Microsomes Prepared from Eluted Enterocytes Yield High Cytochrome P450 and UGT Activities that are…

Microsomes Prepared from Eluted Enterocytes Yield High Cytochrome P450 and UGT Activities that are…

Published:  8/25/2004 12:00:00 AM

Microsomes Prepared from Eluted Enterocytes Yield High Cytochrome P450 and UGT Activities that are Stable over Multiple Freeze/Thaw Cycles


Abstract
Enterocytes in the upper region of the small intestine play a significant role in the first-pass metabolism (pre-systemic clearance) of many orally ingested xenobiotics. For this reason, functionally active and stable intestinal subcellular fractions are required to assess the first-pass metabolism of drugs by cytochromes P450, UDP-glucuronosyltransferases and other drug-metabolizing enzymes. The present study summarizes enzymatic activity data from individual and pooled animal and human intestinal microsomes that were prepared from fresh duodenum/jejunum based on an enterocyte elution method with EDTA and various protease inhibitors. All samples were analyzed for their ability to catalyze testosterone 6ß-hydroxylation, 4-methylumbelliferone glucuronidation, and NADPH-cytochrome c reduction. Microsomes prepared from chemically eluted enterocytes had substantially greater CYP3A activity than those prepared from small intestinal samples subjected to mechanical scraping. Freezing/thawing small intestinal microsomes for up to 5 cycles did not cause significant loss of CYP3A, NADPH-cytochrome c reductase or UDP-glucuronosyltransferases (UGTs) activity. These results suggest that our elution method for processing small intestines is well suited to preserving microsomal enzymatic activities.

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