Journal of Pharmacy Research Vol.4.Issue 4. April 2011 Nahla Nour El Din Ahmed Salama et al. / Journal of Pharmacy Research 2011,4(4),956-959 956-959 Research Article ISSN: 0974-6943 Available online through http://jprsolutions.info * Corresponding author. Nahla Nour El Din Ahmed Salama National Organization for Drug Control and Research (NODCAR), Giza, Egypt, Raw Materials Department ,6 Abou Hazem Street, Pyramids Ave, P.O. Box 29, Cairo, Egypt Validated Spectrophotometric and Spectrofluorimetric Stability-Indicating Methods for the Determination of Veralipride through Charge Transfer Complex Formation Nahla Nour El Din Ahmed Salama a , Maissa Yacoub Salem b , Lobna Mohammed Abd El Halim a *, Laila El- Sayed Abdel Fattah c a National Organization for Drug Control and Research (NODCAR), 6 Abu Hazem Street, Pyramids Ave, P.O. Box 29, Giza, Egypt b Analytical Chemistry Department, Faculty of Pharmacy, Cairo University, Cairo, Egypt c Analytical Chemistry Department, Faculty of Pharmacy, Misr University for Science and Technology, 6 th of October, Egypt Received on: 04-01-2011; Revised on: 17-02-2011; Accepted on:16-03-2011 ABSTRACT Simple, rapid and accurate spectrophotometric and spectrofluorimetric stability indicating methods for analysis of veralipride (Ver) in presence of its hydrolytic and oxidative degradants were developed and validated. The methods depend on charge transfer reaction of Ver as n-electron donor with p-chloranilic acid (p-CA), and 2, 3 – dichloro 5, 6 – dicyano 1, 4 benzoquinone (DDQ) as p- acceptors. On the same principle, a spectrofluorimetric method was developed based on measuring the enhancement of fluorescence intensity of Ver - DDQ charge transfer complex at 450 nm after excitation at 320 nm. Under the optimum reaction conditions, linear relationships with good correlation coefficients (0.9994 - 0.9997) were found between absorbance or the difference in fluorescence intensity of the formed complexes and concentrations of Ver in the range of 10.0 - 400.0 μg mL -1 and 0.5-5.0 μg mL -1 for the spectrophotometric and the spectrofluorimetric methods, respectively. The molar absorptivities and association constants for the colored complexes were evaluated using the Benesi-Hildebrand equation. The standard free energy was also calculated. The methods were successfully applied for the determination of Ver in drug substance, drug product and in laboratory prepared mixtures containing different percentages of hydrolytic and oxidative degradants. Key words: Veralipride; p-CA; DDQ; Charge transfer complex; Stability indicating; Benesi-Hildebrand equation I NTRODUCTI ON 1. Introduction Veralipride (Ver) is substituted benzamide used as anti-psychotic drug. It has been used in 2.3. Standard solutions (1 mg mL -1 ) Veralipride standard solution, 1 mg mL -1 was prepared in acetonitrile. This stock solution was subsequently used for preparation of working standards in concentration ranges 100.0 - 1000.0 2.4. Veralipride degradant solutions (1 mg mL -1 ) Acid, alkaline and oxidative degradants were laboratory prepared according to the method mentioned for Ver [6] . 2.5. General analytical procedures 2.5.1. Spectrophotometric methods 2.5.1.1. Construction of calibration curves Into a series of 10-mL volumetric flasks, aliquots of Ver working standard solutions equiva- lent to 400.0 - 4000.0 μg for p-CA while 100.0 - 1000.0 μg for DDQ were transferred separately, 2.5 mL of 0.2 % (w/v) of p-CA and 1 mL of 0.1 % (w/v) of DDQ were added. The volumes were completed to the mark with acetonitrile and the reactions were allowed to proceed at room temperature. The reactions were achieved instantaneously at ambient temperature (25 ± 2 ° C). The absorbance of the resulting solutions was measured at the wavelengths of maximum absorbance (520 and 588 nm for p-CA and DDQ respectively) against reagent blanks treated similarly. Each measurement was plotted against its corresponding concentration and the regression equations were calculated. 2.5.1.2. Determination of stoichiometry of the charge transfer complex by Job?’s method of continuous variation Equimolar solutions of Ver (2.6 × 10 -3 M) and each p- acceptor, p-CA and DDQ were prepared in acetonitrile. A series of solutions drug and the respective acceptor was made up comprising different complementary proportions (1:9, 2:8, 3:7…, 9:1) in 10-mL volumetric flasks. The absorbances of the resulting solutions were measured at the wavelength of maximum absor- bance against reagent blanks treated similarly. 2.5.1.3. Determination of stability constant, molar absorbitivity and standard free energy change Aliquots volume equivalent to 0.2 - 1.0 mL of solutions of standard Ver solution in acetoni- trile (2.6 x 10 -2 M) were separately transferred into two series of 10-mL volumetric flasks. One milliliter and 0.2 mL of p-CA or DDQ solutions (2.6 x 10 -3 M) were separately transferred to the first and second series respectively. The procedures were followed as mentioned under 2.5.1.1. 2.5.2. DDQ spectrofluorimetric charge transfer method 2.5.2.1. Construction of calibration curve Into a series of 10-mL volumetric flasks, aliquots of Ver working standard solutions equivalent to 5.0 - 50.0 μg were quantitatively transferred and the proposed procedure under 2.5.1.1 was followed. The difference in fluorescence intensity at 450 nm after excitation at 320 nm was recorded at ambient temperature (25 ± 2 ° C) against blank treated similarly. Each measurement was plotted against its corresponding concentration and the regression equation was calcu- lated. for p-CA while, 400.0 - 4000.0 and 5.0 - 50.0 μg mL -1 for DDQ spectrophotometric and spectrofluorimetric methods respectively. The stock solution was kept in refrigerator at 4 o C, and was stable for at least five days. Germany) was prepared as 0.2 % solution (w/v) in acetonitrile. 2, 3 – Dichloro 5, 6 – dicyano – 1, 4 – benzoquinone (DDQ; Acros, New Jersey, USA) was prepared as 0.1 % solution (w/v) in acetonitrile. Acetonitrile (Sigma Aldrich, St. Louis, USA), 1, 4-dioxane (BDH Chemicals Ltd., England), ethanol, methanol, isopropanol, 1, 2-dichloroethane (Fisher scientific - UK) and acetone (lab. Scan – Ireland) are the solvents used. 2, 5-Dichloro-3, 6- dihydroxy-1, 4-benzoquinone, (p-Chloranilic acid, p-CA; Merck, Munich, The molecular interactions between electron donors and electron acceptors are generally associated with the formation of intensely colored charge-transfer complexes, which absorb radiation in the visible region [7] . A variety of electron donating compounds have been reported to yield charge-transfer complexes leads to their utility in the development of simple and convenient spectrophotometric and spectrofluorimetric methods [8-13] . To the best of our knowl- edge, no charge-transfer reactions have been reported for the analysis of Ver, therefore the aim of the present study was directed to investigate this reaction. 2. MATERIAL AND METHODS 2.1. Instruments Spectrophotometer: Shimadzu UV–1601 PC, dual-beam UV-vis spectrophotometer (Japan), with matched 1 cm quartz cells, was used for spectrophotometric study. Spectrofluorimeter: Shimadzu RF–1501 spectrofluorimeter with 1-cm quartz cells (Japan) was used for the spectrofluorimetric study. 2.2. Materials and reagents Veralipride was kindly supplied from Memphis Co. Cairo, Egypt. Its purity was found to be 99.70 % according to the manufacturer’s UV spectrophotometric method [14] . Agreal capsules, labeled to contain 100 mg of veralipride and manufactured by Memphis Co., Cairo, Egypt under license of Synthelabo laboratories, France were purchased from the local market. Batch No. 200501. treatment of cardiovascular and psychological symptoms associated with the menopause [1-3] . It is N - [(1- Allyl – 2- pyrrolidinyl) Methyl] -5- Sulphamoyl -2- Veretramide [4]. Ver is not official in any pharmacopoeia. Few analytical methods were reported for its determination. These include UV spectrophotometry [5] , TLC-densitometry and High Performance Liquid Chromatographic (HPLC) methods [6] . Therefore, the goal of the present study was directed to the development of simple, rapid and inexpensive spectrophotometric and spectrofluorimetric stability indicating methods for the selective analysis of Ver in drug substance, drug product as well as in the presence of its hydrolytic and oxidative degradants.