In-injection port thermal desorption (TD) was evaluated for determination of particulate phase oxygenated-polycyclic aromatic hydrocarbons (O-PAHs) collected on filters coupled with a gas chromatography/mass spectrometry (GC/MS). The analytical parameters were optimized with standard testing and ambient samples. Ten of the most abundant O-PAHs in ambient air were included in the demonstration. A desorption temperature of 275 °C for 9 min was sufficient to transfer entire target compounds to the analytical system. Good linearity (R2 >0.99) on the calibrations for each analyte was achieved. The limit of detection (LOD) ranged from 21 to 269 pg per loading. Method precisions, determined by replicate analyses of calibration standards and ambient samples, was less than 10% for target compounds. Comparisons were conducted on batches of 28 ambient aerosol filter samples using our TD and the traditional SE methods. Reasonably good agreement (R2 = 0.98) by the two methods was demonstrated for most of O-PAHs. The in-injection port TD can improve laboratory efficiency and reduce solvent-based costs for the measurement of O-PAHs.