JOURNAL OF MASS SPECTROMETRY J. Mass Spectrom. 2005; 40: 289–299 Published online 15 February 2005 in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/jms.753 Metabolism of xanthohumol and isoxanthohumol, prenylated flavonoids from hops (Humulus lupulus L.), by human liver microsomes Dejan Nikolic, Yongmei Li, Lucas R. Chadwick, Guido F. Pauli and Richard B. van Breemen * Department of Medicinal Chemistry and Pharmacognosy, UIC/NIH Center for Botanical Dietary Supplements Research, College of Pharmacy, University of Illinois at Chicago, 833 S. Wood Street, Chicago, Illinois 60612-7231, USA Received 1 July 2004; Accepted 23 September 2004 The female flowers of hops (Humulus lupulus L.) used to flavor beer contain the prenylated flavonoids xanthohumol (XN) and isoxanthohumol (IX). IX is moderately estrogenic in vitro and XN has pharmacological properties that might make it useful as a cancer chemopreventive agent. The metabolism of these dietary flavonoids was investigated in vitro using human liver microsomes. Hydroxylation of a prenyl methyl group was the primary route of oxidative metabolism forming either cis or trans hydroxylated metabolites of IX but only the trans isomer of XN. The double bond on the prenyl group of both compounds formed an epoxide which was opened by an intramolecular reaction with the neighboring hydroxyl group. The potent phytoestrogen 8-prenylnaringenin (8-PN) was detected as a demethylation product of IX. However, the analogous demethylation reaction was not observed for XN. Since XN can be converted to IX through acid-catalyzed cyclization in the stomach, XN might contribute to the in vivo levels of estrogenic 8-PN following consumption of hops extracts. Copyright  2005 John Wiley & Sons, Ltd. KEYWORDS: hops; xanthohumol; isoxanthohumol; metabolism; cytochrome P450 INTRODUCTION The female flowers of hops (Humulus lupulus L.) are used in the brewing industry as a flavoring agent for beer. For many years there has been a suspicion that hops are estrogenic, and recently, interest in this possibility has been increasing. There is anecdotal evidence of menstrual disturbances among female pickers of hops, and in Germany hops baths have been used to reduce hot flushes in menopausal women. 1,2 Furthermore, hop extracts are the major constituent of many commercial preparations marketed for breast enhancement. 3 Although some studies have reported that hop extracts are not estrogenic, 4 others have found a high estrogenic effect. 5 Because of on-going exposure of the human population to hop products, we are investigating the estrogenicity of hop extracts and their active compounds. Among the possible active constituents, prenylated flavonoids have received the most attention and may L Correspondence to: Richard B. van Breemen, Department of Medicinal Chemistry and Pharmacognosy, UIC/NIH Center for Botanical Dietary Supplements Research, College of Pharmacy, University of Illinois at Chicago, 833 S. Wood Street, Chicago, Illinois 60612-7231, USA. E-mail: breemen@uic.edu Contract/grant sponsor: Office of Dietary Supplements; Contract/grant number: P50AT00155. Contract/grant sponsor: National Institute of General Medical Sciences. Contract/grant sponsor: National Center for Complementary and Alternative Medicine. Contract/grant sponsor: Office for Research on Women’s Health. be divided into two major groups, prenylated chalcones and prenylated flavanones. The prenylated flavanone 8- prenylnaringenin (8-PN) is notable because it is a potent estrogen. 1 In hop cones, the most abundant prenylated chalcone is xanthohumol (XN; see structure in Fig. 1) which accounts for ¾82–89% of the prenylated flavonoids in this plant. 6 XN is not estrogenic but exhibits strong antiproliferative activity against breast, colon and ovarian cancer cell lines. 7 In addition, XN is an inhibitor of cytochrome P450 isozymes 8 and a potent inducer of quinone reductase induction activity. 9 Gerh¨ auser et al. 10 identified several possible chemopreventive properties of XN using a panel of bioassays and found that XN exhibits a broad spectrum of inhibition of various stages of carcinogenesis. Recently, Zhao et al. 11 found that XN can inhibit nitric oxide production by suppressing the expression of inducible nitric oxide synthase. Because of these activities, XN has been advocated as an agent for cancer chemoprevention. Isoxanthohumol (IX; see structure in Fig. 1) may be regarded as a cyclization product of XN and, as a result, the levels of IX in hops preparations vary according to the conditions used to extract the cones and store the extracts. 12,13 The biological activities of IX have not been thoroughly investigated, but IX has been shown to be moderately estrogenic in vitro. 1 Because XN and IX are abundant hop constituents, it is of interest to determine their human metabolism especially if metabolic transformation might convert them to more potent estrogenic compounds. However, little is known about the Copyright  2005 John Wiley & Sons, Ltd.