Methyl Orange Degradation in Bioelectrochemical System

In the methyl orange degradation experiments, methyl orange solution was introduced to the anode electrolyte. After a stable operation period of 400 h, various concentrations of methyl orange solution (50, 100, 200 mg L⁻¹) were incorporated into the anode solution, operating under different pH conditions (5, 7, 9). Subsequently, the spectrophotometer measured the methyl orange concentration in the anode solution at 465 nm.^{32 (link)} Upon completion of the operation, the anode biofilm was collected and sent to Yanqu Information Technology Co., Ltd (Hangzhou, China). The 16S V3–V4 region was amplified through polymerase chain reaction (PCR) to establish the biofilm library, which underwent high-throughput sequencing. Furthermore, QIIME was employed for data analysis, and statistical tests and data visualization were conducted using the R language. Lastly, the amount of flora on 0.5 cm × 1 cm anode was assessed using the bicinchoninic acid (BCA) protein assay.