TY - JOUR
T1 - Production of Ah Receptor Ligands in Rat Fecal Suspensions Containing Tryptophan or Indole-3-Carbinol
AU - Perdew, Gary H.
AU - Babbs, Charles F.
N1 - Funding Information:
The authors thank Clayton Hollenback for excellent technical assistance. This work was supported in part by Grant No. ES-04869 from the National Institute of Environmental Health Science (Bethesda, MD), Institutional Grant No. IN-17 from the American Cancer Society (Atlanta, GA), and Grant No. CA-38144 from the National Cancer Institute, National Institutes of Health (Bethesda, MD). This is Technical Paper No. 12,320, Indiana Agricultural Experiment Station (West Lafayette, IN). Address reprint requests to Dr. G. H. Perdew, Dept. of Foods and Nutrition, Purdue University, West Lafayette, IN 47907.
PY - 1991/1/1
Y1 - 1991/1/1
N2 - 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.
AB - 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.
UR - http://www.scopus.com/inward/record.url?scp=0026355818&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0026355818&partnerID=8YFLogxK
U2 - 10.1080/01635589109514159
DO - 10.1080/01635589109514159
M3 - Article
C2 - 1663613
AN - SCOPUS:0026355818
SN - 0163-5581
VL - 16
SP - 209
EP - 218
JO - Nutrition and cancer
JF - Nutrition and cancer
IS - 3-4
ER -