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Glycoside hydrolase family 94 (GH94) contains enzymes that reversibly catalyze the phosphorolysis of β-glycosides. We conducted this study to investigate a GH94 protein (PBOR_13355) encoded in the genome of Paenibacillus borealis DSM 13188 with low sequence identity to known phosphorylases. Screening of acceptor substrates for reverse phosphorolysis in the presence of α-d-glucose 1-phosphate as a donor substrate showed that PBOR_13355 utilized d-glucuronic acid and p-nitrophenyl β-d-glucuronide as acceptors. In the reaction with d-glucuronic acid, 3-O-β-d-glucopyranosyl-d-glucuronic acid was synthesized. PBOR_13355 showed a higher apparent catalytic efficiency to p-nitrophenyl β-d-glucuronide than to d-glucuronic acid, and thus, PBOR_13355 was concluded to be a novel glycoside phosphorylase, 3-O-β-d-glucopyranosyl β-d-glucuronide phosphorylase. PBOR_13360, encoded by the gene immediately downstream of the PBOR_13355 gene, was shown to be β-glucuronidase. Collectively, PBOR_13355 and PBOR_13360 are predicted to work together in the cytosol to metabolize oligosaccharides containing the 3-O-β-d-glucopyranosyl β-d-glucuronide structure released from bacterial and plant acidic carbohydrates.

The immunomodulating effect of phlorotannin was investigated in mice stimulated by ovalbumin. When analyzing the main components of phlorotannin concentrate (PTC) from Eisenia nipponica, seven phlorotannins [eckol, 6,6'-bieckol, 6,8'-bieckol, 8,8'-bieckol, dieckol, phlorofucofuroeckol (PFF)-A, and PFF-B] were detected. These phlorotannins accounted for approximately 80% of PTC. Oral administration of PTC to mice daily for 21 days reduced serum immunoglobulin E (IgE) and total IgG

Titanbicus (TB), a hybrid of Hibiscus moscheutos × H. coccineus (Medic.) Walt., has potential to be used as an edible flower. In this study, proximate nutritional content, anthocyanin content, total polyphenol content (TPC), and antioxidant activities in vitro and in vivo were investigated. Three cultivars of TB, namely Artemis (AR), Rhea (R), and Adonis (AD), were used as materials. Protein and carbohydrates were the primary macronutrients, while crude fat and ash were detected in trace amounts. Cyanidin 3-glucoside (Cy3-G) and cyanidin 3-sambubioside (Cy3-Sam), were identified in all TBs. The highest anthocyanin content was observed in AD (47.09 ± 1.45 mg/g extract), followed by R and AR (6.04 ± 0.20 and 2.72 ± 0.11 mg/g extract, respectively). The TPC of AD (225.01 ± 1.97 mg/g extract) was greater than that of AR and R (185.41 ± 3.24 and 144.10 ± 1.71 mg/g extract, respectively). AD exhibited the strongest in vitro antioxidant activity in hydrophilic oxygen radical absorbance capacity, compared to the other two TBs. In addition, AD extract suppressed the generation of reactive oxygen species in caudal fin of wounded zebrafish. Antioxidant activities of AD appeared to be related to its total anthocyanin content, Cy3-G, Cy3-Sam, and TPC. Our findings indicate that TB, particularly the AD cultivar, would be an attractive source of bioactive compounds with antioxidant activities, and can improve both nutritional value and appearance of food.

Functional foods that inhibit α-amylase and α-glucosidase activity are effective for regulating the blood glucose level and preventing hyperglycemia. Extracts of adzuki beans (ABs, Vigna angularis), widely eaten in East Asia, can inhibit α-amylase and α-glucosidase activity. In this study, we identified and evaluated the components in an AB water extract (ABWE) after boiling, which is an essential process for cooking ABs. The ABWE before boiling inhibited α-amylase and α-glucosidase activity and the boiled ABWE showed slightly stronger inhibitory effects. High-performance liquid chromatography, liquid chromatography-mass spectrometry, and nuclear magnetic resonance analyses identified (+)-catechin 7-O-β-d-glucopyranoside (C7G), (+)-epicatechin 7-O-β-d-glucopyranoside (E7G), and (+)-catechin as the bioactive components in boiled ABWE. Interestingly, the quantity of E7G significantly increased after boiling (from 0% to 17.1 ± 1.3%). E7G showed stronger inhibition of α-amylase and α-glucosidase than C7G; the IC50 values for α-amylase were 0.74 ± 0.04 mg/mL (C7G) and 0.40 ± 0.09 mg/mL (E7G), and for α-glucosidase the IC50 values were 0.085 ± 0.032 mg/mL (C7G) and 0.051 ± 0.007 mg/mL (E7G). Our findings suggest that C7G and E7G are the main active components in ABWE as they inhibit α-amylase and α-glucosidase and their inhibitory effect is not lost after boiling. These results support the effectiveness of boiled ABs in the promotion of health. PRACTICAL APPLICATION: We identified (+)-catechin 7-O-β-d-glucopyranoside (C7G), (+)-epicatechin 7-O-β-d-glucopyranoside (E7G), and (+)-catechin in adzuki bean extracts and commercially available boiled adzuki bean products. Interestingly, the E7G content was increased by boiling, and this compound showed strong inhibitory activity toward α-amylase and α-glucosidase. These results support the consumption of boiled adzuki beans to prevent acute rises in blood glucose level.

Osteoporosis is a systemic skeletal disease that causes bone weakness and fragility. Consuming bone-beneficial nutrients through diet can prevent and treat osteoporosis. Acer palmatum (Japanese maple) leaves are used to make tea, but there have been few reports of their health benefits, especially regarding bone homeostasis. In this study, we evaluated the effects of A. palmatum hot water extract (APE) on osteoclastogenesis and osteoblastogenesis in cultured cells. APE suppressed the number of tartrate-resistant acid phosphatase-positive multinucleated osteoclasts in RANKL induced RAW264.7 cells. Furthermore, APE facilitated Alkaline phosphatase activity and calcium deposition during osteoblast differentiation in MC3T3-E1 cells. High-performance liquid chromatography analysis was performed to investigate the effective components of APE, and four flavonoids orientin, isoorientin, vitexin, and isovitexin were identified with the LC-MS analysis. Treatment with fractionated APE suppressed osteoclastogenesis and facilitated osteoblastogenesis in cultured cells. These findings suggest that APE contains antiosteoporotic compounds; thus, APE might have health promoting effects that help prevent osteoporosis by inhibiting osteoclastogenesis and facilitating osteoblastogenesis.

The zebrafish obesogenic test (ZOT) is a powerful tool for identifying anti-adipogenic compounds for in vivo screening. In our previous study, we found that

Various natural products (NPs) have been used to treat obesity and related diseases. However, the best way to fight obesity is preventive, with accurate body weight management through exercise, diet, or bioactive NPs to avoid obesity development. We demonstrated that green tea extract (GTE) is an anti-obesity NP using a zebrafish obesity model. Based on a hypothesis that GTE can prevent obesity, the objective of this study was to assess GTE’s ability to attenuate obesity development. Juvenile zebrafish were pretreated with GTE for seven days before obesity induction via a high-fat diet, adult zebrafish were pretreated with GTE for two weeks before obesity induction by overfeeding. As a preventive intervention, GTE significantly decreased visceral adipose tissue accumulation in juveniles and ameliorated visceral adiposity and plasma triglyceride levels in adult zebrafish obesity models. RNA sequencing analysis was performed using liver tissues from adult obese zebrafish, with or without GTE administration, to investigate the underlying molecular mechanism. Transcriptome analysis revealed that preventive GTE treatment affects several pathways associated with anti-obesity regulation, including activation of STAT and downregulation of CEBP signaling pathways. In conclusion, GTE could be used as a preventive agent against obesity.

We previously found a lipophilic fraction of the methanol/chloroform extract of a brown alga, Eisenia nipponica, that had an antiallergic effect in a murine ear swelling test. In this study, we purified the active component from the lipophilic fraction using high performance liquid chromatography and analyzed the mass and nuclear magnetic resonance spectra. This uncovered the phlorotannin dieckol, which exhibited antiallergic effects in an ear swelling test using mice sensitized by arachidonic acid, 12-O-tetradecanoylphorbol-13-acetate, and oxazolone. Mechanistic investigations indicated that dieckol suppressed degranulation, chemical mediator release, and the expression of mRNA such as cyclooxygenase-2, interleukin-6, and tumor necrosis factor-α in rat basophilic leukemia-2H3 cells. In summary, we isolated dieckol from E. nipponica and demonstrated its antiallergic mechanisms. PRACTICAL APPLICATIONS: As the incidence of allergies increases worldwide, so too does the demand for food components with antiallergic and anti-inflammatory properties. Given this trend, we focused on a brown alga that displays a variety of bioactivities. Here, we have isolated dieckol from the antiallergic lipophilic fraction of E. nipponica and found that it possesses diverse physiological activities that may prevent lifestyle-related diseases. Consequently, dieckol or the alga containing this phlorotannin could be used as a health food ingredient to combat not only allergies, but also variety of disorders including the undesirable effects of aging.

Ruminiclostridium josui Fae1A is a modular enzyme consisting of an N-terminal signal peptide, family-1 carbohydrate esterase module (CE1), family-6 carbohydrate-binding module (CBM6), and dockerin module in that order. Recombinant CE1 and CBM6 polypeptides were collectively and separately produced as RjFae1A, RjCE1, and RjCBM6. RjFae1A showed higher feruloyl esterase activity than RjCE1 towards insoluble wheat arabinoxylan, but the latter was more active towards small synthetic substrates than the former. This suggests that CBM6 in RjFae1A plays an important role in releasing ferulic acid from the native substrate. RjCBM6 showed a higher affinity for soluble wheat arabinoxylan than for rye arabinoxylan and beechwood xylan in native affinity polyacrylamide gel electrophoresis. Isothermal titration calorimetry analysis demonstrated that RjCBM6 recognized a xylopyranosyl residue at the nonreducing ends of xylooligosaccharides. Moreover, it showed exceptional affinity for 23-α-l-arabinofuranosyl-xylotriose (A2XX) among the tested branched arabinoxylooligosaccharides. Fluorometric titration analysis demonstrated that xylobiose and A2XX competitively bound to RjCBM6, and both bound to the same site in RjCBM6. RjCBM6's preference for the xylopyranosyl residue at the nonreducing end of xylan chains explains why the positive effect of CBM6 on RjFae1A activity was observed only during short incubation but not after extended incubation.

Ruminiclostridium josui Abf62A-Axe6A is a modular enzyme comprising (in order from the N-terminus): an N-terminal signal peptide, a glycoside hydrolase family 62 (GH62) catalytic module, a family 6 carbohydrate binding module (CBM6), a dockerin module and an additional carbohydrate esterase family 6 catalytic module (CE6). In this study, three Abf62A-Axe6A derivatives were constructed, overexpressed in Escherichia coli, purified, and biochemically characterized: RjAbf62A-Axe6A, containing all four modules but lacking the signal peptide; RjAbf62A-CBM6, containing the GH62 and CBM6 modules; and RjAxe6A, containing only CE6. RjAbf62A-Axe6A was highly active toward arabinoxylan and moderately active toward sugar beet arabinan, and released mainly arabinose. Analysis of the arabinoxylooligosaccharide hydrolysis products revealed that RjAbf62A-Axe6A released α-1,2- and α-1,3-linked arabinofuranose from both singly and doubly substituted xylosyl residues. Furthermore, RjAbf62A-Axe6A exhibited a weak activity toward linear 1,5-α-l arabinan and arabinooligosaccharides, indicating that it is capable of cleaving α-1,5-linkage. Surprisingly, RjAbf62A-Axe6A also demonstrated an endoxylanase activity toward birchwood and beechwood xylans and xylooligosaccharides. Although RjAbf62A-CBM6 exhibited a similar substrate specificity to RjAbf62A-Axe6A, RjAbf62A-CBM6 showed lower activities toward soluble arabinoxylans, birchwood and beechwood xylans and arabinoxylooligosaccharides but not toward insoluble arabinoxylan. RjAbf62A-Axe6A is the first reported GH62 enzyme with α-l-arabinofuranosidase and endoxylanase activities. Although both RjAbf62A-Axe6A and RjAxe6A had acetylxylan esterase activities, RjAbf62A-Axe6 exhibited a higher activity toward insoluble wheat arabinoxylan compared with RjAxe6.

Because the number of people suffering from allergies has significantly increased, improved ways of treating these conditions by medical, pharmaceutical, and dietary means are required. Large numbers of studies on allergy have been conducted, and many anti-allergic compounds have been found. Phenolic compounds from terrestrial plants, including catechins and flavonoids, possess anti-allergic properties. Although polyphenols are present in some brown algae, their anti-allergic activities were not studied in detail before the 1990s. The focus was on the algal polyphenols, collectively called phlorotannins (eg., eckol, 6,6′-bieckol, 8,8′-bieckol, dieckol, and phlorofucofuroeckol-A), and research was conducted to clarify their anti-allergic activities. This review summarizes the anti-allergic effects of phlorotannins isolated from the brown alga, Eisenia nipponica, and related reports by other research groups.

Here, we investigated the antiallergic effects of brown algae (Ecklonia cava subsp. stolonifera) harvested on the coast of Nishinoshima Town, Oki Islands, Shimane Prefecture, Japan. Specifically, we investigated whether the active components of E. cava ssp. stolonifera vary seasonally and whether such variation correlates with seasonal variation in the antiallergic effects of the algae. High-performance liquid chromatography, liquid chromatography–mass spectrometry, and nuclear magnetic resonance spectra were used to analyze 80% methanol (80M) extract from E. cava ssp. stolonifera and to isolate eckol, 6,6′-bieckol, 8,8′-bieckol, dieckol, and phlorofucofuroeckol-A as the major active compounds in the species' phlorotannins. Monthly measurements of phlorotannin content and the isolated phlorotannins in the 80M extract showed that the crude phlorotannin and isolated phlorotannin content was higher in the summer–autumn period than in the winter–spring period. The antiallergic effects of the monthly 80M extracts and the isolated phlorotannins were confirmed by measuring inhibition of inflammation-related enzymatic activity, suppression of degranulation in rat basophile leukemia-2H3 cells, and effects on allergic inflammation in Institute of Cancer Research strain mice. There was partial seasonal variation in the effect of 80M extract in combination with the amount of isolated phlorotannin and the antiallergic effects. When phlorotannin was removed from the extracts using polyvinylpolypyrrolidone, the antiallergic effects were no longer observed; thus, phlorotannins are the active components. Overall, these results show that correlations exist between the seasonal variations in the suppressive effect of 80M extract on enzymatic activities, allergic inflammation in mice, and the amount of isolated phlorotannins.

Grifola gargal, an edible mushroom originating in southern Argentina and Chile, is known to have such beneficial health effects as antioxidant and anti-inflammatory activities. A low molecular weight fraction from Grifola gargal (GLF) decreased fasting blood glucose levels in high-fat diet-induced diabetic mice. This implicated Grifola gargal as a functional food to prevent type 2 diabetes mellitus (T2DM). To address the unknown underlying molecular mechanisms, we investigated, using L6 myotubes, whether GLF promoted GLUT4 translocation to the plasma membrane and its related signalling pathways. In addition, adenosine was isolated and identified the active compound in GLF to increase glucose uptake and promote GLUT4 translocation in L6 myotubes. Adenosine activated PI3K/Akt signalling and phosphorylation of AMPK, by binding to the adenosine receptor A1. Thus, Grifola gargal is a promising functional food to prevent post-prandial hyperglycaemia and T2DM by promoting glucose uptake in muscle.

Osteoporosis is a systemic skeletal disease that causes bone weakness and fragility. Consuming bone-beneficial nutrients through diet can prevent and treat osteoporosis.

Lignocresol was isolated from softwood with p-cresol using sulfuric acid and phase separation. An alkaline treatment of the lignocresol, followed by acidification, selectively yielded a guaiacyl coumaran, G1, in the acid-soluble fraction. With further alkaline treatment of G1 in 0.5 M of NaOH solution at 170 °C for 60 min, it was strongly suggested that a guaiacyl benzofuran derivative, G2, was obtained by the elimination of formaldehyde, based on analytical data of the reaction mixture. The process is very unique and well-designed based on the reactivity of Cα-ethers, or Cα-OH, Cβ-aryl-ethers, and Cγ-OH of lignin, although condensation reactions via formaldehyde occurred in parallel to give condensed products with a diarylmethane structure. Because these phenolic dimers, G1 and G2, were recovered from the guaiacyl unit linked with the neighboring guaiacyl units via two Beta-aryl-ether bonds, they are promising lignin-derived chemicals that are obtainable in a high yield.

The anti-allergic and anti-inflammatory effects of phlorotannins (seaweed polyphenols) from the brown alga Eisenia arborea using in vitro experiments has already been reported. Therefore, in this study, these effects were examined in vivo using mice. When ear edema was induced in ICR mice by three sensitizers (arachidonic acid (AA), 12-O-tetradecanoylphorbol-13-acetate (TPA) and oxazolone (OXA)), four phlorotannins tested (eckol, 8,8’-bieckol, phlorofucofuroeckol (PFF)-A and PFF-B) inhibited the ear edema after spreading of 0.01 or 0.1 mg onto the mouse ear. The effects of the phlorotannins on the ear edema after AA and OXA treatments tended to be stronger than after TPA treatment, and the inhibitory effects against each sensitizer were similar to epigallocatechin gallate, a typical inhibitor. This is the first report that phlorotannins exhibit anti-allergic and anti-inflammatory effects in in vivo experiments, and inhibit OXA-induced delayed-type (type IV) allergic reactions.

The genome sequence of Bacillus licheniformis SVD1, that produces a cellulolytic and hemi-cellulolytic multienzyme complex, was partially determined, indicating that the glycoside hydrolase system of this strain is highly similar to that of B. licheniformis ATCC14580. All of the fifty-six genes encoding glycoside hydrolases identified in B. licheniformis ATCC14580 were conserved in strain SVD1. In addition, two new genes, xyn30A and axh43A , were identified in the B. licheniformis SVD1 genome. The xyn30A gene was highly similar to Bacillus subtilis subsp. subtilis 168 xynC encoding for a glucuronoarabinoxylan endo-1,4-β-xylanase. Xyn30A, produced by a recombinant Escherichia coli , had high activity toward 4- O -methyl- d -glucurono- d -xylan but showed definite activity toward oat-spelt xylan and unsubstituted xylooligosaccharides. Recombinant Axh43A, consisting of a family-43 catalytic module of the glycoside hydrolases and a family-6 carbohydrate-binding module (CBM), was an arabinoxylan arabinofuranohydrolase (α- l -arabinofuranosidase) classified as AXH-m23 and capable of releasing arabinosyl residues, which are linked to the C-2 or C-3 position of singly substituted xylose residues in arabinoxylan or arabinoxylan oligomers. The isolated CBM polypeptide had an affinity for soluble and insoluble xylans and removal of the CBM from Axh43A abolished the catalytic activity of the enzyme, indicating that the CBM plays an essential role in hydrolysis of arabinoxylan.

Phlorotannin is the collective term for polyphenols derived from brown algae belonging to the genera Ascopyllum, Ecklonia, Eisenia, Fucus and Sargassum etc. Since the incidence of allergies is currently increasing in the world, there is a focus on phlorotannins having anti-allergic and anti-inflammatory effects. In this study, six purified phlorotannins (eckol, 6,6&prime, bieckol, 6,8&prime, 8,8&prime, phlorofucofuroeckol (PFF)-A and PFF-B) from Eisenia arborea, orally administered to mice, were examined for their suppression effects on ear swelling. In considering the suppression, we also examined whether the phlorotannins suppressed release of chemical mediators (histamine, leukotriene B4 and prostaglandin E2), and mRNA expression and/or the activity of cyclooxygenase-2 (COX-2), using RBL-2H3 cells, a cultured mast cell model. Results showed that the phlorotnannins exhibited suppression effects in all experiments, with 6,8&prime, bieckol, 8,8&prime, bieckol and PFF-A showing the strongest of these effects. In conclusion, orally administered phlorotannins suppress mouse ear swelling, and this mechanism apparently involves suppression of chemical mediator release and COX-2 mRNA expression or activity. This is the first report of the anti-allergic effects of the orally administered purified phlorotannins in vivo. Phlorotannins show potential for use in functional foods or drugs.

The entomogenous ‘Hanasanagitake’ mushroom, Isaria japonica, is used as a folk medicine and as a traditional health food choice in Japan. A search for naturally occurring antioxidative compounds from the mushroom led to the isolation of a novel pseudo-di-peptide, named Hanasanagin, and a possible biogenetic precursor. The structures of the pseudo-di-peptides were determined as (R)-3,4-diguanidinobutanoyl-(S)-DOPA and (R)-3,4-diguanidinobutanoyl-(S)-tyrosine by spectral analysis, chemical synthesis and enzymatic conversion.

Eisenia arborea, an edible brown alga, is occasionally used as a folk medicine due to its anti-allergic effect. In the present study to identify the anti-allergic constituents in the alga, the extract of the alga was purified by partition between solvents and by reversed phase chromatography. Separation of the extract was guided by the inhibitory activity upon β-hexosaminidase release from the rat basophilic leukemia-2H3 cells. HPLC purification afforded six active compounds. Spectral analyses clarified their structures as eckol, 6,6′-bieckol, 6,8′-bieckol, 8,8′-bieckol, phlorofucofuroeckol-A, and phlorofucofuroeckol-B. Most of the phlorotannins exhibited activities similar to or greater than the typical inhibitor, epigallocatechin gallate. Phlorofucofuroeckol-B showed the greatest activity among the tested phlorotannins at 2.8 times greater than epigallocatechin gallate.

15種の香辛料粉末の80%エタノール抽出物についてヒト白血病細胞の増殖抑制率を調べた。(1) タイム,ナツメグ,スターアニス,クローブ,トウガラシ,ターメリック,ジンジャー,ガーリック,シナモン,ブラックペーパー,オールスパイス,ローズマリー,セージ,ローレルの抽出物のうち200μg/mlの濃度で80%以上の細胞増殖抑制率を示したものはタイム,ナツメグ,クローブ,ターメリック,ジンジャー,シナモン,ブラックペーパー,ローズマリー,クミン,ローレルであった。これらの香辛料抽出物の50μg/ml濃度で細胞増殖抑制率が70%以上のものはナツメグ,ターメリック,シナモン,ブラックペーパー.ローズマリー,セージ,ローレルであった。香辛料抽出物の10μg/ml濃度でターメリックは100%,ローレルは95%の高い細胞増殖抑制率を示したのに対して,他のものはいずれも50%以下であった。(2) 細胞増殖抑制率の高かったターメリックとローレルの抽出物の作用機構としてDNA断片化が観察され,アポトーシス誘導によるものと推定した。

Paenibacillus curdlanolyticus B-6 Xyn10C is a single module xylanase consisting of a glycoside hydrolase family-10 catalytic module. The recombinant enzyme, rXyn10C, was produced by Escherichia coli and characterized. rXyn10C was highly active toward soluble xylans derived from rye, birchwood, and oat spelt, and slightly active toward insoluble wheat arabinoxylan. It hydrolyzed xylooligosaccharides larger than xylotetraose to produce xylotriose, xylobiose, and xylose. When rye arabinoxylan and oat spelt xylan were treated with the enzyme and the hydrolysis products were analyzed by thin layer chromatography (TLC), two unknown hydrolysis products, U1 and U2, were detected in the upper position of xylose on a TLC plate. Electrospray ionization mass spectrometry and enzymatic analysis using Bacillus licheniformis α-L-arabinofuranosidase Axh43A indicated that U1 was α-L-Araf-(1→2)-[α-L-Araf-(1→3)]-D-Xylp and U2 was α-L-Araf-(1→2)-D-Xylp, suggesting that rXyn10C had strong activity toward a xylosidic linkage before and after a doubly arabinose-substituted xylose residue and was able to accommodate an α-1,2- and α-1,3-linked arabinose-substituted xylose unit in both the -1 and +1 subsites. A molecular docking study suggested that rXyn10C could accommodate a doubly arabinose-substituted xylose residue in its catalytic site, at subsite -1. This is the first report of a xylanase capable of producing α-L-Araf-(1→2)-[α-L-Araf-(1→3)]-D-Xylp from highly arabinosylated xylan.

We have investigated the effect of extract prepared from skin of red grape (Vitris vinifera) on the growth of human lymphoid leukemia Molt 4B cells. It was found that the extract obtained by extraction with butanol suppressed the growth of Molt 4B cells. Fraction IV obtained by separation with HPLC inhibited the growth of the Molt 4B cells by 86% at the concentration of 600 μM. We determined the structure of compound contained in Fraction IV and identified it as enin, a kind of anthocyanin, by 1H- and 13C-NMR. The enin-treated cells clearly exhibited morphological changes, indicating the blebbing and chromatin condensation which are characteristic of apoptosis. Characteristic oligonucleosomal-sized DNA fragments were observed in the enin-treated cells. These findings suggest that the suppression by enin of growth of Molt 4B cells results from the induction of apoptosis by this compound.

Very fast tryptic degradation of the silkworm diapause hormone was found and the degradation pathway was analyzed by moderating the reaction conditions. It proceeded via cleavage at Arg23 and finally at Arg15 of DH. As the C-terminal structure of DH was essential for exhibiting bioactivity, the first cleavage caused rapid inactivation of the hormone. This tryptic digestion was strongly suppressed by adding VAP-map, a synthetic analog of the cuticular peptide of silkmoths, Bm ACP-6.7 (VAP-peptide), which is a natural synergist of DH. VAP-map suppressed the enzymic reaction by interacting with the substrate, but not with the enzyme.

(1) ゴマ脱脂粕に生育した微生物をAbsidia corymbiferaであると同定した.(2) ゴマ脱脂粕を本菌と接触させると原料に含まれていたセサミノールトリグルコシドが著しく減少し,抗酸化能を持つセサミノールが大量に生成する事を発見した.(3) 生成物の経時変化解析から,セサミノールの生成は菌との接触開始84時間後に最大となる.(4) セサミノール生成はセサミノールトリグルコシドの酵素触媒による脱グルコシル化によるものと推測した.(5) 今回開発した手法によれば,100gのゴマ脱脂粕は抗酸化性を持つ約200mgのセサミノールを与える.(6) 84時間の培養では,セサミンとセサモリンの含量は変化しなかった.

Three thiosulfinates were isolated from oil-macerated garlic extract, and their structures were identified as 2-propene-1-sulfinothioic acid S-(Z,E)-1-propenyl ester [AllS(O)SPn-(Z,E)], 2-propenesulfinothioic acid S-methyl ester [AllS(O)SMe], and methanesulfinothioic acid S-(Z,E)-1-propenyl ester [MeS(O)SPn-(Z,E)]. This is the first report of isolating these thiosulfinates from oil-macerated garlic extract. Antimicrobial activities of AllS(O)SPn-(Z,E) and AllS(O)SMe against Gram-positive and negative bacteria and yeasts were compared with 2-propene-1-sulfinothioic acid S-2-propenyl ester [AllS(O)SAll, allicin] which is well-known as the major thiosulfinate in garlic. Antimicrobial activity of AllS(O)SMe and AllS(O)SPn-(Z,E) were comparable and inferior to that of allicin, respectively. This result suggested that the antimicrobial activity of 2-propene sulfinothioic acid S-alk(en)yl esters were affected by alk(en)yl groups. The order for antimicrobial activity was: allylor = methylpropenyl.

We found that sesaminols, a mixture of sesaminol and its stereoisomers, are potent inhibitors of the oxidation of low-density lipoprotein induced by 2,2'-azobis(2,4-dimethylvaleronitrile). Although sesaminols strongly inhibit lipid peroxidation related to their ability to scavenge free radicals, their antioxidant effects have not been investigated. To confirm the involvement of the phenolic moiety in the sesaminol structure in antioxidant activity, sesaminols were reacted with 2,2'-azobis(2,4-dimethylvaleronitrile). The reaction products were isolated by high-performance liquid chromatography and found to have a 1-cyano-1,3-dimethyl-butyl-peroxyl group in their structures. These chemical structures suggest that the sesaminols reacted with the alkylperoxyl radicals to form four major reaction products that are stereoisomers of each other, although the stereochemistry of each isomer has not yet been confirmed. Further instrumental analyses of the reaction products may increase our understanding of the antioxidant activity of sesaminols.

A compound showing antimicrobial activity was isolated from an oil-macerated garlic extract by silica gel column chromatography and preparative TLC. On basis of the results of NMR and MS analyses, it was identified as Z-4,5,9-trithiadeca-1,6-diene-9-oxide (Z-10-devinylajoene; Z-10-DA). Z-10-DA exhibited a broad spectrum of antimicrobial activity against such microorganisms as gram-positive and gram-negative bacteria and yeasts. The antimicrobial activity of Z-10-DA was comparable to that of Z-ajoene, but was superior to that of E-ajoene. Z-10-DA and Z-ajoene are different in respect of substitution of the allyl group by the methyl group flanking a sulfinyl group. This result suggests that substitution by the methyl group would also be effective for the inhibition of microbial growth.

The ‘Hanasanagitake’ mushroom, Isaria japonica , is a folk medicine and a traditional health food. Fractionation of the 60% ethanol extract of the mushroom, guided by the antioxidant activity test, led to the isolation of a new pseudo-di-peptide, and it was called ‘hanasanagin’. Spectral analysis and chemical transformation determined the structure of hanasanagin as 3,4-diguanidinobutanoyl-DOPA.

Persimmon (Diospyros) contains polyphenol compounds (proanthocyanidin polymers). The content of polyphenol compounds in persimmon extract was estimated to be 27% by the acidified vanillin method. The effect of persimmon extract on a two-stage model of carcinogenesis which consisted of initiation by 7,12-dimethylbenz [a]-anthracene (DMBA) and promotion by 12-O-tetradecanoyl-phorbol-13-acetate (TPA) was investigated. Formation of mouse skin papillomas was remarkably inhibited by persimmon extract. In in vitro experiments, it was found that the [3H]TPA binding to mouse skin particulates and the activity of ornithine decarboxylase (ODC), a marker enzyme of tumor promotion, were inhibited dose-dependently by the persimmon extract, and that DPPH radicals and active oxygens (OH, O2-) were scavenged by persimmon extract. These findings suggest that the suppression of the skin carcinogenesis results from the inhibition of [3H]TPA binding to mouse skin particulate and ODC activity and the scavenging of active oxygens by the persimmon extract.

An ethanol extract obtained from freeze-dried young green barley leaves was fractionated with column chromatography. The fraction exhibiting strongest antioxidative activity was further purified by HPLC. The active component was identified as 2''(3'')-O-glycosylisoviterin by gas chromatography, W, FABMS, 1 H NMR, and 13 C NMR

The exposure of human lymphoid leukemia Molt 4B cells to sesamol, a component of sesame oil led to both growth inhibition and the induction of apoptosis. Morphological change showing apoptotic bodies was observed in the cells treated with sesamol. The fragmentation of DNA by sesamol to oligonucleosomal-sized fragments that are characteristics of apoptosis was observed to be concentration- and time-dependent. On the other hand, in the sesamol-treated normal human lymphocytes morphological change and DNA fragmentation were not observed at all. These findings suggest that growth inhibition of Molt 4B cells by sesamol results from the induction of apoptosis in the cells.

タケノコの皮の酢酸エチル抽出物より2種の抗酸化性物質,TricinおよびTaxifolinをはじめて単離同定した.TricinおよびTaxifolinの抗酸化性はそれぞれα-Tocopherolの約1/10および約1/100程度であった.