Kampo herbal ointments for skin wound healing.

The management of skin wound healing problems is a public health issue in which traditional herbal medicines could play a determining role. Kampo medicine, with three traditionally used ointments, provides interesting solutions for these dermatological issues. These ointments named Shiunko, Chuoko, and Shinsen taitsuko all have in common a lipophilic base of sesame oil and beeswax from which herbal crude drugs are extracted according to several possible manufacturing protocols. This review article brings together existing data on metabolites involved in the complex wound healing process. Among them are representatives of the botanical genera Angelica, Lithospermum, Curcuma, Phellodendron, Paeonia, Rheum, Rehmannia, Scrophularia, or Cinnamomum. Kampo provides numerous metabolites of interest, whose content in crude drugs is very sensitive to different biotic and abiotic factors and to the different extraction protocols used for these ointments. If Kampo medicine is known for its singular standardization, ointments are not well known, and research on these lipophilic formulas has not been developed due to the analytical difficulties encountered in biological and metabolomic analysis. Further research considering the complexities of these unique herbal ointments could contribute to a rationalization of Kampo's therapeutic uses for wound healing.

Ameliorating Effect of the Edible Mushroom Hericium erinaceus on Depressive-Like Behavior in Ovariectomized Rats.

Estrogen deficiency during menopause causes a variety of neurological symptoms, including depression. The edible Lion's Mane mushroom, Hericium erinaceus (Bull.: Fr.) Pers. (HE), is a medicinal mushroom that has the potential for a neuroprotective effect and ameliorating neurological diseases, such as depression, anxiety, and neurodegenerative diseases. HE contains phytoestrogens, including daidzein and genistein. However, the ameliorating effect of HE on menopausal symptoms is not well understood. Here we investigated the impact of methanol extract of the HE fruiting body on depressive-like behavior in postmenopausal model rats. The activation of estrogen receptor alpha (ERα) causes body weight loss and uterine weight gain. Body weight gain and uterine weight loss by estrogen deficiency in ovariectomized (OVX) rats were reversed with 17ß-estradiol (E2) but not with HE. Thus, the phytoestrogens in HE may hardly activate ERα. Estrogen receptor beta (ERß) is expressed in the brain, and activation of ERß ameliorates menopausal depressive symptoms. Notably, depressive-like behavior in OVX rats evaluated in forced swim test was reduced by administration of not only E2 but also HE for 92 d. Long-term activation of ERα increases the risk of breast and uterine cancers. HE, therefore, may be effective in treating menopausal depression without the risk of carcinogenesis caused by ERα activation.

Cajanin Suppresses Melanin Synthesis through Modulating MITF in Human Melanin-Producing Cells.

Despite its classification as a non-life-threatening disease, increased skin pigmentation adversely affects quality of life and leads to loss of self-confidence. Until now, there are no recommended remedies with high efficacy and human safety for hyperpigmentation. This study aimed to investigate anti-melanogenic activity and underlying mechanism of cajanin, an isoflavonoid extracted from Dalbergia parviflora Roxb. (Leguminosae) in human melanin-producing cells. Culture with 50 µM cajanin for 48-72 h significantly suppressed proliferation in human melanoma MNT1 cells assessed via MTT viability assay. Interestingly, cajanin also efficiently diminished melanin content in MNT1 cells with the half maximum inhibitory concentration (IC50) at 77.47 ± 9.28 µM. Instead of direct inactivating enzymatic function of human tyrosinase, down-regulated mRNA and protein expression levels of MITF and downstream melanogenic enzymes, including tyrosinase, TRP-1 and Dct (TRP-2) were observed in MNT1 cells treated with 50 µM cajanin for 24-72 h. Correspondingly, treatment with cajanin modulated the signaling pathway of CREB and ERK which both regulate MITF expression level. Targeted suppression on MITF-related proteins in human melanin-producing cells strengthens the potential development of cajanin as an effective treatment for human hyperpigmented disorders.

Effects of Pueraria candollei var mirifica (Airy Shaw and Suvat.) Niyomdham on Ovariectomy-Induced Cognitive Impairment and Oxidative Stress in the Mouse Brain.

The effects of the phytoestrogen-enriched plant Pueraria mirifica (PM) extract on ovari-ectomy (OVX)-induced cognitive impairment and hippocampal oxidative stress in mice were investigated. Daily treatment with PM and 17ß-estradiol (E2) significantly elevated cognitive behavior as evaluated by using the Y maze test, the novel object recognition test (NORT), and the Morris water maze test (MWM), attenuated atrophic changes in the uterus and decreased serum 17ß-estradiol levels. The treatments significantly ameliorated ovariectomy-induced oxidative stress in the hippocampus and serum by a decrease in malondialdehyde (MDA), an enhancement of superoxide dismutase, and catalase activity, including significantly down-regulated expression of IL-1ß, IL-6 and TNF-α proinflammatory cytokines, while up-regulating expression of PI3K. The present results suggest that PM extract suppresses oxidative brain damage and dysfunctions in the hippocampal antioxidant system, including the neuroinflammatory system in OVX animals, thereby preventing OVX-induced cognitive impairment. The present results indicate that PM exerts beneficial effects on cognitive deficits for which menopause/ovariectomy have been implicated as risk factors.

α-Keto tetrahydrofuran lignan glucosides from the Bangladeshi medicinal plant Terminalia citrina inhibit estradiol (E2) induced proliferation in cancer cells.

EtOAc extract from the leaves of Terminalia citrina collected in Bangladesh were separated, and seven previously undescribed α-keto tetrahydrofuran lignan glucosides (terminalosides Q to W) were isolated and characterized. NOESY analysis of 1H NMR spectra and ECD spectroscopic data analysis revealed the absolute stereochemistry of the tetrahydrofuran ring of the isolated constituents as being a (7S,8R,8'S)- configuration in terminalosides Q to U and a (7R,8R,8'S)- configuration in terminalosides V and W. All of the isolated compounds were evaluated for their estrogenic and anti-estrogenic properties using two types of estrogen-responsive human breast cancer cell lines (MCF-7 and T47D). Terminaloside R, which has a dioxymethylene group in its aromatic ring, inhibited 90% of estradiol-enhanced cell proliferation in T47D and MCF-7 cells at concentrations of 0.01 µM and 0.1 µM, respectively. On the other hand, terminaloside T, the analogous compound which has two oxymethyl groups in place of dioxymethylene, suppressed 90% of cell proliferation selectively in T47D cells at a concentration of 0.01 µM. However, terminaloside W, the 7R-stereoisomer of terminaloside R, only showed moderate activity.

Naphthalene glycosides in the Thai medicinal plant Diospyros mollis.

This study evaluated methanol extracts from the leaves and branches of the Thai medicinal plant Diospyros mollis (Ebenaceae). Seven triterpenes and 22 aromatic compounds, including five new compounds, were isolated, and their structures were determined. The new compounds had the following structures: diospyrol glycoside (makluoside A, 1), 8,8'-di-O-6-ß-D-apiofuranosyl-ß-D-glucopyranosyl-6,6'-dimethyl-2,3'-binaphthalene-1-ol-1',4'- dione (makluoside B, 2), and 3-methyl-1,8- naphthalenediol glycosides (makluosides C-E, 3-5). Makluoside B is the first example of a naphthoquinone glycoside that has both a 3-methyl-1,8-naphthalenediol unit and a 5-hydroxy-7-methyl-1,4-naphthoquinone unit. The hyaluronidase inhibitory activity of the isolates was evaluated, revealing that one of the triterpene derivatives possessed moderate inhibitory activity.

Isolation of a new compound, 2-butanone 4-glucopyranoside 6'-O-gallate and other 8 compounds from the anti-inflammatory leave extracts of Memecylon edule Roxb.

This present study was designed to isolate the compounds of Memecylon edule. The chemical compounds were purified by chromatographic methods and their structures were established on the basis of spectroscopic analyses (UV, MS and NMR). The major isolated compounds were tested for anti-inflammatory activity. The methanolic extracts of M. edule leaves gave a new compound 2-butanone 4-glucopyranoside 6'-O-gallate (1) with eight known compounds, namely, 3,3'-di-O-methylellagic acid 4-O-ß-d-glucopyranoside (2), epigallocatechin-3-O-gallate (EGCG) (3), (2R, 3R)-dihydromyricetin-4'-ß-d-glucopyranoside (4), myricetin-3-O-α-l-rhamnopyranoside (5), benzyl-(6-O-α-L arabinofuranosyl) O-ß-d-glucopyranoside (6), benzyl-(6-O-α-l-rhanmopyranosyl) O-ß-d-glucopyranoside (7), 2-phenylethyl-(6-O-ß-d-apiofuranosyl)-O-ß-d-glucopyranoside (8) and methyl benzoate 2-(6-O-α-l-rhamnosyl)-O-ß-d-glucopyranoside (9). All compounds were isolated for the first time from this plant. The major compounds (2, 3 and 5) exhibited significant anti-inflammatory activity. In conclusion, M. edule was recognised to be a good source for phenolic compounds and these compounds may contribute to anti-inflammatory activity of the extract.

A novel steroid and cytotoxic constituents from Dioscorea membranacea Pierre against hepatocellular carcinoma and cholangiocarcinoma cells.

ETHNOPHARMACOLOGICAL RELEVANCE: The rhizomes of Dioscorea membrancea Pierre have been used in Thai traditional medicine as an ingredient formula for liver cancer and cholangiocarcinoma treatment. AIM OF THE STUDY: To investigate the cytotoxic activity of ethanolic extract and constituents of D. membrancea to support its traditional use. MATERIALS AND METHODS: The SRB assay was used to determine the cytotoxic activity against hepatocellular carcinoma (HepG2), cholangiocarcinoma (KKU-M156) cells and one normal human keratinocyte immortal cells (HaCaT) with its ethanolic extract and isolated compounds. Bioassay guided isolation was used for isolating cytotoxic compounds. RESULTS: The ethanolic extract of D. membranacea rhizome showed weak cytotoxic against KKU-M156 and HepG2 (IC50 at 72h exposure=30.49±0.82 and 38.97±2.04µg/mL respectively). A new steroid [epipanthogenin B (1)], a known steroid [panthogenin B (2)], two napthofuranoxepins [dioscorealide A (3) and dioscorealide B (4)], phenanthraquinone [dioscoreanone (5)] and two phenanthrene [5,6-dihydroxy-2,4-dimethoxy-9,10-dihydrophenanthrene (6) and 2,5,6-trihydroxy-3,4-dimethoxy, 9, 10-dihydrophenanthrene (7)] were isolated from active chloroform fraction. Compound 4 showed the highest cytotoxicity against HepG2 (IC50 at 72h exposure=2.87±0.21µM) and KKU-M156 (IC50 at 72h exposure=1.67±0.10µM) and less toxicity against normal cell line (HaCaT) (IC50 at 72h exposure>100µM). Compound 5 showed selective cytotoxic activity against KKU-M156 (IC50 at 72h exposure=3.46±0.11µM). Compounds 6 and 7 showed weak cytotoxic activity against HepG2 (IC50 at 72h exposure=24.96±2.32 and 51.31±3.52µM). Compounds (1-3) showed no cytotoxic activity against HepG2 and KKU-M156 cell lines (IC50 at 72 h exposure>100µM). CONCLUSION: Seven compounds were isolated from active chloroform fraction of the ethanolic extract of D. membranacea rhizomes. Only dioscorealide B (4) might be served as a good anticancer agent for liver cancer and cholangiocarcinoma cancer because it can kill cancer cell but not toxic on normal cell. This research support Thai traditional medicine use of D. membranacea for liver cancer and cholangiocarcinoma cancer treatment.

Five furofuranone lignan glucosides from Terminalia citrina inhibit in vitro E2-enhanced breast cancer cell proliferation.

Five new polyalkoxylated furofuranone lignan glucosides, terminalosides L-P (1-5), were isolated from EtOAc extracts of the leaves of Terminalia citrina, a Bangladeshi medicinal plant. The structures of the isolates were deduced primarily by NMR spectroscopy, and four of the isolates were found to contain rare tetraoxygenated aryl groups in their structures. The absolute configurations and conformations of the furofuranone ring were confirmed by ECD spectroscopy. All of the isolates were evaluated for their estrogenic and/or antiestrogenic properties using two estrogen responsive breast cancer cell lines, T47D and MCF-7. At a concentration of 10nM, terminaloside L (1) suppressed E2-enhanced T47D cell proliferation by 90%, while terminaloside M (2) showed 90% antiestrogenic activity against MCF-7 cells. Compared to 2, the antiestrogenic activity of terminaloside O (4) and P (5) was weak, possibly due to the different attachment positions of the sugar moiety that they share in common. This is the first report of furofuranone lignans from any Terminalia species, and also of their antiestrogenic activity.

Furofuran Lignan Glucosides with Estrogen-Inhibitory Properties from the Bangladeshi Medicinal Plant Terminalia citrina.

Extracts from the leaves of the Bangladeshi medicinal plant Terminalia citrina were prepared, and 13 new furofuran lignan glucosides, terminalosides A-K (1-4, 6-12), 2-epiterminaloside D (5), and 6-epiterminaloside K (13), were characterized using various spectroscopic techniques. Twelve of the isolates were found to contain rare tetraoxygenated aryl groups in their structures. Analysis of the NMR chemical shifts for the oxymethine signals in the furofuran ring suggested a pragmatic approach to determining the relative configuration of these compounds. The ECD and NOESY spectroscopic data obtained allowed for the deduction of the absolute configurations and conformations of the compounds. The isolates were tested for their estrogenic/antiestrogenic activity using the MCF-7 and T47D estrogen-responsive human breast cancer cell lines. Terminalosides B (2) and G (8) exhibited inhibitory effects for both cell lines, and estradiol-enhanced cell proliferation was suppressed by 90% at concentrations lower than 10 µM. Terminaloside E (6) showed inhibitory activity against the T47D cell line, whereas terminalosides C (3), F (7), and I (10) and 6-epiterminaloside K (13) displayed antiestrogenic activity against MCF-7 cells.

Phenolic constituents of the Bangladeshi medicinal plant Pothos scandens and their anti-estrogenic, hyaluronidase inhibition, and histamine release inhibitory activities.

Extracts from the stem and roots of the Bangladeshi medicinal plant Pothos scandens L. (Araceae) were isolated, and three hemiterpene glucoside aromatic esters, pothobanosides A (1), B (2), and C (3), and a phenylisobutanoid, pothobanol (4), along with 14 known compounds, were characterized. The isolates were tested for their estrogenic/anti-estrogenic activity using the estrogen-responsive human breast cancer cell lines MCF-7 and T47D, and syringoyl derivatives (2, 3, and canthoside B) showed strong inhibitory activity against both cell lines. Their less oxygenated analogs (1, and markhamioside F) were almost inactive. The isolates were also evaluated for hyaluronidase and histamine release inhibitory activities, and pothobanoside A (1) showed significant hyaluronidase inhibitory activity among the isolated compounds, which was similar to that of the positive control rosmarinic acid. Because hyaluronidase produces an angiogenic response that has been implicated in tumor invasiveness and metastasis, 1 could be valuable as an anti-tumor compound with a different mechanism of action from related compounds (2, 3). Pothobanoside C (3) and pothobanol (4) were also found to inhibit histamine release to a similar degree to the positive control epigallocatechin 3-O-(3"-O-methyl)-gallate. The histamine release inhibitory potency of these isolates may support the traditional uses of this plant in folk medicine.

Effect of miroestrol on ovariectomy-induced cognitive impairment and lipid peroxidation in mouse brain.

Miroestrol (MR) is a phytoestrogen isolated from Pueraria candollei var. mirifica (KwaoKrueaKhao), a Thai medicinal plant used for rejuvenation. We examined the effects of MR on cognitive function, oxidative brain damage, and the expression of genes encoding brain-derived neurotrophic factor (BDNF) and cyclic AMP-responsive element-binding protein (CREB), factors implicated in neurogenesis and synaptic plasticity, in ovariectomized (OVX) mice. OVX decreased serum 17ß-estradiol level and uterine weight. OVX also impaired object recognition performance in the novel object recognition test and spatial cognitive performance in the Y-maze test and the water maze test. Daily treatment of MR dose-dependently attenuated OVX-induced cognitive dysfunction. Moreover, OVX mice had a significantly increased level of thiobarbituric acid-reactive substances, and down-regulated expression levels of BDNF and CREB mRNAs in the hippocampus and frontal cortex. MR treatment as well as hormone replacement therapy with 17ß-estradiol significantly reversed these neurochemical alterations caused by OVX. These results suggest that MR ameliorates cognitive deficits in OVX animals via attenuation of OVX-induced oxidative stress and down-regulation of BDNF and CREB mRNA transcription in the brain. Our findings raise the possibility that MR and Pueraria candollei var. mirifica, the plant of origin of MR, may have a beneficial effect on cognitive deficits like AD in which menopause/ovariectomy are implicated as risk factors.

Structure and antioxidant activity relationships of isoflavonoids from Dalbergia parviflora.

The antioxidant activities of 24 isoflavonoids that were previously isolated as pure compounds from Dalbergia parviflora were evaluated using three different in vitro antioxidant-based assay systems: xanthine/xanthine oxidase (X/XO), ORAC, and DPPH. The isolates consisted of three subgroups, namely isoflavones, isoflavanones, and isoflavans, each of which appeared to have diversified substituents, and were thus ideal for the study of their structure-activity relationships (SARs). The SAR analysis was performed using the results obtained from both the inter-subgroup isoflavonoids with the same substitution pattern and the intra-subgroup compounds with different substitution patterns. The inter-subgroup comparison showed that the isoflavones exhibited the highest antioxidant activities based on all three assays. The intra-subgroup analysis showed that the additional presence of an OH group in Ring B at either R3' or R5' from the basic common structure of the R7-OH of Ring A and the R4'-OH (or -OMe) of Ring B greatly increased the antioxidant activities of all of the isoflavonoid subgroups and that other positions of OH and OMe substitutions exerted different effects on the activities depending on the subgroup and assay type. Therefore, based on the structural diversity of the isoflavonoids in D. parviflora, the present study provides the first clarification of the detailed antioxidant SARs of isoflavonoids.

Cytotoxic and immunomodulatory effects of polyhydroxyoctane isolated from Lentinus polychrous mycelia.

A polyhydroxyoctane, 6-methylheptane-1,2,3,4,5-pentaol (MHP), was first isolated from mycelia of the Thai edible mushroom Lentinus polychrous. MHP was evaluated for its cytotoxic and immunomodulatory effects in vitro. MHP was slightly cytotoxic to murine splenocytes but not to RAW264.7 cells or peripheral blood mononuclear cells. MHP decreased nitric oxide and intracellular O2 (-) production from lipopolysaccharide- and phorbol-12-myristate-13-acetate-activated RAW264.7 cells at levels of 78.98 ± 4.72 and 78.48 ± 2.41 % of controls, respectively. The mRNA expression of pro-inflammatory mediators, including iNOS, TNF-α, IL-1ß, IL-6, COX-1 and COX-2, were significantly suppressed by MHP. In addition, MHP significantly increased the proliferation of phytohemagglutinin and pokeweed mitogen-induced splenocytes. These results indicate that MHP is able to modulate inflammatory responses and the proliferation of both T- and B-lymphocyte cells, suggesting that MHP may be a good natural immunomodulator.

Diastereomers of lithospermic acid and lithospermic acid B from Monarda fistulosa and Lithospermum erythrorhizon.

Monardic acids A (1) and B (2), which are (7R,8R) diastereomers of lithospermic acid (LA) and lithospermic acid B, respectively, were isolated from Monarda fistulosa. A (7S,8R) isomer (3) of LA was also isolated from this plant, and a (7R,8S) isomer (7) of LA was obtained from Lithospermum erythrorhizon. The absolute configuration of 1 was confirmed by analysis of its hydrolysates, 7-epiblechnic acid and 2R-3-(3,4-dihydroxyphenyl)-2-hydroxypropanoic acid. The configuration in the dihydrobenzofuran moieties of 2, 3, and 7 was extrapolated by using the phenylglycine methyl ester method and a Cotton effect at approximately 250-260 nm in their electronic circular dichroism spectra. Diastereomers (1-3 and 7) displayed moderate hyaluronidase inhibitory and histamine release inhibitory activities.

Flavonoid glycosides from Botrychium ternatum.

The MeOH extract from dried whole Botrychium ternatum plants yielded 33 compounds, including seventeen new flavonoid glycosides and sixteen known compounds. The structures of new compounds were established using NMR spectroscopic analysis and chemical evidence.

Flavonoid and stilbenoid production in callus cultures of Artocarpus lakoocha.

Callus cultures of Artocarpus lakoocha Roxb., established from seedling explants and maintained on woody plant medium containing 1mg/l 2,4-dichlorophenoxyacetic acid and 1mg/l benzyladenine, were studied for their chemical constituents and biosynthetic potential of secondary metabolites. Four prenylflavones and prenylated stilbenes, along with nine known polyphenolic compounds, were isolated and elucidated for their structures through extensive analysis of their NMR and MS data. Among the 13 isolates, it appeared that seven of them are prenylated derivatives of 5,7,2',4'-tetrahydroxyflavones, and four are prenylated derivatives of 2,4,3',5'-tetrahydroxystilbene (oxyresveratrol), suggesting that the biosynthetic pathways of these two polyphenolic groups and their prenylating enzymes are highly expressed in A. lakoocha callus cultures. A study on the growth-product relationship of the callus cultures showed that the secondary metabolites were all formed simultaneously during the rapid growth phase of the culture cycle, with various prenylflavones, and a prenylated stilbene as major constituents. In assays for DPPH free radical scavenging activity and tyrosinase inhibitory potential, the stilbenoids appeared to possess moderate effects, whereas the flavonoids showed only weak activity.

Constituents from the roots of Taraxacum platycarpum and their effect on proliferation of human skin fibroblasts.

A MeOH extract from the roots of Taraxacum platycarpum has shown significant effects on the proliferation of normal human skin fibroblasts. Chemical analysis of the extract resulted in the isolation of 26 compounds, including eight new triterpenes, one new sesquiterpene glycoside, and seventeen known compounds. The structure of each new compound was established using NMR spectroscopy. Some triterpenes had a significant effect on the proliferation of normal human skin fibroblasts.

8,12;8,20-diepoxy-8,14-secopregnane glycosides from roots of Asclepias tuberosa and their effect on proliferation of human skin fibroblasts.

A pregnane glycoside fraction from the roots of Asclepias tuberosa L. caused normal human skin fibroblasts to proliferate. This fraction contained 21 pregnane glycosides whose structures were established using NMR spectroscopic analysis and chemical evidence. The aglycones of most of these compounds were identified as 8,12;8,20-diepoxy-8,14-secopregnanes, such as tuberogenin or 5,6-didehydrotuberogenin, the same aglycones as constituents of the aerial parts of this plant. Some of these compounds also caused proliferation of skin fibroblasts.

Estrogenic and anti-estrogenic compounds from the Thai medicinal plant, Smilax corbularia (Smilacaceae).

From the rhizomes of Smilax corbularia Kunth. (Smilacaceae), 11 compounds, (2R,3R)-2″-acetyl astilbin, (2R,3R)-3″-acetyl astilbin, (2R,3R)-4″-acetyl astilbin, (2R,3R)-3″-acetyl engeletin, (2R,3S)-4″-acetyl isoastilbin, 2-(4-hydroxyphenyl)-3,4,9,10-tetrahydro-3,5-dihydroxy-10-(3,4-dihydroxyphenyl)-(2R,3R,10R)-2H,8H-benzo [1,2-b:3,4-b'] dipyran-8-one, 2-(4-hydroxyphenyl)-3,4,9,10-tetrahydro-3,5-dihydroxy-10-(3,4-dihydroxyphenyl)-(2R,3R,10S)-2H, 8H-benzo [1,2-b:3,4-b'] dipyran-8-one, 3,4-dihydro-7-hydroxy-4-(3,4-dihydroxyphenyl)-5-[(1E)-2-(4-hydroxyphenyl) ethenyl]-2H-1-benzopyran-2-one, 3,4-dihydro-7-hydroxy-4-(3,4-dihydroxy-phenyl)-5-[(1E)-2-(3,4-dihydroxyphenyl) ethenyl]-2H-1-benzopyran-2-one, 3,4-dihydro-7-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-5-[(1E)-2-(4-hydroxyphenyl) ethenyl]-2H-1-benzopyran-2-one, and 5,7,3',4'-tetrahydroxy-3-phenylcoumarin along with 34 known compounds were isolated and characterized as 19 flavonoids, 14 catechin derivatives, 6 stilbene derivatives, and 6 miscellaneous substances. All isolates had their estrogenic and anti-estrogenic activities determined using the estrogen-responsive human breast cancer cell lines MCF-7 and T47D. The major constituents were recognized as flavanonol rhamnosides by the suppressive effect on estradiol induced cell proliferation at a concentration of 1µM. Meanwhile, flavanonol rhamnoside acetates demonstrated estrogenic activity in both MCF-7 and T47D cells at a concentration of 100µM, and they enhanced the effects of co-treated E2 on T47D cell proliferation at concentrations of more than 0.1µM.