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Although vaccines and antiviral drugs are available, influenza viruses continue to pose a significant threat to vulnerable populations globally. With the emergence of drug-resistant strains, there is a growing need for novel antiviral therapeutic approaches. We found that 18-hydroxyferruginol (1) and 18-oxoferruginol (2) isolated from Torreya nucifera exhibited strong anti-influenza activity, with 50% inhibitory concentration values of 13.6 and 18.3 μM against H1N1, 12.8 and 10.8 μM against H9N2, and 29.2 μM (only compound 2) against H3N2 in the post-treatment assay, respectively. During the viral replication stages, the two compounds demonstrated stronger inhibition of viral RNA and protein in the late stages (12–18 h) than in the early stages (3–6 h). Moreover, both compounds inhibited PI3K-Akt signaling, which participates in viral replication during the later stages of infection. The ERK signaling pathway is also related to viral replication and was substantially inhibited by the two compounds. In particular, the inhibition of PI3K-Akt signaling by these compounds inhibited viral replication by sabotaging influenza ribonucleoprotein nucleus-to-cytoplasm export. These data indicate that compounds 1 and 2 could potentially reduce viral RNA and viral protein levels by inhibiting the PI3K-Akt signaling pathway. Our results suggest that abietane diterpenoids isolated from T. nucifera may be potent antiviral candidates for new influenza therapies.

The inhibition efficacy of an extract from Ecklonia cava (E. cava) was studied to determine whether the extract and compounds exhibited inhibitory activity against VHSV in the fathead minnow (FHM) cell line and following oral administration to the olive flounder. Based on its low toxicity and effective concentration, the E. cava extract (Ext) and compounds (eckol and phlorofucofuroeckol A) were selected for further analysis. In the plaque reduction assay, simultaneous co-exposure of VHSV to Ext, eckol and phlorofucofuroeckol A showed a higher level of inhibition than the pre- and post-exposure groups. The antiviral activity in the FHM cell line was time-dependent and increased with the exposure time with the virus and Ext or the compounds. In the in vivo experiments, different Ext concentrations were orally administered to the olive flounder. In trial I, the relative percent survival (RPS) following oral administration of 500 and 50 μg/g/day of Ext was 31.25% and 12.50%, respectively. In trial II, the RPS for 1000, 500 and 50 μg/g/day of Ext was 31.57%, 0% and 0%, respectively. In trial III, the RPS after 1 and 2 weeks (1000 μg/g/day) of exposure to Ext was 26.31% and 31.57%, respectively. Oral administration of Ext (1000 μg/g/day) significantly induced inflammatory cytokine responses (IL-1β, IL-6 and IFN-γ) at 1 and 2 days post-oral administration (dpa). Additionally, IFN-α/β (7–12 dpa), ISG15 (2, 7 and 10 dpa) and Mx (7–12 dpa) were significantly activated in the olive flounder. In conclusion, we demonstrated an inhibitory ability of the E. cava extract and compounds against VHSV in the FHM cell line. Moreover, oral administration of the E. cava extract to the olive flounder enhanced antiviral immune responses and the efficacy of protection against VHSV, resulting in an anti-viral status in the olive flounder.

Sialidases are key virulence factors that remove sialic acid from the host cell surface glycan, unmasking receptors that facilitate bacterial adherence and colonisation. In this study, we developed potential agents for treating bacterial infections caused by Streptococcus pneumoniae Nan A that inhibit bacterial sialidase using Turmeric and curcumin analogues. Design, synthesis, and structure analysis relationship (SAR) studies have been also described. Evaluation of the synthesised derivatives demonstrated that compound 5e was the most potent inhibitor of S. pneumoniae sialidase (IC50 = 0.2 ± 0.1 µM). This compound exhibited a 3.0-fold improvement in inhibitory activity over that of curcumin and displayed competitive inhibition. These results warrant further studies confirming the antipneumococcal activity 5e and indicated that curcumin derivatives could be potentially used to treat sepsis by bacterial infections.

This study aimed to evaluate the effects of cinnamamides on atopic dermatitis (AD) and the mechanisms underlying these effects. To this end, the actions of two cinnamamides, (E)-3-(4-hydroxyphenyl)-N-phenylethyl acrylamide (NCT) and N-trans-coumaroyltyramine (NCPA), were determined on AD by orally administering them to mice. Oral administration of the cinnamamides ameliorated the increase in epidermal and dermal thickness as well as mast cell infiltration. Cinnamamides suppressed serum immunoglobulin (Ig) levels and expression of T-helper (Th)1/Th2 cytokines. Moreover, cinnamamides suppressed interleukin (IL)-4, which plays a crucial role in preparing naïve clusters of differentiation (CD)4+ T cells, and decreased the cervical lymph node size and weight. Interestingly, in almost all cases, NCPA exhibited higher anti-AD activity compared to NCT. These results strongly indicate that NCPA may have potential as an anti-AD agent, and further mechanistic comparative studies of NCT and NCPA are required to determine the cause of differences in biological activity.

Glucosyl stevioside was synthesized via transglucosylation by dextransucrase from Leuconostoc citreum KM20 (LcDexT), forming α-d-glucosyl stevioside. A production yield of 94% was reached after 5days of LcDexT reaction at 30°C. Glucosyl stevioside induced a 2-fold improved quality of taste and sweetness, compared to stevioside. After 15days of storage at 25°C, 98% of glucosyl stevioside in an aqueous solution was present in a soluble form, compared to only 11% for stevioside or rebaudioside A. Furthermore, glucosyl stevioside exhibited a similar or improved stability in commercially available soft drinks, when compared to stevioside and rebaudioside A. These results suggest that glucosyl stevioside could serve as a highly pure and stable sweetener in soft drinks.

Glucosyl rubusosides were synthesized by two dextransucrases. LcDexT was obtained from Leuconosotoc citreum, that LlDexT was obtained from Leuconostoc lactis. LcDexT and LlDexT regioselectively transferred a glucosyl residue to the 13-O-glucosyl moiety of rubusoside with high yield of 59-66% as analyzed by TLC and HPLC. Evaluation of the sweetness of these glucosyl rubusosides showed that their quality of taste, in particular, was superior to that of rubusoside. These results indicate that transglucosylation at the 13-O-glucosyl moiety of rubusoside by different regioselective dextransucrases can be applicable for increasing its sweetness and quality of taste.

Two new guaiane-type (2, 6) and one new furanogermacrane-type (11) sesquiterpenoids have been isolated along with twelve known compounds from an EtOAc-soluble extract of Curcuma phaeocaulis rhizomes. The structures of the isolated compounds were elucidated using a combination of NMR, MS, and circular dichroism (CD) spectra. The inhibitory effects of each compound on lipopolysaccharide (LPS)-induced Toll-like receptor 4 (TLR4) activation in THP-1-Blue cells were assessed, and compound 4 showed more potent inhibitory activity against LPS-stimulated TLR4 activation.

We previously reported that the SbROMT3syn recombinant protein catalyzes the production of the methylated resveratrol derivatives pinostilbene and pterostilbene by methylating substrate resveratrol in recombinant E. coli. To further study the production of stilbene compounds in E. coli by the expression of enzymes involved in stilbene biosynthesis, we isolated three stilbene synthase (STS) genes from rhubarb, peanut, and grape as well as two resveratrol O-methyltransferase (ROMT) genes from grape and sorghum. The ability of RpSTS to produce resveratrol in recombinant E. coli was compared with other AhSTS and VrSTS genes. Out of three STS, only AhSTS was able to produce resveratrol from p-coumaric acid. Thus, to improve the solubility of RpSTS, VrROMT, and SbROMT3 in E. coli, we synthesized the RpSTS, VrROMT and SbROMT3 genes following codon-optimization and expressed one or both genes together with the cinnamate/4-coumarate:coenzyme A ligase (CCL) gene from Streptomyces coelicolor. Our HPLC and LC-MS analyses showed that recombinant E. coli expressing both ScCCL and RpSTSsyn led to the production of resveratrol when p-coumaric acid was used as the precursor. In addition, incorporation of SbROMT3syn in recombinant E. coli cells produced resveratrol and its mono-methylated derivative, pinostilbene, as the major products from p-coumaric acid. However, very small amounts of pterostilbene were only detectable in the recombinant E. coli cells expressing the ScCCL, RpSTSsyn and SbROMT3syn genes. These results suggest that RpSTSsyn exhibits an enhanced enzyme activity to produce resveratrol and SbROMT3syn catalyzes the methylation of resveratrol to produce pinostilbene in E. coli cells.

Two viral proteases of severe acute respiratory syndrome coronavirus (SARS-CoV), a chymotrypsin-like protease (3CL(pro)) and a papain-like protease (PL(pro)) are attractive targets for the development of anti-SARS drugs. In this study, nine alkylated chalcones (1-9) and four coumarins (10-13) were isolated from Angelica keiskei, and the inhibitory activities of these constituents against SARS-CoV proteases (3CL(pro) and PL(pro)) were determined (cell-free/based). Of the isolated alkylated chalcones, chalcone 6, containing the perhydroxyl group, exhibited the most potent 3CL(pro) and PL(pro) inhibitory activity with IC50 values of 11.4 and 1.2 µM. Our detailed protein-inhibitor mechanistic analysis of these species indicated that the chalcones exhibited competitive inhibition characteristics to the SARS-CoV 3CL(pro), whereas noncompetitive inhibition was observed with the SARS-CoV PL(pro).

The current study was designed to assess the inhibitory activity of Broussonetia papyrifera-derived polyphenols against 3-chymotrypsin-like and papain-like coronavirus cysteine proteases. The isolated compounds were broussochalcone B (1), broussochalcone A (2), 4-hydroxyisolonchocarpin (3), papyriflavonol A (4), 3′-(3-methylbut-2-enyl)-3′,4,7-trihydroxyflavane (5), kazinol A (6), kazinol B (7), broussoflavan A (8), kazinol F (9), and kazinol J (10). All polyphenols were more potent against papain-like protease (PLpro) than against 3-chymotripsin-like protease (3CLpro); therefore, we investigated their structural features that were responsible for this selectivity. Compound 4 was the most potent inhibitor of PLpro with an IC50 value of 3.7 μM. The active compounds displayed kinetic behaviors, and the binding constants of their interaction with PLpro were determined from surface plasmon resonance analysis. Our results suggest B. papyrifera constituents as promising candidates for development into potential anti-coronaviral agents.

The crystal structure of sialidase from C. perfringens, a pathogenic bacterium causing various gastrointestinal diseases, was determined in complex with a potent natural polyphenolic geranylated flavonoid-based inhibitor. The complex structure and comparative kinetic studies revealed that the geranyl group and C3′ hydroxyl group of the flavonoid backbone contribute to inhibition of the bacterial sialidase and generation of the stable enzyme–inhibitor complex., Sialidase catalyzes the removal of a terminal sialic acid from glycoconjugates and plays a pivotal role in nutrition, cellular interactions and pathogenesis mediating various infectious diseases including cholera, influenza and sepsis. An array of antiviral sialidase agents have been developed and are commercially available, such as zanamivir and oseltamivir for treating influenza. However, the development of bacterial sialidase inhibitors has been much less successful. Here, natural polyphenolic geranylated flavonoids which show significant inhibitory effects against Cp-NanI, a sialidase from Clostridium perfringens, are reported. This bacterium causes various gastrointestinal diseases. The crystal structure of the Cp-NanI catalytic domain in complex with the best inhibitor, diplacone, is also presented. This structure explains how diplacone generates a stable enzyme–inhibitor complex. These results provide a structural framework for understanding the interaction between sialidase and natural flavonoids, which are promising scaffolds on which to discover new anti-sialidase agents.

Puerarin (P), an isoflavone derived from kudzu roots, has strong biological activities, but its bioavailability is often limited by its low water solubility. To increase its solubility, P was glucosylated by three dextransucrases from Leuconostoc or Streptococcus species. Leuconostoc lactis EG001 dextransucrase exhibited the highest productivity of puerarin glucosides (P-Gs) among the three tested enzymes, and it primarily produced two P-Gs with a 53% yield. Their structures were identified as alpha-D-glucosyl-(1-->6)-P (P-G) by using LC-MS or (1)H- or (13)C-NMR spectroscopies and alpha-D-isomaltosyl-(1-->6)-P (P-IG2) by using specific enzymatic hydrolysis, and their solubilities were 15- and 202-fold higher than that of P, respectively. P-G and P-IG2 are easily applicable in the food and pharmaceutical industries as alternative functional materials.

Orthoreoviruses infect virtually all mammalian species, causing systemic infections including mild gastrointestinal and respiratory illnesses. However, little is known about the prevalence or genetic diversity of porcine orthoreoviruses in South Korea. We examined 237 diarrheic fecal samples collected from 78 pig farms around the country. RT-PCR utilizing primers specific for the L1 gene of mammalian orthoreoviruses showed that 45 (19.0%) samples were positive. The 10 strains isolated from orthoreovirus-positive samples formed typical perinuclear cytoplasmic inclusion bodies and had an atypical hemagglutination pattern; these are characteristics of type 3 orthoreovirus. Phylogenetic analysis of the S1 gene in these 10 Korean and other strains showed that type 3 orthoreoviruses could be divided into four lineages; the 10 Korean strains were included in porcine lineage IV, along with T3/porcine/Sichuan/2006. Sequence analysis showed that strains in lineage IV had nucleotide identities of 97.0–98.1% and deduced amino acid identities of 96.4–98.2%. Sequence analysis of the σ1 protein, a viral attachment protein, revealed that the amino acid sequences associated with neurotropism (amino acids 198–204, 249I, 350D, and 419E) were highly conserved among the Korean strains, confirming that neural tropism was present. In conclusion, our findings suggest that porcine orthoreovirus infections are endemic in pig farms in South Korea and that the 10 novel Korean porcine orthoreoviruses belong to porcine lineage IV of type 3 orthoreovirus. In addition, sequence analysis of S1 genes encoding the σ1 protein showed that the 9 of 10 Korean porcine orthoreoviruses exhibited neural tropism.

The present study investigated the effects of azuki bean (Vigna angularis) extract (VAE) on the progress of atopic dermatitis (AD)-like skin lesions in NC/Nga mice induced by 1-chloro-2,4-dinitrobenzene. The efficacy of VAE in NC/Nga mice was determined by measuring gross and histological skin lesions, serum IgE levels, eosinophil ratio in peripheral leucocytes, and mRNA expression levels of interleukin (IL)-4, tumour necrosis factor (TNF)-α and interferon (IFN)-γ in splenocytes. Continuous ingestion of VAE inhibited the development of the AD-like skin lesions in a dose-dependent manner. In the VAE-treated mice, the numbers of mast cells in the skin, eosinophil ratio in peripheral leucocytes, relative mRNA expression of inflammatory cytokines in the spleen, and serum IgE levels were significantly reduced. Results suggest that VAE can inhibit the development of AD-like skin lesions in NC/Nga mice by regulating immune mediators and cells, and may be an effective alternative therapy for AD.

TPDex, a putative dextranase from Thermoanaerobacter pseudethanolicus, was purified as a single 70 kDa band of 7.37 U/mg. Its optimum pH was 5.2 and the enzyme was stable between pH 3.1 and 8.5 at 70 degrees C. A half-life comparison showed that TPDex was stable for 7.4 h at 70 degrees C, whereas Chaetominum dextranase (CEDex), currently used as a dextranase for sugar milling, was stable at 55 degrees C. TPDex showed broad dextranase activity regardless of dextran types, including dextran T2000, 742CB dextran, and alternan. TPDex showed the highest thermostability among the characterized dextranases, and may be a suitable enzyme for use in sugar manufacture without decreased temperature.

In this study, twelve neuraminidase (NA) inhibitory compounds 1-12 were isolated from heartwood of Caesalpinia sappan on the basis of their biological activities against three types of viral NAs. Of isolated homoisoflavonoids, sappanone A (2) showed the most potent NAs inhibitory activities with IC(50) values of 0.7 µM [H1N1], 1.1 µM [H3N2], and 1.0 µM [H9N2], respectively, whereas saturated homoisoflavonoid (3) did not show significantly inhibition. This result revealed that α,β-unsaturated carbonyl group in A-ring was the key requirements for viral NAs inhibitory activity. In our enzyme kinetic study, all NA inhibitors screened were found to be reversible noncompetitive types.

The papain-like protease (PL(pro)), which controls replication of the severe acute respiratory syndrome coronavirus (SARS-CoV), has been identified as a potential drug target for the treatment of SARS. An intensive hunt for effective anti-SARS drugs has been undertaken by screening for natural product inhibitors that target SARS-CoV PL(pro). In this study, diarylheptanoids 1-9 were isolated from Alnus japonica, and the inhibitory activities of these compounds against PL(pro) were determined. Of the isolated diarylheptanoids, hirsutenone (2) showed the most potent PL(pro) inhibitory activity, with an inhibitory concentration (IC(50)) value of 4.1 µM. Structure-activity analysis showed that catechol and α,β-unsaturated carbonyl moiety in the molecule were the key requirement for SARS-CoV cysteine protease inhibition.

Inhibitors of cell adhesion molecule-mediated cell adhesion might be novel therapeutic agents for the treatment of various inflammatory diseases. In this study, nine phenolic compounds were isolated from the methanol extracts of Zingiber officinale roots by bioactivity-guided fractionation. The structures of the compounds were determined by spectroscopic analysis ( 1 H, 13 C NMR and MS), to be 6-gingerol ( 1 ), 8-gingerol ( 2 ), 10-gingerol ( 3 ), 6-shogaol ( 4 ), 8-shogaol ( 5 ), 10-shogaol ( 6 ), dehydro-6-gingerdione ( 7 ), dehydro-10-gingerdione ( 8 ) and 6-paradol ( 9 ). Compounds 3 , 4 , 5 and 7 inhibited direct binding between sICAM-1 and LFA-1 of the THP-1 cells in a dose-dependent manner with IC 50 values of 57.6, 27.1, 65.4 and 62.0 μM, respectively. Compounds 4 and 7 had an inhibitory effect on direct binding between sVCAM-1 and VLA-4 of THP-1 cells. These results suggest that the phenolic compounds from Z. officinale roots are good candidates for therapeutic strategies aimed at inflammation.

Neuraminidase is a proven target in anti-viral drug development. It also appears to be important for infection by certain pathogenic bacteria and has been implicated in biofilm formation. Based on activity-guided fractionation, the acetone extract of Amorpha fruticosa roots gave four flavanones 1-4 and three rotenoids 5-7 which were identified as amoradicin (1), amorisin (2), isoamoritin (3), amoricin (4), amorphigeni (5), dalbinol (6), and 6-ketodehydroamorphigenin (7), respectively. All isolated inhibitors showed strong neuraminidase inhibition with IC₅₀s between 0.12 and 22.03 μM. In particular, amorisin 2 exhibited 120 nM IC(₅₀, which is 30-fold more potent than the positive control, quercetin. In addition, this is the first report detailing rotenoids (IC₅₀ = 8.34-16.74 μM) exhibiting neuraminidase inhibition. Kinetic analysis revealed that all inhibitors were noncompetitive. The most active neuraminidase inhibitors (2, 3, 5, 6) were proven to be present in the native root in high quantities by HPLC. Finally, at concentrations where no toxicity was observed, 3 and 6 inhibited Pseudomonas aeruginosa biofilm production. 29.7% and 21.0% inhibition respectively was observed at 25 μΜ.

Myeloid differentiation factor 88 (MyD88) is a universal adaptor protein able to activate nuclear factor-kappa B (NF-κB) through interactions with interleukin-1 receptor (IL-1R) and the Toll-like receptors (TLRs), with the exception of TLR3. Here, we describe the identification of MyD88 from the rock bream fish Oplegnathus fasciatus and its characterization based on GS-FLX™ sequencing. The cDNA of rock bream MyD88 was found to be composed of 1626 bp, with an 867 bp open reading frame that encodes 288 amino acids. The deduced amino acid sequence of MyD88 possessed both a conserved death domain at the amino terminus and a typical Toll-IL-1 receptor (TIR) domain at the carboxyl terminus, similar to that found in other fishes, amphibians, avians, mammals and invertebrates. The mRNA expression pattern of MyD88 in healthy and bacterially challenged rock bream were examined using quantitative real-time polymerase chain reaction (qRT-PCR). MyD88 transcripts were found to be strongly expressed in blood, gill, liver, spleen, head kidney and kidney, moderately expressed in skin, brain and intestine, and weakly expressed in muscle. Expression levels of MyD88 in blood, spleen and head kidney were dramatically up-regulated upon exposure to LPS and the Gram-negative bacteria Edwardsiella tarda, suggesting that MyD88 plays an important role in rock bream defenses against bacterial infection.

The seeds of Psoralea corylifolia were extracted into five different polar solvents: chloroform, 50% ethanol in water, ethanol, methanol and water. All extracts were evaluated for glycosidase inhibitory activity. The chloroform extract (CE) showed the lowest IC 50 values against α-glucosidase (82.9 μg/ml) and α-mannosidase (132 μg/ml). Chromatography of CE yielded nine phenolic compounds which were identified as isovabachalcone ( 1 ), 4′- O -methylbavachalcone ( 2 ), isobavachromene ( 3 ), corylifolin ( 4 ), bavachinin ( 5 ), psoralidin ( 6 ), neobavaisoflavone ( 7 ), corylifol A ( 8 ), and bakuchiol ( 9 ). All isolated compounds, apart from compound 5 , possessed α-glucosidase inhibitory activities. Among them, compounds 6 – 8 exhibited potent inhibition with IC 50 s of 13.7, 27.7 and 11.3 μM, respectively. Furthermore, compounds 2 and 6 showed α-mannosidase inhibitory activity. Mechanistic analysis of their inhibition modes against α-glucosidase showed that compounds ( 6 and 7 ) were noncompetitive, whereas compound 8 was mixed. Furthermore, the most active glycosidase inhibitors ( 2 , 6 – 8 ) were proven to be present in the native seed in high quantities by an HPLC chromatogram.

Oxidized low-density lipoprotein (oxLDL) and reactive oxygen species (ROS) play key roles in the early stage of atherosclerosis. Nitric oxide (NO) and ROS are responsible for regulation of the transcriptional pathways of nuclear Factor-kappaB (NF-kappaB) and mitogen-activated protein kinase (MAPK), key regulators of cellular inflammatory and immune responses. Previously, we examined LDL-antioxidant activities of the nine flavonoids isolated from Sophora flavescens. Among these, two lavandulyl flavonoids, kurarinone (1) and kuraridin (2) inhibited inducible nitric oxide synthase (iNOS)-dependent NO production and ROS generation, and suppressed remarkably the expression of inflammatory cytokines, CCL2, tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta, and iNOS in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages. Moreover, compounds 1 and 2 attenuated NF-kappaB activation by inhibition of IkappaBalpha proteolysis and p65 nuclear translocation, as well as phosphorylation of extracellular signal-regulated kinase (ERK)1/2, c-Jun N-terminal kinase (JNK), and p38 MAP kinases.

Oxidation of low density lipoprotein (LDL) is strongly implicated as a key process in the onset of atherosclerosis. In this study, nine alkylated (C10-C5) flavonoids from Sophora flavescens were examined for their inhibitory effects on copper-induced LDL oxidation. Of the flavonoids tested, sophoraflavanone G (1), kurarinone (2), kurarinol (3), norkurarinol (4), and kuraridin (9) inhibited the generation of thiobarbituric acid reactive substances (TBARS) with IC50s of 7.9, 14.5, 22.0, 26.9, and 17.5 microM, respectively. The most potent inhibitor, compound 1, also demonstrated significant activities in complementary in vitro investigations, such as lag time (130 min at 5 microM), relative electrophoretic mobility (REM) of ox-LDL (80% inhibition at 20 microM), and fragmentation of apoB-100 (inhibition of 71% at 20 microM). Analysis of the structures of these compounds reveals that a resorcinol moiety in the B-ring is strongly correlated with protection of LDL-oxidation.

A series of multi-substituted oximes were prepared and their potencies for inhibiting lipoprotein-associated phospholipase A(2) (Lp-PLA(2)) activity were evaluated in vitro. Among them, compounds 3a, 3b, and 3m were identified to display a micromolar potency for inhibiting Lp-PLA(2) in whole human plasma and isolated human LDL. Based on these results, structure-activity relationship was studied on modification of three parts of R(1), R(2), and R(3) to identify a potent pharmacophore for Lp-PLA(2). In an attempt to introduce various functional groups at R(2) and R(3), we discovered that replacement of less lipophilic groups led to an increase of inhibitory activity. Among the tested oxime derivatives, cyano- and morpholino-substituted analogue 4f at R(2) and R(3) had the highest potency with an IC(50) value of 0.05 microM in whole human plasma.

Diethyl alpha-alkynyl-alpha-methoxymalonates (2a--e) were smoothly hydrolyzed and then decarboxylated under alkaline conditions employing 1 N KOH in EtOH to give conjugated allenyl esters (6a--e) in high yields, and similar alkaline treatment of diethyl alpha-alkynyl-alpha-acetylaminomalonates (5a, b, d, e) furnished unexpectedly the oxazoles (7a, b, d, e) having three substituent groups in excellent yields.

The oxidized low-density lipoprotein (ox-LDL) plays a critical role at the early stages of atherosclerosis. Thus, the prevention of LDL-oxidation by antioxidants may arrest the progression of atherosclerosis. Two quinoline alkaloids, 3,8-dihydroxyquinoline (1) and 2,8-dihydroxy-3,4-dimethoxyquinoline (3), and 2,4-di-tert-butylphenol (2) were isolated from the dried body of Scolopendra subspinipes. Compounds 1-3 exhibited antioxidant activities on copper-mediated (1: IC50=2.6 microM, 2: IC50=8.2 microM, 3: IC50=63.0 microM), AAPH-mediated oxidation (1: IC50=3.9 microM, 2: IC50=9.9 microM, 3: IC50=71.8 microM), and SIN-1-mediated oxidation (1: 70%, 2: 52%, 3: 29% at 5.0 microM) in the TBARS assay. The antioxidant activities of compounds 1-3 were tested with respect to other parameters, such as the lag time of conjugated diene fromation, relative electrophoretic mobility (REM) of ox-LDL, and apoB-100 fragmentation on copper-mediated LDL-oxidation. In addition, compounds 1-3 showed 1,1-diphenyl-2-picrylhydrasyl (DPPH) radical scavenging activity and compound 1 also exhibited metal chelating activity.

A new diarylbutane lignan, 2′-hydroxy dihydroguaiaretic acid (4), and a known 8-O-4′-type neolignan, machilin D (5), were isolated from the ethyl acetate extracts of the underground parts of Saururus chinensis. Compounds 4 and 5 exhibited low-density lipoprotein (LDL)-antioxidant activity in the thiobarbituric acid-reactive substances (TBARS) assay (4: IC50 = 3.3 μM and 5: IC50 = 3.8 μM), the lag time of conjugated diene production, the relative electrophoretic mobility (REM) of ox-LDL, the apoB-100 fragmentation on copper-mediated LDL oxidation and the macrophage-mediated LDL oxidation, and radical DPPH scavenging activity.

Compounds 4a-j and 5 were synthesized by cyclocondensation of 3a-j and hydrazine and showed significant LDL-antioxidant activities in the TBARS assay, the lag time of conjugated diene production, the relative electrophoretic mobility (REM) of ox-LDL, the apoB-100 fragmentation, and the macrophage-mediated LDL oxidation. Among compounds 4a-j and 5, 4a was found to be the most active compound as an inhibitor of LDL oxidation and 4a (IC50 = 0.1 microM) was 6-fold more potent than probucol (IC50 = 0.6 microM) in the TBARS assay.

Acyl-CoA: cholesterol acyltransferase (ACAT) plays an important role in the esterification of cholesterol with its substrates, cholesterol and fatty acyl coenzyme A, to facilitate both intracellular storage and intercellular transport. ACAT-1 is more involved in macrophage foam cell formation and ACAT-2 plays a critical role in the cholesterol absorption process in intestinal enterocytes. Three aliphatic acid amides, beta-sanshool (1), gamma-sanshool (2), and hydroxy-beta-sanshool (3), were isolated by bioassay-guided fractionation of the ethanolic extracts of Zanthoxylum piperitum DC. Compounds 1 and 2 inhibited human ACAT-1 and -2 activities with IC50 values of 39.0 and 79.7 microM for 1 and of 12.0 and 82.6 microM for 2, respectively. However, the hACAT-1 and -2 inhibitory activities of compound 3 having hydroxyl group were relatively less than those of compounds 1 and 2. A semi-synthetic compound 4, which has acetyl residue at 2'-OH of compound 3, exhibited the increased hACAT-1 and -2 inhibitory activities with IC50 values of 28.1 and 87.5 microM, respectively.

Acyl-CoA: cholesterol acyltransferase (ACAT) catalyzes the acylation of cholesterol to cholesteryl ester with long chain fatty acids and ACAT inhibition is a useful strategy for treating hypercholesterolemia or atherosclerosis. Pentacyclic triterpenes, ursolic acid (1), oleanolic acid (2), and betulinic acid (3) were isolated from the methanol extracts of the leaves of Lycopus lucidus TURCZ. by bioassay-guided fractionation. The structures of compounds 1-3 were elucidated by their spectroscopic data analysis. Among them, betulinic acid (3) exhibited more potent human ACAT-1 and ACAT-2 inhibitory activities with IC(50) values of 16.2+/-0.6 and 28.8+/-1.3 microM, respectively.

This study aimed to develop an economically viable enzyme for the optimal production of steviol (S) from stevioside (ST). Of 9 commercially available glycosidases tested, S-producing β-glucosidase (SPGase) was selected and purified 74-fold from Penicillium decumbens naringinase by a three-step column chromatography procedure. The 121-kDa protein was stable at pH 2.3-6.0 and at 40-60 °C. Hydrolysis of ST by SPGase produced rubusoside (R), steviolbioside (SteB), steviol mono-glucoside (SMG), and S, as determined by HPLC, HPLC-MS, and (1)H- and (13)C-nuclear magnetic resonance. SPGase showed higher activity toward steviol mono-glucosyl ester, ST, R, and SMG than other β-linked glucobioses. The optimal conditions for S production (30 mM, 64 % yield) were 47 mM ST and 43 μl of SPGase at pH 4.0 and 55 °C. This is the first report detailing the production of S from ST hydrolysis by a novel β-glucosidase, which may be useful for the pharmaceutical and agricultural areas.

G9 group A rotaviruses (RVAs) are considered important pathogens in pigs and humans, and pigs are hypothesized to be a potential host reservoir for human. However, intestinal and extra-intestinal pathogenicity and viremia of porcine G9 RVAs has remained largely unreported. In this study, colostrum-deprived piglets were orally infected with a porcine G9P[23] or G9P[7] strain. Histopathologically, both strains induced characteristic small intestinal lesions. Degeneration and necrosis of parenchymal cells were observed in the extra-intestinal tissues, but most predominantly in the mesenteric lymph nodes (MLNs). RVA antigen was continuously detected in the small intestinal mucosa and MLNs, but only transiently in cells of the liver, lung, and choroid plexus. Viral RNA levels were much higher in the feces and the MLNs compared to other tissues. The onset of viremia occurred at day post infection (DPI) 1 with the amount of viral RNA reaching its peak at DPI 3 or 5, before decreasing significantly at DPI 7 and remaining detectable until DPI 14. Our data suggest that porcine G9 RVAs have a strong small intestinal tropism, are highly virulent for piglets, have the ability to escape the small intestine, spread systemically via viremia, and replicate in extra-intestinal tissues. In addition, MLNs might act as a secondary site for viral amplification and the portal of systemic entry. These results add to our understanding of the pathogenesis of human G9 RVAs, and the validity of the pig model for use with both human and pig G9 RVAs in further studies.

Hesperetin ester and ether derivatives possessing a long alkyl chain were synthesized for examining their hypocholesterolemic activities in high cholesterol-fed mice. Hesperetin 7-O-lauryl ether (4b) and hesperetin 7-O-oleyl ether (4e) exhibited strong cholesterol-lowering effects.

A recombinant putative dextransucrase (DexT) was produced from Leuconostoc citreum KM20 as a 160 kDa protein, but its productivity was very low (264 U/l). For optimization, we examined enzyme activity in 7 Escherichia coli strains with inducer molecules such as lactose or IPTG. E. coli BL21-CodonPlus(DE3)-RIL exhibited the highest enzyme activity with lactose. Finally, DexT activity was remarkably increased by 12-fold under the optimized culture conditions of a cell density to start induction (OD₆₀₀) of 0.95, a lactose concentration of 7.5 mM, and an induction temperature of 17 degrees C. These results may effectively apply to the heterologous expression of other large DexT genes.

A variety of flavones (8 examples) is isolated by activity-guided fractionation of the methanolic extracts from Morus Ihou stem bark.

Several allenyl ketones possessing an aryl group were submitted to a new intramolecular endo-mode cyclization with BF3·OEt2 or TiCl4 in CH2Cl2 to afford the corresponding 6-, 7-, and 8-membered car...

A convenient preparation of allenyl aryl ketones was achieved by the Weinreb-modified Grignard reaction of N-methoxy-N-methylamides with propargylmagnesium bromide. On treatment with BF3·OEt2, the ...

Lysozyme (muramidase) represents an important defense molecule of the fish innate immune system. Known for its bactericidal properties, lysozyme catalyzes the hydrolysis of β-(1,4)-glycosidic bonds between the N-acetyl glucosamine and N-acetyl muramic acid in the peptidoglycan layer of bacterial cell walls. In this study, the complete coding sequence of a g-type lysozyme (RBgLyz) was identified in the Oplegnathus fasciatus rock bream fish genome by means of multi-tissue normalized cDNA pyrosequencing using Roche 454 GS-FLX™ technology. RBgLyz is composed of 669 bp, with a 567 bp open reading frame that encodes 188 amino acids. Protein motif searches indicated that RBgLyz contains the soluble lytic transglycosylase domain involved in maintaining cell wall integrity. Furthermore, RBgLyz shares significant identity (81.4%) with Chinese perch Siniperca chuatsi. Quantitative real-time RT-PCR analysis results showed that RBgLyz transcripts are constitutively expressed in various tissues from healthy rock breams. In order to determine RBgLyz function in immunity, its expression was analyzed in head kidney following exposure to known immune stimulants or pathogens. RBgLyz transcripts were significantly up-regulated in response to challenge with lipopolysaccharide (LPS) and Edwardsiella tarda, as compared to non-injected control fish. Polyinosinic:polycytidylic acid (poly I:C) dsRNA stimulated a moderate expression of RBgLyz, as did Streptococcus iniae but to a lesser extent. There were no specific time-dependent effects on RBgLyz mRNA expression observed in response to rock bream iridovirus (RBIV) infection. Taken together, the gene expression results indicated that g-type lysozyme plays a role in the innate immune response to LPS, poly I:C, E. tarda and S. iniae in rock bream. Thus, we generated recombinant RBgLyz in an Escherichia coli expression system and characterized its antimicrobial activity. Our results indicated that recombinant RBgLyz had lytic activity against Gram-negative Vibrio salmonicida, Gram-positive Listeria monocytogenes, S. iniae and Micrococcus lysodeikticus. In addition, observations by scanning electron microscope (SEM) confirmed that the cell morphology of M. lysodeikticus was altered in the presence of recombinant RBgLyz.

Various diethyl α-alkynylmalonates (DAM) having potential as cysteine proteinase inhibitors were synthesized by treatment of diethyl acetyliminomalonate (or ketomalonate) with several lithium acetylides. Hydrolytic decarboxylation of the DAM under the mild basic conditions afforded oxazole derivatives or allenyl esters which caused the Michael type reaction with EtSH.

Several 2-benzazepine-1,5-diones were efficiently obtained by treatment of the corresponding N-alkyl-3-hydroxy-3-propargylisoindolin-1-ones with some tentative Lewis acids via intramolecular endo-mode cyclization in the allenyl ketone intermediates generated in situ.

Quinone-methide triterpenes, celastrol (1), pristimerin (2), tingenone (3), and iguesterin (4) were isolated from Triterygium regelii and dihydrocelastrol (5) was synthesized by hydrogenation under palladium catalyst. Isolated quinone-methide triterpenes (1-4) and 5 were evaluated for SARS-CoV 3CL(pro) inhibitory activities and showed potent inhibitory activities with IC(50) values of 10.3, 5.5, 9.9, and 2.6 microM, respectively, whereas the corresponding 5 having phenol moiety was observed in low activity (IC(50)=21.7 microM). As a result, quinone-methide moiety in A-ring and more hydrophobic E-ring assist to exhibit potent activity. Also, all quinone-methide triterpenes 1-4 have proven to be competitive by the kinetic analysis.

Dehydration-hydration products were obtained in good yields by the treatment of propargylic hydroxy γ-lactams with some Lewis acids. Several propargylic and allenylic hydroxy γ-lactams were successfully converted to the corresponding ring-expanded benzazepines in various yields via a tandem decyclization-cyclization process in the presence of (TMS)2NLi, (TMS)2NNa, or n-BuLi under reflux conditions in THF.

3-Acetyloxybut-1-enylamines 3–9 were easily transformed using iodine to pyrrolidine derivatives 3a–9a, precursors for aza sugars, via a diastereoselective iodoamidation.

This paper presents the synthesis of 2-amino-, 2-acetamido- and 2-benzamidothioxanthones and their 10,10-dioxides.

Some 6,7-dimethoxy-1-halobenzyl-1,2,3,4-tetrahydroisoquinolines were synthesized from 2-(3,4-dimethoxyphenyl)ethylamine and halophenylacetic acids in three steps in good yield.