Zero Order and Area Under Curve Spectrophotometric methods for determination of Oxcarbazepine in Pharmaceutical Formulation

Simple, fast and reliable spectrophotometric methods were developed for determination of Oxcarbazepine in bulk and pharmaceutical dosage forms. The solutions of standard and the sample were prepared in Distilled Water. The quantitative determination of the drug was carried out using the zero order derivative values measured at 256 nm and the area under the curve method values measured at 252-258 nm (n=2). Calibration graphs constructed at their wavelengths of determination were linear in the concentration range of Oxcarbazepine using 5-25 μg/ml (r2=0.998 and r2=0.9986) for zero order and area under the curve spectrophotometric method. All the proposed methods have been extensively validated as per ICH guidelines. There was no significant difference between the performance of the proposed methods regarding the mean values and standard deviations. Developed spectrophotometric methods in this study are simple, accurate, precise and sensitive to assay of Oxcarbazepine in tablets.


Apparatus and instrumentation
A Shimadzu1800 UV/VIS double beam spectrophotometer with 1cm matched quartz cells was used for all spectral measurements. Single Pan Electronic balance (CONTECH, CA 223, India) was IJASR|VOL 01|ISSUE 03|2015 www.ssjournals.com used for weighing purpose. Sonication of the solutions was carried out using an Ultrasonic Cleaning Bath (Spectra lab UCB 40, India). Calibrated volumetric glassware (Borosil®) was used for the validation study.

Materials
Reference standard of Oxcarbazepine API was supplied as gift sample by Marksan Pharmaceutical Ltd., Verna, Goa. Tablet sample with label claim 200 mg per tablet were purchased from local market Pune, Maharashtra, India.

Method development 2.3.1 Preparation of Standard and Sample Solutions:-
Stock solution of 20μg/ml of Oxcarbazepine was prepared in Distilled Water, for zero order and area under the curve spectrophotometric analysis. The standard solutions were prepared by dilution of the stock solution with Distilled water in a concentration range of 5, 10, 15, 20, and 25μg/ml with Distilled water for zero order and area under the curve spectrophotometric methods. Distilled water was used as a blank solution.

Area under curve (Area calculation)
Area under curve method involves the calculation of integrated value of absorbance with respect to the wavelength between two selected wavelengths such as λ1 and λ2 representing start and end point of curve region. The area under curve between λ1 and λ2 was calculated using UV probe software. In this study area was integrated between wavelength ranges from 252 to 258 nm.
Where, α is area of portion bounded by curve data and a straight line connecting the start and end point, β is the area of portion bounded by a straight line connecting the start and end point on curve data and horizontal axis, λ1 and λ2 are wavelength range start and end point of curve region [24].

Assay Procedure
Twenty tablets each containing 200 mg of Oxcarbazepine were weighed crushed to powder and average weight was calculated. Powder equivalent to 10 mg of Oxcarbazepine was transferred in 100 ml of volumetric flask. 50 ml of distilled water was added and sonicated for 15 minutes. Then solution was further diluted up to the mark with distilled water. The solution was filtered using Whatmann filter paper no. 41; first 5 ml of filtrate was discarded. This solution was further diluted to obtain 15µg/mL solution with water subjected for UV analysis using distilled water as blank. Appropriate dilutions were made with Distilled water from stock solution for both zero order and area under the curve spectrophotometric methods. www.ssjournals.com

Results and Discussion
The zero order and area under the curve spectra for Oxcarbazepine were recorded at the wavelength of 256nm and 252-258 nm respectively [ Fig. 2 and 3].

Linearity and Range
Under the experimental conditions described, the graph obtained for zero order and area under the curve spectra showed linear relationship.
Regression analysis was made for the slope, intercept and correlation coefficient values. The regression equations of calibration curves were y=0.0321x+0.0565 (r 2 =0.9980) at 256 nm for zero order derivative spectrophotometry and y=0.0019x+0.0009 (r 2 =0.9986) at 252-258 nm for area under the curve spectrophotometry. The range was found to be 5-25μg/ml for both zero order and area under the curve spectrophotometric methods.

Accuracy
To study the accuracy of the proposed methods, and to check the interference from excipients used in the dosage forms, recovery experiments were carried out by the standard addition method. The accuracy for the analytical method was evaluated at 80%, 100% and 120% levels of 20µg/ml standard solution. For Area under curve (AUC) was measured in wavelength range 252-258 nm and For Zero order derivative at 256 nm and results were obtained in terms of percent recovery. Three determinations at each level were performed and % RSD was calculated for each level. *n=3, % RSD = % Relative Standard Deviation.

Precision
To determine the precision of the method, Oxcarbazepine solutions at a concentration of 20μg/ml were analysed each three times for both zero order and area under the curve spectrophotometric methods. Solutions for the standard curves were prepared fresh everyday (Table II).

Sensitivity
The limit of detection (LOD) and limit of quantification (LOQ) were calculated by using the equations LOD = 3xσ/ S and LOQ = 10xσ/S, where σ is the standard deviation of intercept, S is the slope. The LOD and LOQ were found to be 1.3381μg/ml and 4.0550μg/ml respectively for zero order derivative and The LOD and LOQ were found to be 0.9676µg/ml &2.1140µg/ml for area under the curve methods respectively.

Analysis of the Marketed Formulation
There was no interference from the excipients commonly present in the tablets. The drug content was found to be 100.006% and 100.01% zero order and area under the curve spectrophotometric methods respectively. It may therefore be inferred that degradation of Oxcarbazepine had not occurred in the marketed formulations that were analysed by this method. The low % R.S.D. value indicated the suitability of this method for routine analysis of Oxcarbazepine in pharmaceutical dosage form (Table  4). www.ssjournals.com

Conclusion
No UV or Area under Curve spectrophotometric methods exists for the determination of Oxcarbazepine. Therefore simple, fast and reliable derivative spectrophotometric methods were developed for the routine determination of Oxcarbazepine. The developed methods can be concluded as accurate, sensitive and precise and can be easily applied to the pharmaceutical formulation.