Measurements of peptide and amino acid (and other nutrients) absorption and metabolism by tissues of the portal-drained viscera (PDV; gastrointestinal tract, pancreas, spleen and omental fat) can be obtained using chronic indwelling catheters in animals which allows for simultaneous sampling of arterial and venous blood draining the PDV, or sections of the PDV, and measuring blood flow through the same tissues (Reynolds, 2001). Net rates of peptide or amino acid release or removal by the PDV (or tissue of interest) can then be calculated as blood flow multiplied by venous-arterial concentration difference (Huntington et al., 1989). Net portal appearance of peptide-bound or free amino acids is defined as the total amount of amino acids reaching portal blood minus the daily amounts entering the PDV via arterial blood. Peptides and amino acids liberated at the apical membrane or in the cytoplasm can be incorporated into proteins in the enterocyte, metabolized within the epithelium of the small intestine, or transported across the basolateral membrane, ultimately reaching the hepatic portal circulation. In addition, peptides and amino acids are mixed in the lumen and/or enterocyte with amino acids and peptides of endogenous origin during the digestion and absorption process. For these reasons the amino acids present in the hepatic portal vein do not completely reflect the amino acid pattern of the diet (Reynolds, 2001).
As discussed, it is now accepted that peptide absorption is an important physiological process in farm and other animals, and constitutes the primary source of absorbed amino acids. Peptide absorption across the PDV of ruminants has been reported by several researchers (McCormick and Webb, 1982; Seal and Parker, 1991; Koeln et al., 1993; Han et al., 2001), and has been shown to account for 63-92% of total amino acid flux by sheep and cattle. With net flux procedures, the origin of the peptides cannot be determined, but may be from absorption from the lumen, synthesis by tissues of the PDV, breakdown of endogenous protein by intracellular peptidases, or a combination of factors. At any rate, net flux of free amino acids appears to be low compared with peptide-bound amino acids in ruminants (Koeln et al., 1993; Webb et al., 1993; Han et al., 2001). Based on evidence that chemical deproteinization overestimates the peptide amino acid concentration in plasma (Bernard and Remond, 1996; Backwell et al., 1997), it has been suggested that the high flux of peptides might be due to the procedure of sample deproteinization (Neutze et al., 1996; Backwell et al., 1997). Methods for measuring peptides have generally relied on the difference method, where amino acid analysis of deproteinized samples before and after acid hydrolysis has occurred, with the difference being attributed to peptides. A modification of this procedure has included gel filtration after acid depro-teinization of samples to ensure removal of residual protein before acid hydrolysis (Seal and Parker, 1996; Backwell et al., 1997). However, Seal and Parker (1996) reported that, even after the treatment with both chemical deproteinization and physical filtration, peptide-bound amino acid flux still accounted for 63% of the net PDV appearance of total amino acids. The large appearance (438 g day-1) of peptide-bound and free amino acids when calves were deprived of feed for 72 h (Koeln et al., 1993) suggests that a large portion of small peptides might be the degradation products resulting from tissue protein in the gastrointestinal tract (GIT), spleen, pancreas or a combination of these organs, but also supports an increase in PepTl mRNA and protein expression observed during dietary deprivation in rats (Thamotharan et al., 1999a).
Interestingly, work of Webb et al. (1993) indicated that 86-90% of peptide-bound amino acids in the PDV of sheep and calves comes from the stomach tissues (rumen, reticulum, omasum, cranial abomasum, spleen and pancreas). Similar data from Seal and Parker (1996) suggested that 40% of portal peptide-bound amino acids were from the mesenteric-drained viscera (MDV; small intestine, caecum, colon, mesenteric fat and pancreas) suggesting that the remaining 60% were from the stomach tissues. Therefore, tissues of the stomach appear to account for the majority of peptide-bound amino acid release. Similarly, Han et al. (2001) found in yaks that 78% of the absorbed peptide-bound amino acids were from the stomach, whereas net flux of peptide-bound amino acids across the MDV accounted for 22% of the total peptide amino acids. In ruminants, the ability of the forestomach to absorb small peptides has been demonstrated in vitro (Matthews and Webb, 1995) and in vivo (Bernard and Remond, 1999; see below). The lower absorption (10-40%) of peptide amino acids across tissues of the MDV supports the apparent high intracellular peptide hydrolysis observed in the cytoplasm of the small intesti nal enterocyte (Matthews, 1991; Ganapathy et a!., 1994).
Han et a I. (2001) found that the dominant amino acids appearing in the hepatic portal vein in the form of peptide were glutamate, aspartate, leucine, glycine, lysine, proline and serine. These peptide-bound amino acids accounted for 60% of the total peptide-bound amino acids appearing in hepatic portal blood. Han (1998) found similar results in Holstein steers and growing yaks, and similar results have been reported in calves (McCormick and Webb, 1982; Koeln et al., 1993). Wallace et a I. (1993) reported degradation-resistant peptides in rumen fluid of sheep 6 h after feeding. Interestingly, these peptides contained a significant proportion of aspartate, glycine and proline. In addition, several studies (Gardner and Wood, 1989; Daniel et al., 1992; Pan et al., 1996) have suggested that hydrophobic peptides and peptides resistant to mucosal hydrolysis are absorbed faster than hydrophilic and hydrolysis-susceptible peptides. As suggested by Han et al. (2001), resistance to mucosal hydrolysis might explain the larger fluxes of some pep-tide-bound amino acids. Also, the higher net absorption of glutamate, leucine, lysine, and serine in the form of peptides may imply that these amino acids exist more often in the form of peptides that are relatively hydrophobic and resistant to hydrolysis either by microbial peptidases in the rumen, or by apical-membrane and/or cytosolic peptidases of enterocytes.
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