Journal of Pharmaceutical and Biomedical Analysis 29 (2002) 1089–1096 Validation of a RP-LC method for the simultaneous determination of isoniazid, pyrazinamide and rifampicin in a pharmaceutical formulation E. Calleri a, *, E. De Lorenzi a , S. Furlanetto b , G. Massolini a , G. Caccialanza a a Department of Pharmaceutical Chemistry, Uniersity of Paia, Via Taramelli 12, 27100 Paia, Italy b Department of Pharmaceutical Sciences, Uniersity of Florence, Via G. Capponi 9, 50121 Florence, Italy Received 11 July 2001; received in revised form 21 July 2001; accepted 1 August 2001 Abstract A simple and accurate liquid chromatographic method was developed and validated for estimation of isoniazid (ISN), pyrazinamide (PYR) and rifampicin (RIF) in combined dosage forms. Drugs were chromatographed on a reverse phase C18 column using a mobile phase gradient and monitored at the corresponding maximum of each compounds. Peaks were identified with retention time as compared with standards and confirmed with characteristic spectra using diode-array detector. Solution concentrations were measured on a weight basis to avoid the use of an internal standard. The method does not require any specific sample preparation except the use of a guard column. The method is linear (r 2 0.999), precise (RSD%: 0.50% for ISN, 0.12% for PYR and 0.98% for RIF), accurate (overall average recovery yields: 98.55% for ISN, 98.51 for PYR and 98.56% for RIF) and selective. Due to its simplicity and accuracy the method is suitable for routine quality control analysis of antitubercolosis combination dosage form. © 2002 Elsevier Science B.V. All rights reserved. Keywords: Isoniazid; Pyrazinamide; Rifampicin; Validation; High-performance liquid chromatography www.elsevier.com/locate/jpba 1. Introduction The tuberculosis treatment is an increasing worldwide problem due to the fact that the effec- tiveness of modern chemotherapy has been blunted by the high incidence of primary drug resistance, especially in developing countries. A 6-months regimen with rifampicin (RIF) and isoniazid (ISN), supplemented in the initial 2-months intensive phase by pyrazinamide (PYR), is highly effective. This modern short-course regimen can be simplified by the use of a fixed combination of the used drugs [1]. There are several fixed combination preparations containing ISN, PYR and RIF available on the world market and both the International Union Against Tuberculosis and Lung Disease (IU- ATLD) and the World Health Organisation (WHO) recognise the potential benefits of using * Corresponding author. Tel.: +39-382-507788; fax: +39- 382-422975 E-mail address: enrica.calleri@unipv.it (E. Calleri). 0731-7085/02/$ - see front matter © 2002 Elsevier Science B.V. All rights reserved. PII:S0731-7085(02)00150-4