The availability of sorbed hydrophobic organic contaminants (HOCs) to benthic organisms is important for characterizing sediment toxicity. While many studies show a correlation between the rapid desorption HOC pool and bioavailability to benthic organisms, bioavailability of the slow or very slow desorption fraction is still poorly understood. In this study, Chironomus tentans were exposed to phenanthrene (PHE) or permethrin (PM) to derive biota-sediment accumulation factors (BSAFs) in a sediment that was sequentially desorbed with Tenax extraction or amended with a charcoal to modify the distribution of PHE and PM among the rapid (f rapid ), slow (f slow ) and very slow (f vslow ) desorption pools. As the desorption interval was increased, the f rapid quickly decreased to zero and became negligible after 12h desorption for PHE and 48h desorption for PM. However, in samples with depleted f rapid , BSAF values were substantially greater than zero, suggesting availability of f slow and f vslow . A multivariate linear regression model was further used to estimate BSAFs specific to the different desorption pools, i.e., BSAF rapid , BSAF slow or BSAF vslow . The slow desorption pool was found to be readily available to C. tentans, with BSAF slow values ranging from 25.3 to 73.9% of BSAF rapid . In comparison, BSAF vslow ranged from 0 to 5.9% of BSAF rapid , suggesting a lack of availability. Therefore, the kinetically slow desorption fraction is relatively bioavailable and should not be ignored in sediment toxicity assessment.