SIMULTANEOUS SPECTROPHOTOMETRIC ESTIMATION OF CIPROFLOXACIN AND ORNIDAZOLE IN TABLET DOSAGE FORM

SIMULTANEOUS SPECTROPHOTOMETRIC ESTIMATION OF CIPROFLOXACIN AND ORNIDAZOLE IN TABLET DOSAGE FORM

A.S. Grewal*¹, S.K. Patro ², S.K. Kanungo ² and S.K. Bhardwaj ¹

JCDM College of Pharmacy ¹, Sirsa- 125 055, Haryana, India

Institute of Pharmacy & Technology ², Salipur- 754 202, Cuttack, Orissa, India

ABSTRACT

Three simple sensitive, rapid, accurate and economical methods were developed for simultaneous estimation of Ciprofloxacin & Ornidazole in bulk and its pharmaceutical formulations. First method was based on the simultaneous equation, second method was based on Q-Analysis (Absorbance ratio method) and third method was based on multicomponent mode. The absorption maxima for Ciprofloxacin and Ornidazole were 272 nm and 319 nm in 0.1 N NaOH respectively. The iso-absorptive point for Ciprofloxacin Hydrochloride and Ornidazole was 292 nm. The calibration curve was linear over the concentration range 2.5 to 22.5 µg/ml of Ciprofloxacin & Ornidazole. The methods had been validated statistically and by recovery study. It can be used in the simultaneous estimation of Ciprofloxacin and Ornidazole in bulk drugs and its pharmaceutical preparations in a routine manner.

Validation

INTRODUCTION: Ciprofloxacin, chemically, 1-cyclopropyl-6-fluoro-1, 4- dihydro-4-oxo- 7 (piperazin-1-yl) quinoline-3-carboxylic acid1; belongs to the group of synthetic fluoroquinolone antibiotics with broad antimicrobial activity. Literature review reveals spectrophotometry ^1-11, titrimetry ¹², HPLC ^13-18 and HPTLC ¹⁹ for its determination. Ornidazole, chemically, 1-chloro-3-(2-methyl-5-nitro-1H-imidazol-1-yl) propan-2-ol, is an antimicrobial agent.

Ornidazole is used in the treatment of amoebiasis and other protozoal diseases. This drug is under the category antihelmentics. The combination of two drugs is not official in any pharmacopoeia; hence no official method is available for the estimation of ciprofloxacin and ornidazole in their combined dosage form. Literature review reveals that few methods are available for the determination of ornidazole in bulk and solid dosage form based on spectrophotometry ^20-27 and RP-HPLC.

Ciprofloxacin and ornidazole in combined tablet dosage form is available in the market, has gained increasing acceptance in bacterial and protozoal infections. Spectrophotometric ^28-29 and HPLC ³⁰ method have been established for their simultaneous estimation in tablet dosage form. This paper presents three simple, sensitive, accurate and reproducible spectrophotometric methods for simultaneous determination of ciprofloxacin and ornidazole in tablet dosage form.

FIGURE 1: STRUCTURE OF CIPROFLOXACIN AND ORNIDAZOLE

MATERIALS AND METHODS:

Instrument used: A Shimadzu UV-Vis Double beam spectrophotometer (Pharmaspec-1700) with matched quartz cells was used for all spectral measurements.

Materials: The tablets of the said combination were purchased from a local pharmacy (The label claim contained 500 mg of Ciprofloxacin and 500 mg of Ornidazole). Sodium hydroxide was of analytical grade chemical and distilled water was used to prepare all solutions.

Preparation of 0.1N Sodium Hydroxide Solution: 0.1 N NaOH was prepared by dissolving 4 g of sodium hydroxide in sufficient distilled water to produce 1000 ml.

Preparation of Standard Solutions: The standard stock solution of Ciprofloxacin and Ornidazole were prepared by dissolving 50 mg of each drug in 50 ml of 0.1 N NaOH. Stock solutions of Ciprofloxacin and Ornidazole were further diluted in 0.1 N NaOH to get working standard solutions of concentration 100 µg/ml.

Method-I: Simultaneous equation method (Vierodt’s method): If a sample contains two absorbing drugs (X and Y) each of which absorbs at the I_max of the other, it may be possible to determine both drugs by the technique of simultaneous equation (Vierodt’s method) provided that certain criteria apply.

a)      Study of Beer’s-Lamberts law: The standard solutions of Ciprofloxacin Hydrochloride (20 µg/ml) and Ornidazole (20 µg/ml) in 0.1 N NaOH were scanned in the entire UV range to determine λ_max of both the drugs. The λ_max of Ciprofloxacin hydrochloride and Ornidazole were found to be 272 nm and 319 nm, respectively. A series of standard solutions were prepared having concentration range of 2.5 to 22.5 µg/ml were prepared in 0.1 N NaOH using working standard solution. The absorbance of resulting solution was measured at 278 nm and 319 nm for both Ciprofloxacin HCl and Ornidazole, and calibration curves plotted at these wavelengths. Both the drugs obeyed linearity individually within the concentration range 2.5 - 22.5µg/ml. The overlay UV spectrum of Ciprofloxacin Hydrochloride and Ornidazole is shown in Fig. 2.

FIGURE 2: THE OVERLAY UV SPECTRUM OF CIPROFLOXACIN HYDROCHLORIDE AND ORNIDAZOLE

b)      Estimation of Absorptivity (E 1%, 1cm) values at selected wavelengths: The Absorptivity (E 1%, 1cm value) of Ciprofloxacin HCl and Ornidazole drugs were calculated at 272 nm and 319 nm. Two simultaneous equations were formed using these values as given below.

C_X = 0.0145714 A₁ – 0.0050635 A₂ ………….. (1)

C_Y = 0.039518 A₂ – 0.0145311 A₁ ………….... (2)

Where, A₁ and A₂ are the absorbance of sample at 272 nm and 319 nm respectively. C_X and C_Y are the concentration of Ciprofloxacin HCl and Ornidazole in sample respectively.

c)      Analysis of standard mixture: The mixture of pure Ciprofloxacin HCl (10 µg/mL) and Ornidazole (10 µg/mL) was prepared from stock solution in 0.1 N NaOH. Their absorbance value at the two selected wavelength was recorded and quantitative estimation of the drugs was carried out by solving simultaneous equations.

d)     Analysis of Tablet Formulation: Twenty tablets were weighed and finely powdered. An accurately weighed quantity of the powder equivalent to about 50 mg of Ciprofloxacin & Ornidazole; was taken in 50 mL volumetric flask and dissolved in 25 mL of 0.1 N NaOH ; it was further diluted up to the mark with same solvent. The solutions were then filtered and filtrate gets diluted to get final 10 µg/mL concentration of Ciprofloxacin HCl & Ornidazole. The solution was then read at two selected wavelength. The results of tablet analysis are given in Table 1.

Method-II: Q - Analysis (Absorbance Ratio Method): Q-Absorbance method depends on the property that, for a substance which obeys Beer's law at all wavelength, the ratio of absorbances at any two wavelengths is a constant value independent of concentration or path length. In the quantitative assay of two components in a mixture by the absorbance ratio method, absorbances are measured at two wavelengths: One being the λ_max of one of the component (λ₂) and the other being a wavelength of equal absorptivities of the two components i.e. an iso-absorptive point.

a)      Determination of Iso-absorptive point and selection of suitable Wavelength: An iso-absorptive point (a wavelength of equal absorptivity of the two components) was determined by taking overlain spectrum of the solutions ciprofloxacin HCl and Ornidazole (20 µg/ml each) in 0.1 N NaOH in UV range against the solvent blank. From the overlain spectra of the two drugs, it was found that ciprofloxacin showed λ_max at 272 nm and ornidazole showed λ_max at 319 nm. Iso-absorptive point was found out at 292 nm. The two wavelengths 272 nm at which ciprofloxacin exhibits maximum absorption and 292 nm as iso-absorptive point were selected for estimation of drugs simultaneously.

b)     Study of Beer-Lambert’s law: The solutions having concentrations in range 2.5 - 22.5 µg/ml for both ciprofloxacin HCl and ornidazole were prepared in 0.1 N NaOH using working standard solution. The absorbances of resulting solutions were measured at 272 nm and 292 nm. Calibration curves were plotted at these wavelengths. Both the drugs obeyed linearity individually and combination within the concentration range of 2.5 - 22.5 µg/ml for both ciprofloxacin & Ornidazole.

c)      Estimation of Absorptivity (E 1%, 1cm) values at Selected Wavelengths: The Absorptivity (E 1%, 1cm value) of Ciprofloxacin HCl and Ornidazole drugs were calculated at 272 nm and 319 nm.

d)      Analysis of Standard Mixture: The laboratory mixture of Ofloxacin (5 µg/mL) and ornidazole (12 µg/mL) was prepared from stock solution in 0.5 N acetic acid The absorbance value at the two selected wavelength was recorded The contents of Ciprofloxacin and Ornidazole were calculated by substituting values in the formulae given below.

C_x = (Q_M - Q_Y) A₁ / (Q_X - Q_Y) a_x1

C_y = (Q_M – Q_x) A₁ / (Q_y – Q_x) a_x1

Where, C_x=concentration of Ciprofloxacin; C_y=concentration of Ornidazole; Q_x= ratio of absorptivity of ciprofloxacin at 292 nm and 272 nm; Q_y= ratio of absorptivity of Ornidazole at 292 nm and 272; Q_M= absorbance ratio of mixture at 292 nm and 272 nm; a_x1 and a_y1=absorptivity of pure Ciprofloxacin and Ornidazole respectively at iso-absorptive wavelength.; A₁=absorbance of mixture at iso-absorptive point.

e)      Analysis of tablet formulation: Twenty tablets were weighed and finely powdered. An accurately weighed quantity of the powder equivalent to about 50 mg of ciprofloxacin was taken in 50 ml volumetric flask and dissolved in 25 ml of 0.1 N NaOH; it was diluted up to the mark with 0.1 N NaOH. The solutions were then filtered and filtrate gets diluted to get final concentration of ciprofloxacin HCl (10 µg/ml) and Ornidazole (10 µg/ml). The solution was then read at two selected wavelength. The results of tablet analysis are given in Table 1.

Method-III: Multi-component mode: From the overlain spectra of the two drugs, the two wavelengths selected as sampling wavelengths were 272 and 319 nm which are absorbance maxima for Ciprofloxacin HCL and Ornidazole respectively. Now the sampling wavelengths and concentrations of the two components in each of the mixed standards were fed to the instrument using multi-component mode of the instrument and all the seven mixed standards were scanned in the range of 400 to 200nm. The instrument collects and compiles the spectral data from the mixed standards and gets ready for the quantitative analysis of the samples.

Validation of Analytical Methods: As per I.C.H. guidelines.

Linearity and range: 2.5 – 22.5 µg/ml

Precision: For evaluation of precision of the results a concentration of 10μg/ml was evaluated by 4 replicate determinations. S.D. was found to be less than 0.1. Precision study data is given in Table 2.

Accuracy (Recovery study): It was found out by recovery study using standard addition method. Known amounts of standard Ciprofloxacin and Ornidazole was added to pre-analyzed samples at a level from 80% up to 120% and then subjected to the proposed methods. Results of recovery studies are shown in Table 3.

TABLE 1: ANALYSIS OF CIPROFLOXACIN AND ORNIDAZOLE IN TABLETS

*Average of five determinations, CIP-Ciprofloxacin, ORD-Ornidazole

TABLE 2: PRECISION DATA OF CIPROFLOXACIN AND ORNIDAZOLE

TABLE 3: RESULT OF RECOVERY STUDY 

*Average of five determinations, CIP-Ciprofloxacin, ORD-Ornidazole

RESULTS AND DISCUSSION: The proposed methods were found to be simple, accurate and reproducible for routine simultaneous estimation of ciprofloxacin and ornidazole in combination. In 0.1 N sodium hydroxide solution, ciprofloxacin and ornidazole showed λ_max at 272 nm and 319 nm respectively.

As their λ_max differ more than 20 nm; simultaneous equation method tried for their simultaneous estimation in formulation. Also iso-absorptive point was observed at 292 nm so the absorbance ratio method was tried for simultaneous estimation of drugs in formulation at 272 nm (λ_max of ciprofloxacin) and 292 nm (iso-absorptive wavelength). The calibration curve was linear over the concentration range 2.5 to 22.5µg/ml of Ciprofloxacin & Ornidazole. By observing the validation parameters, all the methods were found to be simple, specific, accurate and precise. Recovery experiments indicated the absence of interferences from the commonly encountered pharmaceutical additives and excipients.

All the methods were successfully used to determine the amounts of ciprofloxacin and ornidazole present in the tablets. The results obtained were in good agreement with the corresponding labelled amount. Hence, all the three methods can be employed for the routine analysis of these two drugs in combinations.

REFERENCES:

1. El-Brashy AM, Metwally ME and El-Sepai FA: Spectrophotometric determination of some fluoroquinolone antibacterials through charge-transfer and ion-pair complexation reactions. Bulletin of the Korean Chemical Society 2004; 25(3): 365-372.
2. Sharma R, Pathodiya G, Mishra GP and Sainy J: A novel spectrophotometric method for quantitative determination of ciprofloxacin hydrochloride and tinidazole in tablets using hydrotropic solubilizing agent. Journal of Pharmacy Research 2011; 4(3): 859-861.
3. Mostafa S, El-Sadek M and Alla EA: Spectrophotometric determination of ciprofloxacin, enrofloxacin and pefloxacin through charge transfer complex formation. Journal of Pharmaceutical and Biomedical Analysis 2002; 27(1-2): 133-142.
4. Mahrouse MA and Elkady EF: Validated spectrophotometric methods for the simultaneous determination of ciprofloxacin hydrochloride and metronidazole in tablets. Chemical and Pharmaceutical Bulletin 2011; 59(12): 1485-1493.
5. Djurdjevic P, Todorovic M, Stankov MJ and Odovic J: Spectrophotometry determination of ciprofloxacin in serum using iron(III) ion as chromogenic agent. Analytical Letters 2000; 33(4): 657-665.
6. Fratini L and Schapoval ES: Ciprofloxacin determination by visible light spectrophotometry using iron(III)nitrate. International Journal of Pharmaceutics 1996; 127(2): 279-282.
7. Diao YH: Determination of ciprofloxacin hydrochloride with UV method and quality investigation of its tablets. Chinese Journal of Hospital Pharmacy 1994; 14: 212-214.
8. Nijhu RS, Jhanker YM and Sutradhar KB: Development of an Assay Method for Simultaneous Determination of Ciprofloxacin and Naproxen by UV Spectrophotometric Method. Stamford Journal of Pharmaceutical Sciences 2011; 4(1): 84-90.
9. Abdel-Gawad FM, Issa YM, Fahmy HM and Hussein HM: Spectrophotometric determination of ciprofloxacin in pure form and in tablets through charge-transfer complexation reactions. Microchimica Acta 1998; 130(1-2): 35-40.
10. Pascual-Reguera MI, Parras GP and Diaz AM: Solid-phase UV spectrophotometric method for determination of ciprofloxacin. Microchemical Journal 2004; 77(1): 79-84.
11. Rizk M, Belal F, Ibrahim F, Ahmed S and Sheribah ZA: Derivative spectrophotometric analysis of 4-Quinolone antibacterials in formulations and spiked biological fluids by their cu (II) complexes; Journal of AOAC International 2001; 84(2): 368-375.
12. Basavaiah K, Nagegowda P, Chikkaswamy B, Somashekar and Ramakrishna V: Spectrophotometric and titrimetric determination of ciprofloxacin based on reaction with cerium (IV) sulphate. Science Asia 2006; 32: 403-409.
13. Kamberi M, Tsutsumi K, Kotegawa T, Nakamura K and Nakano S: Determination of ciprofloxacin in plasma and urine by HPLC with ultraviolet detection. Clinical Chemistry 1998; 44(6): 1251-1255.
14. Dincel A, Yildirim A, Caglayan F and Bozkurt A: Determination of ciprofloxacin in human gingival crevicular fluid by high-performance liquid chromatography. Acta Chromatography 2005; 15: 308-314.
15. Wu SS, Chein CY and Wen YH: Analysis of ciprofloxacin by a simple high-performance liquid chromatography method. Journal of Chromatographic Science 2008; 46(6): 490-495.
16. Krol GJ, Beck GW and Benham T: HPLC analysis of ciprofloxacin and ciprofloxacin metabolites in body fluids. Journal of Pharmaceutical and Biomedical Analysis 1995; 14(1-2): 181-190.
17. Khan A, Zafar I, Jamshaid AK, Ghulam SK, Bilal M and Khan TM: The Development and Validation of HPLC-UV method for analysis of ciprofloxacin in serum and aqueous humour. Archives of Pharmacy Practice 2011; 2(3): 116-122.
18. Witte BD, Dewulf J, Demeestere K, Ruyck MD and Langenhove HV: Critical points in the analysis of ciprofloxacin by high-performance liquid chromatography. Journal of Chromatography A 2007; 1140(1-2): 126-130.
19. Singh BK, Parwate DV, Srivastava S and Shukla SK: High performance thin-layer chromatographic selective and stability indicating method for assay of ciprofloxacin in pharmaceuticals. Der Pharma Chemica 2010; 2(4): 178-188.
20. Mubeen G, Prakash V, Somashekar PL and Uvesh K: Spectrophotometric method for determination of ornidazole, International Journal of ChemTech Research 2009; 1(2): 318-321.
21. Maheshwari RK, Srivastav VK, Prajapat RP, Jain A, Kamaria P and Sahu S: New spectrophotometric estimation of ornidazole tablets employing urea as a hydrotropic solubilizing additive. Indian Journal Pharmaceutical Sciences 2010; 72(2): 258-261.
22. Wankhede SB, Prakash A, Kumari B and Chitlange SS: Simultaneous spectrophotometric estimation of norfloxacin and ornidazole in tablet dosage form. Indian Journal of Pharmaceutical Sciences 2009; 71(3): 325-328.
23. Tulasamma P, Govind V and Venkateswarlu P: Spectrophotometric determination of ornidazole in pure and pharmaceutical formulations, International Journal of Pharmaceutical Sciences and Research 2011; 2(1): 44-48.
24. Nanda RK: Estimation of cefixime and ornidazole in its pharmaceutical dosage form by spectrophotometric method, Journal of Pharmacy Research 2009; 2(7): 1264-1266.
25. Maheshwari RK, Bishnoi SR, Kumar D and Muralikrishna: Quantitative spectrophotometric determination of ornidazole tablet formulations using ibuprofen sodium as hydrotropic solubilising agent. Digest Journal of Nanomaterials and Biostructures 2010; 5(1): 97-100.
26. Hemlata T, Pakhale HT, Wate SP, Nimje NM and Oswal RJ: Spectrophotometric methods for determination of ornidazole and gatifloxacin in tablet dosage form. The Pharma review 2009: 136.
27. Reddy TR, Dachinamoorthi D and Chandrasekhar KB: Spectrophotometric determination for nitro-imidazole derivative ornidazole. International Journal of Pharmaceutical Sciences Research 2010; 1(3): 199-202
28. Bhusari KP and Chaple DR: Simultaneous spectrophotometric estimation of ofloxacin and ornidazole in tablet dosage form. Asian Journal of Research in Chemistry 2009; 2(1): 60-62.
29. Patel SA, Patel NM and Patel MM: Simultaneous spectrophotometric estimation of ciprofloxacin and ornidazole in tablets. Indian Journal of Pharmaceutical Sciences 2006; 68(5): 665-667.
30. Singh R, Maithani M, Saraf SK, Saraf S and Gupta RC: Simultaneous estimation of ciprofloxacin hydrochloride, ofloxacin, tinidazole and ornidazole by RP-HPLC. Eurasian Journal of Analytical Chemistry 2009; 4(2): 161-167.
    

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    How to cite this article:

    Grewal AS, Patro SK, Kanungo SK and Bhardwaj SK: Simultaneous Spectrophotometric Estimation of Ciprofloxacin and Ornidazole in Tablet Dosage Form. Int J Pharm Sci Res 2012; Vol. 3(8): 2716-2720.

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