Sustainable single-cell protein production through optimized semi-continuous methanotroph fermentation and resource valorization

Methane oxidizing bacteria named as Methanotrophs, provide a sustainable way to lower greenhouse gas emissions while producing protein-rich microbial biomass. This study focuses on improving semi-continuous fermentation for single-cell protein (SCP) production using a co-culture of Methylophilus methylotrophus and Methylosinus sporium. The work followed a six-phase optimization plan to adjust key operating conditions, including hydraulic retention time (HRT), pH, mixing speed, and temperature, using an ammonium mineral salts medium with high-purity methane and air. Process stability was maintained through daily monitoring of pH, methane uptake, optical density (OD₆₀₀), and dry biomass, with system changes made only after steady-state conditions were reached with 15-20% variation. Protein content and microbial performance were measured using modified Lowry assays and genomic profiling. The SCP values were high, with a peak value of 5181 mg/L. To identify the bacterial content in the medium, the optical density at 600 nm was also evaluated with peak absorbance reaching 1.4 which is regarded as high in the literature and indicating an effective development of methanotrophic culture.The results show that methanotroph-based SCP production can be scaled up and adapted for different applications. This approach supports circular bioeconomy goals and offers a practical route for producing nutrient-rich biomass for food, feed, and biomanufacturing while reducing dependence on synthetic resources.