Systematic Analysis Reveals Two Novel <i>O</i> -Methyltransferase Genes Involved in Flavonoid Biosynthesis in Ginkgo Leaves

Flavonoids are widely distributed in plants, including <i>Ginkgo biloba</i>, a traditional medicinal and edible species. Methylated flavonoids often exhibit enhanced biological activities compared with their unmethylated forms, and flavonoid <i>O</i>-methyltransferases (FOMTs) responsible for this modification have been identified in several plants but remain poorly characterized in ginkgo. In this study, transcriptomic analysis of ginkgo leaves at different developmental stages identified two <i>S</i>-adenosylmethionine-dependent FOMTs, <i>GbFOMT3</i> and <i>GbFOMT12</i>. Enzymatic assays with seven flavonoid substrates showed that both enzymes methylate quercetin, eriodictyol, and luteolin to produce isorhamnetin, hesperetin, and chrysoeriol, respectively, through methylation at the 3'-OH and 4'-OH positions of the B ring. These findings expand the repertoire of ginkgo flavonoid <i>O</i>-methyltransferases and provide insights into flavonoid methylation in ginkgo.