The Journal of Nutrition Vol. 128 No. 12 December 1998, pp. 2526-2527

LETTER TO THE EDITOR:

Trypsin Inhibitors and Endogenous Amino Acids

	LETTER

Dear Dr. Suttie:

We wish to comment on the paper by Caine et al. (1998) concerning the homoarginine (HA) method used to determine the ileal endogenous amino acid losses in pigs. Initially proposed for protein isolate digestibility studies, the method consists in the conversion, by chemical treatment, of the lysine monomers of dietary proteins into the synthetic derivative, HA. The digestion and absorption of the latter are assumed to be similar to those of lysine, but HA is not incorporated into digestive secretions (Schmitz et al. 1991). Thus, the method provides values of true ileal digestibility of dietary lysine and ileal excretion of endogenous lysine.

The guanidination process uniformly converts a representative proportion of the lysine in the whole soybean meal. However, in the present case, the o-methylisourea treatment removed from 240 to 380 g of matter per kg, including part of the proteins, and only 30 to 46% of lysine residues were guanidinated because a portion of the protein was inaccessible. Moreover, the yield of conversion is very sensitive to o-methylisourea and protein concentrations and to the pH (Rutherford and Moughan 1990). Such conditions, especially a constant pH, are difficult to achieve within the core of soybean meal. The high loss of dry matter and the low rate of conversion of lysine make it unlikely that the HA was representative of all the lysine in soybean meal. If only the most accessible proteins were labeled, an overestimation of the true digestibility of lysine from the whole soybean protein would result. The fact that different ileal endogenous losses (footnote Table 3) and true digestibilities (footnote Table 4) were reported for unguanidinated and total (unguanidinated + HA) dietary lysine should have led the authors to question the validity of the underlying hypotheses.

Then, Caine and coworkers applied the method to estimate the ileal excretion of all the endogenous amino acids (AA) and the true ileal digestibility of these AA in the defatted soy flour. They based the calculation of the recovery of endogenous AA on the assumption that, in guanidinated proteins, the ratio of the HA concentration to that of the exogenous AA remains constant in the intestine, as observed during sequential proteolysis in vitro, following a proposal by Siriwan et al. (1994). However, the latter authors showed that the assumption was acceptable for purified proteins, such as protein isolates (casein, soybean protein isolate) but not for complex proteins (meals, cereals, etc). A constant ratio of HA to exogenous AA implies that the AA composition of the exogenous protein remains the same during the digestion process and, hence, that every AA has the same true digestibility, which is contrary to a body of compelling evidence in the literature. As a consequence, the HA method should not be used for the determination of recoveries of ileal endogenous AA other than lysine.

How the authors could recalculate a different true digestibility for each AA from the estimates of apparent digestibility and endogenous loss is unclear. This very contradictory point can be explained by the use of the wrong formula to calculate the recoveries of endogenous AA:

(3)

where (homo) and (aa) are the concentrations in HA and in the individual AA in the digesta or in the diet, respectively, and (AA) flow as the total ileal flow of each AA.

First, the two ratios should have been bracketed together. Second, the estimation of the proportion of endogenous AA in the ileal digesta cannot be based on a difference of ratios but on the ratio of these ratios, i.e., [(homo)digesta/(aa)digesta]/[(homo)diet/(aa)diet]. According to us, the endogenous AA recoveries should have been calculated as follows:

(3')

which is strictly equivalent to the equation:

(3'')

with digestibility expressed per unit and where the true digestibility is the digestibility of HA.

By applying Equation 3' or 3, we obtained a total endogenous AA recovery of 54.6 g/kg dry matter intake for the unprocessed soybean meal and 18.8 g for the autoclaved meal, instead of 27.6 and 4.2 g AA/kg dry matter intake, respectively. Recalculation of the true digestibility, by correcting the apparent value from the correct estimate of the endogenous loss, will evidently restore the HA value for each amino acid (see Eq. 3). Thus, we suggest that the results of endogenous AA recovery and true AA digestibility presented in Tables 3 and 4 are not correct.

Calculations of the correct values of endogenous losses for the unguanidinated and the total lysine from their apparent digestibilities, assuming that for both of them the true digestibility was equal to the apparent digestibility of HA, gave: 2.9 and 1.8 g/kg dry matter intake, respectively. This would confirm different responses of the two lysine pools to the digestive process and that HA did not represent lysine, in the case of the untreated soy flour.

Finally, the conclusion to an "effective approach" is difficult to accept, even with the reservation that it provides only "qualitative differences," in the study of usual feedstuffs.

Pascal Leterme
Faculté Universitaire des Sciences agronomiques,
Unité de Zootechnie
B-5030 Gembloux, Belgium

Bernard Sève
INRA, Station de Recherches Porcines
F-35590 Saint-Gilles, France

	FOOTNOTES

Manuscript received 1 June 1998. Initial reviews completed . Revision accepted 17 August 1998.

	LITERATURE CITED

• Caine W., Sauer W., Verstegen M., Tamminga S., Li Y., Schulze H. Guanidinated protein test meals with higher concentration of soybean trypsin inhibitors increase ileal recoveries of endogenous amino acids in pigs. J. Nutr. 1998; 128:598-609[Abstract/Free Full Text]
• Rutherford S., Moughan P. Guanidination of lysine in selected dietary proteins. J. Agric. Food Chem. 1990; 38:209-211
• Schmitz M., Hagemeister H., Erbesdobler H. Homoarginine labelling is suitable for determination of protein absorption in miniature pigs. J. Nutr. 1991; 121:1575-1580
• Siriwan P., Bryden W., Annison E. Use of guanidinated dietary protein to measure losses of endogenous amino acids in poultry. Br. J. Nutr. 1994; 71:515-529[Medline]

0022-3166/98 $3.00 ©1998 American Society for Nutritional Sciences