SPECTROPHOTOMETRIC DETERMINATION OF METRONIDAZOLE IN PHARMACEUTICAL PREPARATIONS
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1 SPECTROPHOTOMETRIC DETERMIATIO OF METROIDAZOLE I PHARMACEUTICAL PREPARATIOS Saffaj T*., Charrouf M., Abourriche A., Aboud Y., Bennamara A., Maoufoud S., Berrada M. Laboratoire de Chimie Organique Biomoléculaire, Faculté des Sciences Ben M Sik, Avenue Cdt D. El Harti BP 7955, Casablanca, Maroc. saffajt@yahoo.fr, Fax: Abstract- A rapid and sensitive spectrophotometric method is proposed for determination of Metronidazole. The method depends on the reduction of Metronidazole molecule with zinc dust and hydrochloric acid flowed by diazotation and coupling with 8-quinolinol to give colored chromogens easily measured spectrophotometrically which has λ max = 437 nm. The experimental conditions were optimized and berr s law was obyed over the applicable concentration ranges.both techniques were applied successfully to a wide variety of pharmaceutical preparations. Keywords: Metronidazole, Diazotation, 8-quinolinol, spectrophotometry. 1- ITRODUCTIO Metronidazole (2-methyl-5-nitroimidazole-1-ethanol) is used as antiprotozoal, antiamebic and antibacterial drugs. Excellent reviews have been published on the activity and pharmacokinetics of this drug. Several methods have been reported for determination of metronidazole which includes volumetric, gravimetric, polarographic, CPG, TLC, HPLC, voltammetric, derivative spectrophotometry, flow injection analysis, official methods and spectrophotometry. Most of the spectrophotometric methods reported suffer from the disadvantage, like narrow range of determination, requires heating or extraction, long time for the reaction to complete, use of non-aqueous systems, stability of the coloured product formed, etc. This paper describes sensitive and simple spectrophotometric method for the determination of metronidazole in either pure form or in its pharmaceuticals formulations. The method is based on the reduction of metronidazole molecule with zinc dust and hydrochloric acid flowed by diazotation and coupling with 8-quinolinol. The scientific novelty of the present work is that the reagents used in both the method is easily available and the chemistry of these reagents is already well established. The reactions involed with these reagents are simple, rapid and sensitve in their range of determination compared with other established methods. As Metronidazole is important class of imidazole compouds known for their antiamebic and antiprotozoal activity, this determination in pharmaceutical is of great importance.
2 2- MATERIEL AD METHODS 2-1 Instrumentation 2-2 Reagents A Perkin-Elmer 551 UV-Visible spectrophotometer was used. All chemicals used were of analytical-reagent grade. 8-quinolinol was purchased from. sodium nitrite was purchased from. Metronidazole was obtained as gifts from Aventis Pharma. All other reagents and solvents were of analytical-reagent grade. 2-3 Solutions Accurately weighed ( 100 mg) Metronidazole was transferred to a 100 ml beaker. Add 1g of zinc dust along with 20ml 1M hydrochloric acid. Stir well and wait for 1h at room temperature, filter and the filtrate was diluted with ethanol to 100ml in a volumetric flask. The working standard solution of the reduced Metronidazole containing 100µg ml -1 was prepared by further dilution. A 1% 8-quinolinol solution in 1M HCl and a 10% solution of hydroxyde de sodium were kept in amber-glass volumetric flasks. A 1% sodium nitrite solution and a 5% ammonium sulfamate solution were prepared separately in distilled water. 2.4 Procedure aliquots of the working standard solution of reduced Metronidazole were transfered into 25 ml calibrated flasks. 1ml of 1M HCl was added, cool in an ice bath and add 1ml of 1% ao 2, stir the solution for 2 min. Add 2ml of 5% ammonium sulfamate, stir the solution for 3 min and add 1 ml of 1% of 8-quinolinol. After 2min add 2ml of 10% of a and made up to the mark with ethanol. 2.5 Assay of pharmaceutical tablets tweleve tablets were powdered and mixed thoroughly. An amout equivalent to 100 mg of the drug was reduced as mentioned in and the filtrate was made up to 100ml and an aliquot of this solution was treated as described above for pure sample in both the method. 3- RESULTS AD DISCUSSIO The spectrophotometric method for the determination of Metronidazole is based on the reduction of the nitro to an amino group with zinc dust and hydrochloric acid flowed by diazotation and coupling with 8-quinolinol to give colored product. 3-1 Spectral characteristics and reaction mechanism the absorption spectra of the coloured product with λ max = 437 nm is shown in. The reagent blank has pratically negligible absorption at this wavelength. The stochiometric equation derived was shown in scheme 1.
3 O 2 + HCl Zn H 2 H 2 + ao HCl Cl - HO 2 + Cl - + O C scheme 1: Reaction sequence for the formation of azo colored product 3.2 Optimization of reactions conditions the factors affecting color development, reproducibility, sensitivity, and conformity with Beer s law were investigated. It was found that, 1-3 ml of 1M HCl, 1-4 ml of 1% ao 2 solution, 2-5 ml of 5% ammonium sulfamate, 1-3 ml of 1% 8-quinolinol and 1-3 ml of 10% a solution were necessary to achieve maximum colour intensity. The excess of nitrite sodium could be removed by the addition of 2ml of 5% ammonium sulfamate solution. An excess of ammonium sulfamate has no effect on the colour intensity of the product formed. 3.3 Quantification Beer s law is obeyed over the Metronidazole concentration range of 1-10 µg / ml. The proposed procedure is validated by determining various optical parametrs, which are listed in.
4 Table 1: parametrs for the spectrophotometric determination of Metronidazole λ max (nm) 437 Beer s law range (µg ml -1 ) 1-10 Molar absorptivity ( L mol -1 cm -1 ) Regression equation Slop Intercept Correlation coefficient R.S.D.(%) 1.22 a. y= ax + b where x is the concentration of Metronidazole 3.4 analysis of pharmaceutical preparation. Application of the proposed method to the determination of Metronidazole drug in its dosage forms was successfully made; the results are presented in table 2. the excellent recoveries obtained indicated the absence of any interference from the excipients. Table 2: Analysis of Metronidazole in pharmaceutical preparation Commercial Label claim in mg Recovery a, %(± RSD b ) Formulations analyzed Flagyl /tablet 99.5 (± 1.3) Flagyl 500 idazol /tablet 500/tablet 98.2(± 2.5) 101.3(±1.8) a. Average of 5 determination. b. Relative standard deviation. 4- COCLUSIO The method is found to be simple, economical, selective and more sensitive than most of the spectrophotometric methods reported. The statistical parametrs and recovery study data clearly indicate the reproducibility and accuracy of the method. Analysis of the authentic samples containing Metronidazole showed no interference from the common excipients. Hence, this approach could be considered for the determination of Metronidazole in the quality control laboratories. 5- REFERECES [1] VEGA, E., SOLA,., J. Pharm. Biomed. Anal., Vol. 25, 2001, p. 523 [2] BARI VIDDESH, R., DHORDA, U.J., SUDARESA, M., Anal. Chim. Acta., Vol.376, 1998, p.221
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