FMN Phosphatase and FAD Pyrophosphatase in Rat Intestinal Brush Borders: Role in Intestinal Absorption of Dietary Riboflavin

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Abstract
Flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD), are two major coenzyme forms of dietary riboflavin. Yet little attention has been given to the release of the vitamin from its coenzyme forms during the absorptive process. Homogenates from rat intestine catalyze the hydrolysis of these flavin coenzymes. To determine the location of FMN and FAD hydrolases, homogenates of intestinal mucosa were fractioned. FMN and FAD phosphatases were localized in brush border membranes. FAD pyrophosphatase activity was maximal at pH between 6.5 and 8.5 while FMN phosphatase has a pH optimum of 7.5–8.0. FAD pyrophosphatase is more stable to heat. The two enzymes separate on ion exchange chromatography of an isobutanol extract of intestinal brush border membrane fraction. Inhibition of 14C-riboflavin uptake by FMN and FAD in everted rings of rat intestine is directly related to the amount of conversion of these coenzymes to free riboflavin by intestinal enzymes. When FMN and FAD conversion to riboflavin is inhibited by EDTA, competition with 14C-riboflavin for transport was correspondingly decreased. These studies are best explained by a sequential process in which hydrolysis of FMN and FAD by enzymes of the intestinal brush border is followed by absorption of free riboflavin.