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B. Sandhya et al., IJSID, 2011, 1 (3), 441-450ISSN:2249-5347IJSIDInternational Journal of Science Innovations and DiscoveriesAn International peerReview Journal for ScienceResearch ArticleAvailable online through www.ijsidonline.infoRP-HPLC METHOD DEVELOPMENT AND VALIDATION FOR THE ANALYISIS OFSUNITINIB IN PHARMACEUTICAL DOSAGE FORMSB.Sandhya 1 , V.Chinnari Harika 2 , Bikshal Babu.Kasimalla 3 *, Reshma Syed 3 , Kalpana Pammi 31Dept of Life Sciences, Sims College, Guntur, AP, India, 2 Dept of Food and Nutritional sciences, AcharyaNagarjuna University, Guntur, AP, India, 3 R.V.Labs, Guntur, A.P ,India.Received: 09.09.2011Modified: 11.10.2011Published: 29.12.2011*Corresponding AuthorName:Bikshal Babu. KasimallaPlace:Guntur, AP, IndiaE-mail:bikshalbabu@gmail.comABSTRACTA simple, selective, linear, precise and accurate RP-HPLC methodwas developed and validated for rapid assay of SUNITINIB in tabletdosage form. Isocratic elution at a flow rate of 1ml min -1 was employedon a symmetry C18 column at ambient temperature. The mobile phaseconsisted of Acetonitrile: Methonal: OPA 40:30:30(v/v). The UVdetection wavelength was at 230nm.Linearity was observed inconcentration range of 30-90ppm. The retention time for Sunitinib was3.068 min. The method was validated as per the ICH guidelines. Theproposed method can be successfully applied for the estimation ofSunitinib in pharmaceutical dosage forms.INTRODUCTIONKey words: Sunitinib, HPLC Method, Development, 230nm.INTRODUCTIONInternational Journal of Science Innovations and Discoveries, Volume 1, Issue 3, November-December 2011441

B. Sandhya et al., IJSID, 2011, 1 (3), 441-450INTRODUCTIONSunitinib (marketed as Sutent by Pfizer, and previously known as SU11248) is an oral, smallmolecule,multi-targeted receptor tyrosine kinase (RTK) inhibitor that was approved by the FDA for thetreatment of renal cell carcinoma (RCC) and imatinib-resistant gastrointestinal stromal tumor (GIST) onJanuary 26, 2006. Sunitinib was the first cancer drug simultaneously approved for two differentindications. [1]IUPAC NameN-(2-diethylaminoethyl)-5-[(Z)-(5-fluoro-2-oxo-1H-indol-3-ylidene)methyl]-2,4-dimethyl-1Hpyrrole-3-carboxamideSunitinib inhibits cellular signaling by targeting multiple receptor tyrosinekinases (RTKs).These include all receptors for platelet-derived growth factor (PDGF-Rs) and vascularendothelial growth factor receptors (VEGFRs), which play a role in both tumor angiogenesis and tumorcell proliferation. The simultaneous inhibition of these targets therefore leads to both reduced tumorvascularization and cancer cell death, and ultimately tumor shrinkage.Sunitinib also inhibits KIT(CD117), [2] the RTK that (when improperly activated by mutation) drives the majority of gastrointestinalstromal cell tumors. [3] It has been recommended as a second-line therapy for patients whose tumorsdevelop mutations in KIT that make them resistant to imatinib, or who become intolerant to thedrug. [4][5] In addition, sunitinib inhibits other RTKs. [6] These include: like RCC, GIST does not generallyrespond to standard chemotherapy or radiation. Imatinib was the first cancer agent proven effective formetastatic GIST and represented a major development in the treatment of this rare but challengingdisease. However, approximately 20% of patients do not respond to imatinib (early or primaryresistance), and among those who do respond initially, 50% develop secondary imatinib resistance anddisease progression within 2 years. Prior to sunitinib, patients had no therapeutic option once theybecame resistant to imatinib. [7]International Journal of Science Innovations and Discoveries, Volume 1, Issue 3, November-December 2011442

B. Sandhya et al., IJSID, 2011, 1 (3), 441-450Sunitinib offers patients with imatinib-resistant GIST a new treatment option to stop further diseaseprogression and, in some cases, even reverse it. This was shown in a large, Phase III clinical trial in whichpatients who failed imatinib therapy (due to primary resistance, secondary resistance, or intolerance)were treated in a randomized and blinded fashion with either sunitinib or placebo. [7] The study wasunblinded early, at the very first interim analysis, due to the clearly emerging benefit of sunitinib. At thattime, patients receiving placebo were offered to switch over to sunitinib. In the primary endpoint of thisstudy, median time to tumor progression (TTP) was more than 4-fold longer with sunitinib (27 weeks)compared with placebo (6 weeks, P

PrecisionB. Sandhya et al., IJSID, 2011, 1 (3), 441-450Table.3 Linear Regression Data for Calibration curveDrug SUNITINIBConcentration range 70-10ppmSlope (m) 0.002422Intercept (b) 0.103467Correlation coefficient 0.999% RSD Intra day-0.63Interday-0.64Repeatability of the method was checked by injecting replicate injections of 30 ppm of the solutionfor six times on the same day as intraday precision study of SUNITINIB and the RSD was found to be 0.63for intraday and 0.64for interday.AccuracyTable 4: Precision parameters of SUNITINIBINJECTION CONCENTRATION INTRA DAY INTER DAY1 30ppm 12361 124162 30ppm 12413 125123 30ppm 12295 123094 30ppm 12391 124255 30ppm 12420 125096 30ppm 12218 12361RSD 0.63 0.64The accuracy of the method was determined by calculating recovery of SUNITINIB by the methodof standard addition. Known amount of SUNITINIB (10ppm, 20ppm and 30ppm) was added to a prequantified sample solution and the amount of SUNITINIB was estimated by measuring the peak arearatios and by fitting these values to the straight line equation of calibration curve. The recovery studieswere carried out three times over the specified concentration range and amount of SUNITINIB wasestimated by measuring the peak area ratios by fitting these values to the straight line equation ofcalibration curve. From the above determination, percentage recovery was calculated and the averagerecovery was found to be 99.23%SpecificityThe specificity of the method was determined by comparing test results obtained from analysis ofsample solution containing excipients with that of test results those obtained from standard drug.International Journal of Science Innovations and Discoveries, Volume 1, Issue 3, November-December 2011447

B. Sandhya et al., IJSID, 2011, 1 (3), 441-450LOD and LOQFigure 3: Typical chromatogram of SUNITINIBLimit of detection (LOD) and limit of quantification (LOQ) were calculated as 0.1ppm and 0.4ppmrespectively as per ICH guide-lines. Results are shown in table 5.RobustnessTable 5: Results of LOD and LOQ.ParameterMeasuredLOD0.1ppmLOQ0.3ppmTo determine the robustness of the method, two parameters from the optimized chromatographicconditions were varied. Results of Robustness are shown in table 6.Table 6: Robustness resultsParameter Modification Peak Area % of changeStandard No change 21253 ……….M.PHASE Acetonitrile:Methanol:OPA 30:40:70 12457 0.777PH 5.2 12598 1.66WAVELENGTH 237nm 12490 1.044System Suitability Parameter:System suitability tests were carried out on freshly prepared standard stock solutions ofSUNITINIB and it was calculated by determining the standard deviation of SUNITINIB standards byinjecting standards in six replicates at 6 minutes interval and the values were recorded inTable7.International Journal of Science Innovations and Discoveries, Volume 1, Issue 3, November-December 2011448

B. Sandhya et al., IJSID, 2011, 1 (3), 441-450Table7: System suitability parameters of SUNITINIBParametersValuesλ max (nm)230nmBeer’s law limit (μg/ml) 10-705ppmCorrelation coefficient 0.999Retention time 3.068 minTheoretical plates 8284Tailing factor 1.68Limit of detection 0.1ppmLimit of quantification 0.4ppmOptimization of the chromatographic conditionsRESULT AND DISCUSSIONThe nature of the sample, its molecular weight and solubility decides the proper selection of thestationary phase. The drug SUNITINIB being non-polar is preferably analyzed by reverse phase columnsand accordingly C18 column was selected. So the elution of the compound from the column wasinfluenced by polar mobile phase. The concentration of the methanol and Acetonitrile were optimized togive symmetric peak with short run time based on asymmetric factor and peak area obtained. Differentmobile phases were tried but satisfactory separation, well resolved and good symmetrical peaks wereobtained with the mobile phase Acetonitrile, Methanol,OPA 40:30:30 (V/V). The retention time ofSUNITINIB was found to be 3.068 min, which indicates a good base line. The RSD values for accuracy andprecision studies obtained were less than 2% which revealed that developed method was accurate andprecise. The system suitability and validation parameters are given in Table 7. The high percentage ofrecovery of SUNITINIB was found to be 99.23 indicating that the proposed method is highly accurate.Proposed liquid chromatographic method was applied for the determination of SUNITINIB in tabletformulation. The result for SUNITINIB was comparable with a corresponding labelled amount . Theabsence of additional peaks indicates no interference of the excipients used in the tablets.Table 8: Formulation results of SUNITINIBFormulation Dosage Sample concentration %EstimationSutent (capsule) 50mg 30ppm 99.74CONCLUSIONA validated RP-HPLC method has been developed for the determination of SUNITINIB in tabletdosage form. The proposed method is simple, rapid, accurate, precise and specific. Its chromatographicrun time of 3.068 min allows the analysis of a large number of samples in short period of time. Therefore,it is suitable for the routine analysis of SUNITINIB in pharmaceutical dosage form.International Journal of Science Innovations and Discoveries, Volume 1, Issue 3, November-December 2011449

B. Sandhya et al., IJSID, 2011, 1 (3), 441-450REFERENCES1. US Food and Drug Administration (2006). "FDA approves new treatment for gastrointestinal andkidney cancer".2. Hartmann JT, Kanz L (November 2008). "Sunitinib and periodic hair depigmentation due totemporary c-KIT inhibition". Arch Dermatol 144 (11): 1525–6.doi:10.1001/archderm.144.11.1525.3. "Prescribing information for Sutent (sunitinib malate)". Pfizer, Inc, New York NY.4. Demetri GD et al. (2006). "Efficacy and safety of sunitinib in patients with advanced gastrointestinalstromal tumour after failure of imatinib: a randomised controlled trial". Lancet 368 (9544): 1329–1338.5. "Phase II Trial of Sunitinib (SU011248) in Patients With Recurrent or Inoperable Meningioma"6. "Pfizer Scores New Approval for Sutent in Europe". 2 Dec 2010.7. FDA approves Sutent for rare type of pancreatic cancerInternational Journal of Science Innovations and Discoveries, Volume 1, Issue 3, November-December 2011450