Fecalase: a model for activation of dietary glycosides to mutagens by intestinal flora.

Abstract

Many substances in the plant kingdom and in man's diet occur as glycosides. Recent studies have indicated that many glycosides that are not mutagenic in tests such as the Salmonella test become mutagenic upon hydrolysis of the glycosidic linkages. The Salmonella test utilizes a liver homogenate to approximate mammalian metabolism but does not provide a source of the enzymes present in intestinal bacterial flora that hydrolyze the wide variety of glycosides present in nature. We describe a stable cell-free extract of human feces, fecalase, which is shown to contain various glycosidases that allow the in vitro activation of many natural glycosides to mutagens in the Salmonella/liver homogenate test. Many beverages, such as red wine (but apparently not white wine) and tea, contain glycosides of the mutagen quercetin. Red wine, red grape juice, and tea were mutagenic in the test when fecalase was added, and red wine contained considerable direct mutagenic activity in the absence of fecalase. The implications of quercetin mutagenicity and carcinogenicity are discussed.Damage to DNA by environmental mutagens, both natural and man-made, is likely to be a major cause of cancer and other diseases (1, 2). The Salmonella test (3), along with other short-term tests (4), is being used to survey a wide variety of substances in our environment for mutagenicity. The test measures back mutation in several specially constructed mutants of Salmonella bacteria. A homogenate of rat liver (or other mammalian liver) is added to the (Salmonella) test as an approximation of mammalian metabolism (3). By using this system, approximately 85% 1 5% of the organic carcinogens tested have been detected as mutagens (5-9).Recent studies indicate that many naturally occurring glycosides of mutagenic aglycones are not mutagenic in the Salmonella test (10-23). These glycosides are hydrolyzed by the bacteria in the human intestine where the mutagen is liberated. They are not cleaved by the liver or by liver homogenates used in the Salmonella test. For example, cycasin, a f3-D-glucoside of the mutagen methylazoxymethanol, is not mutagenic in the Salmonella test unless 3-glucosidase is added to the standard test (20,21). Cycasin is a carcinogen in rats but is not a carcinogen when tested with germ-free rats (24, 25), which lack the microorganisms that cleave the sugar from the mutagenic moiety.An enormous variety of substances are present as glycosides in the plant kingdom (10-27). It is desirable to have an enzyme preparation for use in mutagenicity tests that will hydrolyze the hundreds of sugars in these glycosides. have developed a cell-free extract of rat cecal bacteria, cecalase, for use in mutagenicity testing. 22) have used hesperidinase, a mold enzyme preparation, for this pur-The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S.

Pharmacology in Health Food: Metabolism of Quercetin In Vivo and Its Protective Effect Against Arteriosclerosis

“…Quercetin is mostly present in the form of glycosides in vegetables and fruits, and dietary glycosides were believed to be converted to the respective aglycones in the large intestine by the glycosidase activity of intestinal bacteria (12). However, it was recently demonstrated that quercetin 3-O-β-D-glucuronide (Q3GA) and quercetin-3′-sulfate are the major quercetin conjugates in human plasma, in which aglycone could not be detected (13 -15).…”

Differences in Enzymatic Activities of Cecal Contents of Rats Fed on Differently Processed Dietary Fibers.

“…Arabinofuranosidase activity Arabinofuranosidase assays were carried out as described by MacKenzie et al ( 1987), but p-nitrophenol release was measured at 400 nm instead of 410nm (Shimadzu UV-1200 UV-VIS spectrophotometer) (Tamura et a/, 1980). Ferulic acid esterase activity Assays for ferulic acid esterase contained I mg of ACD and enzyme at appropriate dilutions in 50 mM phosphate-buffered saline (pH 7.4).…”

How to Determine the Role of the Microbiome in Drug Disposition

“…This method has been used to assay gut microbial enzyme activity towards herbal glycosides generating genotoxic aglycone products (Tamura et al, 1980;Yeo et al, 2012). This technique was also used to demonstrate that gut bacteria can metabolize the cholesterol lowering drug lovastatin (Yoo et al, 2014).…”

“…The final version may differ from this version. Another approach for studying microbial biochemical transformations from a mixed community is by using a "fecalase" preparation (Tamura et al, 1980), which is a cell-free extract of feces including microbial enzymes (typically starting from a stool sample).…”