Identification of nobiletin metabolites in mouse urine

Abstract

Nobiletin, a major component of citrus polymethoxyflavones, has many potential significant health benefits. While the biological activities of nobiletin have been widely reported, its in vitro and in vivo metabolic fate has been rarely studied. To explore the biotransformation mechanism of nobiletin we conducted an investigation into its metabolic profile in mouse urine, by various analytical techniques. Due to sample amount limitations for isolating and characterizing an individual metabolite, two possible nobiletin metabolites were prepared in a similar multi-step organic synthetic route: 3'-hydroxy-5,6,7,8,4'-pentamethoxyflavone (3'-demethylnobiletin) and 4'-hydroxy-5,6,7,8,3'-pentamethoxyflavone (4'-demethylnobiletin). Normal phase (silica gel) and C(18) reverse phase chromatography, as well as liquid chromatography-mass spectrometry-mass spectrometry, were employed in the separation of 3'-demethylnobiletin and 4'-demethylnobiletin, however, without success due to the structural similarities of these mono-demethylated nobiletins. Using a chiral packed column eluted under supercritical fluid chromatography (SFC) conditions, a clear separation was achieved. Thus, by comparing the SFC profiles of metabolite mixtures with the synthesized standard compounds, the major nobiletin metabolite of mouse urine is identified as 4'-demethylnobiletin, whereas 3'-demethylnobiletin is a minor metabolite. In this study, the concentration of 4'-demethylnobiletin in mouse urine is 28.9 microg/mL.

Anti-dementia Activity of Nobiletin, a Citrus Flavonoid: A Review of Animal Studies

“…Notably, analyses using high-performance liquid chromatography (HPLC) and positron emission tomography (PET) suggest nobiletin penetrates the BBB and reaches a maximal level within 3-5 min after peripheral administration 70,71). Additionally, 4'-demethylnobiletin, a major urinary metabolite of nobiletin in rats and mice,72,73) can cross the BBB and exert memory-improving effects in the brain 74)…”

Polymethoxy flavones are responsible for the anti-inflammatory activity of citrus fruit peel

“…Manthey and Guthrie (2002) have also shown that polymethoxy flavones exhibited a higher anti-proliferative activity than had flavanone glycosides and could serve as potent anti-cancer agents. Indeed, the fact that the hydrophobic nature of the polymethoxy flavones enables them to cross the intestinal membrane easily, and makes them absorbed more readily into the human body (Li, Wang, Sang, Huang, & Ho, 2006). These results indicate polymethoxy flavones to be promising therapeutic phytochemicals of citrus by-products (Li, Yu, & Ho, 2006).…”

Comparative Study on Nobiletin Metabolism with Liver Microsomes from Rats, Guinea Pigs and Hamsters and Rat Cytochrome P450

“…By contrast, the time-course study and the immunoinhibition study using antiserum against hamster CYP1A2 indicated that CYP1A1 and CYP1A2 were the most important P450s in the MC-treated animals used in this study. 4Ј-Demethylated (4Ј-OH)-nobiletin has been reported to be the main metabolite in the urine of rats 24) and mice 25) orally administered nobiletin. However, 4Ј-OH-nobiletin (M-1) was not the most abundant metabolite among the five metabolites formed by liver microsomes of untreated rats (Table 1).…”

“…21) On the other hand, there are not many reports on the biotransformation of nobiletin in animals. [22][23][24][25] Yasuda et al found a few demethylated metabolites in 24 h-urine of rats orally administered nobiletin and determined that a major metabolite is 4Ј-demethylated (4Ј-OH) nobiletin. 24) However, the mechanism which the biological activities mentioned above are exerted by a parent compound or its metabolites remains obscure.…”