This project aims to make precision-fermented proteins more sustainable and cost-effective by developing continuous fermentation with in-situ biomass retention. Unlike fed-batch processes, this approach reduces energy and raw material use, avoids downtime, and improves reactor productivity. Controlled cell flocculation enables efficient biomass retention without membranes or centrifugation, supporting scalable, low-emission
production of proteins, such as beta-lactoglobulin.

The objective of the project is to improve the efficiency of the production of alternative proteins (proteins produced by precision fermentation, for example dairy proteins) and to reduce the energy and raw material consumption thereof, using continuous fermentation with in-situ separation of flocculating microbial cells.
The global protein transition, from a heavy reliance on animal-based diet toward more sustainable alternatives, is increasingly necessary to meet growing protein demand while remaining within planetary boundaries. Dairy proteins produced by precision fermentation fit naturally into the protein transition because they deliver the functionality, nutrition, and sensory performance of animal-derived proteins consumers expect while avoiding
the environmental burdens of livestock production. Precision fermented dairy proteins, such as betalactoglobulin (BLG) can replicate the melting, gelling, foaming, and emulsifying properties that make dairy unique, enabling high quality foods without relying on cows, large land areas, or methane emitting systems.
Current processes for producing food proteins through precision fermentation are often too expensive for broad market implementation. The cost price for BLG needs to approach the cost price of animal proteins for largescale adoption, but is often still (far) above it.
The most widely used production process within fermentation is the fed-batch process. Inefficiencies arise in a fed-batch process because the process is repeatedly stopped and restarted. Therefore, the reactor productivity is suboptimal as the reactor stands idle between batches, and at the start of a fed-batch process, only half of the reactor is filled. Additionally, the conversion of substrate to product (raw material efficiency) is suboptimal in a fed-batch process as biomass must be created anew at the start of each batch, for which substrate is consumed that could otherwise have been used for production and is released as heat requiring energy consumption for cooling. Continuous fermentation could solve these issues, however the existing standard for continuous fermentation processes, the chemostat, presents other challenges. In a chemostat reactor biomass continuously leaves the reactor together with the product. Challenges that are encountered are mutations resulting in
productivity loss, risk of contamination and low productivity to reduce oxygen demand and heat generation. Continuous fermentation with biomass retention would result in optimal efficiency. However, using external membranes, centrifuges or settlers, show limited applicability. Fouling and clogging reduces the efficacy of membrane filtration, whereas the shear forces in a centrifuge stresses the cell reducing the viability of the biomass.
This project generates a technological breakthrough by developing a continuous reactor system for continuous precision fermentation to circumvents these issues. A reactor with a small separation compartment inside the reactor, developed for liquid-liquid separation, will be applied in the production of dairy proteins. Flocculation of the cells is required to enhance the liquid-solid separation in the separation compartment and retaining of the biomass in the continuous reactor. So far this has not been reported for strains engineered for the production of dairy or other animal proteins.

Improve the sustainability and cost effectiveness of the production of alternative proteins, e.g. dairy proteins produced by precision fermentation, through continuous fermentation with in-situ separation to retain biomass.