Many species of female moths produce sex pheromones to attract conspecific males. Most moth species utilize Type I pheromones that consist of straight-chain compounds 10–18 carbons in length with a functional group of a primary alcohol, aldehyde, or acetate ester, and usually with several double bonds. Studies over the past three decades have demonstrated that female moths usually produce sex pheromones as multi-component blends in which the ratio of the individual components is precisely controlled, making it possible to generate species-specific pheromone blends. As for the biosynthesis of Type I pheromones, it is well established that they are de novo synthesized in the pheromone gland (PG) through modifications of fatty acid biosynthetic pathways. However, because many of the molecular components within the PG cells (enzymes, proteins, and small regulatory molecules) had not been functionally characterized, the molecular mechanisms underlying sex pheromone production in PG cells remained poorly understood. To address this, we have characterized some of the key molecules involved in the biosynthesis of the sex pheromone bombykol in the silkmoth, Bombyx mori. Characterization of these molecules has facilitated our understanding of the precise mechanisms underlying lepidopteran sex pheromone production.

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The small intestine is an organ responsible for nutrient absorption, barrier functions, signal recognition/transduction, and the production of bioactive compounds. These functions are known to be regulated by such factors as hormones and cytokines, but substances contained in the daily diet are also thought to play roles as major modulators of intestinal functions. Intestinal epithelial cells (IECs), which form a monolayer covering the inside surface of the intestinal tract, are particularly important in this modulation, because they directly interact with intestinal contents, including food substances, their digests, and gut microbial components. Using cell-based in vitro assays, we investigated the food-IEC interactions at the cellular and molecular levels, and found that a variety of food substances affected the transporter activity, tight junction permeability, metabolic enzyme expression, immune functions, and so on. Modulation of the intestinal functions by dietary substances is therefore essential to promote health.

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Unlike many classical signals and hormones, exposure of the cells to electrophilic molecules potentially induces a series of characteristic and wide-ranging biological responses by covalently attaching with macromolecules such as proteins as well as small cellular reductants. In addition to chemicals originated from xenobiotics or lipid peroxidation, electrophiles in foods have recently attracted much attention. These compounds have recently been found to induce expression of cytoprotective proteins that are involved in the elimination or inactivation of oxidative stress and carcinogenic electrophiles implicated in several pathogeneses. The redox-sensitive regulating systems such as Keap1/Nrf2/ARE play a key role in this induction and thus are considered to be the most important target of electrophiles in foods. This review highlights the food-derived electrophiles as promising protectors against various diseases, with an emphasis on possible molecular mechanisms. Current knowledge of isothiocyanates (ITCs), representative electrophile compounds from cruciferous vegetables, is discussed also, with consideration of the chemistry, metabolism, absorption, and factors influencing the biological activities of ITCs. In addition, this review attempts to provide a balanced perspective on the relative beneficial and harmful effects of the food electrophiles.

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C-Reactive protein (CRP) is an acute-phase protein that increases during systemic inflammation and is currently one of the most frequently studied inflammatory markers in epidemiology. We have determined CRP concentration using novel latex reagent with polyclonal antibody. In the present study, we determined the concentration of CRP using monoclonal antibodies, and evaluated the interaction of antigen-antibody reactive sites and latex agglutination to detect low CRP concentrations. We developed four novel monoclonal antibodies that we classified into two major groups, and that were used to prepare the latex reagents. The latex reagents prepared using a cocktail of monoclonal antibodies for different epitopes appeared highly sensitive. The lower limit of CRP detection, which was defined using the mean 3 SD method, was calculated to be 5 ng/ml for the latex reagents when oligoclonal antibodies were utilized. Furthermore, the latex reagents were found to react specifically with CRP in clinical samples.

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An improved analytical method using reverse-phase high performance liquid chromatography following anthranilic acid derivatization for the measurement of each oligosaccharide level in transition (5 to 10 d lactation) and mature (21 to 155 d lactation) milks of sixteen Samoan women is described. The method was applied for the measurement of sialyl as well as neutral oligosaccharide levels. We found that disialyllacto-N-tetraose (DSLNT), sialylacto-N-tetraose c (LSTc), and 6′-sialyllactose (6′-SL) were the most abundant of the sialyl oligosaccharides. In the neutral oligosaccharide fraction, lacto-N-fucopentaose II and III combined (LNFP II+III) were the most dominant, followed by lacto-N-tetraose (LNT) and 3-fucosyllactose (3-FL). 2′-Fucosyllactose (2′-FL) and lacto-N-fucopentaose I (LNFP I) were absent in some and found at low levels in most of the Samoan women. Our study indicates that the milk oligosaccharide composition in Samoan women is similar to that of Japanese women with respect to sialyl but not to neutral oligosaccharides. The differences in neutral oligosaccharides might have a genetic origin.

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In Leonurus sibiricus herb extract (LHE)-supplemented animals, plasma cholesterol decreased and high-density lipoprotein-cholesterol increased, resulting in a lowered atherogenic index. The plasma trolox equivalent antioxidant capacity, levels of hepatic thiobarbituric acid-reactive substances, and protein carbonyl values decreased significantly in LHE-supplemented mice (p<0.05), whereas the hepatic antioxidant indicators were all significantly elevated (p<0.05). In human umbilical vein endothelial cells stimulated with tumor necrosis factor alpha, LHE significantly suppressed intracellular reactive oxygen species, LOX-1, and adhesion molecules. LHE supplementation may modulate the lipoprotein composition and attenuate oxidative stress by elevated antioxidant processes, thus suppressing the activation of inflammatory mediators. This is a possible mechanism of the anti-atherogenic effect.

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Scorpion venoms are composed of a number of peptides, many of which show neurotoxicity. In addition to these neurotoxins, several antimicrobial peptides have also been isolated from the venoms. The scorpion Isometrus maculatus, belonging to the Buthidae family, is found in many tropical regions including Japan, but little attention has been paid to its biological activity and chemical composition. In this study, we isolated a novel insect toxin, Im-1, by bioassay-guided fractionation of the venom of I. maculatus. Rapid and reversible paralysis was observed after injection of Im-1 into crickets. Im-1 consists of 56 amino acids, and is predicted to form an amphipathic α-helix. Since Im-1 shares sequence similarity to an antimicrobial peptide, parabutoporin, we evaluated its effects on several bacterial strains and found that it showed an antimicrobial activity profile similar to parabutoporin. This suggests that Im-1 and parabutoporin exert their antimicrobial effects through similar mechanisms.

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Two highly hydroxylated 1,3-dihydroisobenzofurans, FR198248 (1) and FR202306 (2), were isolated as peptide deformylase (PDF) inhibitors from Aspergillus flavipes. Compounds 1 and 2 inhibited Staphylococus aureus PDF with IC₅₀ values of 3.6 and 2.5 μM, respectively, and also showed antibacterial activity with an MIC value of 25 μg/ml. In contrast, 6-O-methyl derivative 3 of compound 2 was inactive against both PDF and S. aureus.

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The pathway of leucine to mevalonate, which has attracted attention in the study of the biosynthesis of isoprenoid in parasitic protozoa and myxobacterium, was observed in the biosynthesis of the lipid core in halophilic archaea. The involvement of isovaleryl-CoA dehydrogenase was strongly suggested, with stereospecific conversion of the diastereotopic methyl group of leucine to isoprenoidal lipid.

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Two α-N-acetylgalactosaminidases, α-N-acetylgalactosaminidase (α-GalNAcase) I and II, were purified from the digestive organ of starfish. Purified α-GalNAcase I and II gave nearly single protein bands on SDS-polyacrylamide gel electrophoresis, individually. Even the final preparation of α-GalNAcase I contained α-galactosidase activity, while α-GalNAcase II was almost free from that activity with p-nitrophenyl and 4-methylumbelliferyl α-N-acetylgalactosaminides as substrates. α-GalNAcase I and II both hydrolyzed terminal α-N-acetylgalactosaminyl linkages of the natural compounds investigated: Forssman hapten glycolipid, blood group A active oligosaccharide and GalNAc-α1-O-serine. On the other hand, oligosaccharides, and glycolipid containing α-galactosyl terminals were hydrolyzed by α-GalNAcase I but not by α-GalNAcase II. The substrate specificities and other enzymatic properties of α-GalNAcase I were similar to those of human placental α-GalNAcase, but distinct from α-GalNAcase II.

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We examined the functional equivalency between Escherichia coli RNase P protein (C5) and Pyrococcus horikoshii RNase P proteins (PhoPop5, PhoRpp21, PhoRpp29, PhoRpp30, and PhoRpp38) for RNase P activity. The C5 protein and P. horikoshii RNase P proteins were unable to activate non-congnate RNase P RNAs, P. horikoshii RNase P RNA (PhopRNA) and E. coli RNase P RNA (M1 RNA) respectively. Two chimeric RNAs, in which functional C- and S-domains of M1 RNA and PhopRNA were exchanged, were prepared and characterized with respect to the cleavage of P. horikoshii pre-tRNA^Tyr in the presence of C5 or P. horikoshii proteins. The results suggest that PhoPop5 and PhoRpp30 function equivalently to the C5 protein in the E. coli RNase P, being involved in activation of the PhopRNA C-domain. On the other hand, PhoRpp21 and PhoRpp29 are implicated in stabilization of the PhopRNA S-domain.

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Serum transferrin is an iron-binding glycoprotein with a bilobal structure. It binds iron ions in the blood serum and delivers them into target cells via transferrin receptor. We identified structural and functional characteristics of recombinant human transferrin which is produced in the yeast Pichia pastoris. Using the signal sequence of the α factor of the yeast Saccharomyces cerevisiae, high-level secretion was obtained, up to 30 mg/l of culture medium. Correct processing at designed sites was confirmed by N-terminal sequence analysis. Carbohydrate modification was determined by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) analysis after digestion with endo-β-N-acetylglucosaminidase H. Reflecting the secondary structure, the circular dichroism spectrum of the recombinant protein was indistinguishable from that of serum transferrin. Consequently, the recombinant product had an iron binding function just as the serum specimen has: two Fe³⁺ sites existed in a recombinant transferrin molecule, as estimated by titration analysis using visible absorption, fluorescence spectra, and electrophoretic behavior in urea denaturing polyacrylamide gel electrophoresis (PAGE).

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Ceramidase (CDase) catalyzes the hydrolysis of ceramide (Cer) to sphingosine (Sph) and fatty acid. We have reported the molecular cloning and preliminary characterization of the Mycobacterium CDase (MtCDase) (J. Biol. Chem., 274, 36616–36622 (1999)). To determine its function further, MtCDase was expressed in Escherichia coli and purified by Ni-Sepharose and gelfiltration. The purified recombinant enzyme showed a single band and a molecular weight estimated to be 71 kDa on SDS–PAGE. It had a pH optimum at 8.0–9.0 and quite broad specificity for various Cers. Of the Cers of different fatty acid moieties tested, those composed of C6–C24 fatty acids were well hydrolyzed, and Cers with mono unsaturated fatty acids were much more hydrolyzed than those with saturated fatty acids. Using N-dodecanoyl-7-nitrobenz-2-oxa-1,3-4-diazole (NBD)-D-erythro-sphingosine (C12-NBD-Cer) as substrates, the reaction followed normal Michaelis-Menten kinetics. The apparent Km and Vmax values for C12-NBD-Cer were 98.7 μM and 21.1 pmol/min respectively. The purified enzyme also catalyzed the synthesis of Cer in vitro, using NBD-labeled dodecanoic acid and Sph as substrates.

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Artemisia sacrorum Ledeb. (Compositae) (ASL) is a traditional Chinese medicine used to treat different hepatic diseases. However, a hypolipidemic effect of ASL on fatty liver disease has not been reported. Therefore, we investigated whether 95% ethanol eluate (EE), an active part of ASL, would attenuate hepatic lipid accumulation in human HepG2 cells by activating AMP-activated protein kinase (AMPK). Significant decreases in triglyceride levels and increases in AMPK and acetyl-CoA carboxylase (ACC) phosphorylation were observed when the cells were treated with 95% EE. EE down-regulated the lipogenesis gene expression of sterol regulatory element-binding protein 1c (SREBP1c) and its target genes, such as fatty acid synthase (FAS) and stearoyl-CoA desaturase 1 (SCD1), in a time- and dose-dependent manner. In contrast, the lipolytic gene expression of peroxisome proliferator-activated receptor α (PPAR-α) and CD36 increased in a time- and dose-dependent manner. These effects were abolished by pretreatment with compound C, an AMPK inhibitor. However, there were no differences in the gene expression of SREBP2, low density lipoprotein receptor (LDLR), hydroxymethyl glutaryl CoA reductase (HMG-CoA), or glucose transporter 2 (GLUT2). At the same time, 95% EE significantly increased the gene expression of acyl CoA oxidase (ACOX) in a time- and dose-dependent manner. Thus, AMPK mediated 95% EE induced suppression of SREBP1c and activation of PPAR-α respectively. These finding indicate that 95% EE attenuates hepatic lipid accumulation through AMPK activation and may be active in the prevention of serious diseases such as fatty liver, obesity, and type-2 diabetic mellitus.

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Replication protein A (RPA) binds to single-stranded DNA generated during DNA replication and other processes. The roles of RPA in telomere maintenance have been demonstrated in yeasts, but not in telomerase-positive human cells. In this study, we found that expression of mutant RPA70 in human cells caused telomere shortening, suggesting that RPA is required for telomere-length regulation in human cancer cells.

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We have found that coupling between catharanthine and vindoline occurs non-enzymatically in the presence of flavin mononucleotide and manganese ions with near-ultraviolet light irradiation in vitro. The present study found that the concentrations of catharanthine and vindoline in Catharanthus roseus decreased and those of dimeric indole alkaloids increased under near-ultraviolet light at 4 °C. It indicates that this coupling reaction at 4 °C occurs non-enzymatically.

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The activated structure of RNase P RNA (PhopRNA) in Pyrococcus horikoshii OT3 was characterized by circular dichroism (CD) and ultraviolet (UV) absorbance spectra. The results suggested that interaction of four RNase P proteins (PhoPop5, PhoRpp21, PhoRpp29, and PhoRpp30) with PhopRNA results in destabilization of base stacking in PhopRNA, whereas the addition of a fifth protein, PhoRpp38, increases base stacking in PhopRNA.

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A bow with horse tail hair is used to play the violin. New and worn-out bow hairs were observed by atomic force microscopy. The cuticles of the new bow hair were already damaged by bleach and delipidation, however the worn-out bow hairs were much more damaged and broken off by force, which relates to wearing out.

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The mechanisms of eukaryotic cell-cycle regulation are closely linked to cellular tumorigenesis. Compounds that affect the cell cycle are good candidates for developing anti-tumor drugs. We developed a screening method for cell-cycle blockers using a Saccharomyces cerevisiae cdc2-1 rad9Δ strain that can detect the activity of substances by cell growth. We performed screening on culture broth of various microbes, and identified five compounds, borrelidin, mycophenolic acid, UCS15A, copiamycin analog, and fredericamycin A, that were known to possess anti-tumor activity. These results indicate that this screening method is effective as a first-screening system for anti-tumor agents.

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L-Amino acid α-ligase (Lal), catalyzing the formation of α-dipeptides from unprotected L-amino acids in an ATP-dependent manner, is used in cost-effective fermentative production of dipeptides. We searched for novel Lals by in silico screening using Hidden Markov Model-based profile analysis, and identified five novel Lals that showed low similarity and different substrate specificity from known Lals.

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Cellulase hyperproducing mutants derived from the fungus Trichoderma reesei QM9414 were analyzed. They exhibited higher filter-paper degrading activity and a lower growth rate than the wild-type QM9414 strain. Transcription of the cellobiohydrolase I and endoglucanase I genes in the mutants was also greater than that of QM9414, suggesting that cellulase hyperproduction by these mutants was regulated at the transcriptional level.

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A rice ubiquitin-related modifier-1 (Rurm1) gene was cloned and transformed in Escherichia coli. We successfully expressed the RURM1 protein as a glutathione S-transferase (GST)-fusion protein by cultivating the E. coli cells at 16 °C for 16 h. After cleavage of GST, we obtained a single protein of 12 kDa. This protein was identified as the RURM1 protein by western blot analysis.

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An endo-acting family 3 pectate lyase (PL I) gene from Streptomyces thermocarboxydus was cloned and expressed in Escherichia coli. The deduced PL I sequence had highest similarity to a putative pectate lyase of S. coelicolor. Recombinant PL I had significantly lower specific activity, probably a result of incorrect folding, as indicated by circular dichroism spectra analysis.

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We analyzed changes in gene expression in Caco-2 cells cocultured with THP-1 cells over time periods of 0, 1, 3, 6, 24, and 48 h using a DNA microarray. Differentially expressed genes extracted with maSigPro indicated that an early defense response and cell death occured at the same time, causing an inflammatory condition.

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We explored the mechanism of the stabilization of Moloney murine leukaemia virus reverse transcriptase (MMLV RT) by eliminating RNase H activity. Without the template-primer (T/P) poly(rA)-p(dT)₁₅, the temperature reducing initial reverse-transcription activity by 50% over a 10-min incubation of the RNase H activity-deficient variant D524A was higher by 3.7 °C than that of the wild-type enzyme (WT). In the reverse transcription reaction, the K_m values for T/P of WT and D524A were almost the same. These results suggest that elimination of RNase H activity enhanced the intrinsic thermal stability of MMLV RT rather than its affinity toward T/P.

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Cookies containing iron, defatted rice bran, and several oils were prepared, and their oxidative stability evaluated. Oxidation was suppressed by the defatted rice bran, but a limit to the suppressive effect was observed. The maximum peroxide values obtained with defatted rice bran were low, and similar, regardless of the degree of unsaturation of the oils. A chemical analysis suggested that proteins and polyphenols in the defatted rice bran contributed to the suppressive effect. Cookies without the defatted rice bran showed decreasing maximum peroxide value as the relative humidity increased. No such dependence was observed for the cookies containing defatted rice bran. The water sorption isotherm of defatted rice bran indicated that the weak dependence was due to low water sorption.

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CH-19 Sweet is a newly found chili pepper breed bearing much less pungent fruits. Because CH-19 Sweet fruits were found to contain three analogs (capsinoids) of capsaicin, a major component of pungency of hot peppers (the analogs are capsiate or CST, dihydrocapsiate or DCT, and nordihydrocapsiate or NDCT), we assessed in this study the bio-potencies of these three capsinoids by comparing them with capsaicin. The three capsinoids bound to transient potential vanilloid 1 (TRPV1) receptors expressed in cultured cells and activated Ca²⁺ influx in a concentration-dependent manner with similar magnitudes. In contrast to capsaicin, capsinoids at the same concentration induced virtually no nociceptive responses when applied to the eyes or the oral cavities of mice. Intravenous administration of capsaicin or 20-fold increased doses of each capsinoid to rats induced significant increases in plasma catecholamine levels. Orally administered, each capsinoid enhanced oxygen consumption in mice. Based on the present results, capsaicin and these three capsinoids should have similar bio-potency, though capsinoids do not generate pungency or sensory irritation.

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Schiandra chinensis is a well-known Chinese traditional medicine for the treatment of hepatic disease. In this study, we investigated whether the nine major compounds of Schiandra chinensis could be applied to suppress lipopolysaccharide (LPS)-induced inflammatory responses in murine macrophages (Raw 264.7 cells). Among the nine lignans, three, gomisin J, gomisin N, and schisandrin C, were found to reduce nitric oxide (NO) production from LPS-stimulated Raw 264.7 cells. These three lignans showed low cytotoxic effects in Raw 264.7 cells. Pre-treatment of Raw 264.7 cells with gomisin J, gomisin N, or schisandrin C reduced the expression of mRNA and the secretion of pro-inflammatory cytokines. These inhibitory effects were found to be caused by blockage of p38 mitogen-activated protein kinase (MAPK), extracellular signal–regulated kinases 1 and 2 (ERK 1/2), and c-Jun N-terminal kinase (JNK) phosphorylation.

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High-fat diets (HFDs) supplemented with the L-Trp and L-Leu-OEt derivatives of the monascus pigment were fed to mice. Compared to the HFD group, the average body weight gain of the four HFD-pigment groups was decreased by 13.6–50.9%, and the intra-peritoneal adipose tissues weight was reduced by 16.7–30.5%. The derivatives also reduced the respective serum total cholesterol and triglyceride levels of the mice by 9.7–14.4% and 12.5–17.2%. The L-Trp derivative greatly increased the HDL-cholesterol level by 64.8–66.4% and the HTR value by 73.8–81.7%. The L-Leu-OEt and L-Trp derivatives decreased the total cholesterol level in the mice liver by 9.7–24.2% and 36.2–39.9%, respectively. Reductions in the triglyceride level were 21.5–22.4% for the L-Leu-OEt derivative and 17.9–18.8% for the L-Trp derivative. The L-Leu-OEt derivative exhibited higher in vitro inhibitory activities against HMG-CoA reductase and lipoprotein lipase than the L-Trp derivative. An in vivo test with mice showed the L-Trp derivative to have higher anti-obesity effects than the L-Leu-OEt derivative.

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We investigated the anti-allergic effect of a new strain (Pediococcus pentosaceus Sn26, the Sn26 strain) among 59 strains isolated from Japanese fermented vegetable pickles, the Sunki pickle. The Sn26 strain increased Th1 type cytokine (IL-12 and IFN-γ) production of Peyer’s patch (PP) cells in BALB/c mice, improved the Th1/Th2 balance, and inhibited IgE production of splenocytes of ovalbumin (OVA)-induced allergic diarrheic mice. Next we demonstrated, by neutralizing IL-12 and IFN-γ, that the Sn26 strain first induced IL-12, that IL-12 induced IFN-γ, and that decreases in IL-4 and IgE production followed. Furthermore, oral administration of the Sn26 strain decreased serum OVA-specific IgE levels and ameliorated the appearance of diarrhea in OVA-induced allergic diarrheic mice. Based on these results, it was assumed that oral administration of the Sn26 strain ameliorated type-1 allergies through improvement of the Th1/Th2 balance and decreases in IgE production.

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The aim of the present study was to analyze the antibiotic susceptibility of bifidobacterial strains distributed in the Japanese market. A total of 23 strains, including probiotic isolates from foods, supplements, pharmaceuticals and reference strains of each species (or subspecies), were tested for susceptibility to 15 antibiotics by the broth microdilution method and examined for the presence of possible resistant determinants. The strains were susceptible overall to chloramphenicol, ampicillin, vancomycin and linezolid, and were intrinsically resistant to aminoglycoside group agents. Susceptibility to erythromycin, clindamycin, rifampicin, tetracycline and trimethoprim varied among the strains. All strains of Bifidobacterium animalis subsp. lactis were resistant to tetracycline and appeared to harbor tet(W) genes. No risk factor for safety was found for bifidobacterial strains distributed in the Japanese market in respect of their antimicrobial resistance, although the presence of the tet(W) gene in some strains stresses the need for future evaluation.

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Fucoidan exhibits various biological properties. We raised a novel antibody against fucoidan extracted from Cladosiphon okamuranus and developed a sandwich ELISA method to measure fucoidan. The fucoidan antibody was specific and did not cross-react with other polysulfated polysaccharides. Fucoidan recovery from serum and urine by ELISA was 86–113%. Intra- and inter-assay CVs were 1.5–13.4%. Assay linearity was maintained after 3-fold dilution of each sample with phosphate-buffer saline (PBS). In the serum and urine of healthy volunteers (n=10), fucoidan was not detected before administration, and the levels markedly increased 6 and 9 h after oral administration. The molecular weight of the serum fucoidan determined by HPLC gel filtration remained unchanged, whereas that of urine fucoidan was significantly reduced. This is the first ELISA method of measuring serum and urine fucoidan levels after oral administration. The method is simple, reliable, and practical for the analysis of samples, especially urine samples.

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This study aimed to determine whether oral antibiotic administration during infancy is associated with the spontaneous development of atopic dermatitis-like skin lesions by modulating intestinal microbiota. Female NC/Nga mice at 3 weeks of age were orally administered kanamycin or polymyxin B. Clinical symptoms, scratching behavior, and serum antibody levels were evaluated. Changes in intestinal microbiota were determined by culture-independent analysis and cultural analysis. The kanamycin-treated mice showed higher clinical scores and scratching frequency than the control mice. IgE levels were significantly higher in the kanamycin-treated mice than in the control mice. Transient changes in intestinal microbiota were observed under kanamycin treatment. Polymyxin B treatment failed to affect scratching behavior. These results suggest that oral administration of kanamycin during infancy promoted the development of atopic dermatitis-like skin lesions in NC/Nga mice and was associated with a transient change in intestinal microbiota.

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This study examined the effects of the inulin-type fructans known as fructo-oligosaccharide (FOS) and inulin on the growth and persistence of Lactobacillus plantarum No. 14 (LP14) in the gut. Cultivation in medium that contained either FOS or inulin revealed that both fructans supported the growth of LP14. Fecal recovery of viable L. plantarum at 24 and 30 h following administration of LP14 were highest in mice fed inulin, followed by mice fed FOS and the control diet. However, neither of the fructans prolonged the appearance of LP14 in the gut. Cultivation in fecal extract medium prepared from mice fed each of the test diets showed higher growth of LP14 in the mice fed fructans. Histological analysis following fluorescein-stained LP14 administration showed that LP14 was largely localized to the luminal contents. These results suggest that inulin-type fructans support the growth of LP14 in the luminal contents only during their passage through the gut.

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To determine the effects of kaempferol, rat pheochromocytoma cells (PC12) and Institute of Cancer Research (ICR) mice were utilized as neuronal models. Using in vitro assays, kaempferol was shown to have protective effects against oxidative stress-induced cytotoxicity in PC12 cells. Administration of kaempferol also significantly reversed amyloid beta peptide (Aβ)-induced impaired performance in a Y-maze test. Taken altogether, the results reported here suggest that further investigation is warranted of the influence of kaempferol on pathways related to Alzheimer’s disease.

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The effects of two types of Welsh onion, green-leafy and white-sheath types, on hyperlipidemia in rats fed on diets high in fat and sucrose were studied. A significant lowering effect on cholesterol in the plasma and on total lipids, triacylglycerol, and cholesterol in the liver was observed in rats fed on the green, but not white, Welsh onion. These Welsh onions had no significant effect on the hepatic enzyme activities responsible for fatty acid synthesis and degradation. The results suggest that the green Welsh onion might be effective in attenuating hyperlipidemia in a manner other than affecting fatty acid metabolism in the liver. Kaempferol seemed to be one of the components in green Welsh onion acting to lower lipid deposition.

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Kucha (Camellia assamica var. kucha) is an endemic tea that grows in cloudy and foggy highlands in southwest China. Its hypoglycemic effects were studied here. The postprandial blood glucose levels in mice after sucrose and soluble starch loading were significantly reduced by Kucha administration. The glycosidase inhibitory activities were effective both in vitro and in vivo. It is concluded that the hypoglycemic effects of Kucha might be due to inhibition of disaccharidases activities.

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Lactophorin is a heat-stable phosphoglycoprotein, also known as milk glycosylation-dependent cell adhesion molecule 1 (GlyCAM-1). Bovine 18 kDa lactophorin was purified by heparin affinity chromatography from cow’s milk whey. Its N-glycans were obtained by proteomic techniques, including two-dimensional polyacrylamide gel electrophoresis (2D-PAGE), followed by in-gel digestion with peptide-N⁴-(N-acetyl-β-glucosaminyl)-asparagine amidase (PNGase F). The released N-glycans were derivatized with 2-aminopryridine (PA) and analyzed by matrix-assisted laser desorption ionization quadruple ion trap time of flight mass spectrometry (MALDI-QIT-TOF MS). Among the MS analyzed peaks, 15 peaks were found to be N-glycan molecules as detected by MS² analysis. These glycans consisted of mono-sialylated bi-, tri-, and tetra-antennary complex-type N-glycans carrying Gal-GlcNAc (LacNAc) or GalNAc-GlcNAc (LacdiNAc) with and without core-fucose.

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pCAR1 endows carbazole-degrading ability on its host. Three different host Pseudomonas (P. fluorescens Pf0-1, P. resinovorans CA10dm4, and P. chlororaphis IAM1511) were prepared to compare their behaviors in artificial microcosms (sterile soil and water supplemented with carbazole) with previous results for P. putida SM1443 (Shintani et al., Appl. Microbiol. Biotechnol., 80, 485–497 (2008)). We monitored carbazole degradation, bacterial survival, and conjugative transfer of pCAR1. Carbazole degradations were not detected with any of three hosts in the soil microcosms, but were in most of the water microcosms. Degradative efficiency varied host by host. SM1443 is an appropriate host for carbazole degradation in soil microcosms, while CA10dm4 is for water microcosms. pCAR1 transfers were detected only in the water microcosms inoculated with SM1443 or IAM1511 as the host. This is the first report to compare the behaviors of various hosts of the same plasmid in these microcosms.

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In infection cultures of Spodoptera frugiperda (Sf-9) insect cells with a recombinant baculovirus, vhpR, carrying human preprorenin cDNA in the polyhedrin locus of Autographa californica multiple nuclear polyhedrosis virus (AcMNPV), the expressed inactive recombinant human (rh)-prorenin is reported to be proteolytically processed to yield active rh-renin in the very late phase of culture (Takahashi et al., Biosci. Biotechnol. Biochem., 71, 2610–2613 (2007)). To identify the enzyme that catalyzes the processing of rh-prorenin, referred to as prorenin processing enzyme (PPE), we purified potential PPE from virus-infected Sf-9 culture supernatant by the use of an internally quenched fluorescent (IQF) substrate for PPE. The 32-kDa protein band agreed well with PPE activity on the final Mono Q FPLC. By N-terminal amino acid sequence analysis, the protein was revealed to be a cysteine protease encoded by the AcMNPV gene. Enzyme activity was inhibited by cysteine protease inhibitors but not by other protease inhibitors. When the purified rh-prorenin was incubated with the 32-kDa protein, renin activity appeared concomitant with the disappearance of rh-prorenin. The N-terminal amino acid sequence of the activated product was identical to that of the rh-renin that had accumulated in the infection cultures. These results indicate that the 32-kDa cysteine protease derived from the AcMNPV gene is the enzyme PPE of virus-infected Sf-9 cells.

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Extracellular polysaccharide (EPS) production by Lactobacillus confusus in liquid and solid state fermentation was carried out using coconut water and sugarcane juice as renewable wastes. High concentrations of EPS of 62 (sugarcane juice) and 18 g/l of coconut water were produced in solid state fermentation when nitrogen sources were reduced 5-fold from the original medium.

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The effects of Cl⁻ on the antibiotics productivity of Pseudomonas sp. YGJ3 were investigated. YGJ3 produced pyoluteorin and 2,4-diacetylphloroglucinol, depending on the concentration of Cl⁻ in the culture medium. Cl⁻ stimulated the biosynthesis of pyoluteorin, thereby repressing 2,4-diacetylphloroglucinol biosynthesis. The cell-free extract from the bacteria grown without Cl⁻ showed high activity of monoacetylphloroglucinol acetyltransferase, an essential enzyme in 2,4-diacetylphloroglucinol biosynthesis.

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