Rats were treated with vicianin 23.5 mg/100 g diet.

beta-Vicianosidase activity for Vicianin hydrolase was measured with the natural substrate vicianin. Enzyme activity was determined by measuring the liberation of aglycones from each glycoside. Each reaction mixture (50 mkl) contained 10mM substrate, 20mM citrate buffer (pH 6.0) and 10 mkl of the enzyme solution. A mixture without the enzyme was pre-incubated at 37C, and the reaction was started by adding the enzyme and stopped by the addition of 50 mkl of 1M Na2CO3. To determine the activity of a reaction mixture with vicianin, amygdalin and prunasin, each 20 ml of reaction mixture was subjected to HPLC, and benzaldehyde, which is a product of mandelonitrile liberated from the substrates, was detected at 250 nm. One unit was defined as the amount of enzyme liberating 1 mkmol benzaldehyde/min under the assay conditions. In the reactions with pNP b-glycosides, the activity was determined spectrophotometrically by measuring the liberated p-nitrophenol at 405 nm. For the reactions with furcatin, 20 mkl of reaction mixture was subjected to HPLC, and p-allylphenol (retention time, 7.6 min) liberated from the substrate was detected at 277 nm.