First and Second Derivative Spectrophotometric Methods for Determination of Paliperidone in Pharmaceutical Formulation

Simple, fast and reliable derivative spectrophotometric methods were developed for determination of paliperidone in bulk and pharmaceutical dosage forms. The solutions of standard and the sample were prepared in methanol. The quantitative determination of the drug was carried out using the first derivative values measured at 248 nm and the second derivative values measured at 246 nm (n=6). Calibration graphs constructed at their wavelengths of determination were linear in the concentration range of paliperidone using 2-10 μg/ml for first and second derivative spectrophotometric method. The calibration graphs constructed at their wavelength of determination were found to be linear for UV and derivative spectrophotometric methods. 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, specific, sensitive, and reproducible and can be directly and easily applied to pharmaceutical dosage form.


INTRODUCTION:
Paliperidone is the major active metabolite of risperidone which is a widely used as atypical antipsychotic ((Sherje and Londhe 2015)). Paliperidone is a benzisoxazole derivative and approved for the treatment of schizophrenia ((Mathew, Joshi Chintankumar, and Jonils 2011)) and other psychiatric disorders . Chemically it is (±)-3-[2-[4-(6-fluoro-1,2benzisoxazol-3-yl)-1-piperidinyl]ethyl]-6,7,8,9tetrahydro-9-hydroxy-2-methyl-4Hpyrido[1,2-a]pyrimidin-4-one with empirical formula C23H27FN4O3 and molecular weight 426.49 ((Nageswara Rao, Ganapaty, and Lakshmana Rao 2013). Specific mechanism is not know. Its therapeutic effect may be due to combination of D2 and 5-HT receptor antagonism (Barbosa et al. 2012)). Paliperidone has an antagonistic effect on α1 and α2 adrenergic receptors and H1 histaminic receptors ( (Mathew,  Other methods for determination of paliperidone:-Reverse phase HPLC method: -This was developed and validated for the assay of paliperidone in pure and tablets. Analysis was carried out on Shimadzu HPLC model equipped with inertsil, LC-10AT pump, inertsil ODS C-18 column variable wavelength programmable UV-Detector SPD-10AVP and rheodyne injector with 20 µL fixed loop. Acetonitrile and methanol in the ratio 10:90 v/v at a flow rate of 1.0 mL/min was employed for elution and the components were detected at 235 nm at ambient temperature. The method was validated as per the ICH guidelines. The retention time for paliperidone was found to be 5.67 min. Linearity was observed in the concentration range of 20-120 ppm with correlation coefficient 0.9990. The percent of relative standard deviation of six replicate measurements was found to be 1.25 indicate that the proposed method was precise. Recovery studies were conducted at three different concentration levels within the linearity limits and the average percent of recovery was 99.56. The amount of paliperidone drug content present in tablet dosage form was determined and found to be 99.80%. Therefore, the developed method was simple, precise and accurate and can be successfully applied for the estimation of paliperidone in pharmaceutical dosage forms. ). The eluted compounds were monitored at 238 nm, the flow rate was 0.5 mL/min, and the column oven temperature was maintained at 35°C. The resolution of Paliperidone palmitate and eight (potential, bi-products and degradation) impurities was greater than 2.0 for all pairs of components. The high correlation coefficient (r2 > 0.999) values indicated clear correlations between the investigated compound concentrations and their peak areas within the test ranges. The repeatability and intermediate precision, expressed by RSD, were less than 10.0%. The accuracy and validity of the method were further ascertained by performing recovery studies via a spike method. The accuracy of the method expressed as relative error was satisfactory. The drug was subjected to the International Conference on Harmonization (ICH) prescribed hydrolytic, oxidative, photolytic, and thermal stress conditions. The performance of the method was validated according to the present ICH guidelines for specificity, limit of detection, limit of quantification, linearity, accuracy, precision, ruggedness, and robustness. PREPARATION OF STANDARD SOLUTIONS: Stock solution of 1000 μg/ml of paliperidone was prepared in methanol, for first and second derivative spectrophotometric analysis. The standard solutions were prepared by dilution of the stock solution with methanol in a concentration range of 2, 4, 6, 8 and 10 μg/ml for first and second derivative spectrophotometric methods. Methanol was used as a blank solution.

ASSAY PROCEDURE:
A total of 20 tablets of paliperidone were accurately weighed and powdered. An amount of tablet triturate equivalent to label claim of paliperidone was weighed and transferred in 10 ml calibrated volumetric flask, diluted with methanol stirred for about 45 min and then volume made up with methanol. This solution was filtered to remove any insoluble matter. The filtrate was collected in a clean volumetric flask. Appropriate dilutions were made to obtain 10 μg/ml with methanol from stock solution for both UV and derivative spectrophotometric methods ( Table 7).

VALIDATION:
The proposed method was validated according to the International Conference on Harmonization (ICH) guidelines 8 .

LINEARITY:
From the standard stock solution of paliperidone, appropriate aliquots were pipetted out in to A series of volumetric flasks and volume was made up with methanol to get concentrations of 2-10µg/ml. Absorbances of these solutions was measured at 279 for zero order spectroscopy, 248 for first order derivative spectroscopy and at 246 nm for second order spectroscopy.
Standard graphs of linearity for proposed methods Results were shown in Table 1, Table 2 and Table 3.

PRECISION:
To evaluate the precision of the method, pure drug solution (Within the Linearity range of 2-10µg/ml) was analyzed and being repeated six times. The relative error (%) and relative standard deviation (%) were less than 2.0 and indicate the high precision for the proposed methods (Table  4).

L.O.Q and L.O.D
The limits of detection and quantification of the proposed method was also analysed and reported (Table 5).

ACCURACY:
To assure the reliability of the above method recovery studies were carried out by mixing a known quantity of the standard drug with the formulation sample. The recovery studies were carried out at three different levels i.e. 80 %, 100% and 120 % levels. The contents were analyzed by the proposed method for UV, first and second derivative spectroscopy (Table 6).

Linearity:
The UV, first and second derivative spectra for paliperidone were recorded at the wavelength of 279nm, 248nm, 246 nm respectively (Figure3, Figure 5 and Figure  7). The proposed method was found that the drug obeys linearity with in concentration range of 2-10μg/ml ( Table  1, Table 2 and Table 3) and (Figure 2, Figure 4 and Figure  6) Table 1: Linearity of UV for Paliperidone     From the precision studies it was found that the % RSD was less than 2% which indicates that the method has good reproducibility. To determine the precision of the method, paliperidone solution of concentration 6 μg/ml was analyzed six times for UV, first and second derivative spectrophotometric methods (Table 4).  Accuracy: The % recovery values of indicates that the method is accurate and reveals that commonly used excipients and additives present in the pharmaceutical formulations did not interfere in the proposed method. (Table 6)  Due to high sensitivity and simple sample preparation, the methods described can be used for undergraduate studies. Moreover simple spectrophotometric methods have obvious advantages over sophisticated instrumental analysis such as HPLC. This method was validated for precision, linearity and accuracy as per ICH guidelines. All the above parameters lead to the conclusion that the proposed method is accurate, precise, simple, sensitive and cost effective and can be applied successfully for the routine estimation of paliperidone in bulk and pharmaceutical formulation.