A three-phase hollow fiber liquid-phase microextraction (HF-LPME) coupled either with capillary electrophoresis (CE) or high performance liquid chromatography (HPLC) with UV detection methods was successfully developed for the determination of trace levels of the anti-diabetic drug, rosiglitazone (ROSI) in biological fluids. The analyte was extracted into dihexyl ether that was immobilized in the wall pores of a porous hollow fiber from 10 mL of aqueous sample, pH 9.5 (donor phase), and was back extracted into the acceptor phase that contained 0.1M HCl located in the lumen of the hollow fiber. Parameters affecting the extraction process such as type of extraction solvent, HCl concentration, donor phase pH, extraction time, stirring speed, and salt addition were studied and optimized. Under the optimized conditions (extraction solvent, dihexyl ether; donor phase pH, 9.5; acceptor phase, 0.1M HCl; stirring speed, 600 rpm; extraction time, 30 min; without addition of salt), enrichment factor of 280 was obtained. Good linearity and correlation coefficients of the analyte was obtained over the concentration ranges of 1.0-500 and 5.0-500 ng mL(-1) for the HPLC (r(2)=0.9988) and CE (r(2)=0.9967) methods, respectively. The limits of detection (LOD) and limits of quantitation (LOQ) for the HPLC and CE methods were (0.18, 2.83) and (0.56, 5.00) ng mL(-1), respectively. The percent relative standard deviation (n=6) for the extraction and determination of three concentration levels (10, 250, 500 ng mL(-1)) of ROSI using the HPLC and CE methods were less than 10.9% and 13.2%, respectively. The developed methods are simple, rapid, sensitive and are suitable for the determination of trace amounts of ROSI in biological fluids.