High concentration powder carrier bio-fluidized bed (HPB) process has demonstrated significant advantages in municipal wastewater treatment. However, the influent C/N ratio as an important factor has not yet been explored in HPB system. In this study, eight parallel sequential batch reactors (SBR) were established to investigate the effect of influent C/N ratio on HPB process, using diatomite powder and FeC powder as carriers, respectively. Process performance, sludge characteristics, and functional microbes under different C/N ratio (2–8) were explored. Results showed that TN removal efficiency reached 48.2 %, 64.3 %, 76.8 %, and 83.3 %, respectively, in diatomite group, and it was stable at 64.0 %, 73.6 %, 80.2 %, and 83.1 %, respectively, in FeC group. The increase in influent C/N ratio improved the denitrification in anoxic period while also enhancing the simultaneous nitrification and denitrification in aerobic period. A higher sludge concentration and a bigger particle size were observed under a higher C/N ratio. EPS content also showed an upward tread with increase in C/N ratio, and EPS content in biofilm was always higher than that in floc sludge in both diatomite and FeC groups. Microbial analysis suggested the activity rate and abundance of denitrifying bacteria was enhanced with increase in C/N ratio, while it declined in nitrifying and anammox bacteria. The rich pore and groups of diatomite promoted the enrichment of nitrifying bacteria. And denitrification process was enhanced by FeC carrier as electron donors and its micro-electrode functions. These results will provide a theoretical basis for operation optimization and engineering practice of the HPB process.