**2. Protein and peptide digestion**

Before a discussion of protein digestibility evaluation methods, it seems worthwhile to introduce a reader to the current understanding of protein and peptide digestion process in order to provide him with a holistic picture of what could happen to an ingested protein. That is why this section will give a short description of digestive tract and an evaluation of enzymes involved in protein digestion in stomach and intestine. A special emphasis will be laid on characterization of intestinal wall peptidases (surface as well as intracellular) with description of their specificities and their role in overall protein digestion.

Methods of Protein Digestive Stability Assay – State of the Art 213

The core properties of the luminal proteases are summarized in table 1, to give the reader an understanding of the processes, which take place at different stages of digestion. It should be noted that most digestive enzymes are present not as a single entity, but rather as a group of slightly different molecules with quite similar substrate specificity and kinetic properties. Some authors suggest it to be a sign of evolutionary importance of these enzymes (Silk et al. 1976). A detailed discussion of these enzymes is, however, beyond the scope of this chapter, and we refer a curious reader to the excellent reviews available (Freeman & Kim 1978;

Besides the common luminal enzymes, which are secreted by specialized organ – pancreas, a significant amount of intestinal wall proteases is also present in the lumen. The sources of these enzymes are desquamated mucosal cells resulting from active replacement of the lining epithelium. An approximate quantity of these enzymes is about 30 g per day

Chemical assay data, thermostability studies, and examination of electrophoretic mobilities of luminal peptide hydrolases indicate that aforementioned jejunal enzymes originate predominantly from the cytoplasm of intestinal mucosal cells, whereas the brush border of mucosal cells is a major source of the enzymes in the ileum. All of these enzymes recognize primarily short peptides and are aminopeptidases; they are discussed in more detail in the

The products of luminal proteolysis are free amino acids and small peptides having a chain length of two to six amino acid residues. Analysis of post-prandial intestinal contents aspirated from human jejunum reveals that approximately only one-third of the total amino acid content exists in the free form (Silk et al. 1985). Some of the short peptides are directly absorbed by specific transport systems. However, to increase the overall efficiency of the assimilation process the mucosal cells of intestinal wall provide an additional source of proteolytic enzymes. As these cells line the intestinal wall and are intensively shed into lumen, they contribute both to luminal and parietal digestion. A profound characteristic of mucosal cells is high aminopeptidase activity. This peptidase activity is located in two main subcellular fractions: the cytoplasm and the brush border membrane. The brush border membrane fraction is associated with less than 10% of the total hydrolytic cellular activity for dipeptides but as much as 60% for tripeptides (Freeman & Kim 1978; Sterchi & Woodley

The whole digestion process is quite rapid in man, as well as in rat. So, according to the data of Curtis *et al.*, 44.9 % of the meal had left the stomach of a rat in 30 min, and 84.7 % of the meal had left the stomach in 60 min. At 30 min and thereafter, negligible amounts of 14Clabeled protein, which was included in the meal, were detected in intestinal contents of proximal third of small intestine. At 30 min and 1 hr, 14C accumulated in medial and distal thirds of small intestine. At 2, 3 and 4 hr small quantities of 14C were detected in intestinal contents of distal third of small intestine and the colon. The absorption of ingested protein was virtually complete 2 hr after administration of the meal (Curtis et al. 1978). In human, the situation is quite the same: digestion and absorption of dietary protein are completed in the jejunum, the whole process of digestion and absorption reaches its maximum at about 45 min or possibly earlier, and absorption may be largely complete within 75 to 90 min. However, the data of some studies show that the remnants of dietary protein can be recovered from the ileum as late as 4 hr after a meal (Adibi 1976). Nevertheless, it appears

Turner 1968; Whitcomb & Lowe 2007).

(Freeman & Kim 1978; Gropper et al. 2009).

next section.

1980a).
