Dual role of citric acid: facilitate the bioconversion of CO to CH and enhance bioavailability of zero-valent iron.

Studies on the bioconversion of CO to CH using Fe have reported that a fraction of CO is converted to FeCO, forming a passivation layer on the Fe surface, which reduces its reactivity and limits process efficiency. To mitigate FeCO precipitation and enhance CO-to-CH conversion using Fe, this study evaluates the role of citric acid (CA) as a ligand to Fe, which helps maintain iron in a soluble form and prevents its rapid conversion to insoluble iron carbonate (FeCO). Batch experiments with 30 mM Fe, anaerobic granular sludge (AGS), and NaHCO with CA as a ligand showed that citrate biodegraded into acetic acid and CO gas. To prevent citrate degradation and elucidate its effect on Fe-mediated methanogenesis, 5 % v/v of antibiotic antimycotic solution (100x) (10,000 units penicillin, 10 mg streptomycin and 25 μg amphoctericin per ml) were introduced as a research tool, selectively inhibiting bacterial activity without affecting hydrogenotrophic methanogens. Methane production was notably enhanced only in the sample with NaHCO, CA, antibiotics, and Fe, reaching 38 ml by day 50, whereas the corresponding sample without Fe produced only 22 ml, resulting in a net methane production of 16 ml. The results indicate that CA enhances methane production by maintaining iron solubility and inhibiting FeCO precipitation, thereby facilitating continuous iron oxidation and sustained electron release, which supports the microbial reduction of CO to CH. While antibiotics provided a controlled environment to uncover these mechanisms, their use is not a viable long-term solution. Based on the findings of this study, future work may explore short-term CA exposure (<12 h) or alternative ligands to minimize biodegradation without relying on antibiotics.