APPLICATION OF THE STEREOSPECIFIC INHIBITOR L-PHENYLALANINE TO THE ENZYMORPHOLOGY OF INTESTINAL ALKALINE PHOSPHATASE

Abstract

The intestine-specific inhibitor of alkaline phosphatase, l-phenylalanine, has been employed in the study of alkaline phosphatase in rat intestine. Five substrates were used, β-glycerophosphate, o-carboxyphenylphosphate, α-naphthyl acid phosphate, naphthol AS-BI phosphate and naphthol AS-TR phosphate. The hydrolytic rate and specific l-phenylalanine inhibition was most marked in the case of the first three substrates. When formol-Ca fixed, gum-sucrose treated sections of rat intestine were incubated in these substrates in the presence separately of 0.05 Md- and l-phenylalanine and Gomori's calcium-cobalt method was employed, a striking inhibition by the l-isomer was regularly found. Azo dye methods were applied under different conditions of time, temperature and cation concentration in some instances. The contrast was marked but not dramatic in the case of α-naphthyl acid phosphate, and it was evident but not marked in the case of naphthol AS phosphate substrates. These findings have been discussed. The alkaline phosphatase of intestinal epithelial cells (human) grown in tissue culture exhibited great sensitivity to l-phenylalanine. Rat kidney and leukocyte alkaline phosphatase were relatively insensitive under the same conditions. Morphologically, the l-phenylalanine-sensitive alkaline phosphatase in rat intestine is largely confined to the striated border of the epithelial cells. In animals maintained on a high fat diet, azo dye methods demonstrated the presence of fine granules in the region of the Golgi apparatus and terminal web in the presence of d-phenylalanine. These reactions were absent in companion sections incubated in l-phenylalanine-containing substrate solutions. The possibility of comparing the morphology of a tissue in which both test and control solutions are identical except for the configuration of one component may contribute to a more precise interpretation of the results obtained in the test section.