An assay system was developed to test whether bacteria in the gastrointestinal tract are capable of metabolizing tryptophan to compounds that are able to bind to the aryl hydrocarbon (Ah) receptor. Tryptophan (1 mM) was added to feces diluted 1:1,000 in phosphate-buffered saline and incubated at 37°C overnight. The suspensions were extracted with chloroform to obtain the hydrophobic compounds. To test for the presence of Ah receptor ligands, a competition binding assay using [2-125I]iodo-7,8-dibromodibenzo-p-dioxin and Hepa 1c1c7 cytosol was employed; it was capable of detecting picogram levels of a competing ligand with similar affinity. Fecal suspensions in the presence of I mM tryptophan and oxygen are capable of producing greater than 60% inhibition of radioligand binding per 10 µg of feces. In contrast, oxygen-equilibrated fecal suspensions without tryptophan and argon-equilibrated fecal suspensions with tryptophan exhibited 10% inhibition of radioligand binding per 10 µg of feces in the competition binding assay. Other indolylic compounds and amino acids were similarly tested. Histidine, tyrosine, phenylalanine, glycine, indole-3-acetic acid, and tryptamine were all negative in this assay. Indole-3-carbinol was capable of forming compounds that bind to the Ah receptor under a variety of conditions: in fecal suspensions with or without oxygen, in 50 mM HCl for 80 minutes, and in neutral pH buffer overnight at 37°C. Addition of oxygenated tryptophan-fecal incubation extracts to Hepa 1 and Hepa c4 mutant (defective Ah receptor) cell cultures resulted in the induction of ethoxyresorufin O-deethylase activity in Hepa 1 cells, but no induction was observed in Hepa c4 cells. These results suggest that bacteria in the gastrointestinal tract under the proper conditions are able to metabolize tryptophan to compounds that are Ah receptor ligands.
All Science Journal Classification (ASJC) codes
- Medicine (miscellaneous)
- Nutrition and Dietetics
- Cancer Research