Carbon Dioxide Flushing and Zero-Valent Iron Addition during Anaerobic Digestion: Biogas Upgrading Does Not Necessarily Indicate Enhanced Biodegradation

Carbon dioxide (CO2) supplementation and zero-valent iron (ZVI) addition have been proposed as strategies for enhancing methane production during anaerobic digestion. In this study, eight semibatch anaerobic digesters (four treatments operated in duplicate) treating piggery waste were operated for 100 days to evaluate the combined effects of CO2 flushing strategy and ZVI addition on methane production, process stability, and organic matter conversion. ZVI-amended reactors achieved substantially higher methane contents, reaching up to 98% CH4 under single CO2 flushing and approximately 90% CH4 under periodic CO2 flushing. Periodic CO2 supplementation promoted sustained methane generation, resulting in the highest cumulative methane production when combined with ZVI. Preliminary experiments further showed that CO2 flushing can effectively reduce pH under alkaline conditions, whereas ZVI addition enhances hydrogen generation and counteracts excessive acidification. Despite their superior methane production and methane contents, the ZVI-amended reactors exhibited lower organic matter conversion than the corresponding reactors without ZVI. The ZVI + CO2 once-flushed reactors showed the highest residual TOC and soluble COD concentrations (approximately 2104 mg L–1 and 5129 mg L–1, respectively) due to the high pH, whereas the greatest organic matter removal was achieved in the periodically flushed reactors without ZVI. Thermodynamic calculations further demonstrated that increasing bicarbonate concentrations selectively favored hydrogenotrophic methanogenesis while reducing the thermodynamic driving force for acetoclastic methanogenesis and protein (alanine) biodegradation. These findings indicate that higher methane production in ZVI-amended systems did not necessarily reflect greater organic matter degradation.