Plant cell cultures are widely used in the micro-propagation of clonal plants, especially virus-free plants, and in the production of useful metabolites such as paclitaxel. On the other hand, the use of plant cell cultures for the more basic characterization of plant functions is rather limited due to the difficulties associated with functional differentiation in cell cultures. In this review, I overview our experience with functionally differentiated cultured plant cells and their characteristics, especially with regard to photoautotrophism and secondary metabolism. I emphasize the high potential of functionally differentiated cell cultures, as well as some of the pitfalls, in the characterization of plant functions and biotechnological applications.

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Circadian clocks enable organisms to define subjective time, that is, to anticipate diurnal day and night cycles. Endogenous circadian rhythms regulate many aspects of an organism's physiological and morphological growth and development. These daily oscillations are synchronized to the environment by external cues such as light and temperature, resulting in enhanced fitness and growth vigor in plants. Recent findings concerning biochemical properties of central oscillators in Arabidopsis thaliana have advanced our understanding of circadian clock function. Central oscillators are composed of three classes of transcriptional repressors. The interactions among them include a repressilator structure. Output from the circadian clock is transduced through regulating transcription of downstream genes directly by the oscillator components. The essential role of the output pathway in the circadian system is to make different elementary steps responsible for daily cellular processes exert maximum effects at specific times of the day. Recently, significant progress was made in defining the mechanisms by which plant growth on a day-to-day basis is activated at specific times of the day in a manner dependent on photoperiod and temperature conditions. Plant growth is controlled by the clock through interactions with light and phytohormone signaling. This review focuses on the node that connects clock output to light and phytohormone signaling that coordinates plant growth with rhythmic changes in the environment.

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Im-3 was isolated from the venom of the scorpion Isometrus maculatus through several steps of HPLC fractionation based on the insect paralytic activity. Injecting Im-3 into crickets induced paralysis, but no toxicity was apparent in mice after an intracerebroventricular injection. Im-3 shares sequence similarity to scorpion β-toxins that specifically affect insect sodium channels.

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The full-length cDNA of the gene PoLOX1 encoding a lipoxygenase (LOX) and its corresponding genomic DNA were isolated from the basidiomycete mushroom Pleurotus ostreatus strain H1. The deduced amino acid sequence of PoLOX1 showed similarity to a valencene dioxygenase of Pleurotus sapidus, putative LOX-like proteins from ascomycete, basidiomycete, and deuteromycete fungi, and known LOXs from plants, animals, and bacteria. PoLOX1 also contained the LOX iron-binding catalytic domain in the C-terminal region, but not the polycystin-1, lipoxygenase, alpha-toxin (PLAT) domain, which is usually found in the N-terminal region of eukaryotic LOXs. Genomic sequence analysis revealed that PoLOX1 was interrupted by one intron, and that the promoter region included TATA and CAAT boxes. Southern blot analysis indicated that PoLOX1 is a member of a small gene family comprising highly similar genes. Northern blot analysis revealed that it is transcribed more abundantly in the stipes of the fruit bodies than in the caps.

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Considering the growing evidence of the presence of antioxidant compounds in plant extracts, the objectives of this study were to identify antioxidant compounds in Lindera obtusiloba Blume (Lauraceae) and to evaluate their antimelanogenic activities on B16F10 melanoma cells. Organic solvent fractions were separated from L. obtusiloba extracts (LOE). The ethyl acetate fraction (LOE-E) was significantly active against oxidative damage induced by tert-butyl hydroperoxide in primary rat hepatocytes. Two single purified compounds, quercitrin (quercetin-3-O-α-L-rhamnopyranoside) and afzelin (kaempferol-3-O-α-L-rhamnoside), were identified by HPLC and NMR. These compounds were evaluated for antioxidant activities by 1,1-diphenyl 2-picrylhydrazyl (DPPH) radical scavenging assay and ferric reducing antioxidant power (FRAP) assay, and for their antimelanogenic activities by tyrosinase inhibitory assay melanin formation inhibition assay and Western bolt analysis for the signaling pathway. The significant effects of quercitrin on antioxidant and antimelanogenic activities, and signal modulation of ERK and MITF in B16F10 melanoma cells were observed. This is the first report to identify quercitrin in L. obtusiloba and its whitening effect.

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We synthesized a novel series of (E)-2-((substituted phenyl)diazenyl)phenyl 4-methylbenzenesulfonate derivatives (2 and 3) and (E)-2-((substituted phenyl)diazenyl)phenol derivatives (4 and 5), and conducted an evaluation in order to determine their inhibitory effects on mushroom tyrosinase, with the aim of discovering a tyrosinase inhibitor. Most of the compounds (3–5) exhibited higher inhibitory effects than kojic acid (IC₅₀ = 49.08 µM), a representative tyrosinase inhibitor. A novel synthesized compound, (E)-2-((2,4-dihydroxyphenyl)diazenyl)phenyl 4-methylbenzenesulfonate (3), showed the best results with an IC₅₀ of 17.85 µM, and showed competitive inhibition on Lineweaver-Burk plots, as further confirmed by the docking results. In addition, active compounds 3–5 were not cytotoxic to cultured B16F10 cells at the concentrations tested, and inhibited both tyrosinase and melanin synthesis. Therefore the active compounds (3–5) might be considered excellent candidates for use in the development of therapeutic agents for diseases associated with hyperpigmentation.

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The recently cloned β-galactosidase from Bacillus circulans ATCC 31382, designated BgaD, contains a multiple domain architecture including a F5/8 type C domain or a discoidin (DS) domain in the C-terminal peptide region. Here we report that the DS domain plays an essential role in repressing the production of galactooligosaccharides (GOSs). We prepared deletion mutants and point-mutated forms of rBgaD-A (deletion of the BgaD signal peptide) to compare their reaction behaviors. The yields of GOSs for all of the point-mutated forms as well as the deletion mutants of rBgaD-As increased as compared to rBgaD-A. In particular, W1540A mutant BgaD-A (rBgaD-A_W1540A) produced much more GOSs than rBgaD-A. Surface plasmon resonance experiments indicated that both the wild-type and the W1540A mutant DS domains showed high affinity for galactosyllactose. rBgaD-A, which has a wild-type DS domain, showed high hydrolytic activity toward galactosyllactose, while the hydrolytic activities of rBgaD-D, without a DS domain, and rBgaD-A_W1540A, with a mutant DS domain were extremely low. The findings obtained in this study indicate that the wild-type DS domain of rBgaD-A has a function that aids galactosyllactose molecules to be properly oriented within the active site, so that they can be hydrolyzed efficiently to produce galactose/glucose by inhibiting the accumulation of GOSs.

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Treatment of Bacillus licheniformis α-amylase (BLA) with guanidine hydrochloride (GdnHCl) produced both denatured and aggregated forms of the enzyme as studied by circular dichroism, fluorescence, UV difference spectroscopy, size exclusion chromatography (SEC), and enzymatic activity. The presence of CaCl₂ in the incubation mixture produced significant recovery in spectral signals, being complete in presence of 10 mM CaCl₂, as well as in enzymatic activity, which is indicative of protein stabilization. However, the SEC results obtained with GdnHCl-denatured BLA both in the absence and the presence of 10 mM CaCl₂ suggested significant aggregation of the protein in the absence of CaCl₂ and disaggregation in its presence. Although partial structural stabilization with significant retention of enzymatic activity was observed in the presence of calcium, it was far from the native state, as reflected by spectral probes. Hence, spectral results as to BLA stabilization should be treated with caution in the presence of aggregation.

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The nucleotide sequence of the gene encoding Protobothrops elegans (Crotalinae) pancreatic phospholipase A₂ (PLA₂), abbreviated PePancPLA₂, was determined by means of inverted PCR techniques. Since its deduced amino acid sequence contains a pancreatic loop and shows high similarity to that of Laticauda semifasciata (Elapinae) group IB pancreatic PLA₂, PePancPLA₂ is classified into group IB PLA₂. The nucleotide sequences of the PePancPLA₂ gene, the L. semifasciata group IB pancreatic PLA₂ gene, and the L. semifasciata group IA venom PLA₂ gene are similar to one another but greatly dissimilar to those of Protobothrops genus (Crotalinae) group II venom PLA₂ genes, suggesting that the Elapinae group IB PLA₂ gene and the group IA PLA₂ gene appeared after Elapinae was established, and that the Crotalinae group II venom PLA₂ genes came into existence before Elapinae and Crotalinae diverged. A phylogenetic analysis of their amino acid sequences confirms this.

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Hepcidin, a cysteine-rich antimicrobial peptide, is widespread in fish and shows multiple activities, including antimicrobial, antivirus, and antitumor. Here, a new four-cysteine hepcidin isoform gene, EC-hepcidin3, was cloned from the marine-cultured orange-spotted grouper (Epinephelus coioides). The complete cDNA sequence consisted of 603 bases with an open reading frame (ORF) of 270 bases. The genomic DNA sequence was composed of two introns and three exons, and its 312-bp upstream region had multiple putative transcription factor binding sites. Soluble recombinant protein EC-proHep3 containing a His-tag at the C-terminus was obtained from expression plasmid pET-28a/EC-proHep3 in Escherichia coli Rosetta. It was purified by immobilized metal affinity chromatography (IMAC), and it showed antibacterial activity in vitro. Kinetic studies indicated that recombinant EC-proHep3 has strong, rapid activity against Staphylococcus aureus and Pseudomonas stutzeri. The results indicate that EC-hepcidin3 might be an effective component in the innate immune system of groupers.

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The facultative photosynthetic bacterium Rhodobacter sphaeroides 2.4.1 has a nitric oxide-response transcriptional regulator, NnrR, and nitric oxide reductase (NOR), although it is incapable of denitrification. To investigate at the genomic level the physiological response to nitrosative stress of R. sphaeroides, the transcriptome profiles of strain 2.4.1 and its NnrR mutant were analyzed before and after exposure to nitrosating agents, S-nitrosoglutathione (GSNO) and sodium nitroprusside (SNP), under microaerobic conditions. GSNO and SNP affected the expression of different but overlapping sets of genes. Only a limited number of these genes, including the genes for NOR, were under the control of NnrR, and those genes were significantly upregulated by GSNO and by SNP. The oxygen-responsive regulator FnrL and a predicted iron-sensing regulator were perhaps also involved in the transcriptome response to reactive nitrogen species. Some genes, including hemN for heme biosynthesis, were subject to dual regulation by NnrR and FnrL.

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To our knowledge, there is no report that directly shows an inhibitory effect of ginsenoside on melanin synthesis in B16 melanoma cells. Hence, we investigated whether the aglycone of Rh₄ (A-Rh4) inhibits melanin synthesis in B16 melanoma cells, and determined the mechanism of melanin inhibition. We isolated 12 ginsenoside compounds from leaves of Panax ginseng and tested them in B16 melanoma cells. It significantly reduced melanin content and tyrosinase activity under alpha-melanocyte stimulating hormone- and forskolin-stimulated conditions. It significantly reduced the cyclic AMP (cAMP) level in B16 melanoma cells, and this might be responsible for the regulation down of MITF and tyrosinase. Phosphorylation of a downstream molecule, a cAMP response-element binding protein, was significantly decreased according to Western blotting and immunofluorescence assay. These data suggest that A-Rh4 has an anti-melanogenic effect via the protein kinase A pathway.

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There are controversial reports on the relationship between helminthic infection and allergic diseases. Although IgE cross-reactivity between nematode Ascaris antigens and house dust-mite allergens in allergic patients have been reported, whether Ascaris or the mite is the primary sensitizer remains unknown. Here we found that immunization of naïve animals with Ascaris lumbricoides (Al) antigens induced production of antibodies cross-reactive to mite antigens from Dermatophagoides farinae (Df). Sera from Bangladeshi children showed IgE reactivity to Ascaris and mite extracts. IgG from rabbits immunized with Al extract exhibited reactivity to Df antigens. Treatment of the anti-Al antibody with Df antigen-coupled beads eliminated the reactivity to Df antigens. In immunoblot analysis, an approximately 100-kDa Df band was the most reactive to anti-Al IgG. The present study is the first step towards the establishment of animal models to study the relationship between Ascaris infection and mite-induced allergic diseases.

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A bacteriocin-producing (11,000 AU mL⁻¹) strain was isolated from the rhizosphere of healthy Algerian plants Ononis angustissima Lam., and identified as Brevibacillus brevis strain GM100. The bacteriocin, called Bac-GM100, was purified to homogeneity from the culture supernatant, and, based on MALDI-TOF/MS analysis, was a monomer protein with a molecular mass of 4375.66 Da. The 21 N-terminal residues of Bac-GM100 displayed 65% homology with thurincin H from Bacillus thuringiensis. Bac-GM100 was extremely heat-stable (20 min at 120 °C), and was stable within a pH range of 3–10. It proved sensitive to various proteases, which demonstrated its protein nature. It was also found to display a bactericidal mode of action against gram-negative (Salmonella enteric ATCC 43972, Pseudomonas aeruginosa ATCC 49189, and Agrobacterium tumefaciens C58) and gram-positive (Enterococcus faecalis ENSAIA 631 and Staphylococcus aureus ATCC 6538) bacteria, and a fungistatic mode of action against the pathogenic fungus Candida tropicalis R2 CIP 203.

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The thallus, the gametophyte body of the liverwort Marchantia polymorpha, develops clonal progenies called gemmae that are useful in the isolation and propagation of isogenic plants. Developmental timing is critical to Agrobacterium-mediated transformation, and high transformation efficiency has been achieved only with sporelings. Here we report an Agrobacterium-mediated transformation system for M. polymorpha using regenerating thalli. Thallus regeneration was induced by cutting the mature thallus across the apical-basal axis and incubating the basal portion of the thallus for 3 d. Regenerating thalli were infected with Agrobacterium carrying binary vector that contained a selection marker, the hygromycin phosphotransferase gene, and hygromycin-resistant transformants were obtained with an efficiency of over 60%. Southern blot analysis verified random integration of 1 to 4 copies of the T-DNA into the M. polymorpha genome. This Agrobacterium-mediated transformation system for M. polymorpha should provide opportunities to perform genetic transformation without preparing spores and to generate a sufficient number of transformants with isogenic background.

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Fibroblast growth factor 4 (FGF4) is considered a crucial gene in the development of mammalian embryos. Here we identified common amino acid sequences predicted from coding exons of the FGF4 gene in five pigs of two breeds, and HispFGF4, a 6× histidine-tagged porcine FGF4, was produced in Escherichia coli. HispFGF4 was purified efficiently from the supernatant of cell lysate by heparin column chromatography. In a porcine embryonic fibroblast cell line, HispFGF4 showed significant mitogenic activities at concentrations as low as 0.001 nM (p<0.01). To the best of our knowledge, this is the first report describing the complete nucleotide sequence of coding exons for the porcine FGF4 protein in two breeds, together with the production of a recombinant, bioactive porcine FGF4 derivative.

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Molecular diversity within six viroid species and different molecular variants, in each species infecting fruit trees was first estimated by the single-strand conformation polymorphism (SSCP) technique and then by direct sequencing analysis. The different variants studied are to three Australian grapevine viroids(AGVd), four citrus dwarfing viroids (CDVd), eleven grapevine yellow speckle viroids type-1 (GYSVd-1), four hop stunt viroids (HSVd), seven peach latent mosaic viroids (PLMVd), and eight pear blister canker viroids (PBCVd). Polyacrylamide gel electrophoresis (PAGE) conditions were compared and optimized to improve the sensitivity of the existing SSCP parameters. The relationships among the various SSCP profiles observed and the variation in nucleotide sequences was studied. The results indicate that the variations of some parameters of electrophoresis for each species allowed higher resolution and hence detection of single nucleotide variations among clones initially clustered into the same group.

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Cellobiose 2-epimerase (CE), found mainly in anaerobes, reversibly converts D-glucose residues at the reducing end of β-1,4-linked oligosaccharides to D-mannose residues. In this study, we characterized CE-like proteins from various aerobes (Flavobacterium johnsoniae NBRC 14942, Pedobacter heparinus NBRC 12017, Dyadobacter fermentans ATCC 700827, Herpetosiphon aurantiacus ATCC 23779, Saccharophagus degradans ATCC 43961, Spirosoma linguale ATCC 33905, and Teredinibacter turnerae ATCC 39867), because aerobes, more easily cultured on a large scale than anaerobes, are applicable in industrial processes. The recombinant CE-like proteins produced in Escherichia coli catalyzed epimerization at the C2 position of cellobiose, lactose, epilactose, and β-1,4-mannobiose, whereas N-acetyl-D-glucosamine, N-acetyl-D-mannosamine, D-glucose, and D-mannose were inert as substrates. All the CEs, except for P. heparinus CE, the optimum pH of which was 6.3, showed highest activity at weakly alkaline pH. CEs from D. fermentans, H. aurantiacus, and S. linguale showed higher optimum temperatures and thermostability than the other enzymes analyzed. The enzymes from D. fermentans, S. linguale, and T. turnerae showed significantly high k_cat and K_m values towards cellobiose and lactose. Especially, T. turnerae CE showed a very high k_cat value towards lactose, an attractive property for the industrial production of epilactose, which is carried out at high substrate concentrations.

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Four genes of Takifugu rubripes, tentatively designated Tr1–Tr4, encoding homologs of pufferfish saxitoxin- and tetrodotoxin-binding protein, were identified by BLAST search and 3′-RACE. RT-PCR and MALDI-TOF mass spectrometry allowed the identification and discrimination of Tr isoforms from the non-toxically cultured specimens. The expression of Tr1 and Tr3 mRNAs exclusively in the liver and the presence of their products as 120-kDa plasma proteins were confirmed.

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The putative α-L-rhamnosidase gene from Streptomyces avermitilis was cloned and expressed. The recombinant enzyme released L-rhamnose from p-nitrophenyl α-L-rhamnoside, Citrus flavonoids such as naringin, rutin, and hesperidin, and gum arabic which is an arabinogalactan-protein. Calcium ions increased L-rhamnose production by the enzyme from gum arabic, whereas enzyme activity was not affected by any metal ions.

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This study describes increases in extraction efficiency and the bioconversion of catechins after treatment with several commercial enzymes. Tannase was also used to improve the anti-radical activities of green tea extracts. Enzymatic treatment with various commercial enzymes was introduced to improve the extraction efficiency of polyphenols. The total polyphenol, flavonoid, and catechin contents and the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity of the green tea extract treated with Viscozyme (VG) were significantly higher than those treated with other commercial enzymatic extractions (p<0.05). More than 95% of the epigallocatechingallate (EGCG) and of the epicatechingallate (ECG) was hydrolyzed to epigallocatechin (EGC) and to epicatechin (EC) in successive 20 min treatments with Viscozyme and tannase (TG). Due to its hydrolytic activity, treatment involving tannase resulted in a significant release of gallic acid (GA), EGC, and EC, leading to greater radical scavenging activities. Regarding the IC₅₀ values of the DPPH and 2,2-azino-di-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radicals, the green tea extract treated with TG showed values of 131.23 and 28.83 µg/mL, VG showed values of 224.70 and 32.54 µg/mL, and normal green tea extract (NG) showed values of 241.11 and 66.27 µg/mL, respectively. These results indicate that successive treatment with Viscozyme and tannase improves the extraction efficiency of polyphenols and increases radical scavenging activities.

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We incubated 11 strains of Staphylococcus aureus in a brain heart infusion broth at 10–37 °C with two inoculum sizes and examined their enterotoxin A (SEA) production by a Western blot analysis to clarify the effect of incubation temperature on SEA production. Although SEA was detected in the exponential phase at 15–37 °C, it was also detected in the stationary or death phase at 10 °C. The maximal SEA concentrations of most strains increased as the temperature was increased, although some strains produced as much at 15 °C and 20 °C as they did at 37 °C. The maximal SEA concentration was definitely lowest at 10 °C, and as the temperature was increased, the production rate increased. However, a relationship between the production rates at the two different temperatures was not apparent. Some strains produced more SEA at 10–20 °C with a smaller inoculum size than with a larger one. SEA production therefore did not necessarily depend on the incubation temperature, and it would be difficult to predict at 10 °C and 15 °C from the production at 37 °C.

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This study investigated the effects of dietary supplementation with burdock powder and Aspergillus awamori-fermented burdock powder at 5% on the intestinal luminal environment and body fat in rats fed a high-fat (HF) diet. Food intake and growth were unaffected by dietary manipulation. Consumption of the burdock and fermented burdock diets significantly elevated fecal IgA and mucins (indices of intestinal immune and barrier functions) and reduced fecal lithocholic acid (a risk factor for colon cancer) (p<0.05). The fermented burdock diet markedly elevated cecal Bifidobacterium and organic acids, including lactate, acetate, propionate, and butyrate, and reduced fecal deoxycholic acid (a risk factor for colon cancer) and perirenal adipose tissue weight (p<0.05), but the burdock diet did not. These results suggest that consumption of fermented burdock improves the intestinal luminal environment and suppresses obesity in rats fed a HF diet.

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Arg-specific gingipain (Rgp) is a major pathogenic determinant of Porphyromonas gingivalis which is a major pathogen in periodontal disease. We prepared protein extracts with Rgp-inhibitory activity from polished rice (Oryza sativa) and evaluated the effects of these extracts on the growth and pathogenicity of P. gingivalis. The extracts inhibited the proteolytic degradation of human proteins by P. gingivalis proteinases, and repressed the growth and homotypic biofilm formation of P. gingivalis. The disruption of adhesion of epithelial cells by P. gingivalis was also restricted by the rice protein extracts. Our results suggested that the rice protein extracts suppressed the pathogenicity and growth of P. gingivalis by inhibiting the bacterial proteinase activities, implying that the Rgp-inhibitory proteins prepared from rice may be potentially valuable as nutraceutical agents for preventing periodontal diseases.

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The influence of different extraction solutions on the proteins extracted from rice grains was investigated. The largest amounts of salt-soluble proteins were extracted with solutions supplemented with Tris–HCl at pH 8.0. Rice allergens were analyzed by multiplex immunodetection. Except for α-globulin extracted with the solutions at pH 8.0, which showed a low-molecular-weight band besides the main band, no significant solution-dependent difference among the allergens was found. Total proteins were extracted with four kinds of solution. The extraction of the basic subunit of glutelin was found to be SDS-dependent, and more protein was obtained with extraction solutions supplemented with SDS. The contents of α-globulin and α-amylase/trypsin inhibitors were higher in the extracts without SDS than with SDS. We conclude from the present data that, in order to obtain comparable data from rice grain salt-soluble and total protein analyses, differences in the protein extraction efficiency of solutions used should be taken into consideration.

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CD36 is an integral membrane protein that mediates the cellular uptake of oxidized low-density lipoprotein (oxLDL) through recognition of the oxidized glycerophospholipids (oxPLs) formed during LDL oxidation. We aimed to devise an assay system to detect binding between CD36 and oxLDL/oxPL without using recombinant proteins. A peptide corresponding to amino-acid residues 149–168 of mouse CD36 with biotin at its N-terminus (named biotin-CD36_149–168) and variants of it were synthesized and immobilized onto streptavidin-coated plates. oxLDL labeled with Alexa-Fluor-488 bound specifically and saturably to immobilized biotin-CD36_149–168, but poorly or not at all to the variants, such as that with a scrambled amino-acid sequence. The binding of fluorescence-labeled oxLDL to biotin-CD36_149–168 was inhibited efficiently by an oxPL species, but not by a nonoxidized glycerophospholipid. This assay system using biotin-CD36_149–168 provides a convenient means not only of characterizing binding profiles between CD36 and oxLDL/oxPL but also of finding competitors for the binding.

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The effects of 16 aliphatic aldehydes with 3–10 carbons on the growth and patulin production of Penicillium expansum were examined. When P. expansum spores were inoculated into apple juice broth, some alkenals, including 2-propenal, (E)-2-butenal, (E)-2-pentenal, and (E)-2-hexenal, inhibited fungal growth and patulin production. Their minimal inhibitory concentrations were 5, 50, 80, and 80 µg/mL respectively. Vital staining indicated that these alkenals killed mycelia within 4 h. Treatment of the spores with these aldehydes also resulted in rapid loss of germination ability, within 0.5–2 d. On the other hand, aliphatic aldehydes with 8–10 carbons significantly enhanced patulin production without affecting fungal growth: 300 µg/mL of octanal and 100 µg/mL of (E)-2-octenal increased the patulin concentrations in the culture broth by as much as 8.6- and 7.8-fold as compared to that of the control culture respectively. The expression of the genes involved in patulin biosynthesis in P. expansum was investigated in mycelia cultured in apple juice broth containing 300 µg/mL of octanal for 3.5, 5, and 7 d. Transcription of the msas gene, encoding 6-methylsalicylic acid synthase, which catalyzed the first step in the patulin biosynthetic pathway was remarkably high in the 3.5-d and 5-d-old cultures as compared with the control. However, octanal did not any increase the transcription of the msas in the 7-d-old culture or that of the other two genes, IDH and the peab1, in culture. Thus the enhanced patulin accumulation with supplementation with these aldehydes is attributable to the increased amount of the msas transcript.

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The aim of this study was to investigate the effects of the essence of chicken on brain function by near-infrared spectroscopy. Twelve healthy elderly subjects took the essence of chicken or a placebo for 7 d in a double-blind cross-over design study. Changes in oxy-hemoglobin concentrations in the bilateral prefrontal areas of the brain were measured while the subjects performed the simple reaction task, the Groton Maze Learning Test, and the working memory task. In the latter case, there were significant interactions in the changes in oxy-hemoglobin concentrations between treatment and period of intake according to two-way repeated ANOVA. The changes in oxy-hemoglobin concentrations significantly increased in several regions of the prefrontal areas of the brain in those taking essence of chicken for 7 d. These results suggest that essence of chicken is useful as a nutritional supplement to enhance or maintain brain function in the elderly.

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We have previously reported that phytoceramide and phytosphingosine (PHS) stimulated the transcriptional activity of peroxisome proliferator-activated receptor γ (PPARγ) in cells. PPARγ is a therapeutic target for type 2 diabetes. We found in this study that an oral administration of PHS improved diet-induced glucose intolerance in mice. Since PHS is highly expressed in yeast, PHS in fermented foods may improve diabetes.

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We investigated the effects of equol on adipogenesis by measuring lipid accumulation and analyzing the change in adipocyte-related gene expression in MC3T3-L1 cells. Treatment with 10 µM equol tended to increase adipocyte-related gene expression, whereas 100 µM equol reduced lipid accumulation and suppressed the expression of these genes and proteins. Our results suggest that equol regulated adipogenesis in a bi-phasic fashion.

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Statistical experimental designs were applied to optimize cold-active lipase production by the psychrophilic bacterium Moritella sp. 2-5-10-1. First, a Plackett–Burmen design (PBD) was used to evaluate the significant effects of various fermentation parameters. The results indicated that soybean meal, temperature, and Tween-80 had significant influences on lipase production. The levels of these variables were optimized subsequently using central composite design (CCD). A quadratic regression model of cold-active lipase production was built, and verification experiments confirmed its validity. On subsequent scale-up in a 10-L bioreactor using optimized conditions, cold-active lipase production (30.56 U/mL) was obtained. The results clearly indicated that the model was adequate even on a large scale. To our knowledge, this is the first report of statistical optimization of cold-active lipase production by a psychrophilic bacterium.

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This study investigated temporal variations in the potential maximum quantum yield of photosystem II (F_v/F_m ratio) and growth-phase dependent cellular protein expressions of Chattonella antiqua under laboratory conditions. Despite the culture conditions, significant positive correlations between the F_v/F_m ratio and daily growth rate were observed. Threshold F_v/F_m ratios associated with positive cell growth were calculated to be >0.44, >0.44, and >0.37, and those associated with active cell growth (growth rate >0.5 div. d⁻¹) were >0.58, >0.60, and >0.49 under control culture, low nutrient and intense light conditions, respectively. Proteome profiles obtained by two-dimensional gel electrophoresis (2-DE) indicated that 42 protein spots were differentially expressed at various growth phases of C. antiqua, which indicates changes in cellular physiological status throughout the growth cycle, and suggests that oxygen evolving enhancer 1 and 2-cysteine peroxiredoxin play roles in maintaining the positive growth of C. antiqua.

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The objective of this study was to develop an efficient production system for cellulase preparation with a high level of xylanolytic enzymes using soluble carbon sources. When xylose and arabinose were simultaneously fed with glucose and cellobiose, a mutant of Trichoderma reesei, M3-1, showed sufficient levels of cellulolytic and xylanolytic activities, indicating that xylose and arabinose are good inducers for the production of xylanolytic enzymes. In a continuous feeding experiment using glucose/cellobiose and glucose/xylose/cellobiose, cellulase preparations with various levels of xylanolytic enzymes were obtained by altering the feeding solutions and the timing of their addition. The volumetric production rates for xylanolytic activities at the glucose/xylose/cellobiose-feeding phase were significantly higher than at the glucose/cellobiose-feeding phase, while those for cellulolytic activities were comparable under the two conditions. Thus the composition of the enzyme preparation produced by the mutant was readily controlled by varying the inducers and the pattern of their addition, facilitating the tailored production of enzymes in a diversity of bioconversion processes.

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Pseudomonas aeruginosa has two nitric oxide (NO)-detoxification enzymes, NO reductase and flavohemoglobin, which catalyze the reduction and oxygenation of NO respectively. In this study, the NO reductase-deficient mutant showed decreased virulence against the silkworm Bombyx mori, but the flavohemoglobin-deficient mutant did not, indicating that NO-reduction is important to the full virulence of P. aeruginosa against B. mori.

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