As part of the CHEERS project, AINIA is applying advanced in vitro technologies to characterise the next generation of protein ingredients obtained through sustainable and circular bioprocesses. One of our latest developments is a range of single-cell protein (SCP) ingredients designed for use in pet food.

Simulating canine digestion in the lab

To evaluate the nutritional value of these novel protein sources, AINIA uses models of simulated canine gastrointestinal digestion, including the DigestSim multicompartment system. This dynamic platform replicates the key physiological processes of the dog’s digestive tract under controlled conditions:

  • Gastric and intestinal pH profiles.
  • Temperature and transit times.
  • Gradual release of digestive enzymes and secretions.
  • Solubilisation and release of amino acids.

These in vitro simulations allow the digestibility of proteins and the bioaccessibility of amino acids to be assessed, providing valuable insights at an early stage that would otherwise require in vivo studies for this preliminary ingredient.

Canine macrophage models to study biological effects

In addition to assessing digestibility, AINIA employs a canine macrophage in vitro model (DH82 cell line) to examine the biological activity of digested protein fractions. Through this approach, we can:

  • Confirm the biocompatibility of the ingredient.
  • Evaluate potential immune-modulating properties.
  • Analyse changes in key cytokines involved in inflammatory balance (e.g., IL‑1β, IL‑6 and IL‑10)

These cellular assays provide a deeper understanding of how the ingredient may interact with immune pathways relevant to canine health.

Why in vitro models matter

Integrating digestion models and cell-based assays offer multiple advantages:

  • Rapid and ethically responsible screening of new ingredients.
  • Mechanistic insight into nutrient behaviour and bioactivity.
  • Early prediction of nutritional and functional performance.
  • Scientific support for developing safe, high-quality, circular protein sources for the pet food industry.
CHEERS: boosting innovation through circular bio-based proteins

By developing these advanced analytical tools, the CHEERS partners are driving innovation in zero-waste biorefinery strategies and contributing to the EU’s transition toward more sustainable protein sources.

AINIA’s in vitro platforms are crucial for accelerating the optimisation of new ingredients and ensuring their suitability for real-world applications.

In vitro Dynamic digestion and immune system – dog

This diagram shows a step-by-step workflow to evaluate single-cell protein (SCP) ingredients for pet food using in vitro methods. First, SCP is run through a simulated canine gastrointestinal digestion model to generate a “digest.” That output is used to assess protein digestibility and amino acid bioaccessibility. Next, the digested fractions are tested in a canine macrophage cell model (basal vs. inflamed conditions), and the response is read out via lab analysis to understand the potential effects on the canine immune system.

A Faster, More Ethical Path to Better Proteins

By combining in vitro digestion with immune-response cell assays, researchers can screen next-generation, circular protein ingredients faster, earlier, and more ethically. This integrated approach strengthens evidence on digestibility, safety, and potential functionality, helping accelerate innovation while supporting the development of high-quality, sustainable proteins for the pet food industry.