Impacts of subsurface biogeochemical processes over time have always been a concern for the long-term performance of zero valent iron (Fe0) based Permeable Reactive Barriers (PRB’s). To evaluate the biogeochemical impacts, laboratory experiments were performed using flow-through glass columns for 210 days at controlled temperature (20 °C). Two different particle size of Fe0 were used in the columns, and to simulate indigenous microbial activity, extra carbon source was provided in two columns (biotic columns) and remaining two columns were kept abiotic. Heavy metals (Zn, As) were removed efficiently in all the columns, and no exhaustion of treatment capability or clogging were observed during our experimental duration. Newly formed Fe- mineral phases and precipitates were characterized using XRD, SEM-EDX and micro-XRF techniques in solid phase at the end of the experiment. In addition, 16S rRNA gene extraction was used for microbial community identification in biotic columns. During the incubation, microbial population shifted in favour of Desulforsporosinus species (sulfate-reducing bacteria) from initial dominance of Acidithiobacillus ferrooxidans in sediments. Dominant mineral phases detected in biotic columns were mackinawite (FeS) and sulfate green-rust while in abiotic columns magnetite/maghemite phases were more prevalent.