl)-6-methyl 1,4-dihydropyridine 3,5-dicarboxylate-benzene sulphonate, is shown in Fig. 1a. It is a calcium channelblocker used as an anti-hypertensive and in the treatment of angina. As othercalcium channel blockers, amlodipine acts by relaxing the smooth muscle in thearterial wall, decreasing peripheral resistance and hence reducing blood pressure; inangina it increases blood flow to the heart muscle. Olmesartan medoxomil (OLM),

Received 24 January 2009; accepted 12 August 2009.

The authors would like to thank Aristo Pharmaceuticals Ltd. (Bhopal, India) for providing a gift

sample of standard amlodepine besylate and olmesartan. The authors would like to thank Dr. K. R.

Mahadik, Principal, Poona College of Pharmacy, Pune, India for providing the necessary facilities to carry

out the work. The authors would like to thank AICTE for providing financial support for carrying out

research work.

Address correspondence to Sunil R. Dhaneshwar, Department of Pharmaceutical Chemistry,

Bharati Vidyapeeth University, Poona College of Pharmacy, Pune, Maharashtra 411038, India. E-mail:

[email protected]

Analytical Letters, 43: 251–258, 2010

Copyright # Taylor & Francis Group, LLC

ISSN: 0003-2719 print=1532-236X online

DOI: 10.1080/00032710903325906

251

Dow

nloa

ded

by [

Purd

ue U

nive

rsity

] at

02:

57 0

8 Ju

ne 2

013

chemically 4-(1-Hydroxy-1-methylethyl)-2-propyl-1-[20-(1H-5-tetrazolyl)biphenyl-4-ylmethyl]imidazole-5-carboxylic acid 5-methyl-2-oxo-1, 3-dioxol-4-ylmethyl ester,(Fig. 1b) is used as antihypertensive agents (Kamat and Chaturvedi 2005).

Literature review reveals that methods have been reported for analysis of AML(Amlodipine) and OLM (Olmesartan) by LC-MS in human plasma (Streel et al.2002; Vaidya et al. 2008), stability indicating HPLC method for amlodipine in tablet(Kamat and Chaturvedi 2005), RP-HPLC method for determination of amlodipinein combination with other drugs (Naidu, Kale, and Shingare 2005; Chaudhari, Patel,and Shah 2007), RP-HPLC method for determination of OLM in combination withother drugs (Sagirli et al. 2007; Chintan et al. 2007; Shah et al. 2007), and HPTLCmethod for quantitation of AML in combination with other drugs (Mahadik,Aggrawal, and Kaul 2004; Meyyanathan and Suresh 2005).

To date, there have been no published reports about the simultaneous quanti-tation of AML and OLM by HPLC and HPTLC in bulk drug and in tablet dosageform. This present study reports for the first time the simultaneous quantitation ofAML and OLM by HPLC and HPTLC in bulk drug and in tablet dosage form.The proposed method is validated as per ICH guidelines.

Figure 1. (a) Structure of amlodipine besylate (b) structure of olmesartan medoxomil.

252 A. Y. KAMBLE ET AL.

Dow

nloa

ded

by [

Purd

ue U

nive

rsity

] at

02:

57 0

8 Ju

ne 2

013

EXPERIMENTAL

Materials

Aristo Pharmaceuticals Ltd, Bhopal, India, kindly supplied a pure drug sampleof AML as a gift sample of Batch No.: PC-76DFG and OLM of Batch No.OM12DRT. It was used without further purification and certified to contain98.55% (w=w) on dried basis. All chemicals and reagents used were of HPLC gradeand were purchased from Merck Chemicals, India.

Instrumentation

The HPLC system consisted of a Pump (model Jasco PU 2080), Intelligent LCpump with sampler programmed at 20 mL capacity per injection was used. Thedetector consisted of UV=VIS (Jasco UV 2075) model operated at a wavelength of254 nm. Data was integrated using Jasco Borwin version 1.5, LC-Net II=ADCsystem. The column used was PerfectSil-100 ODS-3–C18(250mm� 4.6mm, 5.0 m)from MZ-Analysetechnik GmbH, Germany.

The samples were spotted in the form of bands 6mm width with a Camag 100microlitre sample (Hamilton, Bonaduz, Switzerland) syringe on silica gel precoatedaluminum plate 60 F254, (20� 10 cm) with 250 mm thickness; E. Merck, Darmstadt,Germany, supplied by Anchrom Technologists, Mumbai) using a Camag LinomatV (Switzerland). The plates were prewashed by methanol and activated at 110�Cfor 5min prior to chromatography. A constant application rate of 0.1 mL=s wasemployed and the space between two bands was 5mm. The slit dimension was keptat 5mm� 0.45mm and 10mm=s scanning speed was employed. The monochroma-tor bandwidth was set at 20 nm, each track was scanned thrice and the baselinecorrection was used. The mobile phase consisted of n-butanol: acetic acid: water(5.0:1.0:0.1, v=v=v), and 10mL of mobile phase was used per chromatography.Linear ascending development was carried out in a 20 cm� 10 cm twin trough glasschamber (Camag, Muttenz, Switzerland) saturated with the mobile phase. The opti-mized chamber saturation time for mobile phase was 30min at room temperature(25�C� 2) at relative humidity of 60%� 5. The length of chromatogram run was8 cm. Subsequent to the development, HPTLC plates were dried in current of airwith the help of air dryer in a wooden chamber with adequate ventilation. The flowrate in the laboratory was maintained unidirectionally (laminar flow, towardsexhaust). Densitometric scanning was performed on a Camag HPTLC scanner IIIin the reflectance-absorbance mode at 254 nm and operated by CATS software (V3.15, Camag). The source of radiation utilized was deuterium lamp emittingcontinuous UV spectrum between 190 and 400 nm. Concentrations of the compoundchromatographed were determined from the intensity of diffuse light. Evaluationwas via peak areas with linear regression.

Preparation of Standard Stock Solutions

Standard stock solutions of a concentration of 1mg=mL of AML and4mg=mL of OLM were prepared separately using methanol. From the standardstock solution, the mixed standard solution was prepared using the methanol to

SIMULTANEOUS METHOD DEVELOPMENT AND VALIDATION 253

Dow

nloa

ded

by [

Purd

ue U

nive

rsity

] at

02:

57 0

8 Ju

ne 2

013

contain 0.1mg=mL of AML and 0.4mg=mL of OLM. The stock solution was storedat 2–8�C, protected from light.

Optimization of HPLC and HPTLC Method

The HPLC and HPTLC procedure was optimized with a view to develop asimultaneous assay method for AML and OLM respectively. The mixed standardstock solution (0.1mg=mL of AML and 0.4mg=mL of OLM) injected in HPLCand spotted on to HPTLC plates and run in different solvent systems.

Optimization of the HPTLC method was very critical in this case as both drugshad opposite polarity. The AML was highly polar and OLM was non-polar.Initially, toluene: ethyl acetate: methanol (5:4:1 v=v=v) was selected but AML dueto polar nature not moved from the application position. Hence to increase thepolarity of the system, toluene volume decreased and ethyl acetate increased, but stillAML did not run. Hence, more polar system n-butanol: acetic acid: water in differ-ent ratios were tried to get the optimum RF for AML and OLM. First, n-butanol:acetic acid: water (5:1:1) was tried; in this mobile phase the AML spot was run(RF 0.5) but the OLM spot was over-run due to high polarity of the system. Then,the water volume was decreased; finally, the mobile phase consisting of n-butanol:acetic acid: water in the ratio 5:1:0.1 v=v=v was found to be optimum (RF 0.30 forAML and RF 0.72 for OLM). In order to reduce the neckless effect, the HPTLCchamber was saturated for 20min using saturation pads. The mobile phase wasrun up to a distance of 8 cm, which takes approximately 20min for complete devel-opment of the HPTLC plate.

For the HPLC method optimization, different ratios of acetonitrile and acetatebuffer at different pH were tried but it was found that acetonitrile: acetate buffer(30mM) in the ratio 55:45 v=v, pH-3 adjusted with glacial acetic acid at flow rate1mL=min gave acceptable retention time (tR 5.8min for AML and tR 7.2min forOLM), plates, and good resolution for AML and OLM.

Validation of the Method

Validation of the optimized HPLC and HPTLC method was carried out withrespect to the following parameters.

Linearity and range. From the mixed standard stock solution (0.1mg=mL ofAML and 0.4mg=mL of OLM), 1 to 10 ml solution was spotted on the HPTLC plateto obtain the final concentration 100–1000 ng=spot for AML and 400–4000 ng=spotfor OLM. Each concentration was applied three times to the HPTLC plate. Theplate was then developed using the previously described mobile phase and the peakareas were plotted against the corresponding concentrations to obtain the calibrationcurves.

The mixed standard stock solution (0.1mg=mL of AML and 0.4mg=mL ofOLM) further diluted to get AML and OLM concentration in the range of10–100 mg=mL and 40–400 mg=mL, respectively. Linearity of the method was studiedby injecting six concentrations of the drug prepared in the mobile phase in triplicate

254 A. Y. KAMBLE ET AL.

Dow

nloa

ded

by [

Purd

ue U

nive

rsity

] at

02:

57 0

8 Ju

ne 2

013

into the LC system keeping the injection volume constant. The peak areas wereplotted against the corresponding concentrations to obtain the calibration graphs.

Precision. The precision of the method was verified by repeatability and inter-mediate precision studies. Repeatability studies were performed by analysis of threedifferent concentrations by HPLC (10, 40, 60 mg=mL for AML and 40, 160,360 mg=mL for OLM) and HPTLC (100, 400, 600 ng=spot for AML and 400,1600, 3600 ng=spot for OLM) of the drug in six times on the same day. The inter-mediate precision of the method was checked by repeating studies on three differentdays.

Limit of detection and limit of quantification. Limits of detection (LOD)and quantification (LOQ) represent the concentration of the analyte that would yieldsignal-to-noise ratios of 3 for LOD and 10 for LOQ, respectively. The LOD andLOQ were determined by measuring the magnitude of analytical background byspotting a blank and calculating the signal-to-noise ratio for AML and OLM byspotting a series of solutions until the S=N ratio 3 was obtained for the LOD and10 for the LOQ. To determine the LOD and LOQ, serial dilutions of mixed standardsolution of AML and OLM were made from the standard stock solution in the rangeof 1–10 ng=spot. The samples were injected in the LC system and applied to theHPTLC plate and the chromatograms were run and a measured signal from thesamples was compared with those of blank samples.

Robustness of the method.

For HPTLC method. Following the introduction of small changes in themobile phase composition (�0.1mL for each component), the effects on the resultswas examined. Mobile phases having different compositions, e.g., n-butanol: aceticacid: water (5.1:01:0.1 v=v=v), (4.9:01:0.1 v=v=v), (5:1.1:0.1 v= v=v), (5:0.9:0.1 v=v=v),were tried and chromatograms were run. The amount of mobile phase was variedover the range of � 5%. The plates were prewashed with methanol and activatedat 60�C for 2, 5, and 7min, respectively, prior to chromatography. The time fromspotting to chromatography and from chromatography to scanning was varied from�10min. The robustness of the method was determined at three different concen-tration levels 100, 400, and 600 ng=spot and 400, 1600, and 3600 ng=spot for AMLand OLM, respectively.

For HPLC method. To evaluate the robustness of a HPLC method, fewparameters were deliberately varied. The parameters included variation of flow rate,percentage of methanol in the mobile phase, and pH of mobile phase. The resolutionof drug in a mixture of stressed samples was studied by performing the analyses on adifferent chromatographic system. Robustness of the method was done at threedifferent concentration levels 10, 40, 60 mg=mL and 40, 160, 360 mg=mL for AMLand OLM, respectively. Also, robustness was verified by studying the resolution ofthe drug in a mixture of degraded samples on a different chromatographic systemon a different day.

Specificity. The specificity of the method was determined by analyzingstandard drug and test samples. The spot for AML and OLM in the samples wasconfirmed by comparing the RF and spectrum of the spot with that of a standard.

SIMULTANEOUS METHOD DEVELOPMENT AND VALIDATION 255

Dow

nloa

ded

by [

Purd

ue U

nive

rsity

] at

02:

57 0

8 Ju

ne 2

013

The peak purity of AML and OLM was determined by comparing the spectrum atthree different regions of the spot, i.e., peak start (S), peak apex (M) and peak end(E). Effect of excipients of formulation (maize starch, crosspovidone, MCC pH102,dicalcium phosphate, and aerosil and magnesium stearate) studied whether it inter-fered with the assay.

Accuracy. Accuracy of the method was carried out by applying the method tothe drug sample (AML and OLM combination tablet) to which a known amount ofAML and OLM standard powder corresponding to 80, 100, and 120% of label claimhad been added (Standard addition method), mixed, and the powder was extractedand analyzed by running chromatogram in an optimized mobile phase.

Analysis of a marketed formulation. To determine the content of AML ofOLM in a conventional tablet (Brand name: OLMEZEST AM, Label claim: 10mgamlodepine besylate and 40mg olmesartan medoxomil per tablet), twenty tabletswere weighed, their mean weight determined and finely powdered. The weight ofthe tablet triturate equivalent to 10mg of AML and 40mg VAL was transferredinto a 25mL volumetric flask containing 10mL methanol, sonicated for 30min,and diluted to 25mL with methanol. The resulting solution was centrifuged at3000 rpm for 5min and the drug content of the supernatant was determined (0.4and 1.6mg=mL for AML and OLM, respectively). Then 2.5mL of the aforemen-tioned filtered solution was diluted to produce a concentration of 100 and400 mg=mL for AML and OLM, respectively, and 2 mL of this solution (200 and800 ng=spot for AML and OLM, respectively) was applied to a HPTLC plate whichwas developed in an optimized mobile phase. The analysis was repeated in triplicate.The possibility of excipient interference with the analysis was examined.

Results and Discussion

The results of validation studies on the simultaneous estimation methoddeveloped for AML and OLM in the current study involving n-butanol: acetic acid:water (5:1:0.1, v=v=v) as the mobile phase for HPTLC and acetonitrile: 30mMacetate buffer (55:45, v=v) are given as follows.

Linearity. The AML and OLM showed a good correlation coefficient(r2¼ 0.9984 for AML and 0.9979 for OLM) in the given concentration range(100–1000 ng=spot for AML and 400–4000 ng=spot for OLM) by HPTLC andHPLC (r2¼ 0.9992 for AML and 0.9983 for OLM).

Precision. The repeatability and intermediate precision RSD (%) values bythe HPTLC method for AML was found to be in the range of 0.637–1.310 and0.653–1.527, respectively, and the RSD (%) values for OLM were found to be inthe range of 0.119–0.442 and 0.144–0.302, respectively. The repeatability and inter-mediate precision RSD (%) values by the HPLC method for AML was found to bein the range of 0.264–0.826 and 0.416–1.285, respectively, and the RSD (%) valuesfor OLM were found to be in the range of 0.185–1.113 and 0.363–1.314, respectively.The developed method was found to be precise as the RSD values for repeatabilityand intermediate precision studies were <2%, respectively, as recommended by ICHguidelines.

256 A. Y. KAMBLE ET AL.

Dow

nloa

ded

by [

Purd

ue U

nive

rsity

] at

02:

57 0

8 Ju

ne 2

013

LOD and LOQ. Signal-to-noise ratios of 3:1 and 10:1 were obtained for theLOD and LOQ, respectively. The LOD and LOQ were found to be 50 ng=spotand 100 ng=spot for AML and OLM by HPTLC and 5 mg=mL and 10 mg=mL forAML and OLM by HPLC, respectively.

Robustness of the method. The standard deviation peak of the areas wascalculated for each parameter and the % RSD was found to be less than 2% bythe HPLC and HPTLC methods. The low values of the % RSD indicated robustnessof the method.

Specificity. The peak purity of AML and OLM was assessed by comparingtheir respective spectra at the peak start, apex, and peak end positions of the spot,i.e., r (S, M)¼ 0.9983 and r (M, E)¼ 0.9996. A good correlation (r¼ 0.9997) was alsoobtained between the standard and sample spectra of AML and OLM, respectively.Also, excipients from formulation were not interfering with the assay.

Recovery studies. Good recoveries of the AML and OLM in the range from97 to 99% were obtained at various added concentrations by the HPLC and HPTLCmethod.

Analysis of a formulation. Experimental results of the amount of AML andOLM in tablets, expressed as a percentage of label claims were in good agreementwith the label claims thereby suggesting that there is no interference from any ofthe excipients which are normally present in tablets. The drug content was foundto be 97.1% of AML and 98.3% of OLM by HPTLC; 98.4% of AML and 99.3%of OLM by HPLC. Two different lots of AML and OLM combination tablets wereanalyzed using the proposed procedures.

CONCLUSION

Introducing HPTLC into pharmaceutical analysis represents a major step interms of quality assurance. Today, HPTLC is rapidly becoming a routine analyticaltechnique due to its advantages of low operating costs, high sample throughput, andthe need for minimum sample preparation. The major advantage of HPTLC is thatseveral samples can be run simultaneously using a small quantity of mobilephase-unlike HPLC; thus reducing the analysis time and cost per analysis.

The developed HPTLC and HPLC technique is precise, specific, and accurate.Statistical analysis proves that the method is suitable for the analysis of AML andOLM as a bulk drug and in pharmaceutical formulation without any interferencefrom the excipients. It may be extended to study the degradation kinetics of AMLand OLM and also for its estimation in plasma and other biological fluids.

REFERENCES

Chaudhari, B. G., N. M. Patel, and P. B. Shah. 2007. Stability indicating RP-HPLC methodfor simultaneous determination of atorvastatin and amlodipine from their combinationdrug products. Chem. Pharm. Bull. 55: 241–46.

Chintan, V. P., C. V. Patel, A. P. Khandhar, A. D. Captain, and K. T. Patel. 2007. Absorptionfactor spectrophotometric and reversed-phase high-performance liquid chromatographic

SIMULTANEOUS METHOD DEVELOPMENT AND VALIDATION 257

Dow

nloa

ded

by [

Purd

ue U

nive

rsity

] at

02:

57 0

8 Ju

ne 2

013

methods for the determination of ramipril and olmesartan medoxomil in pharmaceuticalformulations. Eurasian J. Anal. Chem. 2(3): 159–171.

ICH. 1996. Topic Q2 (R1): Validation of Analytical Procedures, Test and Methodology(CPMP=ICH=381=95). Available at http://www.ich.org/LOB/media/MEDIA417[1].pdf

Kamat, K., and S. C. Chaturvedi. 2005. Stability indicating assay method for amlodipinetablets. Indian J. Pharm. Sci. 67(2): 236–239.

Liu, D., P. Hu, N. Matsushima, X. Li, L. Li, and J. Jiang. 2007. Quantitative determination ofolmesartan in human plasma and urine by liquid chromatography coupled to tandem massspectrometry. J. Chromator. B 856(1–2): 190–197.

Mahadik, K. R., H. Aggrawal, and N. Kaul. 2004. Simultaneous HPTLC estimation ofamlodipine besylate and losartan potassium in tablet dosage form. Indian Drugs 41(1):

32–35.Meyyanathan, S. N., and B. Suresh. 2005. HPTLC method for the simultaneous determi-

nation of amlodipine and benazepril in their formulations. J. Chromatogr. Sci. 43(2): 73–75.Naidu, K. R., U. N. Kale, and M. S. Shingare. 2005. Stability indicating RP-HPLC method

for simultaneous determination of amlodipine and benazepril hydrochloride from theircombination drug product. J. Pharm. Biomed. Anal. 39(1–2): 147–155.

Sagirli, O., A. Anal, S. E. Toker, and D. Sensoy. 2007. Simultaneous HPLC analysis ofolmesartan and hydrochlorothiazide in combined tablets and in vitro dissolution studies.Chromatographia 66(3–4): 213–218.

Shah, N. J., B. N. Suhagia, R. R. Shah, and N. M. Patel. 2007. Development and validation ofa simultaneous HPTLC method for the estimation of olmesartan medoxomil and hydro-chlorothiazide in tablet dosage form. Indian J. Pharm. Sci. 69(6): 834–836.

Streel, B., C. Laine, C. Zimmer, R. Sibenalerand, and A. Ceccato. 2002. Enantiomericdetermination of amlodipine in human plasma by liquid chromatography coupled totandem mass spectrometry. J. Biochem. Biophys. Methods 54(1–3): 357–368.

Vaidya, V., S. M. Roy, S. S. Joshi, S. M. Yetel, and S. A. Parekh. 2008. LC–MS–MSdetermination of olmesartan in human plasma. Chromatographia 67(1–2): 147–150.

258 A. Y. KAMBLE ET AL.

Dow

nloa

ded

by [

Purd

ue U

nive

rsity

] at

02:

57 0

8 Ju

ne 2

013