Development and Validation of a HPLC Method for Determination of Pefloxacin in Tablet and Human Plasma

Document Type : Original Article

Authors

Department of Pharmaceutics, Faculty of Pharmacy, University of Karachi, Karachi, Pakistan

Abstract

Objective(s)
Developing and validating a simple, efficient, reproducible and economic reversed phase high performance liquid chromatographic (RP-HPLC) method for the quantitative determination of pefloxacin in bulk material, tablets and in human plasma.
Materials and Methods
A shim-pack CLC-ODS column and a mobile phase constituting acetonitrile: 0.025 M phosphoric acid solution (13:87 v/v, pH 2.9 adjusted with KOH) were used. The flow rate was 1 ml/min and the analyses performed using ultraviolet (UV) detector at a wavelength of 275 nm using acetaminophen as an internal standard.
Results
The developed method showed good resolution between pefloxacin and acetaminophen. It was selective to pefloxacin and able to resolve the drug peak from internal standard and from formulation excipients. The percentage of coefficient variation (CV) of the retention times and peak areas of pefloxacin from the six consecutive injections were 0.566% and 0.989%, respectively. The results showed that the peak area responses are linear within the concentration range of 0.125 qg/ml-12 qg/ml (R2= 0.9987). The limits of detection (LOD) and limits of quantitation (LOQ) for pefloxacin were 0.03125 qg/ml and 0.125 qg/ml. The intra-day and inter-day variation, RSD were 0.376-0.9056 and 0.739-0.853 respectively; also, inter-day variation with relative standard deviation (RSD) were 0.1465-0.821 in plasma. The accuracy results of 70%, 100%, and 130% drugs were 100.72%, 100.34%, and 100.09%, respectively.
Conclusion
The method is linear, quantitative, reproducible and could be used as a more convenient, efficient and economical method for the trace analysis of drug in biological fluids, in raw material and tablets.

Keywords


1.Mitsuhashi S. Comparative antibacterial activity of new quinolone-carboxylic acid derivatives. Rev Infect Dis 1989; 10:S27-31.
2.Yourassowsky E, Van der Linden MP, Crokaert F, Gluyczynski Y. In vitro activity of pefloxacin compared to other antibiotics. J Antimicrob Chemother 1986; 17:S19-28.
3.Frydman AM, LeRoux Y, Lefevre MA, Djebbar F, Fourtillan JB, Faillot J. Pharmacokinetics of pefloxacin after repeated intravenous and oral administration (400 mg bid) in young healthy volunteers. J Antimicrob Chemother 1986; 17:65-79.
4.Gerding DN, Hitt JA. Tissue penetration of the new quinolones in humans. Rev Infect Dis 1989; 11:S1046-1057.
5.Webster A, Gaya H. Quinolones in the treatment of serious infections. Rev Infect Dis 1989; 10: S225-230.
6.Lauwers S, Vincken W, Naessens A, Peirard D. Efficacy and safety of pefloxacin in the treatment of severe infections in patients hospitalized in intensive care units. J Antimicrob Chemother 1986; 17:111-115.
7.Raghow G, Meyer MC. High-performance liquid chromatographic assay of tolbutamide & carboxytolbutamide in human plasma. J Pharm Sci 1981;70:1166-1167.
8.Ni Y, Wang Y, Kokot S. Multicomponent kinetic spectrophotometric determination of pefloxacin and norfloxacin in pharmaceutical preparations and human plasma samples with the aid of chemometrics. Spectrochim Acta A Mol Biomol Spectrosc 2008;70:1049-1059.
9.Basavaiah K, Prameela HC, Somashekar BC. Spectrophotometric determination of pefloxacin mesylate in pharmaceuticals. Acta Pharm 2007; 57:221-230.
10.Mostafa S, El-Sadek M, Alla EA.Spectrophotometric determination of enrofloxacin and pefloxacin through ion- pair complex formation. J Pharm Biomed Anal 2002; 28:173-180.
11.Sun SW, Chen LY. Quantitative determination of gatifloxacin, levofloxacin, lomefloxacin and pefloxacin. J Chromatogr A 1997; 766:215-224.
12.Fierens C, Hillaert S, Van den Bossche W. The qualitative and quantitative determination of quinolones of first and second generation by capillary electrophoresis. J Pharm Biomed Anal 2000; 22:763-772.
13.Flurer CL. Analysis of antibiotics by capillary electrophoresis. Electrophoresis 1997; 18:2427.
14.Santoro MI, Kassab NM, Singh AK, Kedor-Hackmam ER. Quantitative determination of gatifloxacin, levofloxacin, lomefloxacin and pefloxacin fluoroquinolonic antibiotics in pharmaceutical preparations by high- performance liquid chromatography. J Pharm Biomed Anal 2006; 40:179-184.
15.Samanidou VF, Demetriou CE, Papadoyannis IN. Development of a validated HPLC method for the determination of four penicillin antibiotics in pharmaceuticals and human biological fluids. Anal Bioanal Chem 2003; 375:623-629.
16.Joshi S. Quantitative determination of ciprofloxacin and norfloxacin in pharmaceutical preparations by high performance liquid chromatography. J Pharm Biomed Anal 2002; 28:795-809. 
17.Sanzgiri D, Knaub SR, Riley CM. Quantitative determination of gatifloxacin, levofloxacin, lomefloxacin and pefloxacin fluoroquinolonic antibiotics in pharmaceutical preparations by high-performance liquid chromatography. Anal Profiles Drug Subst Excipients 1994; 23:325-369.
18.Groeneveld AJ, Brouwers JR. Quantitative determination of ofloxacin, ciprofloxacin, norfloxacin and pefloxacin in serum by high performance liquid chromatography. Pharm Weekbl Sci 1986; 8:79-84.
19.Montay G, Blain Y, Roquet F, Le Hir A. High-performance liquid chromatography of pefloxacin and its main active metabolites in biological fluids. J Chromatogr 1983; 272:359-365.
20.Beltagi AM. Determination of the antibiotic drug pefloxacin in bulk form, tablets and human serum using square wave cathodic adsorptive stripping voltammetry. J Pharm Biomed Anal 2003; 31:1079-1088.
21.Wang PL, Chen L, Fan YF. Selective separation and simultaneous determination of trace levels of five types of fluorinated quinolone drugs by thin-layer chromatography /fluorescence densitometry. J AO AC Int 2001; 84:684-688.
22.Fratini L, Schapoval EES. In sensitive spectrophotometer methods for the determination of amoxycillin, ciprofloxacin. Int J Pharm 1996; 127:279-282.
23. Jelikic-Stankov M, Veselinovic D, Malesev D, Radovic Z. Spectrophotometric determination of pefloxacin in pharmaceutical preparations. J Pharm Biomed Anal 1989; 7:1571-1577.
24.Belal F, Al-Majed AA,Al-Obaid AM. Methods of analysis of 4-quinolone antibacterials. Talanta 1999; 50:765¬786.
25.United States Pharmacopeia.28th ed. United States Pharmacopeial Convention, Rockville: 2005.
26.Global Quality Guideline, Validation of analytical procedures 2002; Number: G-6.9, Version 1.0.
27.ICH Q2B, Guidance for Industry, Validation of analytical procedures-methodology, 1996.
28.Reviewer guidance validation of chromatographic methods, Center for drug evaluation and research, food and drug administration 1994.
29.AL-Obaidy, Po SS, McKiernan ALWPJ, Glasgow JFT, Millership J. Assay of paracetamol and its metabolites in urine, plasma and saliva of children with chronic liver disease. J Pharm Biomed Anal 1995; 13:1033-1039.
30.United States Pharmacopeia, XXV11. Chromatography, 2004.
31.Mohammad Y, Gunawan I. Validation of chromatographic methods of analysis. Profiles of drug substances, excipients, and related methodology, Vol 32, 2005.
32.Garofolo F. Bioanalytical method validation, In: Chan CC, Lam H, Lee YC, Zhang XU, editors. Analytical method. Validation and instrument performance verification. Wiley-Interscience; 2004. p.105-138.
33.Hartmann C, Smeyers-Verbeke J, Massart Dl, McDowall RD. Validation of bioanalytical chromatographic methods. J Pharm Biomed Anal 1998; 17:193-218.
34.Guidances for Industry, Bioanalytical method validation, US department of health services, food and drug administration, center for drug evaluation and research, center for veterinary medicine.
35.Dadgar D, Burnet PE, Choc MG, Gallicano K, Hooper JW. Application issues in bioanalytical method validation, sample analysis and data reporting. J Pharm Biomed Anal 1995; 13:89-97.
36.Abanmi N, Zaghloul I, EI Sayed N, Al-Khamis KI. Determination of pefloxacin and its main active metabolite in human serum by high performance liquid chromatography. Ther Drug Monit 1996; 18:158-163.
37.Wilson, Gisvold’s. Textbook of organic and medicinal and pharmaceutical chemistry.10th ed. 1998.
38.Brkich GE, Arzamastsev AP, Kaz'mina EM, Mikhalev AV, HPLC determination of pefloxacin and ciprofloxacin in blood serum, Pharm Chem J 1999; 33:219-221.