In this work, Tenax consecutive extractions of polycyclic aromatic hydrocarbons (PAHs) were conducted in two spiked sediments to investigate the influence of different Tenax addition amounts and desorption times on the rapidly desorbing fraction of PAHs, and to determine a reliable method for estimating PAHs bioavailability. The results indicated that a large Tenax addition amount has a positive effect on the desorption of PAHs from sediments. The desorption amounts of target PAHs compounds (3-ring phenanthrene and 4-ring fluoranthene) increased as the Tenax: sediment ratios increased from 0.25 to 2 in two spiked sediments. The highest desorption percentages of phenanthrene and fluoranthene were 48.91% and 34.70% for Jialing industrial park sediment, and 43.36% and 33.24% for Huanghuayuan bridge sediment, respectively. The results of desorption kinetics were suitably fitted with first order three-compartment model to estimate the rapidly desorbing fraction, Moreover, the Tenax: sediment ratio of 1 and desorption time of 24 h were found to be suitable for the desorption of phenanthrene and fluoranthene from sediments. The PAHs in sediments were biodegraded well by the bacterial strain J1-q. Comparing the maximum biodegraded amount of target PAHs in 30 days and the desorbed fraction over 400 h, the results showed that Tenax had better correlation with the high molecular weight fluoranthene than with the low molecular weight phenanthrene.