Comparing nutritional and digestibility aspects of sustainable proteins using the INFOGEST digestion protocol

Simulated gastro-intestinal digestion is widely employed in many fields of food and nutritional sciences, as conducting human trials are often costly, resource intensive, and ethically disputable. As a consequence, in vitro alternatives that determine endpoints such as the bioaccessibility of nutrients and non-nutrients or the digestibility of macronutrients (e.g. lipids, proteins and carbohydrates) are used for screening and building new hypotheses. Various digestion models have been proposed, often impeding the possibility to compare results across research teams. For example, a large variety of enzymes from different sources such as of porcine, rabbit or human origin have been used, differing in their activity and characterization. Differences in pH, mineral type, ionic strength and digestion time, which alter enzyme activity and other phenomena, may also considerably alter results. Other parameters such as the presence of phospholipids, individual enzymes such as gastric lipase and digestive emulsifiers vs. their mixtures (e.g. pancreatin and bile salts), and the ratio of food bolus to digestive fluids, have also been discussed at length. In the present consensus paper, within the COST Infogest network, we propose a general standardised and practical static digestion method based on physiologically relevant conditions that can be applied for various endpoints, which may be amended to accommodate further specific requirements. A frameset of parameters including the oral, gastric and small intestinal digestion are outlined and their relevance discussed in relation to available in vivo data and enzymes. This consensus paper will give a detailed protocol and a line-by-line, guidance, recommendations and justifications but also limitation of the proposed model. This harmonised static, in vitro digestion method for food should aid the production of more comparable data in the future.

Developing a mechanistic understanding of the impact of food structure and composition on human health has increasingly involved simulating digestion in the upper gastrointestinal tract. These simulations have used a wide range of different conditions that often have very little physiological relevance, and this impedes the meaningful comparison of results. The standardized protocol presented here is based on an international consensus developed by the COST INFOGEST network. The method is designed to be used with standard laboratory equipment and requires limited experience to encourage a wide range of researchers to adopt it. It is a static digestion method that uses constant ratios of meal to digestive fluids and a constant pH for each step of digestion. This makes the method simple to use but not suitable for simulating digestion kinetics. Using this method, food samples are subjected to sequential oral, gastric and intestinal digestion while parameters such as electrolytes, enzymes, bile, dilution, pH and time of digestion are based on available physiological data. This amended and improved digestion method (INFOGEST 2.0) avoids challenges associated with the original method, such as the inclusion of the oral phase and the use of gastric lipase. The method can be used to assess the endpoints resulting from digestion of foods by analyzing the digestion products (e.g., peptides/amino acids, fatty acids, simple sugars) and evaluating the release of micronutrients from the food matrix. The whole protocol can be completed in ~7 d, including ~5 d required for the determination of enzyme activities.

The protein content in foodstuffs is estimated by multiplying the determined nitrogen content by a nitrogen-to-protein conversion factor. Jones' factors for a series of foodstuffs, including 6.25 as the standard, default conversion factor, have now been used for 75 years. This review provides a brief history of these factors and their underlying paradigm, with an insight into what is meant by "protein." We also review other compelling data on specific conversion factors which may have been overlooked. On the one hand, when 6.25 is used irrespective of the foodstuff, "protein" is simply nitrogen expressed using a different unit and says little about protein (s.s.). On the other hand, conversion factors specific to foodstuffs, such as those provided by Jones, are scientifically flawed. However, the nitrogen:protein ratio does vary according to the foodstuff considered. Therefore, from a scientific point of view, it would be reasonable not to apply current specific factors any longer, but they have continued to be used because scientists fear opening the Pandora's box. But because conversion factors are critical to enabling the simple conversion of determined nitrogen values into protein values and thus accurately evaluating the quantity and the quality of protein in foodstuffs, we propose a set of specific conversion factors for different foodstuffs, together with a default conversion factor (5.6). This would be far more accurate and scientifically sound, and preferable when specifically expressing nitrogen as protein. These factors are of particular importance when "protein" basically means "amino acids," this being the principal nutritional viewpoint.