Oxidative in Vitro Metabolism of Liquiritigenin, a Bioactive Compound Isolated from the Chinese Herbal Selective Estrogen β-Receptor Agonist MF101

Abstract

ABSTRACT:Liquiritigenin [2,3-dihydro-7-hydroxy-2-(4-hydroxyphenyl)-(S)-4H-1-benzopyran-4-one] is one of the major active compounds of MF101, an herbal extract currently in clinical trials for the treatment of hot flashes and night sweats in postmenopausal women. MF101 is a selective estrogen receptor ␤ agonist but does not activate the estrogen receptor ␣. Incubation with pooled human liver microsomes yielded a single metabolite. Its structure was elucidated using tandem mass spectrometry in combination with analysis of the fragmentation patterns. The metabolite resulted from the loss of two hydrogens and rearrangement to the stable 7,4-dihydroxyflavone. The structure was also confirmed by comparison with authentic standard material. Maximum apparent reaction velocity (V max ) and Michaelis-Menten constant (K m ) for the formation of 7,4-dihydroxyflavone were 32.5 nmol/g protein/min and 128 M, respectively. After correction for protein binding (free fraction ‫؍‬ 0.84), the apparent intrinsic clearance (CL int ) for 7,4-dihydroxyflavone formation was 0.3 ml/g/min. Liquiritigenin was almost exclusively metabolized by CYP3A enzymes. Comparison of liquiritigenin metabolism in human liver microsomes isolated from 16 individuals showed 9.5-fold variability in metabolite formation (3.4-32.2 nmol/g protein/min). An estrogen receptor luciferase assay indicated that the metabolite was a 3-fold more potent activator of the estrogen receptor ␤ than the parent compound and did not activate the estrogen receptor ␣.Flavonoids have been recognized for their anticarcinogenic, antioxidant, and anti-inflammatory properties. Over 4000 flavonoids have been isolated and identified from many types of fruits, vegetables, and herbs. Chemically, they can be categorized into flavonols, flavones, flavanones, isoflavones, catechins, anthocyanidins, and chalcones. Liquiritigenin is the 7,4Ј-dihydroxyflavanone and is found most notably in licorice. Licorice is one of the oldest and most commonly used Chinese herbal medicines and is used in many different medications. Usually, it is found concomitantly with liquiritin, which is the 4Ј-glycoside of liquiritigenin (Wang and Nixon, 2001;Moon et al., 2006).Liquiritigenin is an active ingredient of MF101, an herbal extract currently in clinical trials for the relief of postmenopausal symptoms in women. MF101 is an ethanol/aqueous extract of 22 herbal species used in traditional Chinese medicine (Cvoro et al., 2007). Because of its effects on the estrogen receptor, MF101 belongs to a class of compounds referred to as selective estrogen receptor modulators. Most clinically used estrogen receptor agonists or antagonists similarly affect both estrogen receptor ␣ and ␤. Selective estrogen receptor modulators discriminate between estrogen receptor ␣ and ␤ (Riggs and Hartmann, 2003). MF101 is a potent agonist of estrogen receptor ␤ but does not activate estrogen receptor ␣ (Cvoro et al., 2007). Most importantly, it is not implicated in tumor formation as a result of estrogen receptor ␣ activation (Cvoro e...

C5-Hydroxylation of liquiritigenin is catalyzed selectively by CYP1A2

“…The metabolite (M1) was observed as negative molecule [M-H] − at m/z 253 ( = 255 − 2, consistent with a loss of two hydrogens). The fragmentation patterns and exact masses of the parent and fragments were confirmed with 7,4-dihydroxyflavone ( Figure 3C), which was in accord with the CYP3A4-catalyzed metabolite described by Kupfer et al (2008). The metabolite (M2) was observed as negative molecule [M-H] − at m/z 271 ( = 255 + 16, consistent with a direct hydroxylation).…”

“…In recently published study of the oxidative metabolism of liquiritigenin by using HLMs, Kupfer et al (2008) found only one metabolite (7,4′-dihydroxyflavone). In vitro metabolism of liquiritigenin with RLMs, however, yielded six metabolites (7,3′,4′-trihydroxyflavone, a hydroxyl quinine metabolite, two A-ring dihydroxymetabolites, 7,4′-dihydroxyflavone, and 7-hydroxychromone) (Nikolic & van Breemen, 2004).…”

“…Recently, one oxidative metabolite of liquiritigenin (7,4′-dihydroxyflavone) was detected in HLMs and identified as CYP3A4-mediated reaction (Kupfer et al, 2008). However, no information is available to date about any other oxidative metabolites of liquiritigenin and which isozyme of CYP involved in these reactions in human.…”

Liquiritigenin Attenuates Alzheimer’s-Like Neuropathology in an Amyloid Protein Precursor Transgenic Mouse Model and the Underlying Mechanisms

“…Third, we have found that liquiritigenin does not induce the proliferation of MCF-7 or T47D breast cancer cells (unpublished observations), which is consistent with the results of other studies (Gustafsson et al, 2003). Finally, pharmacokinetics data has demonstrated that liquiritigenin is absorbed well by the intestine amongst rats, with an in vitro blood-brain barrier penetration rate of (29.7±6.8)% within 90 min, similar to that of chloramphenicol (34.0±4.9)%, a known blood-brain barrier and highly penetrative drug (Kupfer et al, 2008;Lu et al, 2008;Kang et al, 2009). Taken together, these observations suggest that liquiritigenin may be a potentially effective therapy for AD.…”

“…Liquiritigenin (7, 4'-dihydroxyflavanone) is a flavonoid extracted from the radix of Glycyrrhiza, an herbal that is frequently used to treat injury or swelling, or for detoxification in traditional Oriental medicine. Liquiritigenin is also one of the major active compounds of MF101 (Kupfer et al, 2008), an herbal extract currently used in clinical trials for the treatment of hot flushes and night-sweats in post-menopausal women. Our interest in liquiritigenin is based upon the following observations.…”