•Novel approach for high-strength pharmaceutical wastewater by PD/A was confirmed.•PD/A biofilm was developed rapidly and had high tolerance to real wastewater.•Higher LB-EPS was produced with protein 3-turn helices structure was obtained.•Relatively stable TN removal efficiency of 81.2% was achieved with increasing load.•Thauera was highly enriched and dominated for partial NO3− reduction to NO2−. Biological nitrogen removal from pharmaceutical wastewater has drawn increasing attention due to biotoxicity and inhibition. In this study, for the first time, a novel approach integrating partial-denitrification with anaerobic ammonia oxidation (PD/A) in a sequencing biofilm batch reactor (SBBR) was proposed and demonstrated to be efficient to treat the bismuth nitrate and bismuth potassium citrate manufacturing wastewater, containing ammonia (NH4+-N) and nitrate (NO3−-N) of 6300±50 mg L − 1 and 15,300±50 mg L − 1. The maximum anammox activity was found at the shock effect of influent total nitrogen (TN) of 100 mg L − 1 with NO3−-N/NH4+-N of 1.0. Long-term operation demonstrated that the PD/A biofilm was developed rapidly after 30 days using synthetic influent, with TN removal efficiency increasing from 40.9% to 80.8%. Significantly, the key bacteria for PD/A had high tolerance and adapted rapidly to pharmaceutical wastewater, achieving a relatively stable TN removal efficiency of 81.2% with influent NH4+-N and NO3−-N was 77.9 ± 2.6 and 104.1 ± 4.4 mg L − 1 at a relatively low COD/NO3−-N of 2.6. Anammox pathway contributed to TN removal reached 83.6%. Significant increase of loosely-bound extracellular polymeric substances was obtained with increasing protein of 3-turn helices structure as response to the inhibitory condition. High-throughput sequencing analysis revealed that the functional genus Thauera was highly enriched in both biofilms (9.5%→43.6%) and suspended biomass (15.5%→57.5%), which played a key role in high NO2−-N accumulation. While the anammox bacteria decreasing from 7.8% to 1.6% in biofilm, and from 1.8% decreased to 0.1% in the suspended sludge. Overall, this study provides a new method of high-strength pharmaceutical wastewater treatment with low energy consumption and operation cost, as well as a satisfactory efficiency. Display omitted