In this study, a novel application of polyetheretherketone (PEEK) tubing for solid phase extraction (SPE) of urinary hydroxylated metabolites of polycyclic aromatic hydrocarbons (OH-PAHs) is presented. The use of PEEK tubing for extracting nine OH-PAHs (2-5 rings) from different matrixes (e.g., urine, acid/enzymatic hydrolysis solution) was demonstrated; a facile method for fast (<2 min) quantification of urinary 1-hydroxypyrene (1-OHPyr) was also developed by the use of PEEK tubing extraction coupled to electrospray ionization tandem mass spectrometry (ESI-MS/MS). Although no optimization was performed for the extraction process, a limit of detection (LOD) as low as 0.01 μg L(-1) was obtained, and the ratio of signal intensity of 1-OHPyr to that of 1-OHPyr-d9 (internal standard) was linearly related with the 1-OHPyr concentration over the range of 0.05-5.00 μg L(-1) (R(2) = 0.9995). Satisfactory recoveries (87-91%) were achieved, and less than 2 min was required to carry out the whole analytical procedure including sample pretreatment and mass spectrometric detection. In a biomonitoring study, the PEEK tubing extraction based method was successfully applied to the quantification of 1-OHPyr in eight human urine samples, further confirming the potential of PEEK tubing for SPE of organic compounds.