DETERMINATION AND IDENTIFICATION OF LORATADINE BY VARIOUS ANALYTICAL METHODS USING UV - VISIBLE, FT - IR, AND HPLC CHROMATOGRAPHIC TECHNIQUES

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1 Page2899 Indo American Journal of Pharmaceutical Research, 2015 ISSN NO: DETERMINATION AND IDENTIFICATION OF LORATADINE BY VARIOUS ANALYTICAL METHODS USING UV - VISIBLE, FT - IR, AND HPLC CHROMATOGRAPHIC TECHNIQUES Mohammad Aftab Mansury *1, Rajesh Kumar Jatav 2, Rakesh K. Jatav 3 Department of Pharmaceutics, Central India Institute of Pharmacy Indore, M.P., India. ARTICLE INFO Article history Received 17/09/2015 Available online 30/09/2015 Keywords Loratadine, Sparingly Soluble, PH, PKA, 0.1 N Hcl, HPLC, FT-IR. ABSTRACT Loratadine, histamine H1 receptor antagonist used in the treatment of allergic rhinitis and urticaria Administration of Loratadine through oral route is a challenge in children, who have not yet learned to swallow tablets. In the present study pharmaceutical and analytical investigation carried out proceeding and supporting formulation development efforts of the dosage form of the drug substance. Loratadine is sparingly soluble in water but freely soluble in the ph of 1.2 to 2.0. The Pka of Loratadine is 4.3. It is extensively absorbed from the gastric ph. A reduction in gastric transit time may cause the drug to move into the alkaline ph of the intestine. standard curve of Loratadine is prepared in 0.1 N HCl absorbance measured at max of 280 nm. Standard curve was plotted between conc. ( g/ml) on x-axis and absorbance on y-axis. Various spectroscopic methods are used for the identification and analysis of Loratadine using HPLC, FT-IR and other spectroscopic techniques. development of various analytical techniques a new HPLC method for the determination of impurities in Loratadine drug substance and its validation. The method was found to be selective, sensitive, precise and accurate for the determination of impurities and degradation products. This method can be used for the routine determinations in pharmaceutical quality control laboratories and recommend future research. Corresponding Author Mohammad Aftab Mansury Department of Pharmaceutics Central India Institute of Pharmacy, Indore, M.P., India aftab_cool86@rediffmail.com, aftab86@gmail.com Please cite this article in press as Mohammad Aftab Mansury et al. Determination and Identification of Loratadine by Various Analytical Methods Using UV-Visible, FT-IR, and HPLC Chromatographic Techniques. Indo American Journal of Pharmaceutical Research.2015:5(09). Copy right 2015 This is an Open Access article distributed under the terms of the Indo American journal of Pharmaceutical Research, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

2 Page2900 INTRODUCTION We have proposed to develop various rapid and sensitive analytical techniques of identification for the separation of qualitative and quantitative determination of a second-generation antihistamine (Loratadine). The method will be used for applications in pharmaceuticals and biological environments recommend future research. Loratadine is long acting anti-histamine with selective peripheral histamine antagonistic activity, which lacks CNS depressant effects used in the treatment of allergic skin disorder, specially atopic dermatitis and urticaria, [1] allergic rhinitis, acute coryza, ocular allergies at the dose of 10 mg once a day in adult and 5 mg (if, 30 kg) or 10 mg (if, 30 kg) in 2 12 years children [2]. Administration of drugs through oral route is the most common and the easiest way to administer a drug. Loratadine is a piperidine derivative chemically [3,4] known as ethyl-4-(8-chloro-5,6 dihydro-11hbenzo[5.6] cyclohepta [1,2 b] pyridine-11-ylidene)-1-piperidine carboxylate is a long acting non-sedating antihistamine with no significant of antimuscarinic activity. Loratadine is orally administered drug and well absorbed from gastrointestinal tract,it has rapid firstpass metabolism, it is metabolized by isoenzymes of the cytochrome P450 system, [5] Loratadine is almost bound 97-99% to plasma proteins. Its metabolite is largely responsible for the antihistaminergic effects. It binds to plasma proteins by 73-76%. Figure 1: Structure of Loratadine. Figure 2: 3D view of Chemical Structure of Loratadine. Identification of Loratadine: Loratadine was identified by organoleptic evaluation, solubility studies, melting point, UV Spectroscopy, FT-IR spectroscopy [6] and HPLC Chromatographic Technique [7]. MATERIAL AND METHOD All chemicals and reagents used for this study were of analytical grade. Loratadine and other chemicals were obtained as a gift sample from Plethico pharmaceuticals Indore, M.P. Experimental Work: [7] Identification by organoleptic evaluation: The colour, odour, and taste of the drug were characterized visually.

3 Page2901 Table 1: Physical Appearance. S. No. Physical Appearance Standard Observed 1 Colour Off white powder White powder 2 Odour Odourless Odourless 3 Taste Slightly Bitter Slightly Bitter Identification by solubility studies: The spontaneous interaction of two more substances to form a homogenous molecular dispersion is known as solubility. The sample was qualitatively tested for its solubility in various solvents. For this test excess quantity of substance (in each solvent) and a maximum of 30 ml of each solvent are necessary Method: Weigh 100 mg finally powdered substance in a stoppered tube (16 mm in internal diameter and 160 mm long) add 0.1 ml of solvent, if the substance is completely dissolve, it is very soluble. If not completely dissolve add 0.9 ml of solvent, if substance is completely dissolve it is freely soluble. If not completely dissolve add 2 ml of solvent, if substance is completely dissolve it is soluble. If not completely dissolve add 7 ml of solvent if substance is completely dissolve it is sparingly soluble. If not completely dissolve add 10 ml of solvent, if substance is completely dissolve it is slightly soluble. If not completely dissolve weigh 1 mg of finely substance in stoppered tube, add 10 ml of the solvent it is very slightly soluble. Table 2: Solubility profile of Loratadine. Sr. No. Solvents Standard Observed 1 Methanol Freely soluble Freely soluble 2 Acetone Freely soluble Freely soluble 3 Toluene Freely soluble Freely soluble 4 Chloroform Freely soluble Freely soluble 5 PBS ph 7.4 Soluble Soluble 6 HCl Soluble Soluble 7 Water Insoluble Insoluble Identification by melting point: [7] A small quantity of powder was placed into a capillary tube which was sealed at one end. That tube was placed in the melting point determining apparatus. The temperature of the apparatus was gradual increased automatically and read the temperature at which powder started to melt and the temperature when all the powder gets melted. The melting point was determined in between C Table 3: Melting Point. S. No. Parameter Standard Observed 1 Melting Point C C 2 Loss on Drying Not more than 0.5% w/w 0.2 % w/w Determination of Moisture Content and Loss on Drying: [7] The moisture in a solid can be expressed on a wet-weight or dry-weight basis. On a wet-weight basis, the water content of material is calculated as percentage of the wet solid, whereas on the dry weight basis, the water is expressed as a percentage of the weight of the dry solid. Procedure: Weigh the container and add 1 gm of Loratadine as weight of wet sample, after weighing them kept into the oven at 120 C for 4 hours. After 4 hour weight of the sample was taken and then this process was repeated every 30 minute until the constant weight not obtained. Record the weight as the weight of dry sample. Determination of absorbance maxima (λ max ): [7] Weigh accurately 1 mg of Loratadine dissolve in and dilute up to 100 ml with the help of 0.1 N HCl: This resulted in preparation of solution of 10 g/ml. The solution was kept in cuvette of fused silica cell and scan the sample in the range of 200 nm to 400 nm and compare for the maxima and minima. The spectrum obtained from the test solution should be concordant with the spectrum obtained from standard solution.

4 Page2902 Figure 3: Determination of absorption maxima (λ max ) of Loratadine. Table 4: Determination of absorption maxima of Loratadine. Wavelength(nm) Interpretation Inference Scanning range Drug absorption maxima (λ max ) Highest peak nm of loratadine Calibration curve of Loratadine in 0.1 N HCl at λ max 280 nm: [7] The drug sample was weighed accurately 100mg and dissolved in minimum quantity of 0.1 N HCl and volume was made up to 100 ml using 0.1 N HCl. This resulted in preparation solution of 1000 g/ml. Withdrawn 10 ml of this solution and transfer in 100 ml volumetric flask and volume was made up to 100 ml using 0.1 N HCl. This resulted in preparation solution of 100 g/ml. Withdrawn, 0.2, 0.4, 0.6, 0.8, and 1.0 ml of sample separately and further diluted up to 10 ml with the 0.1 N HCl Absorbance of samples was taken at max of 280 nm. Standard curve was plotted between conc. ( g/ml) on x-axis and absorbance on y-axis Preparation of Calibration curve of Loratadine: Correlation coefficient (R 2 ) = Equation of regressed line; y = 0.113x Figure 4: Calibration curve of Loratadine in 0.1 N HCL. Where x = Value of concentration y = Value of absorbance Slope of regressed line= 0.113

5 Page2903 Table 5: Calibration curve of Loratadine. Sr. No. Concentration (µg/ml) Absorbance FT-IR Study: [7] Fourier transform spectrophotometers used polychromatic radiation reaction & calculate the spectrum in frequency domain from the original data by fourier transformation. Preparation of standard: Weighed mg of Loratadine & 0.1 g of KBr Preparation of sample: Weighed mg of sample & 0.1 g of KBr Infrared Spectra of Loratadine: Figure 5: IR spectrum of Loratadine (Standard). Figure 6: IR spectrum of Loratadine (Sample).

6 Page2904 Figure 7: Purity index of Loratadine. Table 6: Interpretation of FTIR spectra of Loratadine. Sr. No. Bonds Standard (cm-1) Observed (cm-1) 1 O-H stretch C=O stretch C-O stretch C=C stretch Chromatographic technique: High Performance Liquid Chromatography (HPLC) High Performance Liquid Chromatography is effective quantitative and qualitative method generally used the perpose for estimation of pharmaceutical and biological study. HPLC is the fastest, safest, dependable and most versatile chromatographic technique for purity and identification of drug components. HPLC: Agilent 1260 quats pump VL series Column: C 8, 150 X 4.6 mm 5µ inertsil or equivalent Flow rate: 1 ml/min Detector: UV at 254 nm. Column temp.: 25 0 C Injection volume: 15 µl Mobile phase: Mix 0.01 M dibasic potassium phosphate, methanol and acetonitrile in the ratio of 7:6:6 and adjust the ph 7.2 with 10% phosphoric acid. Filter with 0.45µ membrane & degas the solution. Diluent: Transfer 400 ml of 0.05 N HCI in 1000 ml volumetric flask, add 80 ml 0.6 M Dibasic potassium phosphate, and make up the volume with a mixture of methanol and acetonitrile (1:1) and mix. Standard preparation: Weigh and transfer accurately about 40 mg of Loratadine working standard in 100 ml volumetric flask, add 50 ml sonicate to dissolve the content and make up the volume with diluent. Sample preparation: Weigh and transfer accurately about 40 mg of Loratadine sample in 100 ml volumetric flask, add 50 ml diluent, sonicate to dissolve the content and make up the volume with diluent. Procedure: Separately inject about 15 µl of the blank, standard solution in 5 replicates and the test solution in duplicates in to the chromatographic system and record the chromatogram and measure the responses for the major peak.

7 Page2905 Graphical Observations: High Performance Liquid Chromatographs: Figure 8: HPLC curve of Blank solution injection no.1. Figure 9: HPLC curve of Blank solution injection no.2. Figure 10: HPLC curve of Loratadine (Standard) injection no. 1.

8 Page2906 Figure 11: HPLC curve of Loratadine (Standard) injection no. 2. Figure 12: HPLC curve of Loratadine (Standard) injection no. 3. Figure 13: HPLC curve of Loratadine (Standard) injection no. 4.

9 Page2907 Figure 14: HPLC curve of Loratadine (Standard) injection no. 5. Figure 15: HPLC curve of Loratadine (Standard) injection no. 6. Figure 16: HPLC curve of Loratadine (Sample) injection no. 1.

10 Page2908 Figure 17: HPLC curve of Loratadine (Standard) injection no. 2. Table 7: Loratadine std. observation (HPLC). Inj. No. Name Retention Time Area (in min.) (µv* sec.) USP Plate Count USP Tailing 1 Loratadine Loratadine Loratadine Loratadine Loratadine Loratadine Mean Std. Dev % RSD RESULTS AND DISCUSSION Loratadine found to be white to off-white crystalline powder, tasteless, odorless in organoleptic evaluation. Melting point of Loratadine was found to be C. Loss on Drying was observed 0.2 % w/w, Result of solubility studies Loratadine is freely soluble in methanol, acetone, toluene and chloroform, soluble in PBS ph 7.4 and HCl, and insoluble in water. UV spectroscopy study showed that the maximum absorbance at 280 nm after baseline correction in the UV spectrophotometer, when 10 μg/ml solution of Loratadine in 0.1 N HCl was scanned between 200 to 400 nm. FT-IR study concludes that characteristic peaks of Loratadine standard and sample were comparable (Figure No. 5, 6 and 7) in the range of cm -1 (Table No.6). HPLC studies concludes the retention time of the major peak in the chromatogram of the assay preparation corresponds to that in the chromatogram of the standard preparation as obtained in the assay (Figure No. 8-17) (Table No.7). CONCLUSION The present paper describes the various ways of determination and identification of new drug molecule development of various analytical techniques a new HPLC method for the determination of impurities in Loratadine drug substance and its validation. The method was found to be selective, sensitive, precise and accurate for the determination of impurities and degradation products. This method can be used for the routine determinations in pharmaceutical quality control laboratories recommend future research. ACKNOWLEDGEMENT The authors wish to thank the management of Plethico Pharmaceutical Ltd, for supporting this work. Authors wish to acknowledge the process research group for providing the samples for our research. Authors would also like to thank colleagues in separation science division of Formulation and Development department of Plethico Pharmaceutical Limited, Indore. Authors Statements: The authors declare no conflict of interest.

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