The distribution of micropollutants in biotic phases of horizontal sub-surface flow (HSSF) constructed wetlands was investigated. 88 diverse micropollutants (personal care products, pharmaceuticals and illicit drugs) were monitored for in full-scale HSSF steel slag and gravel beds to assess their fate and behaviour during tertiary wastewater treatment. Of the studied micropollutants 54 were found in receiving and treated wastewaters. Treatment reduced concentrations of several micropollutants by >50% (removal range −112% to 98%) and resulted in changes to the stereo-isomeric composition of chiral species. For example, stereo-selective changes were observed for 3,4-methylenedioxymethamphetamine (MDMA) and atenolol during HSSF constructed wetland treatment for the first time. Analysis of sludge present within the HSSF beds found 37 micropollutants to be present. However, concentrations for the majority of these micropollutants were not considered high enough to suggest partitioning into sludge was a contributing mechanism of removal. Nevertheless the preservative methylparaben was found at 2772 mg bed−1. Its daily removal from wastewater of 3.4 mg d−1 indicates partitioning and accumulation in sludge contributes to its removal. Other micropollutants found at high levels in sludge (relative to their overall removals) were the antidepressants sertraline and fluoxetine, and the metabolite desmethylcitalopram. Furthermore, process balances indicated uptake and metabolism by Phragmites australis (Cav.) Trin. ex Steud did not contribute significantly to micropollutant removal. However analysis of plant tissues evidenced uptake, metabolism and accumulation of recalcitrant micropollutants such as ketamine and carbamazepine. It is considered that the rate of uptake was too slow to have a notable impact on removal at the 14 h hydraulic retention time. Despite evidence of other removal mechanisms at play (e.g., partitioning into sludge and plant uptake), findings indicate biodegradation is the dominant mechanism of micropollutant removal in HSSF constructed wetlands.