Estrogenic Effect of Main Components Kakkalide and Tectoridin of Puerariae Flos and Their Metabolites

Abstract

Most herbal medicines are orally administered as decoctions. The components of these herbal medicines are therefore inevitably brought into contact with intestinal microflora in the alimentary tract. Some components may be transformed by intestinal bacteria before absorption from the gastrointestinal tract. Studies on the metabolism of such components by human intestinal microflora are of great importance to understanding their biological effects. 1,2)The flower of Pueraria thunbergiana (family Leguminosae), which contains isoflavones such as kakkalide and tectoridin as main components, is frequently used for counteracting symptoms associated with drinking alcohol, liver injury, and menopause in the traditional medicine clinic.3-6) Lee et al. and Han et al. reported that kakkalide isolated from the flower of P. thunbergiana (Puerariae Flos, PF) is metabolized to irisolidone by human intestinal microflora, and that irisolidone shows hepatoprotective activity. 7,8) Park et al. reported that tectoridin isolated from PF is metabolized to tectorigenin, which showed more potent antiallergic activity than tectoridin. 9) Many researchers have reported that isoflavones exhibit estrogenic effect. [10][11][12] Nevertheless, the estrogenic effects of kakkalide and tectoridin have not been studied.Therefore we isolated kakkalide and tectoridin from PF as well as their metabolites by human intestinal microflora and measured their estrogenic effects. MATERIALS AND METHODSMaterials Sulforhodamine B (SRB), NP40, 17b-estradiol, p-nitrophenyl-b-D-xylopyranoside (PNX), Dulbecco's modified Eagle medium (DMEM), fetal bovine serum (FBS), and charcoal dextran-stripped FBS (CD-FBS) were purchased from Sigma Chem. Co. (U.S.A.). General anaerobic medium (GAM) was purchased from Nissui Pharmaceutical Co., Ltd., (Tokyo, Japan). Protein assay reagent was purchased from Bio-Rad Laboratories (U.S.A.). The enhanced chemiluminescence (ECL) kit was purchased from Amersham Co. (U.S.A.). Progesterone receptor (PR) was purchased from Santa Cruz Co. (U.S.A.). Protease inhibitor cocktail was purchased from Roche Co. (Germany). All other chemicals were of analytical reagent grade, and all solutions used after redistillation.Kakkalide, kakkalidone, tectoridin, and ginsenoside Ra1 were isolated according to our previously reported methods. 7,8,13) Preparation of Specimens Fresh fecal specimens from volunteers (male adults in their twenties) (3 g) were septically collected in plastic cups, carefully mixed by a spatula, and suspended with 27 ml of cold saline. The fecal suspension was centrifuged at 100ϫg for 5 min. The supernatant was then centrifuged at 10000ϫg for 20 min. The resulting precipitates (about 0.3 g) were used as metabolic enzyme source for the assay of kakkalide-or tectoridin-hydrolyzing activity.Isolation and Culture of Kakkalide-Hydrolyzing Bacteria Bacterial strains previously isolated from feces of healthy men and some lactic acid bacteria were cultured in GAM broth and assayed for kakkalide-transforming activity. Isolated bacteria wer...

Preliminary Research for the Anti-obesity Effect of Puerariae Flos Extract in Humans

“…1,2) In East Asia, this flower has also recently been used in herbal medicines for the management of menopausal symptoms. 4,5) Puerariae thomsonii is known to contain 7 isoflavones (4 isoflavone glucosides: tectoridin, tectorigenin 7-O-xylosylglucoside, 6-hydroxygenistein-6,7-diglucoside, and glycitin; and 3 aglycones: tectorigenin, glycitein, and genistein) and 3 saponins (soyasaponin I, kaikasaponin III, and kakkasaponin I). 6) It has been reported that isoflavones such as genistein affect lipid metabolism in the liver and inhibit adipocyte differentiation, 7,8) and that saponins have an inhibitory effect on lipase activity.…”

Isolation and Identification of Urinary Metabolites of Kakkalide in Rats

“…Ohwi, where kakkalide is a predominant isoflavone, and Pueraria thomsonii Benth, where tectoridin is a major isoflavone ). Pharmacologically, kakkalide, irisolidone-7-O-␤-D-xylopyranosyl-(136)-␤-D-glucopyranoside, shows a wide spectrum of bioactivities such as hepatoprotection (Yamazaki et al, 1997;Han et al, 2003;Lee et al, 2005a), estrogenic effect (Shin et al, 2006), anti-inflammation (Park et al, 2007), antihyperlipidemia (Min and Kim, 2007), antioxidation , and others. The major metabolite in the blood of rats orally given kakkalide was its aglycone irisolidone, but not kakkalide itself.…”

“…Han et al (2003) isolated irisolidone and kakkalidone (irisolidone-7-O-␤-D-glucopyranoside) from an anaerobic medium containing kakkalide and human fecal bacteria, and they found that kakkalide exerts the protective activity against ethanol-induced mortality and hepatic injury in mice only by oral route. On the other hand, the activity of irisolidone was stronger than that of kakkalide whether it was given orally or intraperitoneally (Yamaki et al, 2002;Min and Kim, 2007;Shin et al, 2006). In addition, some isoflavone aglycone metabolites such as tectorigenin, glycitein, and genistein showed more potent activity than their glycoside precursors (Yamaki et al, 2002).…”

“…Thus, peoples consuming phytochemicals containing Puerariae Flos may be exposed to high levels of kakkalide. As a phytoestrogen, the metabolic forms of an isoflavone would be responsible for its activity and side effects (Shin et al, 2006). Thus, it is necessary to understand the metabolic fate of kakkalide.…”

Analysis of Xanthine Oxidase Inhibitors fromPuerariae flosUsing Centrifugal Ultrafiltration Coupled with HPLC-MS

“…Ohwi. In China, P. flos was traditionally used to treat diabetes mellitus and alcoholic intoxication because of its activities including antioxidant, 17 detoxification of alcohol, 18 hepatoprotective 19 and anticancer. 20 The aqueous extract of P. flos contained various isoflavonoids and triterpenoid saponins possessing pharmacological activity.…”