Abstract Bioretention systems are a low-impact development (LID) measure to effectively control stormwater runoff and reduce pollutant concentrations. In this paper, three groups of bioretention cells with different filling materials (1# bioretention soil media (BSM), 2# BSM + 5% biochar, and 3# BSM +5% biochar +biological filler) were constructed to analyze the pollutant removal characteristics and microbial action under different simulated rainfall conditions. Results showed that the overall pollutant removal capacity of systems 2# and 3# was higher than that of system 1#, with system 3# having the lowest effluent concentrations of 2.71 mg/L for total nitrogen (TN) and 64.3 mg/L for chemical oxygen demand (COD). The load reduction effect for heavy metals of the three systems was ranked as 2# > 1# > 3#, and average load reduction rates were 80.3, 75.1, and 84.8% for Cu, Pb, and Zn in 2#. Microbial community analysis indicated that Proteobacteria and Firmicutes were the absolute dominant bacteria of the three bioretention systems, and the dominant genera included Bacillus, Hyphomicrobium, Micrococcaceae, and Nitrospira. In addition, the total number of denitrifying functional bacteria genera in systems 2# and 3# was increased by 1.39 and 52.1% compared to system 1#.