CRUDE ALGAL EXTRACTS FOR ENHANCED BIOMASS PRODUCTION IN DIGESTATE-FED MICROALGAL CULTURES

Introduction

Microalgae cultivation on wastewater and anaerobic digestate enables simultaneous biomass production and nutrient recovery while reducing the need for mineral fertilisers. The resulting biomass can be valorised as biostimulants, biofertilisers, feed ingredients, bio-based materials, or energy products (Berden-Zrimec et al., 2024). However, biomass productivity in digestate-fed systems is often lower than expected from nutrient availability alone. Previous studies have shown that additional CO₂ supplementation can significantly improve algal growth, indicating that inorganic carbon availability may become limiting under cultivation conditions (Berden-Zrimec et al., 2026). While effective, continuous CO₂ injection increases operational costs.

Recent laboratory studies demonstrated that crude algal extracts prepared from Scenedesmus-dominated biomass stimulate growth in digestate-based cultures, suggesting improved utilisation of inorganic carbon already present in the medium (Berden-Zrimec et al., 2026). The present study evaluated whether a similar effect could be achieved in pilot-scale digestate-fed raceway ponds.

Materials and methods

Experiments were conducted in identical raceway ponds (7.5 m², approximately 1250 L) cultivated on biochar-filtered anaerobic digestate from a food-waste biogas plant (Ljubljana, Slovenia). The ponds (Figure 1) were operated under the same conditions with digestate addition of approximately 10 L day^-1 and automatic pH control through CO₂ injection. The dominant species were Scenedesmus dimorphus and S. quadricauda (Figure 1).

Figure 1. Raceway pond culture fed with anaerobic digestate and dominated by Scenedesmus dimorphus and S. quadricauda.

Crude algal extract (CAE) was prepared from harvested pond biomass by drying and mechanical disruption and was directly added to the selected pond (100 g). Biomass growth was monitored by optical density (OD680) and dry weight measurements. Two approaches for lipid stimulation were applied after the growth phase to explore the possibility of digestate-grown biomass valorisation for fuels.

Results and discussion

CAE supplementation increased biomass accumulation in both pilot-scale experiments (Figure 2). After 11 days of the first trial , the CAE-treated pond reached more than twice the biomass concentration of the untreated control. The first experiment was affected by intermittent power outages and interrupted digestate feeding between days 4 and 6. Despite these disturbances, the CAE-treated pond maintained substantially higher biomass accumulation than the control, suggesting improved culture robustness under suboptimal cultivation conditions. The second experiment was performed later in the season (Figure 2b) in stable operation conditions, exhibiting difference between treatments only in the first days after supplementation.

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Figure 2. Experiments results: (left) First study, (right) Second study.

The lipids production was not stimulated by lowering the digestate addition and adding phosphate. On the other hand, removing 75% of the medium and replacing it with water with phosphate, resulted in alteration of the composition of total lipids content. Although the total lipids were rather constant, their composition in phospholipids was decreased suggesting an enrichment of total lipids with glycerides. The free fatty acids produced by the degradation of phospholipids can be utilized for the synthesis of glycerides.

The findings indicate that the stimulatory effect of CAE is reproducible and pronounced shortly after addition. While CAE initially stimulated biomass accumulation in both experiments, the effect was stronger and more persistent under interrupted feeding and other suboptimal cultivation conditions. CAE supplementation may represent a practical tool for improving biomass productivity in digestate-fed cultures by enhancing the utilisation of nutrients and inorganic carbon already available in the cultivation medium.

Acknowledgements

This work was supported by the Horizon Europe projects CRONUS (101084405) and FUELPHORIA (101118286).

References

Berden-Zrimec, M.; Lazar, D.; Trontelj, D.; Barampouti, E.M.; Mai, S.; Malamis, D.; Reinhardt, R. Crude Microalgae Extract for Increased CO2 Capture and Higher Biomass Production in Algal Cultivation Systems. Phycology 2026, 6, doi:10.3390/phycology6010023

Berden-Zrimec, M.; Reinhardt, R.; Rossi, S.; Ficara, E.; Rodero, M.R.; Muñoz, R. Algae and biogas plants: Digestate remediation and nutrient recycling with algal systems. In: Algae Mediated Bioremediation: Industrial Perspectives 2024, Wiley, doi: 10.1002/9783527843367.ch12