Development and Validation of a RP-HPLC Method for Estimation of Montelukast Sodium in Bulk and in Tablet Dosage Form

Indian Journal of Pharmaceutical Sciences 235 March April 2010 3. Bindoli A, Rigobello MP, Musacchio E, Scuri R, Rizzoli V, Galzigna L. Protective action of a new benzofuran derivative on lipid peroxidation and sulphydryl groups oxidation. Pharmacol Res 1991;24:369-75. 4. Lu Z, Gregory R. Potent, selective, orally bioavailable inhibitors of tumor necrosis factor-α converting enzyme (TACE): Discovery of indole, benzofuran, imidazopyridine and pyrazolopyridine P1′ substituents. Bioorg Med Chem Lett 2008;18:1958-62. 5. Kirilmis C, Ahmedzade M, Servi S. Synthesis and antimicrobial activity of some novel derivatives of benzofuran: Part 2, The synthesis and antimicrobial activity of some novel 1-(1-benzofuran-2-yl)-2mesitylethanone derivatives. Eur J Med Chem 2008;43:300-8. 6. Rizzo S, Rivière C, Piazzi L, Bisi A, Gobbi S, Bartolini M. Benzofuran-based hybrid compounds for the inhibition of cholinesterase activity, beta amyloid aggregation, and abeta neurotoxicity. J Med Chem 2008;51:2883-6.

Montelukast sodium working standard was obtained from Vitalife Laboratories (A division of Arch Pharma lab) Gurgaon, Haryana, India.Montair Tablets (montelukast sodium tablets, 10 mg) was purchased from a local pharmacy store.All chemicals and reagents used were of HPLC grade and purchased from E. Merck Chemicals Corporation Ltd.Mumbai, India.A waters HPLC system containing 600 controller, retention time obtained by HPLC for montelukast is about 3.4 min as shown in fi g.1.
Linearity was studied by preparing standard solutions of montelukast at different concentration levels.The linearity ranges were found in the range of 1-100 µg/ml.The standard calibration curve was generated using regression analysis with Microsoft excel.The assay was judged to be linear as the correlation coeffi cient was greater than 0.995 by the least-square method as shown in Table 1.
Recovery studies of the drug were carried out for the accuracy parameter at three different concentration levels i.e. multiple level recovery studies.A known amount of montelukast standard was added into preanalysed sample and subjected to the proposed HPLC method.Percentage recovery was found to be within the limits as listed in Table 2.
inline degasser, 717 plus auto-sampler, 486 tunable absorbance detector and temperature control device with operating software Millennium 32 were used during the study.A sunfi re C 18 column (250×4.6 mm, 5 µm particle size) was used as stationary phase.The mobile phase was optimized with acetonitrile and 1 mM sodium acetate buffer (adjusted to pH 6.3 with acetic acid), in the proportion of 90:10 v/v, UV detection was carried out at 285 nm with a fl ow rate of 1.5 ml/min.About 10 mg of montelukast sodium standard was weighed accurately and transferred to 100 ml volumetric fl ask.The volume was made up to mark with the mobile phase to obtain a concentration of 100 µg/ml.Further dilutions were made to obtain the concentration in the range of 1-100 µg/ml of montelukast sodium.For analysis in tablet dosage form, twenty tablets were weighed.The tablets were finely powdered and powder equivalent to 10 mg of montelukast sodium was accurately weighed and transferred into a 100 ml volumetric fl ask.The volume was made up to mark with the mobile phase to obtain a concentration of 100 µg/ml.Further dilutions were made to obtain a concentration of 10 µg/ml and fi ltered through 0.45 µm membrane fi lter.
The system suitability was checked by injecting 20 µl of standard solution and found the results within the range.The relative standard deviation on fi ve replicate injections was obtained 0.5%, tailing factor 1.25, and the column effi ciency 1278 theoretical plates.
Twenty microlitres of standard and sample solutions were separately injected on HPLC system.From the peak area of montelukast the amount of drugs in the sample were computed.The analysis was repeated in triplicate.The % assay of the bulk was found to be 100.3±0.60 and for tablet dosage form 99.51±1.76 of the labelled claim.
The developed method was validated in terms of specifi city, linearity, accuracy, limit of detection, limit of quantifi cation, intra-day and inter-day precision and robustness for the assay of montelukast sodium as per ICH guidelines [17] .Specificity was studied for the examination of the presence of interfering components.Montelukast standard solution of 10 µg/ml was injected and none of the impurities were interfering in its assay.The   Precision was studied to fi nd out intra and inter day variations in the test methods of montelukast in the concentration range of 5-50 µg/ml for three times on the same day and different day.Precision was determined by analysing corresponding standard daily for a period of three days.The inter-day and intra-day precision obtained was % RSD (<2) indicates that the proposed method is quite precise and reproducible as shown in Table 3.
The Limit of Detection (LOD) and Limit of Quantitation (LOQ) was calculated based on the standard deviation (SD) of the response and the slope (S) of the calibration curve at levels approximating the LOD and LOQ, LOD= 3.3 (SD/S) and LOQ= 10 (SD/S) is shown in Table 1.Robustness was done by small changes in the chromatographic conditions like mobile phase, flow rate etc. and found to be unaffected.
A simple, accurate, fast and precise isocratic reverse phase high performance liquid chromatographic method has been developed for the determination of montelukast sodium in bulk and in tablet dosage form.The developed method was found to be simple and have short run time which makes the method rapid.The results of the study indicate that the proposed HPLC method is simple, precise, accurate and less time consuming.

TABLE 2 : RECOVERY STUDIES OF MONTELUKAST IN TABLET Label claim mg/ tablet Total amount added (mg)
*Each value is a mean±standard deviation of three determinations