The urgent need for technological innovation to combat water pollution underscores the significance of biosorption as a potential solution. The success of biosorption hinges on the careful selection of a suitable biosorbent. Biofilms, composed of microbial communities, emerge as a promising alternative due to their expansive adsorption capacity and ready availability. In practical applications, biosorption is often executed at pollutant concentrations lethal to microbes. Consequently, comprehending the biosorption potential of biofilms with deceased microbes becomes imperative. Notably, biofilms with deceased microbes offer the added advantage of minimizing the risk of pathogenic microbial contamination. Despite this, studies are scarce comparing biosorption between intact biofilms and those with deceased microbes. This comparative analysis could enhance the feasibility of biofilms in biosorption as an eco-aquatic technology for alleviating aquatic pollution. This study aims to scrutinize the biosorption characteristics of intact biofilm (with living microbes) and autoclaved biofilm (with deceased microbes). The methods employed for analyzing biosorption characteristics encompass examining electric charge properties, FTIR spectra analysis, ion adsorption, and ion desorption. The model ions chosen for this study are K⁺ (monovalent ion) and Mg²⁺ (divalent ion). Results indicate that the biofilm's electric charge properties and adsorption capacity remain relatively unchanged post-autoclaving. Based on these findings, it can be concluded that biofilms, whether intact or autoclaved, present substantial potential as biosorbents in the advancement of eco-aquatic technology for mitigating water pollution.