In enzymology, a riboflavinase (EC 3.5.99.1) is an enzyme that catalyzes the chemical reaction

riboflavin + H₂O ${\displaystyle \rightleftharpoons }$ ribitol + lumichrome

Thus, the two substrates of this enzyme are riboflavin and H₂O, whereas its two products are ribitol and lumichrome.

This enzyme belongs to the family of hydrolases, those acting on carbon-nitrogen bonds other than peptide bonds, specifically in compounds that have not been otherwise categorized within EC number 3.5. The systematic name of this enzyme class is riboflavin hydrolase. This enzyme participates in riboflavin metabolism.

• Foster JW, Yanagita T (August 1956). "A bacterial riboflavin hydrolase". The Journal of Biological Chemistry. 221 (2): 593–607. PMID 13357454.