Metabolism of flavonoids mediated by cytochrome P450 2C subfamily of enzymes

U okviru ovog diplomskog rada provedena je analiza 11 flavonoida koji su prethodnim istraživanjima na humanim jetrenim mikrosomima pokazali metabolizam putem citokroma P450. Flavonoidi su polifenolni spojevi široko rasprostranjeni u biljkama koji se zbog svojeg antioksidirajućeg, protuupalnog, kardioprotektivnog te antitrombotičkog djelovanja nalaze u središtu ovog rada. Primjenom tekućinske kromatografije spregnute sa spektrometrijom masa odredili smo koji se od 11 flavonoida metabolizira preko određenog citokroma P450 2C potporodice enzima. Analizom kromatograma izdvojenog iona dobili smo podatke o masi, strukturi te molekulskoj formuli novonastalih metabolita, te smo iz razlika u masi odredili o kojem se metabolitu i reakciji radi. Od svih analiziranih spojeva, samo su dva flavonoida pokazala metabolizam putem citokroma P450 2C potporodice enzima, galangin i 7-hidroksiflavon. Oba navedena flavonoida metaboliziraju se putem citokroma 2C19 prilikom čega dolazi do aromatske hidroksilacije te nastaju metaboliti sakuranetin (iz galangina) i 4',7-dihidroksiflavon (iz 3-hidroksiflavona). Međutim, 4',7-dihidroksiflavon nastaje u izuzetno maloj količini, stoga su za a metabolizam 7-hidroksiflavona primarno odgovorni drugi enzimi, poput citokroma 3A4. Ovim radom upotpunili smo dosadašnje spoznaje o metabolizmu flavonoida te pomogli u budućoj prevenciji mogućih interakcija flavonoida i lijekova biotransformiranih citokromom 2C19.In this thesis, an analysis of 11 flavonoids was carried out. These flavonoids had shown, in earlier reasearch on human liver microsomes, metabolism mediated by cytochrome P450. Flavonoids are polyphenol compounds, widely found in plants, that are at the heart of this work because of its antioxidant, anti-inflammatory, cardioprotective and antithrombotic activity. Using liquid chromatography coupled with mass spectrometry, we determined which of the 11 flavonoids were metabolised by a specific cytochrome P450 2C subfamily of enzymes. Analysis of ion isolated chromatograph yielded data on the mass, structure and molecular formula of the new metabolites, and from the mass difference we determined which metabolite was formed and by which reaction. Of all the samples tested, only two flavonoids showed metabolism via cytochrome P450 2C subfamily of enzymes, galangin and 7-hydroxyflavone. Both flavonoids are metabolised by cytochrome 2C19 in a reaction called aromatic hydroxylation during which metabolites sakuranetin and 4',7-dihydroxyflavon are formed. However, the 4',7-dihydroxyflavon is produced in an extremely small amount, so other enzymes, such as CYP3A4, are primarily responsible for the 7-hydroxyflavone metabolism. With this work, we have sumplemented our understanding of flavonoid metabolism and assisted in the future prevention of possible interactions between flavonoids and other drugs biotransformed by cytochrome 2C19