Journal of Chromatography B, 810 (2004) 313–318 Bioanalysis of tobramycin for therapeutic drug monitoring by solid-phase extraction and capillary zone electrophoresis Humphrey Fonge a , Eliangiringa Kaale a,1 , Cindy Govaerts a , Koenraad Desmet b , Ann Van Schepdael a,∗ , Jos Hoogmartens a a Laboratory of Pharmaceutical Chemistry and Drug Analysis, K.U. Leuven, E. Van Evenstraat 4, B-3000 Leuven, Belgium b Department of Clinical Pathology, UZ-K.U. Leuven Gasthuisberg, Herestraat 49, B-3000 Leuven, Belgium Received 27 May 2004; accepted 16 August 2004 Abstract A method based on solid-phase extraction (SPE) and capillary zone electrophoresis (CZE) for the analysis of tobramycin in human serum is presented. An off-line SPE employing a carboxypropyl bonded phase (CBA) cartridge was used for the extraction of tobramycin from human serum. Adsorbed tobramycin was eluted from the CBA cartridge using a mixture of NH 3 (25%, w/v)–methanol (30:70, v/v). After evaporation, the analyte was reconstituted and derivatized with o-phthaldialdehyde (OPA)/3-mercaptopropionic acid (MPA). The resulting tobramycin–OPA/MPA derivative was purified, and then identified by mass spectrometry. The tobramycin–OPA/MPA derivative was then analysed by CZE with a background electrolyte (BGE) comprising of 30 mM sodium tetraborate pH 10.0-acetonitrile (ACN) (80:20, v/v) with ultraviolet detection at 230 nm. A linear response was observed in the range of 0.3–30 g/ml with r 2 = 0.992. The sensitivity of the method was determined by its limit of quantitation (LOQ) and limit of detection (LOD) of 0.3 g/ml and 0.1 g/ml, respectively. SPE recovery ranged from 68 to 79% at the trough levels to 98% at the peak levels found in serum. Furosemide has been added as internal standard (IS) to improve precision. For the therapeutic range of tobramycin in serum (2–10 g/ml) the relative standard deviation (R.S.D.) was less than 11% for the entire SPE/CE process. The method demonstrated excellent selectivity as shown by the lack of interference from a total of 20 drugs investigated. The method was then used in therapeutic drug monitoring of patients receiving the drug. © 2004 Elsevier B.V. All rights reserved. Keywords: Tobramycin; Bioanalysis; Solid-phase extraction 1. Introduction Tobramycin is a broad spectrum aminoglycoside antibi- otic produced by Streptomycestenebrarius [1]. It is indicated for the treatment of aerobic Gram-positive and some aerobic Gram-negative bacteria for which less toxic antibiotic are ineffective or contra-indicated. Like all other aminoglyco- sides, its narrow therapeutic index is implicated by its oto- ∗ Corresponding author. Tel.: +32 16 323443; fax: +32 16 323448. E-mail address: ann.vanschepdael@pharm.kuleuven.ac.be (A. Van Schepdael). 1 Current address: Department of Medicinal Chemistry, School of Phar- macy, MUCHS, P.O. Box 65013, Dar Es Salaam, Tanzania. and nephrotoxicity especially when therapy is prolonged [2]. Careful monitoring of this substance is therefore required in order to detect elevated and appropriate levels in serum for therapy so as to improve efficacy. Quantitative methods currently used for the analy- sis of tobramycin in biological matrices [3,4] include radioimmunoassay (RIA) [5], high-performance liquid- chromatography (HPLC) [6–10], fluorescence polarization immunoassay (FPIA) [11], microbiological assays [5,12], radiochemical assay [13] and enzyme immunoassay (EIA) [14,15]. These methods each have merits and demerits. The use of microbiological assay (as described by United States Pharmacopeia Drug Information (USPDI) [16]) for therapeutic drug monitoring and pharmacokinetic studies of 1570-0232/$ – see front matter © 2004 Elsevier B.V. All rights reserved. doi:10.1016/j.jchromb.2004.08.020