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official method for the determination of trazodone hydrochloride (TRH) is potenti- .... continuous variation.18 The method is simple and widely used for ...
J. Serb. Chem. Soc. 71 (7) 829–837 (2006) JSCS – 3474

UDC 543.4/.5:546.131+547.979.7:615 Original scientific paper

Sensitive extraction spectrophotometric methods for the determination of trazodone hydrochloride in pure and pharmaceutical formulations K. HARIKRISHNA, R. SUDHIR KUMAR, J. SEETHARAMAPPA* and D. H. MANJUNATHA Department of Chemistry, Karnatak University, Dharwad-580 003, India (e-mail: [email protected]) (Received 20 July, revised 25 October 2005) Abstract: Two simple, rapid and sensitive extraction spectrophotometric methods have been developed for the assay of trazodone hydrochloride (TRH) in pure and pharmaceutical formulations. These methods are based on the formation of chloroform soluble ion-association complexes of TRH with bromocresol green (BCG) and with methyl orange (MO) in a KCl–HCl buffer of pH 1.5 (for BCG) and in a NaOAc–HCl buffer of pH 3.29 (for MO) with absorption maximum at 415 nm and at 422 nm for BCG and MO, respectively. The reaction conditions were optimized to obtain the maximum colour intensity. The absorbance was found to increase linearly with increasing concentration of TRH, which was corroborated by the calculated correlation coefficient values (0.9992 and 0.9994). The systems obeyed the Beer law in the range of 0.9–17 and 1–20 mg/ml for BCG and MO, respectively. Various analytical parameters were evaluated and the results were validated by statistical data. No interference was observed from common excipients present in pharmaceutical formulations. The proposed methods are simple, accurate and suitable for quality control applications. Keywords: sprectrophotometric determination, trazodone hydrochloride. INTRODUCTION

Trazodone hydrochloride, 2-{-[4-(3-chlorophenyl)-1-piperazinyl]propyl}-1,2,4-triazolo-[4,3-a] pyridin-3-(2H)-ne monohydrochloride, is an anti-depressant (Fig. 1). It has been shown to be effective in patients with major depressive disorders and other subsets of depressive disordes. It is generally more useful in depressive disorders associated with insomnia and anxiety. This drug does not aggravate psychotic symptoms in patients with schizophrenia or schizoaffective disorders. The official method for the determination of trazodone hydrochloride (TRH) is potentiometric non-aqueous titration with perchloric acid1 and HPLC using an octadecyl *

Corresponding author.

doi: 10.2298/JSC0607829H

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Fig. 1. Structure of trazodone hydrochloride.

silane column and methanol–0.01 M ammonium phosphate buffer pH 6.0 (60:40) as the mobile phase.2 Analytical methods that are reported for the determination of TRH in pharmaceutical formulations include UV absorption measurement at 246 nm,3 ion-selective electrode,4,5 voltammetry6,7 and HPLC.2,8 Various chromatographic methods have been reported for the determination of TRH in biological fluids, including HPLC,9,10 capillary gas chromatography,11 gas chromatography mass spectrometry12 and instrumental thin layer chromatography.13 Though modern methods of analysis (HPLC, GLC, NMR and Mass spectrometry) for purity assay of any drug afford simplicity, speed, good specificity and excellent precision and accuracy. However, they involve sophisticated equipments, which are not within the reach of most laboratories and small-scale industries. Moreover, they pose maintenance problems. For single component preparations, the simplest assay method involves the direct measurement of the UV absorption at the wavelength of maximum absorbance. TRH is a relatively weak UV absorbing compound, therefore, direct UV absorbance measurements at low concentrations will be unreliable. Recently, spectrophotometric, spectrofluorimetric and LC determination of TRH have been reported.14 The sensitivity, detection limits and stability of the spectrophotometric methods were not discussed. Moreover, the effect of common excipients was not investigated by the spectrophotometric method. This prompted us to develop simple, sensitive and accurate spectrophotometric methods for the determination of TRH in pure and pharmaceutical formulations. These sensitive methods are based on the formation of chloroform soluble ion-association complexes of TRH with BCG and with MO in a KCl–HCl buffer of pH 1.5 for BCG and in NaOAc–HCl buffer of pH 3.29 for MO. EXPERIMENTAL Apparatus A Hitachi UV-visible spectrophotometer model U-2001 with 1 cm matched quartz cells was used for the absorbance measurements. The pH measurements were made on a Schott Gerate CG 804 pH meter.

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Reagents All employed chemicals were of analytical or pharmaceutical grade and quartz distilled, high-purity water was used throughout. Trazodone hydrochloride was obtained as a gift sample from Protec, Mumbai, India. Aqueous solutions of BCG and MO (each of 0.1 %) were prepared separately in high purity water. Series of buffer solutions of KCl–HCl (pH 1.0–2.2), NaOAc–HCl (pH 1.99–4.92), NaOAc–AcOH (pH 3.72–5.57) and potassium hydrogen phthalate–HCl (pH 2.2–3.6) were prepared by standard methods. A stock solution of TRH containing 250 mg/ml was prepared in distilled water. The solution is stable at room temperature. Commercial tablets of TRH were obtained from different firms. Assay procedure for the pure drug Aliquot of a solution containing 9–170 mg (for BCG) or 10–200 mg (for MO) of TRH were transferred into a series of 125 ml separating funnels. A volume of 3 ml of KCl–HCl buffer of pH 1.5 (for BCG) or 5 ml of NaOAc–HCl buffer of pH 3.29 (for MO), and 3 ml of BCG or 2 ml MO were added. Chloroform (10 ml) was added to each of the separating funnels, the contents were shaken well and left at room temperature for a minute. The two phases were allowed to separate and the chloroform layer was passed through anhydrous sodium sulphate. The absorbances of the yellow coloured complexes were measured at 415 and 422 nm for BCG and MO, respectively, against the corresponding reagent blank. A calibration graph was plotted. Assay procedure for tablets Six tablets were powdered and weighed. An amount of the powder equivalent to 100 mg of TRH was weighed into a 100 ml volumetric flask containing about 75 ml of distilled water. It was shaken thoroughly for about 15–20 min, filtered through a Whatman filter paper No. 40 to remove the insoluble matter and diluted to the mark with distilled water. A volume of 25 ml of the filtrate was diluted to 100 ml and a suitable aliquot was analyzed using the procedure given above. RESULTS AND DISCUSSION

Extraction spectrophotometric procedures are popular because of their sensitivity in the assay of drugs and hence, ion-pair extractive spectrophotometry has received considerable attention for the quantitative determination of many pharmaceutical compounds.15–17 TRH reacts with BCG and with MO in acidic buffer to give chloroform soluble ion-association complexes, which exhibit absorption

Fig. 2.A. Absorption spectrum of a) Reagent blank and b) ion-association complex of TRH (15 mg/ml) with BCG.

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Fig. 2.B. Absorption spectrum of a) Reagent blank and b) ion-association complex of TRH (15 mg/ml) with MO.

maxima at 415 and 422 nm for BCG and MO, respectively (Figs. 2.A and 2.B). Under the experimental conditions, the reagents blank showed negligible absorbance thereby permitting good analytical conditions for the quantitative determination of TRH. The drug–dye stoichiometric ratio was determined by the Job’s method of continuous variation.18 The method is simple and widely used for elucidating the composition of complexes and is based on the variation of both the drug and the reagent (BCG/MO) of equal molar concentrations, keeping the total volume of the drug and the reagent constant. A plot of absorbance against the mole fraction of the drug in the mixture shows a maximum at the composition of the complex. In the present study, the drug and dye concentrations were each 10–3 M, and a total volume of 10 ml was maintained. The drug–dye stoichiometric ratio as determined by the Job’s method was found to be 1:1 with both BCG and MO. The method of Foley and Anderson19 modified by Dey and Mukherji20 was applied to determine the stability constants of the drug–dye complex using the equation, x K= (a - x )(b - x ) where K is the stability constant of the complex, a is the initial concentration of the drug, b is the initial concentration of the reagent and x is the initial concentration of the complex. The stability constant of the drug–dye complex was found to be 0.68 ´ 103 and 0.44 ´ 103 LM–1 for BCG and MO, respectively. Optimization of the reaction conditions The optimum reaction conditions for the quantitative determination of the ion-pair complexes were established via a number of preliminary experiments. It was observed that the effective extraction of the complex depends on the type of buffer used and its pH. The effect of pH was studied by extracting the coloured complexes in the presence of various buffers, such as KCl–HCl (pH 1.0–2.2),

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NaOAc–HCl (pH 1.99–4.92), NaOAc–AcOH (pH 3.72–5.57) and potassium hydrogen phthalate–HCl (pH 2.2–3.6). It was noticed that the maximum colour intensity and constant absorbances were observed in KCl–HCl buffer (Clark & Lubs) of pH 1.5 for BCG and in NaOAc–HCl buffer (Walpole) of pH 3.29 for MO. Further, 3 ml of KCl–HCl buffer for BCG and 5 ml of NaOAc–HCl buffer for MO gave maximum absorbances and reproducible results. Low absorbance values were observed at pH values higher or lower than 1.5 for BCG and 3.29 for MO in the respective buffer medium. Hence, Clark & Lubs buffer of pH 1.5 for BCG and Walpole buffer of pH 3.29 for MO were selected for all subsequent measurements. The effects of the reagents were studied by measuring the absorbances of solutions containing a fixed concentration of TRH and varied amounts of the reagent separately. The maximum colour intensity of the complex was achieved with 3 ml of 0.1 % BCG or with 2 ml of 0.1 % MO. Although a larger volume of the reagent had no pronounced effect on the complex formation, the absorbances increased slightly due to background of the coloured reagent. Several organic solvents, viz., chloroform, carbon tetrachloride, ethyl acetate, xylene, diethylether, butyl acetate, toluene, dichloromethane and chlorobenzene, were tried for effective extraction of the coloured species from the aqueous phase. Only partial extraction of the complex was achieved with solvents other than chloroform. It was observed that only one extraction was adequate to achieve a quantitative recovery (99.68–100 %) of the complex with chloroform. Shaking times of 0.5 to 2 min produced constant absorbances and hence a shaking time of 1 min was maintained throughout. There was no appreciable change in the absorbance or colour of the product even if the order of addition of the reactants was varied. Effect of temperature on the coloured complexes The effect of temperature on the coloured complexes was studied at different temperatures. It was found that the coloured complexes were stable up to 35 ºC. At higher temperatures, the drug concentration was found to increase due to the volatility of the chloroform. As a result, the absorbances of the coloured complexes increased. However, the complexes wre stable for more than 8 h at room temperature. Detection and quantification limits According to the Analytical Methods Committee,21 the deterction limit (LOD) is the concentration of TRH corresponding to a signal equal to the blank mean (YB) plus three times the standard deviation of the blank (SB). The quantification limit (LOQ) is the concentration of TRH corresponding to the blank mean plus ten times the standard deviation of the blank. The LOD values were found to be 4.88 and 7.44 ng/ml for TRH with BCG and with MO, respectively. The LOQ values were observed to be 16.25 and 24.77 ng/ml for TRH with BCG and with MO, respectively. These values indicate that the BCG method is more sensitive than the MO method.

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Quantification The limits of the Beer law, the molar absorptivity and the Sandell’s sensitivity values were evaluated and are given in Table I. Regression analyses of the Beer law plots at their respecvite lmax values revealed a good correlation. Graphs of absorbances versus concentration showed zero intercept, and are described by the regression equation, Y = bX + c (where Y is the absorbance of a 1 cm layer, b is the slope, c is the intercept and X is the concentration of the drug in mg/ml) obtained by the least-squares method. The results are summarized in Table I. TABLE I. Optical characteristics, precision and accuracy data Parameter

Values BCG

MO

lmax/nm

415

422

Beer law limits/(mg/ml)

0.9–17

1–20

Molar absorptivity/(1 mol-1 cm-1)

1.66 ´ 104

1.37 ´ 104

Sandell’s sensitivity/(ng

cm–2)

24.57

29.59

Stability/h

8.5

8.25

Correlation coefficient/R

0.9992

0.9994

Slope, b

0.0387

0.0336

Intercept, c

0.0191

0.0032

Regression equation/(Y)a

Relative standard

deviation/%d

% Range of errord (95 % confidence limit)

a

0.94

0.89

0.87

0.98

Limit of detection/ng ml-1

4.88

7.44

Limit of quantification/ng ml-1

16.25

24.77

Y = bX + c, where X is the concentration of drug in mg/ml; dAverage of six determinations

Validation of the method The validity of the methods for the assay of TRH was examined by determining the precision and accuracy. These were determined by analyzing six replicates of the drug within the Beer law limits. The low values of the relative standard deviation (R.S.D.) indicate good precision of the methods. To study the accuracy of the methods, recovery studies were carried out by the standard addition method. For this, known quantities of pure TRH were mixed with definite amounts of pre-analyzed formulations and the mixtures were analyzed as before. The total amount of the drug was then determined and the amount of the added drug was calculated by difference. The results are given in Table II. The average percent recoveries obtained were quantitative (99.23 ± 0.66 to 99.38 ± 0.84 %), indicating good accuracy of the methods.

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TABLE II. Analysis of tablet, recovery and ruggedness of the assay of TRH by the proposed methods and their comparison with the Official method2 Sample

Drug Found* ± SD; % and their comparison with Official method present Official method BCG method MO method (mg)

Commercial tablet

100

100.4 ± 0.78

99.49 ± 0.55 F = 2.01, t = 1.49

99.61 ± 0.81 F = 1.07, t = 1.97

Recovery

100



99.38 ± 0.84

99.23 ± 0.66

Between-day analysis

100



99.12 ± 0.96

100.11 ± 0.79

Within-day analysis

100



99.58 ± 0.67

99.34 ± 0.82

* Average of six determinations

Interference studies The effects of common excipients and additives were tested for their possible interferences in the assay of TRH. It was observed that talc, glucose, starch, lacotose, dextrose, gum acacia and magnesium stearate did not interfere in the determination at the levels normally found in dosage forms. Ruggedness To ascertain the ruggedness of the methods, four replicate determinations at different concentration levels of the drugs were carried out. The within-day RSD values were less than 1 %. The values of between-day RSD for different concentrations of drugs obtained from the determinations are given in Table II, and indicate that the proposed method has reasonable ruggedness. Analysis of pharmaceutical formulations, and statistical comparison of the results with an Official method2 The proposed methods were successfully applied to the analysis of TRH in commercial tablets. The results of analysis of pharmaceutical formulations (Table II) were compared statistically by the Student t-test and by the variance ratio F-test with those obtained by the Official method. The Student t-values at 95 % confidence level did not exceed the theoretical value, indicating that there was no significant difference between the proposed and the Official method. It was also observed that the variance ratio F-values calculated for p = 0.05 did not exceed the theoretical value, indicating that there was no significant difference between the precision of the proposed and the Official method. CONCLUSION

Unlike gas chromatographic and HPLC procedures, the instrumentation is simple and not of high cost. The importance lies in the chemical reactions upon which the procedures are based rather than upon the sophistication of the instrument. This aspect of spectrophotometric analysis is of major interest in analytical

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pharmacy since it offers a distinct possibility in the assay of a particular component in complex dosage formulations. The reagents utilized in the proposed methods are cheap, readily available and the procedures do not involve any critical reaction conditions or tedious sample preparation. The method is unaffected by slight variations in the experimental conditions, such as pH, reagent concentration. Moreover, the methods are free from interference by common additives and excipients. The wide applicability of the new procedures for routine quality control was well established by the assay of TRH in pure form and in pharmaceutical preparations. Acknowledgements: The authors express their sincere thanks to Protec, Mumbai, India, for supplying the gift sample of trazodone hydrochloride. Thanks are also due to the authorities of Karnatak University, Dharwad, for providing the necessary facilities.

IZVOD

OSETQIVA SPEKTROFOTOMETRIJSKA METODA ZA ODRE\IVAWE ^ISTOG TRAZODON HIDROHLORIDA U FARMACEUTSKIM PREPARATIMA K. HARIKRISHNA, R. SUDHIR KUMAR, J. SEETHARAMAPPA i H. MANJUNATHA Department of Chemistry, Karnatak University, Dharwad-580 003, India

Predlo`ene su dve jednostavne, brze i osetqive spektrofotometrijske metode, pogodne u kontroli kvaliteta, za odre|ivawe trazodon hidrohlorida (TRH), ~istog i u farmaceutskim preparatima. Metode se zasnivaju na formirawu, u hloroformu rastvornog, kompleksa TRH sa bromkrezol zelenim (BCG) i metiloran`om (MO) u KCl–HCl puferu pH 1,5 (BCG) i u NaOAc–HCl puferu pH 3,92 (za MO), sa apsorpcionim maksimumom na 415 nm za BCG odnosno na 422 nm za MO. Uslovi reakcije su optimizovani u ciqu dobijawa boje maksimalnog intenziteta. Na|eno je da apsorbancija raste sa pove}awem koncentracije TRH. Sistem sledi Lambert–Beer-ov zakon o upsegu od 0,9 – 17 mg/mL za BCG odn. 1 – 20 mg/mL za MO. Evaluirani su i validirani analiti~ki parametric od zna~aja. Nisu registrovane smetwe sa uobi~ajenim matriksima kori{}enim u farmaceutskim preparatima. (Primqeno 20. jula. revidirano 25. oktobra 2005)

REFERENCES 1. British Pharmacopoeia, Her Majesty’s Stationery Office London, 1998, p. 1318 2. The United States Pharmacopoeia, 24 revision, Asian Edition, United States Pharmacopeial Convention, Inc., Twinbrook Parkway, Rockville, MD, 2000, p. 1681 3. S. N. Dhumal, P. M. Dikshit, I. I. Ubharay, B. M. Mascarenhas, C. D. Gaitonde, Indian Drugs 28 (1991) 565 4. S. Khalil, Analyst 124 (1999) 139 5. H. Suzuki, K. Akimoto, H. Nakagawa, I. Sugimoto, J. Pharmacol. Sci. 78 (1989) 62 6. D. Dogrukol-Ak, V. Zaimoglu, M. Tuncel, Eur. J. Pharmacol. Sci. 7 (1999) 215 7. J. M. Kauffmann, J. C. Vire, G. J. Patriarche, L. J. Nunez-Vergara, J. A. Squella, Electrochim. Acta 32 (1987) 1159 8. R. T. Sane, V. R. Nurerkar, R. V.Tendolkar, D. P. Gangal, P. S. Mainkar, S. N. Dhumal, Indian Drugs 27 (1990) 251 9. T. Ohkubo, T. Osanai, K. Sugawara, M. Ishida, K. Otani, K. Mihara, N. Yasui, J. Pharm.

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Pharmacol. 47 (1995) 340 10. G. T. Vatassery, L. A. Holden, D. K. Hazel, M. W. Dysken, Clin. Biochem. 30 (1997) 149 11. O. Andriollo, C. Lartigue-Mattei, J. L. Chabard, H. Bargnoux, J. Petit, J. A. Berger, J. F. Pognat, J. Chromatogr. 575 (1992) 301 12. R. E. Gammans, E. H. Kerns, W. W. Bullen, R. R. Covington, J. W. Russell, J. Chromatogr. 339 (1985) 303 13. T. J. Siek, J. Anal. Toxicol. 11 (1987) 225 14. A. El-Gindy, B. El-Zeany, T. Awad, M. M. Shabana, J. Pharm. Biomed. Anal. 26 (2001) 211 15. Julie Milano, Simone Goncalves Cardoso, J. Pharm. Biomed. Anal. 37 (2005) 639 16. E. Regulska, M. Tasaseiwicz, H. Puzunowska-Taraseiwicz, J. Pharm. Biomed. Anal. 29 (2002) 335 17. B. G. Gowda, M. B. Melwanki, J. Seetharamappa, J. Pharm. Biomed. Anal. 25 (2001) 1021 18. P. Job, Ann. Chim. France. 9 (1928) 19. R. T. Foley, R. C. Anderson, J. Am. Chem. Soc. 70 (1948) 1195 20. A. K. Mukherji, A. K. Dey, Anal. Chim. Acta 18 (1958) 324 21. Analytical Methods committee, Analyst 112 (1987) p. 199.