The Golgi apparatus of the eukaryotic cell is an essential organelle at the center of the network of vesicular transport delivering proteins and lipids to the correct locations in the cell. There are several Golgi compartments that have distinct resident proteins and functions, but the mechanism creating and maintaining the differences has long been an unsolved mystery in cell biology. After the discovery and molecular characterization of the transport vesicles and their coat proteins, we realized that the Golgi is an extremely dynamic organelle existing as repeating cycles of appearance, maturation, and disappearance. In this review, we describe essential findings as to the Golgi apparatus uncovered by work on an excellent model microorganism, the yeast Saccharomyces cerevisiae, with special reference to the results of our studies.

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This review summarizes our research into the synthesis and structure-activity relationships of epolactaene, neoechinulin A, plakevulin A, pseudodeflectusin and ustusorane C. These natural products are attractive in view of their apoptosis-inducing activity, cytoprotective activity against peroxynitrite, inhibitory activity against DNA polymerases, and cytotoxicity in cancer cells.

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Human mature milk and colostrum contain 12–13 g/L and 22–24 g/L of milk oligosaccharides respectively, and the structures of least 115 human milk oligosaccharides (HMOs) have been characterized to date. By way of comparison, bovine colostrum collected immediately post partum contains only around 1 g/L of oligosaccharides, and this concentration rapidly decreases after 48 h. It was recently recognized that HMOs have several biological functions, and this study area has become very active, as illustrated by a recent symposium, but it appears that advances in studies on the milk oligosaccharides of domestic farm animals, including cows, have been rather slow compared with those on HMOs. Nevertheless, studies on bovine milk oligosaccharides (BMOs) have progressed recently, especially in regard to structural characterization, with the development of methods termed glycomics. This review is concerned with recent progress in studies on the milk oligosaccharides of domestic farm animals, especially of BMOs and bovine glycoproteins, and it discusses the possibility of industrial utilization in the near future.

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We developed and validated a new high-performance liquid chromatographic analysis for electrochemically detecting guaiacol and vanillin as important components in vanilla extract. Separation was achieved with Capcell Pak C-18 MG, the potential of the working electrode being set at +1000 mV. The respective calibration curves for guaiacol and vanillin were linear in the range of 1.60–460 µg/L and 5.90–1180 µg/L. The respective limits for the quantities of guaiacol and vanillin were 1.60 µg/L and 2.36 µg/L. The related standard deviations of the intra- and inter-day precision of the retention time and peak area were all less than 4%. The recovery of guaiacol and vanillin was both more than 97%, all of the validation data being within an acceptable range. This analysis method is well suited for the simultaneous and convenient analysis of guaiacol and vanillin in a vanilla extract to evaluate the quality of the vanilla extract.

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Scutellariae radix (SR, from the roots of Scutellaria baicalensis Georgi) is thought to regulate blood pressure. In this study, HPLC-based purification coupled with MS, NMR analysis revealed that baicalin, a major flavone in SR, stimulates endothelial NO generation, suggesting its potential as an ingredient in medicinal food and beverage to treat hypertension.

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As part of our chemical screening program for new microbial secondary metabolites, we discovered a new compound, JBIR-107 (1), from the culture of Streptomyces tateyamensis NBRC 105047 isolated from a marine sponge sample. Extensive NMR and MS spectroscopic data enabled the structure of 1 to be determined as 5-acetamido-6-(4-(methyl(2-oxo-3-phenylpropyl)amino)phenyl)-4-oxohexanoic acid.

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The spirocyclic pyrrole alkaloid, acortatarin A, which had been isolated from a Chinese medicinal plant, was synthesized from a known olefinic compound by a concise six-step sequence involving N-alkylation of a pyrrole derivative with an α-bromo ketone intermediate as the key step.

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Abscisic acid-, stress-, and ripening-induced (ASR) protein play important roles in protecting plants from abiotic stress. The functions of some ASR proteins are known to be modulated by binding to metal ions. In this study, we demonstrated that the non-tagged full-length soybean (Glycine max) ASR protein (GmASR) can bind Fe³⁺, Ni²⁺, Cu²⁺, and Zn²⁺. The direct binding properties of GmASR to Fe³⁺ and Zn²⁺ were further confirmed by intrinsic fluorescence assays. The GmASR protein was found to have three Fe³⁺ binding sites but only two Zn²⁺ binding sites. Natively disordered in aqueous solution, GmASR remained disordered in the presence of Fe³⁺, but was found to aggregate in the presence of Zn²⁺. The aggregated GmASR protein was partially resolubilized after Zn²⁺ was chelated by EDTA. GmASR exhibited Fe³⁺-binding-dependent antioxidant activity in vitro. We speculate that GmASR thus protects against oxidation damage by buffering metal ions, thus alleviating metal toxicity in plant cells under stressed conditions.

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Kuromoji (Lindera umbellata) essential oil (KEO) has long been used in Japan as a traditional medicine. It contains linalool (C₁₀H₁₈O), a naturally occurring small terpenoid. For this study, we investigated the anti-inflammatory effect of KEO in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. Mouse macrophage-like RAW 264.7 cells were stimulated with LPS. Then they were treated with 25 or 50 µg/mL of KEO for 24 h. KEO suppressed LPS-induced pro-inflammatory cytokine production such as that of nitric oxide (NO), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) in a dose-dependent manner. In addition, inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2) mRNA expression and protein levels were suppressed by treatment with KEO cells. In addition, by treatment with 25 or 50 µg/mL of linalool showed the same anti-inflammatory effect. The results suggest that KEO and linalool can be regarded as a natural resource for use in anti-inflammatory therapeutic products.

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Because sucrose stored in mature stalks (in excess of 40% of stalk dry weight) can be wholly mobilized to supply carbon for the growth of heterotrophic tissues, we propose that sucrose mobilization requires a net sink-to-source transition that acts in toto within sett internode storage parenchyma. Based on our data we propose that mobilization of sucrose from culm storage parenchyma requires minimal investment of metabolic resources, and that the mechanism of sucrose mobilization is metabolically neutral. By magnetic resonance spectroscopy and phloem-specific tracer dyes, strong evidence was found that sucrose is mobilized from sett storage parenchyma via phloem to the growing shoot tissue. An analysis of the enzyme activities involved in sucrose metabolism and glycolysis suggested that sucrose synthase activity is downregulated due to the effects of sucrose mobilization. Overall, metabolism in storage parenchyma shifts from futile cycling to a more quiescent state during sucrose mobilization.

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Lysosomal hexosaminidases are glycosyl hydrolases that remove the terminal hexosamine residues of glycoconjugates. Though mammalian hexosaminidases are well characterized, the biochemical nature of these enzymes among invertebrates remains elusive. In this study, we purified two thermostable N-acetyl β-D-hexosaminidases (hex A and B) to homogeneity from soluble extracts of whole Unio animal tissue by a combination of chromatographic procedures. Purified hex A and hex B migrated as a single protein species on native PAGE and exhibited enzyme activity. However on SDS–PAGE, hex A dissociated into two subunits of molecular masses about 75 kDa and 30 kDa respectively, while hex B showed a molecular mass of 40 kDa. Hex A and B were recognized by the affinity purified mannose 6-phosphate receptor 46 on ligand blot analysis. This specific interaction was similar to what is known for the vertebrate receptors and lysosomal enzymes. The enzymes showed different K_M values with respect to the substrates p-nitrophenyl N-acetyl-β-D-glucosaminide and p-nitrophenyl N-acetyl-β-D-galactosaminide. The enzymes were thermally stable up to 80 °C and showed pH optima between 5.0 and 6.0. This is the first report on the purification of two forms of hexosaminidases from Unio.

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D-rhamnose (D-Rha) residue is a major component of lipopolysaccharides (LPSs) in strains of the phytopathogen Pseudomonas syringae pathovar glycinea. To investigate the effects of a deficiency in GDP-D-rhamnose biosynthetic genes on LPS structure and pathogenicity, we generated three mutants defective in D-Rha biosynthetic genes, encoding proteins GDP-D-mannose 4,6-dehydratase (GMD), GDP-4-keto-6-deoxy-D-mannose reductase (RMD), and a putative α-D-rhamnosyltransferase (WbpZ) in P. syringae pv. glycinea race 4. The Δgmd, Δrmd, and ΔwbpZ mutants had a reduced O-antigen polysaccharide consisting of D-Rha residues as compared with the wild type (WT). The swarming motility of the Δgmd, Δrmd, and ΔwbpZ mutant strains decreased and hydrophobicity and adhesion ability increased as compared with WT. Although the mutants had truncated O-antigen polysaccharides, and altered surface properties, they showed virulence to soybean, as WT did.

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Two wheat (Triticum aestivum) cultivars, freeze-tolerant Dongnongdongmai 1 and freeze-sensitive Jimai 22, were used in this study. They were grown under field conditions, the leaves were sprayed with 1 × 10⁻⁵ mol/L ABA and distilled water as control respectively at the three-true-leaf stage. Application of exogenous ABA increased the accumulation of carbohydrates in both cultivars, mainly in sucrose and fructose, except that the fructose content decreased in the leaves of Jimai 22. Exogenous ABA also enhanced the expression of key enzyme genes, especially in Dongnongdongmai 1 above 0 °C and in Jimai 22 below 0 °C. The carbohydrate contents and expression levels of the genes encoding key enzymes showed co-regulation in some, but not all of the pathways examined. Taken together, the higher sugar accumulation and the upregulation in expression of key enzymes in freeze-tolerant Dongnongdongmai 1 due to exogenous ABA suggest that active carbohydrate metabolism is mainly associated with the freeze tolerance of Dongnongdongmai 1.

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The glycosylphosphatidylinositol (GPI) attachment is a most important post-translational modification of proteins that plays essential roles in promoting the biochemical activities of eukaryotic cells. Described here is an analysis of the amino acid properties of mammalian GPI-anchored proteins (GPI-APs) and the development of an innovative method of detecting them. GPI-APs are characterized by two high-hydropathy regions: the signal peptide, located inside the Endoplasmic Reticulum (ER), and the GPI attachment signal, a sequence adjacent to the GPI-anchoring site (the ω-site). Especially in sequence analysis of known GPI-APs, there were some distinct aspects of the amino acid propensities around the ω-sites. Therefore, a method of detecting GPI-APs was developed based on hydropathy profiles and a position-specific scoring matrix (PSSM) calculated by position-specific amino acid propensities. First, sequences of GPI-APs and negative controls, determined by screening based on hydropathy and residue volume profiles, were aligned based on residue volume profiles in the C-terminal region, and the position-specific amino acid propensities of each group were calculated according to their alignment positions. Then, a PSSM was devised using the amino acid propensities of GPI-APs and negative controls, and discrimination scores were estimated for each dataset. Based on these scores at a threshold was fixed for each dataset. GPI-APs were detected with 81.1% sensitivity and a 0.818 success rate in an optimized calculation region determined by adjusting the window size of this region using a 5-fold dataset. The results indicate that a PSSM around the ω-site can effectively discriminate GPI-APs.

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The ascomycete Trichoderma reesei is known as one of the most prolific producers of plant biomass-degrading enzymes. While several mutant strains have been developed by mutagenesis to improve enzyme productivity for a variety of industrial applications, little is known about the mechanical basis of these improvements. A genomic sequence comparison of mutant and wild-type strains was undertaken to provide new insights in this regard. We identified a number of single-nucleotide polymorphisms (SNPs) after sequencing the genome of a hyper-cellulolytic T. reesei strain, PC-3-7, with a next-generation sequencer. Of these, the SNP detected in cre1, the carbon catabolite repressor gene, was found to be responsible for increased cellulase production. Further comparative genomic analysis enabled the identification of an SNP that correlated well with high cellulase production in a T. reesei mutant. These results provide a better understanding of the genetic changes induced by classical mutagenesis and how they correlate with desirable phenotypes in filamentous fungi.

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Transcript levels of the polyubiquitin gene McUBI1 had been reported to be constant during Crassulacean acid metabolism (CAM) induction in the facultative CAM plant, Mesembryanthemum crystallinum. Here, we report the sequences of the full-length cDNA of McUBI1 and its promoter, and validation of the McUBI1 promoter as an internal control driving constitutive expression in transient assays using the dual-luciferase system to investigate the regulation of CAM-related gene expression. The McUBI1 promoter drove strong, constitutive expression during CAM induction. We compared the activities of this promoter with those of the cauliflower mosaic virus (CaMV) 35S promoter in detached C3- and CAM-performing M. crystallinum and tobacco leaves. We confirmed stable expression of the genes controlled by the McUBI1 promoter with far less variability than under the CaMV 35S promoter in M. crystallinum, whereas both promoters worked well in tobacco. We found the McUBI1 promoter more suitable than the CaMV 35S promoter as an internal control for transient expression assays in M. crystallinum.

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Ile58 of hen egg-white lysozyme (HEL) is buried in the interior of the molecule and is considered to participate in sugar residue binding at subsite C through hydrophobic interaction. The contribution of Ile58 to lysozyme function and stability was investigated by replacement of Ile58 with less hydrophobic residues, Val (I58V) and Ala (I58A). Replacement of Ile58 with Ala decreased substrate binding ability to an N-acetylglucosamine trisaccharide, (GlcNAc)₃, and a GlcNAc polymer, chitin, whereas replacement with Val had little effect. Similar results were obtained as to enzymatic activity toward both the bacterial cell substrate and glycol chitin. Kinetic analysis by substrate (GlcNAc)₅ revealed that replacement of the Ile residue reduced the sugar residue affinity at subsite C and the rate constant of glycosidic bond cleavage. The rate constant of glycosidic cleavage for mutant I58A was about one-third of that for the wild-type. Guanidine hydrochloride unfolding experiments showed that mutants I58V and I58A were less stable than the wild-type, by 1.88 and 2.88 kcal/mol respectively. Moreover, the stability of the protein inserted at this position decreased linearly with decreasing hydrophobicity of the inserted residue. It appears that the hydrophobicity of Ile58 is an important factor in the efficient substrate binding, enzymatic reaction, and structural stability of HEL.

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In the present study, 27 mitochondrial genomes of diverse avian supra-orders were collected from the Genbank database and their genes were aligned separately. From the alignments, the conserved sequences were selected to design novel conserved primers for amplification and sequencing of the different mitochondrial genes. The reproducibility of these primers was tested in the amplification and sequencing of diverse avian supra-order mitochondrial genomes and was confirmed. This method helped in designing a new set of primers to accelerate both the amplification and the sequencing of bird mitogenomes. It also aids in building mitogenome markers in studying the genetic framework of endemic birds as a preliminary strategy for conservation management of them.

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Habu serum factor (HSF) and HSF-like protein (HLP) are fetuin family proteins isolated from Protobothrops flavoviridis (habu snake) serum with different physiological activities. A comparison of their cDNAs and intronic sequences revealed that nucleotide substitutions were primarily in protein-coding regions, and the substitution patterns indicated accelerated evolution of these proteins. Genomic DNA fragment analysis, including intron 1, revealed a 6.6-kb insertion homologous to the full-length mammalian LINE1 (L1) retrotransposable element (PfL1) only in the HLP gene. This segment retains an open reading frame (ORF) that encodes a reverse transcriptase (RT)-like protein (PfRT). We further found that a large number of homologous segments have dispersed in the habu snake genome, although we could not determine the enzymatic activities of their products. Moreover, an analysis of habu snake liver RNA indicated active transcription of the PfRT genes, suggesting that high levels of RT activity in this snake have driven the evolution of unique phenotypes of venom enzymes and serum inhibitors of them.

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Remarkable progress has been made in genome science during the past decade, but understanding of genomes of eukaryotes is far from complete. We have created DNA flexibility maps of the human, mouse, fruit fly, and nematode chromosomes. The maps revealed that all of these chromosomes have markedly flexible DNA regions (We named them SPIKEs). SPIKEs occur more frequently in the human chromosomes than in the mouse, fruit fly, and nematode chromosomes. Markedly rigid DNA regions (rSPIKEs) are also present in these chromosomes. The ratio of the number of SPIKEs to the total number of SPIKEs and rSPIKEs correlated positively with evolutionary stage among the organisms. Repetitive DNA sequences with flexible and rigid properties contribute to the formation of SPIKEs and rSPIKEs respectively. However, non-repetitive flexible and rigid sequences appear to play a major role in SPIKE and rSPIKE formation respectively. They might be involved in the genome-folding mechanism of eukaryotes.

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Ebp2 has an essential role in the biogenesis of 60S ribosomal subunits. Synthetic-sick alleles with the ebp2-14 mutation were screened. The mutations were localized to the ARD1 and NAT1 genes, which encode the catalytic subunit and the auxiliary subunit of N^α-acetyltransferase NatA respectively. Polysome analyses revealed that ard1Δ and nat1Δ caused a synergistic defect with ebp2-14 in the assembly of 60S ribosomal subunits. To identify the proteins that functionally interact with NatA, we designed mutants in which the second amino acid was substituted for proline in Ebp2 and functionally related proteins: Brx1, a partner of Ebp2 in ribosome biogenesis, and the ribosomal protein L36a/b, overexpression of which suppresses a growth defect in ebp2-14. Among these, only brx1-S2P exhibited a synthetic defect with ebp2-14. These results suggest that optimal NatA function is important to the cooperative function of Brx1 with Ebp2 in 60S ribosomal subunit biogenesis.

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The cytotoxic effects of various oxysterols on several culture cells were examined. Ozonolysis products of cholesterol, secosterols (3β-hydroxy-5-oxo-5,6-secocholestan-6-al) and its aldolization product (3β-hydroxy-5β-hydroxy-B-norcholestane-6β-carboxaldehyde) and their keto alcohol and acid derivatives, were found to have potent cytotoxic activities, as compared with major endogenous oxysterols such as 5β,6β-epoxycholesterol, 7β-hydroxycholesterol, 7-ketocholesterol, and 25-hydroxycholesterol. Secosterols might play important roles in tissue damage and inflammation-associated diseases.

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The mechanism underlying proteoglycan (PG) absorption in the intestine is not clear. Hence we analyzed the transport of salmon PG in the rat jejunum, ileum, and colon by the everted-sac method. The jejunum showed the largest capacity for PG transport. Jejunal transport of PG was also greater than that of chondroitin A and C. An inhibitor of clathrin-mediated endocytosis reduced jejunal PG transport. We conclude that intestinal PG transport is highest in the jejunum, and is partially dependent on clathrin-mediated endocytosis.

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We established a new method of transient expression using tomato fruit protoplasts. The method showed that L-ascorbic acid (AsA) content in tomato protoplasts was increased by transient expression of the L-galactose-1-phosphate phosphatase gene. This system provides a means of rapid analysis to clarify the function of AsA biosynthetic enzymes and AsA roles in tomato fruit.

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Hemolytic lectin CEL-III isolated from the sea cucumber Cucumaria echinata forms transmembrane pores by self-oligomerization in target cell membranes. It also formed soluble oligomers in aqueous solution upon binding with specific carbohydrates under conditions of high pH and a high salt concentration. The size of the soluble CEL-III oligomers decreased when treated with detergents such as Triton X-100 and SDS. Small-angle X-ray scattering measurements suggested that the dissociated unit of the oligomer was a tightly associated CEL-III heptamer. Without detergents in solution, these heptamers further assembled into larger 21mer oligomers, comprising three heptamers held together by relatively weak hydrophobic interactions.

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Soybean oil-in-water (O/W) emulsion-agar gel samples were prepared and their digestibility evaluated by using an in vitro gastrointestinal digestion model. Emulsion-agar sols were obtained by mixing the prepared O/W emulsions with a 1.5 wt % agar solution at 60 °C, and their subsequent cooling at 5 °C for 1 h formed emulsion-agar gels. Their gel strength values increased with increasing degree of polymerization of the emulsifiers, and the relative gel strength increased in the case of droplets with an average diameter smaller than 700 nm. Flocculation and coalescence of the released emulsion droplets depended strongly on the emulsifier type; however, the emulsifier type hardly affected the ζ-potential of emulsion droplets released from the emulsion-agar gels during in vitro digestion. The total FFA content released from each emulsion towards the end of the digestion period was nearly twice that released from the emulsion-agar gel, indicating that gelation of the O/W emulsion may have delayed lipid hydrolysis.

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This study investigated the anti-obesity effect of Adenophora triphylla root ethanol extract (ATREE). C57BL/6 mice were divided into five groups, a normal diet group (N), a control group fed only high-fat diet (HFD) (C), a positive control group fed HFD with 0.5% catechin (PC), and groups fed HFD with 0.5% (E1) or 1% (E2) ATREE. The body weight gain, hematological and serum biochemistry data, and anti-oxidative index in the liver and epididymal adipose tissue were improved significantly in the E group (E1, E2), as compared to the C group. As for histological findings, adipocyte size was reduced by ATREE. E group (E1, E2) showed significant increases in adiponectin, AMPK, and PPAR-α, and significant decreases in TNF-α, GPDH, and PPAR-γ, as compared to the C group. The above findings indicate that ATREE might have an anti-obesity effect through antioxidant and anti-inflammatory action. It is considered ATREE can be used as a natural treatment for obesity and metabolic syndrome induced by oxidative stress.

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Lactoferrin (LF) is a multifunctional glycoprotein found in mammalian milk. We have shown in a previous clinical study that enteric-coated bovine LF tablets decreased visceral fat accumulation. To address the underlying mechanism, we conducted in vitro studies and revealed the anti-adipogenic action of LF in pre-adipocytes. The aim of this study was to assess whether LF could increase the lipolytic activity in mature adipocytes. Pre-adipocytes were prepared from rat mesenteric fat and differentiated into mature adipocytes for assays of lipolysis. The addition of LF significantly increased the glycerol concentration in the medium in a dose-dependent manner, whereas pepsin-degraded LF did not. A DNA microarray analysis demonstrated that LF decreased the expression of perilipin and affected the cAMP pathway. These findings are supported by the results of quantitative RT-PCR of perilipin and assays of cAMP. These data collectively indicate that visceral fat reduction by LF may result from the promotion of lipolysis and the additional anti-adipogenic activity of LF.

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The effect of Lactobacillus pentosus strain S-PT84 (S-PT84) on postprandial hypertriacylglycerolemia was investigated in rats. S-PT84 dose-dependently inhibited the hydrolysis of triacylglycerols by pancreatic lipase in vitro. Intragastric administration of S-PT84 significantly reduced the lymphatic recovery of ³H-trioleoylglycerol up to 8 h. The oral administration of a fat emulsion, with or without S-PT84, resulted in the concentration of plasma triacylglycerol 2 h and 3 h after administration being significantly lower in the S-PT84 group than in the group without S-PT84 (control group). These results suggest that S-PT84 alleviated postprandial hypertriacylglycerolemia by delaying triacylglycerol absorption in the intestine through the inhibition of pancreatic lipase.

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The key odorants of Tahitian vanilla beans (Vanilla tahitensis) were characterized by a sensory evaluation, aroma extract dilution analysis (AEDA), quantification, and aroma reconstitution. Vanillin and anisaldehyde were identified in the same highest flavor dilution (FD) factor as the most characteristic odor-active compounds in Tahitian vanilla beans, followed by anisyl alcohol and anisyl acetate. Vanillin and anisyl alcohol were by far the most abundant odorants present with the highest concentration in the beans, followed by acetic acid, anisaldehyde, and anisyl acetate. A sensory evaluation of Tahitian vanilla beans and its reconstitute aroma concentrate characterized both samples as similar. These results indicated vanillin, anisaldehyde, anisyl alcohol, and anisyl acetate to be the key odorants in Tahitian vanilla beans. 3-Methylnonane-2,4-dione were identified for the first time in vanilla beans. β-Damascenone and phenylacetic acid were identified for the first time in Tahitian vanilla beans.

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The odor-active volatiles in Madagascar vanilla beans (Vanilla planiforia) of two grades, red whole beans as standard quality and cuts beans as substandard quality, were characterized by instrumental and sensory analyses. The higher contents of vanillin and β-damascenone in red whole beans than in cuts beans respectively contributed to significant differences in the sweet and dried fruit-like notes, while the higher contents of guaiacol and 3-phenylpropanoic acid in cuts beans than in red whole beans respectively contributed to significant differences in the phenolic and metallic notes. A sensory evaluation to compare red whole beans and their reconstituted aroma characterized both samples as being similar, while in respect of the phenolic note, the reconstituted aroma significantly differed from the reconstituted aroma with guaiacol added at the concentration ratio of vanillin and guaiacol in cuts beans. It is suggested from these results that the concentration ratio of vanillin and guaiacol could be used as an index for the quality of Madagascar vanilla beans.

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The defatted rice bran extracts obtained by being treated with various subcritical aqueous fluids at 230 °C for 5 min were further subjected to an acetone-solubilization treatment. The 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity depended on the total phenolic content of a sample, the acetone-soluble fraction of the extract obtained with 40% (v/v) aqueous acetone showing the highest activity. Based on the UV spectrometric and HPLC analyses, the substances with higher absorbance around 280 nm and higher hydrophobicity were found solubilizing out from the extracts into the acetone during fractionation. A gel permeation chromatographic analysis suggested that the extracts comprised monomeric or oligomeric substances. The antioxidative activity was evaluated by a DPPH radical scavenging activity analysis, rancidity test, and autoxidation analysis, revealing that it would be most likely to exert a radical scavenging effect more effectively during the initial stage of lipid oxidation than during the propagation stage.

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This study determined the influence of γ-aminobutyric acid (GABA) on brain protein synthesis via the mammalian target of rapamycin (mTOR) pathway. Experiments were carried out on three groups of 6-wk-old male rats with 0%, 0.5%, and 1% GABA. The percentage-phosphorylated S6K1 and growth hormone (GH) concentration was significantly increased by the GABA administration. The insulin level was not significantly changed, while the insulin-like growth factor 1 (IGF-1) level was significantly decreased by the GABA administration.

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We investigated the degradation of betaine in the rumen by a continuous supply of betaine to dairy cows. After 24 h of betaine incubation using rumen fluid from dairy cows, more than 80% of the betaine remained undegraded. Furthermore, the continuous supply of betaine for about 70 d did not influence the in vitro degradation of betaine.

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A gene encoding L-ribulose 3-epimerase (L-RE) from Mesorhizobium loti, an important enzyme for rare sugar production by the Izumoring strategy, was cloned and overexpressed. The enzyme showed highest activity toward L-ribulose (230 U/mg) among keto-pentoses and keto-hexoses. This is the first report on a ketose 3-epimerase showing highest activity toward keto-pentose. The optimum enzyme reaction conditions for L-RE were determined to be sodium phosphate buffer (pH 8.0) at 60 °C. The enzyme showed of higher maximum reaction a rate (416 U/mg) and catalytic efficiency (43 M⁻¹ min⁻¹) for L-ribulose than other known ketose 3-epimerases. It was able to produce L-xylulose efficiently from ribitol in two-step reactions. In the end, 7.2 g of L-xylulose was obtained from 20 g of ribitol via L-ribulose at a yield of 36%.

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The aim of the present study was to develop a strain-specific polymerase chain reaction (PCR) primer set for the detection of Bifidobacterium bifidum OLB6378 (OLB6378) that can serve as suitable probiotics for infants. The random amplified polymorphic DNA (RAPD)-PCR technique was used to obtain OLB6378-specific PCR products. One OLB6378-specific RAPD-PCR product was obtained after testing 97 RAPD primers, and was sequenced. Thirteen PCR primer sets were designed from the sequence. One PCR primer set was found to amplify one PCR product when genomic DNA of OLB6378 was used as template. The primer set did not amplify any PCR product when the other genomic DNA was used as template. The primer set was tested with 47 strains of B. bifidum and 20 strains of the other Bifidobacterium species. As a result, we developed an OLB6378-specific primer set, one that should be useful not only for the detection of OLB6378 but also for the quantification of OLB6378.

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An enzymatic production method for synthesizing L-alanyl-L-glutamine (Ala-Gln) from L-alanine methyl ester hydrochloride (AlaOMe) and L-glutamine (Gln) was developed in this study. The cultivation conditions for an Escherichia coli strain overexpressing α-amino acid ester acyltransferase from Sphingobacterium siyangensis AJ 2458 (SAET) and reaction conditions for Ala-Gln production were optimized. A high cell density culture broth prepared by fed-batch cultivation showed 440 units/mL of Ala-Gln-producing activity. In addition, an Ala-Gln-producing reaction using intact E. coli cells overexpressing SAET under optimum conditions was conducted. A total Ala-Gln yield of 69.7 g/L was produced in 40 min. The molar yield was 67% against both AlaOMe and Gln.

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Bacillus circulans KA-304 α-1,3-glucanase (Agl-KA) includes an N-terminal discoidin domain (DS1), a carbohydrate binding module family 6 (CB6), threonine and proline repeats (TPs), a second discoidin domain (DS2), an uncharacterized conserved domain (UCD), and a C-terminal catalytic domain. Domain deletion enzymes lacking DS1, CB6, and DS2 exhibited lower α-1,3-glucan-hydrolyzing and -binding activities than the wild type, Agl-KA. An α-1,3-glucan binding assay with fluorescent protein fusion proteins indicated that DS1, CB6, and DS2 bound to α-1,3-glucan and fungal cell walls, and that binding efficiency was increased by their combined action. In contrast, UCD did not exhibit any α-1,3-glucan-binding activity. A dramatic decrease in protoplast formation in the Schizophyllum commune mycelium was observed given only a DS1 deletion. An Agl-KA with deletion DS1, CB6, and DS2 produced no protoplasts. These results indicate that the combined actions of DS1, CB6, and DS2 contributed to increased cell-wall binding and were indispensable for efficient Agl-KA cell-wall degradation.

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A more rapid and simpler method than those currently used was developed to screen conjugated linoleic acid (CLA)-producing bacteria isolated from cow milk. The screening of 500 strains was completed in 10 d and the screening efficiency was 10%. One strain resembling a Lactobacillus paracasei strain and two resembling L. helveticus strains converted free linoleic acid to total CLA ≥85%.

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Algal biomass serves as a fuel for electricity generation in microbial fuel cells. This study constructed a model consortium comprised of an alga-digesting Lactobacillus and an iron-reducing Geobacter for electricity generation from photo-grown Clamydomonas cells. Total power-conversion efficiency (from Light to electricity) was estimated to be 0.47%.

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