Curcuma zedoaria 50% methanol extracts increase adiponectin secretion by enhancing PPARγ mRNA expression in 3T3-L1 cells.

Curcuma zedoaria is a characteristic species of its genus that contains little to no curcuminoid. Here, we demonstrate that C. zedoaria extracts with 50% methanol increases adiponectin secretion into the media by enhancing PPARγ mRNA expression in 3T3-L1 cells. These results indicate that C. zedoaria may be useful for preventing/improving lifestyle-related diseases such as diabetes and atherosclerosis.

Influence of particle size on the in vitro and in vivo anti-inflammatory and anti-allergic activities of a curcumin lipid nanoemulsion.

The polyphenolic compound, curcumin, is a natural yellow pigment component of turmeric. It exerts various biological effects, such as anti-inflammatory effects, and we have previously demonstrated that curcumin is a specific inhibitor of DNA polymerase λ. Curcumin is characterized by poor bioavailability as it is water-insoluble, is poorly absorbed and is systemically eliminated. In order to increase the bioavailability of curcumin, in this study, we produced a curcumin-loaded lipid nanoemulsion of various particle sizes (50, 100 and 200 nm). The curcumin lipid nanoemulsion was prepared by a modified thin-film hydration method followed by sonication. To identify the optimal particle size which exhibits the strongest physiological activity, we investigated the inhibitory effects of the obtained nanoemulsions against inflammatory and allergic activities. In in vitro cell culture experiments, the 100-nm curcumin lipid nanoemulsion showed the most prominent inhibitory effect on the production of tumor necrosis factor-α (TNF-α) induced by lipopolysaccharide (LPS) in RAW264.7 murine macrophages, and on the release of ß-hexosaminidase induced by the calcium ionophore, A23187, in rat basophilic leukemia RBL-2H3 cells. In an in vivo experiment, in which mice were administered the curcumin-loaded lipid nanoemulsion of various particle sizes, the 100-nm curcumin lipid nanoemulsion showed the most prominent anti-inflammatory and anti-allergic effects, inhibiting 12-O-tetradecanoylphorbol-13-acetate-induced inflammatory ear edema and immunoglobulin E (IgE)-induced passive cutaneous anaphylactic (PCA) reaction. The effects of particle size on serum curcumin absorption were also assessed in mice, and the 100-nm lipid nanoemulsion showed the greatest absorption. The results from our study suggest that the physiological activities of curcumin lipid nanoemulsions differ depending on particle size. Our data indicate that the curcumin lipid nanoemulsion with a particle size of 100 nm has potential for use in enhancing the bioavailability and medical value of curcumin.

5-O-Acyl plumbagins inhibit DNA polymerase activity and suppress the inflammatory response.

We previously found that vitamin K3 (menadione, 2-methyl-1,4-naphthoquinone) inhibits the activity of human mitochondrial DNA polymerase (pol) γ. In this study, we focused on plumbagin (5-hydroxy-2-methyl-1,4-naphthoquinone), and chemically synthesized novel plumbagins conjugated with C2:0 to C22:6 fatty acids (5-O-acyl plumbagins). These chemically modified plumbagins displayed enhanced mammalian pol inhibition, with plumbagin conjugated to docosahexaenoic acid (C22:6-acyl plumbagin) exhibiting the strongest inhibition of pol λ among the ten 5-O-acyl plumbagins synthesized. C22:6-acyl plumbagin selectively inhibited the activities of mammalian pol species, but did not influence the activities of other pols or DNA metabolic enzymes tested. The inhibition of pol λ, a DNA repair/recombination pol, by these compounds was significantly correlated with both their suppression of lipopolysaccharide (LPS) induced tumor necrosis factor-α (TNF-α) production by mouse RAW264.7 macrophages and the reduction of 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced inflammation in the mouse ear. These data indicate that 5-O-acyl plumbagins act as anti-inflammatory agents by inhibiting mammalian pol λ. These results further suggest that C22:6-acyl plumbagin is a promising anti-inflammatory candidate and that acylation could be an effective chemical modification to improve the anti-inflammatory activity of vitamin K3 derivatives, such as plumbagin.

Bioactive dihydronaphthoquinone derivatives from Fusarium solani.

New dihydronaphthoquinone derivatives, karuquinone A (1), karuquinone B (2), and karuquinone C (3), were isolated from a fungal culture broth of Fusarium solani. The structures were determined by interpretation of spectroscopic data (1D/2D NMR, MS, and IR). Three known compounds, javanicin (4), 2,3-dihydro-5-hydroxy-8-methoxy-2,4-dimethylnaphtho[1,2-b]furan-6,9-dione (5), and 5-hydroxydihydrofusarubin C (6), were also isolated. The six isolated compounds were tested for cytotoxicity against three human cancer cell lines and a human umbilical vein endothelial cell (HUVEC) line. Of these, karuquinone A exhibited the strongest cytotoxic activity. Karuquinone B did not affect the proliferation of the cancer cell lines but did inhibit the proliferation of HUVEC. Additionally, we demonstrated that karuquinone A induces apoptosis in cancer cells through the generation of reactive oxygen species (ROS).

Inhibition of mammalian DNA polymerases and the suppression of inflammatory and allergic responses by tyrosol from used activated charcoal waste generated during sake production.

The components adsorbed onto activated charcoal following the fermentation process of the Japanese rice wine "sake" have been studied with the aim of identifying suitable applications for this industrial food waste product. The absorbed materials were effectively extracted from the charcoal, and inhibited the activity of several mammalian DNA polymerases (pols). Subsequent purification of the extract afforded tyrosol [4-(2-hydroxyethyl)phenol] as the active component, which selectively inhibited the activity of 11 mammalian pols with IC50 values in the range of 34.3-46.1 µM. In contrast, this compound did not influence the activities of plant or prokaryotic pols or any of the other DNA metabolic enzymes tested. Tyrosol suppressed both anti-inflammatory and antiallergic effects in vivo, including 12-O-tetradecanoylphorbol-13-acetate-induced inflammatory mouse ear edema, and immunoglobulin E-induced passive cutaneous anaphylactic reaction in mice. These results suggested that this byproduct formed during the sake-brewing process could be used as an anti-inflammatory and/or antiallergic agent.

Suppression of allergic and inflammatory responses by essential oils derived from herbal plants and citrus fruits.

The aim of the present study was to investigate the biological activity of 20 essential oils (EOs) derived from herbal plants and citrus fruits. The in vitro anti-allergic and anti-inflammatory activities of these oils were investigated, and the EO which was found to have the strongest activity of the 20 EOs examined, was investigated further to identify its components and bioactive compounds. The in vitro anti-allergic activity was determined by measuring the release of ß-hexosaminidase from rat basophilic leukemia (RBL-2H3) cells treated with the calcium ionophore, A23187. The in vitro anti-inflammatory activity was determined by measuring the production of tumor necrosis factor-α (TNF-α) in RAW264.7 murine macrophages treated with lipopolysaccharide. Among the EOs examined, lemongrass [Cymbopogon citratus (DC.) Stapf] elicited the strongest anti-allergic and anti-inflammatory effects. A principal component of this EO is citral (3,7-dimethyl-2,6-octadien-1-al) (74.5%), a mixture of the stereoisomers, geranial (trans-citral, 40.16%) and neral (cis-citral, 34.24%), as determined by chromatography-mass spectrometry analysis. The activities of citral and geranial are similar to those of lemongrass EO. These compounds elicited significant in vivo anti-allergic and anti-inflammatory effects, suppressing an immunoglobulin E (IgE)-induced passive cutaneous anaphylactic reaction in mice and a 12-O-tetradecanoylphorbol-13-acetate-induced inflammatory mouse ear edema, respectively. Our data demonstrate that lemongrass EO and its constituents, citral and geranial, may be a therapeutic candidate for allergic and inflammatory diseases.

Anti-tumor effects of novel 5-O-acyl plumbagins based on the inhibition of mammalian DNA replicative polymerase activity.

We previously found that vitamin K3 (menadione, 2-methyl-1,4-naphthoquinone) inhibits the activity of human mitochondrial DNA polymerase γ (pol γ). In this study, we focused on plumbagin (5-hydroxy-2-methyl-1,4-naphthoquinone), and chemically synthesized novel plumbagins conjugated with C2:0 to C22:6 fatty acids (5-O-acyl plumbagins). These chemically modified plumbagins enhanced mammalian pol inhibition and their cytotoxic activity. Plumbagin conjugated with chains consisting of more than C18-unsaturated fatty acids strongly inhibited the activities of calf pol α and human pol γ. Plumbagin conjugated with oleic acid (C18:1-acyl plumbagin) showed the strongest suppression of human colon carcinoma (HCT116) cell proliferation among the ten synthesized 5-O-acyl plumbagins. The inhibitory activity on pol α, a DNA replicative pol, by these compounds showed high correlation with their cancer cell proliferation suppressive activity. C18:1-Acyl plumbagin selectively inhibited the activities of mammalian pol species, but did not influence the activities of other pols and DNA metabolic enzymes tested. This compound inhibited the proliferation of various human cancer cell lines, and was the cytotoxic inhibitor showing strongest inhibition towards HT-29 colon cancer cells (LD50 = 2.9 µM) among the nine cell lines tested. In an in vivo anti-tumor assay conducted on nude mice bearing solid tumors of HT-29 cells, C18:1-acyl plumbagin was shown to be a promising tumor suppressor. These data indicate that novel 5-O-acyl plumbagins act as anti-cancer agents based on mammalian DNA replicative pol α inhibition. Moreover, the results suggest that acylation of plumbagin is an effective chemical modification to improve the anti-cancer activity of vitamin K3 derivatives, such as plumbagin.

Inhibition of DNA polymerase λ and associated inflammatory activities of extracts from steamed germinated soybeans.

During the screening of selective DNA polymerase (pol) inhibitors from more than 50 plant food materials, we found that the extract from steamed germinated soybeans (Glycine max L.) inhibited human pol λ activity. Among the three processed soybean samples tested (boiled soybeans, steamed soybeans, and steamed germinated soybeans), both the hot water extract and organic solvent extract from the steamed germinated soybeans had the strongest pol λ inhibition. We previously isolated two glucosyl compounds, a cerebroside (glucosyl ceramide, AS-1-4, compound ) and a steroidal glycoside (eleutheroside A, compound ), from dried soybean, and these compounds were prevalent in the extracts of the steamed germinated soybeans as pol inhibitors. The hot water and organic solvent extracts of the steamed germinated soybeans and compounds and selectively inhibited the activity of eukaryotic pol λ in vitro but did not influence the activities of other eukaryotic pols, including those from the A-family (pol γ), B-family (pols α, δ, and ε), and Y-family (pols η, ι, and κ), and also showed no effect on the activity of pol ß, which is of the same family (X) as pol λ. The tendency for in vitro pol λ inhibition by these extracts and compounds showed a positive correlation with the in vivo suppression of TPA (12-O-tetradecanoylphorbol-13-acetate)-induced inflammation in mouse ear. These results suggest that steamed germinated soybeans, especially the glucosyl compound components, may be useful for their anti-inflammatory properties.

Formosusin A, a novel specific inhibitor of mammalian DNA polymerase ß from the fungus Paecilomyces formosus.

Variotin (1) and three novel compounds, formosusin A (2), B (3), and C (4), were isolated from the cultures of the fungus Paecilomyces formosus, and their structures were determined by spectroscopic analyses. Compound 2 is (6Z,8E,10E)-variotin, a new cis-olefin analog of compound 1. Compound 2 selectively inhibited the activity of mammalian DNA polymerase ß (pol ß) in vitro, with an IC50 of 35.6µM. By contrast, compounds 1, 3, and 4 did not influence the activity of pol ß. These four compounds showed no effect on the activities of other 10 mammalian pols (i.e., pols α, γ, δ, ε, η, ι, κ, λ, and µ, and terminal deoxynucleotidyl transferase). These compounds also did not inhibit the activities of fish, insect, plant, and prokaryotic pols and other DNA metabolic enzymes tested. These results suggested that compound 2 could be a selective inhibitor of mammalian pol ß. The compound 2-induced inhibition of rat pol ß activity was competitive and non-competitive with respect to the DNA template-primer substrate and the dNTP substrate, respectively. On the basis of these results, the relationship between the three-dimensional structure and pol ß inhibitory mechanism of compound 2 is discussed.

Synthesis, structure, and cytotoxicity studies of some fungal isochromanes.

Ustusorane D and penicisochromans B-D are natural isochromans isolated from Aspergillus ustus 094102 and Penicillium sp. PSU-F40, respectively. Herein, we report the syntheses of (-)-ustusorane D and (+)-penicisochroman B and the structures of penicisochromans C and D. The relative configuration of natural ustusorane D and the absolute configuration of natural penicisochroman B were determined. Two plausible structures for penicisochroman C were evaluated through synthesis, but their ¹H and ¹³C NMR data were not in agreement with those of the natural product. The structural revision and the determination of the absolute configuration of natural penicisochroman D were achieved. Structure-activity relationship studies of the synthetic compounds as well as a series of related isochromans indicated that the enone of the furanone moiety was essential for the cytotoxicity of these compounds toward HCT116 human colon cancer cells. Pseudodeflectusin, the related natural isochroman, suppressed cell growth and induced apoptosis in HCT116 cells.

Inhibitory effects of a major soy isoflavone, genistein, on human DNA topoisomerase II activity and cancer cell proliferation.

The inhibitory activity of 3 soy isoflavones (daidzein, genistein and glycitein) and their glycosides (daidzin, genistin and glycitin) on mammalian DNA polymerases (pols) and topoisomerases (topos) was investigated. Of the compounds tested, only genistein selectively inhibited human topo II activity and had an IC50 value of 37.5 µM. These isoflavones had no effect on the activity of human topo I; mammalian pols α, ß, γ and κ; or on any other DNA metabolic enzyme tested. Thermal transition analysis indicated that genistein did not influence the direct binding to double-stranded DNA. Genistein prevented the proliferation of HCT116 human colon carcinoma cells with an LD50 of 94.0 µM and it halted the cell cycle in G2/M phase. These results suggest that decreases in cell proliferation due to genistein may result from the inhibition of cellular topo II and that genistein, a major soy isoflavone, may be an anticancer food component. The relationship between the structures and these bioactivities of soy isoflavones is discussed.

Inhibitory effects of α-mangostin on mammalian DNA polymerase, topoisomerase, and human cancer cell proliferation.

We found that the ethanol extract of mangosteen (Garcinia mangostana L.) fruit rind had a strong inhibitory effect on mammalian DNA polymerase (pol) activity and isolated α-mangostin as a potent pol inhibitor from the extract. In this study, the inhibitory activities against mammalian pols by α-mangostin and its related five compounds, 3-isomangostin, xanthone, 9,10-anthraquinone, 9-anthracenecarboxylic acid, and anthracene, were investigated. α-Mangostin was the most potent inhibitor of the mammalian pol species among the tested compounds, with IC50 values of 14.8-25.6 µM. This compound also inhibited human DNA topoisomerases (topos) I and II activities with IC50 values of 15.0 and 7.5 µM, respectively, but did not inhibit the activities of other DNA metabolic enzymes tested. α-Mangostin also did not directly bind to double-stranded DNA as determined by thermal transition analysis. α-Mangostin was found to suppress human colon HCT116 carcinoma cell proliferation with an LC50 of 18.5 µM, inhibit the activity of cellular topos, halt cell cycle in the G2/M phase, and induce apoptosis. These results suggest that decreased proliferation by α-mangostin may be a result of the inhibition of cellular topos rather than pols, and α-mangostin might be an anticancer chemotherapeutic agent.

Inhibitory effects of low molecular weight polyphenolics from Inonotus obliquus on human DNA topoisomerase activity and cancer cell proliferation.

Low molecular weight (LMW) polyphenolics containing a polyhydroxylated benzyl moiety are abundant in medicinal plants. In the present study, we report on the activities of seven LMW polyphenolics isolated from Inonotus obliquus, a medicinal mushroom. The isolated compounds included caffeic acid (CA), 3,4-dihydroxybenzalacetone (DBL), gallic acid, syringic acid, protocatechuic acid, 3,4-dihydroxybenzaldehyde and 2,5-dihydroxyterephthalic acid. We analyzed their inhibitory effects on DNA polymerase (pol) and DNA topoisomerase (topo), and their effects on human cancer cell growth. All isolated compounds inhibited human topo II activity; the most potent were DBL and CA, which contain a catechol propanoid moiety. CA and DBL inhibited the activity of human topo I, whereas other compounds had no effect. No compound modulated the activities of 11 mammalian pol species or other DNA metabolic enzymes, including T7 RNA polymerase, mouse IMP dehydrogenase (type II), T4 polynucleotide kinase and bovine deoxyribonuclease I. CA and DBL markedly suppressed the proliferation of human colon HCT116 carcinoma cells with an LD50 of 70.0 and 49.4 µM, respectively, and halted the cell cycle in the G2/M phase. The suppressive effect of these compounds on cancer cell growth correlated with their ability to inhibit topo II. These results suggest that CA- and DBL-dependent decreases in cell proliferation are due to the inhibition of cellular topo II. The mechanism of action of these catechol propanoid compounds and the implication for their use as anticancer agents are discussed.

Cycloartenyl trans-ferulate, a component of the bran byproduct of sake-brewing rice, inhibits mammalian DNA polymerase and suppresses inflammation.

During the screening of selective DNA polymerase (pol) inhibitors, we isolated cycloartenyl trans-ferulate (CAF), which is a major component of γ-oryzanol, which is a byproduct formed during the production of Japanese rice wine "sake". CAF selectively inhibited the activity of mammalian A, B, and X pol families, but Y family pols were not affected. CAF did not influence the activities of plant or prokaryotic pols, nor the activity of other DNA metabolic enzymes tested. Individual chemical components of CAF, including cycloartenol (CA) and ferulic acid (FA), did not inhibit pol enzyme activities. CAF suppressed TPA (12-O-tetradecanoylphorbol-13-acetate)-induced inflammation in the mouse ear, but CA and FA did not. The ability to inhibit mammalian pol enzymes in vitro was positively correlated with their propensity to suppress inflammation in vivo. These results suggest that this byproduct formed during the sake-brewing process is useful as an anti-inflammatory agent.

Inhibitory effects of myricetin on mammalian DNA polymerase, topoisomerase and human cancer cell proliferation.

In this study, the inhibitory activities against mammalian DNA polymerases (pols) of 16 major bioflavonoids were investigated. Myricetin (3,3',4',5,5',7-hexahydroxyflavone) was the most potent inhibitor of pols amongst the compounds tested, with IC50 values of 21.3-40.9 µM. This compound did not affect the activities of plant (cauliflower) pol α or prokaryotic pols. Myricetin also inhibited human DNA topoisomerase II (topo II) activity with an IC50 value of 27.5 µM, but did not inhibit the activities of other DNA metabolic enzymes tested. Myricetin also did not influence the direct binding to double stranded DNA as determined by thermal transition analysis. It was found to prevent the proliferation of human colon HCT116 carcinoma cells with an LD50 of 28.2 µM, halt the cell cycle in G2/M phase, and induce apoptosis. These results suggest that the decrease of proliferation may be a result of the inhibition of cellular topoisomerase (topo) II rather than pols.

Inhibitory effect of novel somatostatin peptide analogues on human cancer cell growth based on the selective inhibition of DNA polymerase ß.

The present study was designed to investigate the anticancer activity of novel nine small peptides (compounds 1-9) derived from TT-232, a somatostatin structural analogue, by analyzing the inhibition of mammalian DNA polymerase (pol) and human cancer cell growth. Among the compounds tested, compounds 3 [tert-butyloxycarbonyl (Boc)-Tyr-Phe-1-naphthylamide], 4 (Boc-Tyr-Ile-1-naphthylamide), 5 (Boc-Tyr-Leu-1-naphthylamide) and 6 (Boc-Tyr-Val-1-naphthylamide) containing tyrosine (Tyr) but no carboxyl groups, selectively inhibited the activity of rat pol ß, which is a DNA repair-related pol. Compounds 3-6 strongly inhibited the growth of human colon carcinoma HCT116 p53(+/+) cells. The influence of compounds 1-9 on HCT116 p53(-/-) cell growth was similar to that observed for HCT116 p53(+/+) cells. These results suggest that the cancer cell growth suppression induced by these compounds might be related to their inhibition of pol. Compound 4 was the strongest inhibitor of pol ß and cancer cell growth among the nine compounds tested. This compound specifically inhibited rat pol ß activity, but had no effect on the other 10 mammalian pols investigated. Compound 4 combined with methyl methane sulfonate (MMS) treatment synergistically suppressed HCT116 p53(-/-) cell growth compared with MMS alone. This compound also induced apoptosis in HCT116 cells with or without p53. From these results, the influence of compound 4, a specific pol ß inhibitor, on the relationship between DNA repair and cancer cell growth is discussed.

Profiles of lipid components, fatty acid distributions of triacylglycerols and phospholipids in Jack beans (Canavalia gladiata DC.).

Endogenous tocopherols in extracted lipids from Jack beans (Canavalia gladiata DC.) were determined by high-performance liquid chromatography (HPLC), and were investigated in relation to the fatty acids (FA) distribution of triacylgycerols (TAG) and phospholipids (PL). The dominant tocopherols were (δ)-tocopherol (78.9-96.5mg%) and (γ)-tocopherol (42.1-56.1mg%) with much smaller amounts of (α)-tocopherol (1.1-1.3mg%). The lipids of Jack beans comprised mainly TAG (34.6-38.6 wt.%) and PL (54.8-57.4 wt.%), and other components were also detected in minor proportions (0.3-3.8 wt.%). The PL components included phosphatidyl choline (46.2-48.7 wt.%), phosphatidyl inositol (23.4-29.6 wt.%) and phosphatidyl ethanolamine (18.5-21.2 wt.%). Comparison of these different beans showed, with a few exceptions, no significant differences (P>0.05) in FA distribution. The FA distribution of TAG among the five beans was evident in the Jack beans: unsaturated FA (93.3-95.3 wt.%) were predominantly concentrated at the sn-2 position and saturated FA (33.6-34.4 wt.%) primarily occupying the sn-1 position or sn-3 position. The results obtained from this work would provide useful information to both producers and consumers for manufacturing functional foods or beverages in Japan and elsewhere.

Inhibitory effects of catechin derivatives on mammalian DNA polymerase and topoisomerase activities and mouse one-cell zygote development.

In this study, the inhibitory activities against DNA polymerases (pols) and DNA topoisomerases (topos) by eight major green tea catechin derivatives (flavan-3-ols) were investigated. Some catechins inhibited mammalian pols (α and ß) and human topos (I and II), with (-)-epigallocatechin gallate (EGCg) the strongest inhibitor of both enzyme types, showing IC(50) values of 3.8-21.5 and 2.0-20.0 µM, respectively. EGCg did not affect the activities of plant (cauliflower) pol α or prokaryotic pols and showed no effect on the activities of other DNA metabolic enzymes tested. Next, a method was established for assay of mouse one-cell zygote development inhibition, the catechin derivatives screened for bioactivity, and the inhibition was assessed and their effects ranked as: EGCg > GCg > Cg >> others. In the mouse one-cell zygote assay, EGCg at 50 µM increased abnormal cells and 75 µM of EGCg-induced apoptosis. The observed ranking of catechin derivative inhibition effects against mouse one-cell zygote development in vivo was similar to their ranking by topo inhibition in vitro rather than by pol inhibition; therefore, topo inhibition might have been effecting zygote development inhibition. These results suggested that catechin derivatives indeed reached the nuclear DNA where topo inhibition can occur, thus causing the observed cellular effects. From these findings, this zygote development inhibition assay will be useful as an anti-pregnant agent screening.

Effects of essential oils from herbal plants and citrus fruits on DNA polymerase inhibitory, cancer cell growth inhibitory, antiallergic, and antioxidant activities.

In this study, the biological activity of 20 essential oils (EOs) from herbal plants and citrus fruits were investigated in terms of mammalian DNA polymerase (pol) inhibitory activity, cancer cell (human colon carcinoma, HCT116) growth inhibitory activity, antiallergic activity, as anti-ß-hexosaminidase release activity in rat basophilic leukemia RBL-2H3 cells treated with calcium ionophore A23187, and antioxidant activity by a lipophilic-oxygen radical absorbance capacity method. These EOs showed patterns of inhibition of pol α, a DNA replicative pol, similar to their cancer cell growth inhibitory activity, and their inhibitory activity on pol λ, a DNA repair/recombination pol, by the EOs showed correlation with anti-ß-hexosaminidase release activity. Among these EOs, chamomile (Matricaria chamomilla L.) was the strongest inhibitor of pols α and λ and showed significant effects on both cancer cell growth and mast cell degranulation. On the basis of these results, chamomile EO can be recommended as a potentially useful, bioactive candidate for therapeutic applications.

Antifungal properties of Japanese cedar essential oil from waste wood chips made from used sake barrels.

In this study, we prepared essential oil (EO) from waste wood chips made from used sake barrels (USBs) of Japanese cedar (i.e., EO-USB) by steam distillation. We found that EO-USB and three commercially purchased EOs derived from xylem tissue of Japanese woods, such as Japanese cedar (Cryptomeria japonica), Japanese cypress (Chamaecyparis obtusa) and false arborvitae (Thujopsis dolabrata), suppressed fungal growth activity against Trichophyton rubrum, which is the cause of tinea disease. The magnitude of the suppressive effects of the EOs ranked as follows: T. dolabrata > USB = C. japonica > C. obtusa. These EOs also inhibited the activity of DNA polymerase in an extract from T. rubrum mycelia with the following ranking: T. dolabrata > USB = C. japonica > C. obtusa. In addition, 50 µg/ml of EO-USB showed antifungal properties, killing T. rubrum mycelia at 27-42˚C in 20 min. By gas chromatography/mass spectrometry analysis, the main sesquiterpenes in EO-USB were δ-cadinene (25.94%) and epi-cubenol (11.55%), and the composition of EO-USB was approximately the same as that of EO-C. japonica. Three prepared sesquiterpenes, δ-cadinene, epi-cubenol and ß-eudesmol, inhibited the fungal growth and DNA polymerase activities of T. rubrum, and epi-cubenol showed the strongest inhibition among the compounds tested. These sesquiterpenes had no inhibitory effects on the activities of other DNA metabolic enzymes, such as DNA topoisomerase II, IMP dehydrogenase, polynucleotide kinase and deoxyribonuclease from T. rubrum. Taken together, these results suggest that EO-USB containing epi-cubenol may be useful for its anti-tinea disease properties, which are based on DNA polymerase inhibition.