The chain conformation of an arabinoxylan isolated from the rice endosperm cell wall, which was composed of the linear backbone of a 1, 4-linked β-xylose residue highly substituted at the C3 position with α-L-arabinofuranose, was studied by X-ray fiber diffraction and a conformational analysis. From the X-ray results the polysaccharide was suggested to take an extended left-handed, three-fold helical structure which is not a desirable conformation to make a complex firmly associated with cellulose or xyloglucan that was elucidated to be present in the cell wall. A conformational analysis, using the PFOS force field for the O-α-L-arabinofuranose-(1→3)-β-D-xylose residue and O-α-L-arabinofuranose-(1→3)-[(1→4)-β-D-xylose residue]₃, in which the arabinose residue was linked to the C3 position of the middle residue of the xylotriose residue, and using the PS79 computer program for the arabinoxylan indicated that the backbone xylan chain could not take a two-fold helix similar to that of cellulose because of significant steric interaction between the arabinose side chain and the backbone. Thus, the left-handed, three-fold helix was found to be reasonable.

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A chemiluminescence probe specific for the superoxide anion, 2-methyl-6-phenyl-3, 7-dihydroimidazo-[1, 2-a]pyrazine-3-one, was used to directly demonstrate the O·^^-__2 generation in rice blade protoplasts with the purified blast-fungus elicitor (proteoglucomannan). A hyperbolic relationship between the relative photon emission and the amount of elicitor applied, implied the presence of a putative receptor for the elicitor. Pretreatment of the elicitor with α-D-mannosidase completely crossed out the stimulative activity to generate O·^^-__2. The presence of mannostatins, inhibitors of α-D-mannosidase, also inhibited O·^^-__2 generation. Plasma membrane turbulence was observed by the quenching of fluorescence from a membrane turbulence reporter, bis-(3-propyl-5-oxo-isoxazol-4-yl) pentamethine oxonol, when the protoplasts were stimulated with the elicitor. Although treating rice protoplasts with IAA alone did not stimulate O·^^-__2 generaion, protoplasts pretreated with IAA markedly activated the O·^^-__2 generation capability when stimulated by the elicitor. The O·^^-__2 generation activated by stimulating the elicitor was abruptly blocked by the addition of IAA.

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Four semi-synthetic and fourteen quassinoids were tested for their antifeedant and insecticidal activity against 3rd instarlarvae of the diamondback moth (Plutella xylostella). In this quassinoid series, isobrucein-B was the most potent compound in both assays. Chemical conversion of the methoxy and/or methylenedioxy groups in the A and C rings to hydroxy groups among these quassinoids resulted in decreased activity.

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Lipophilicity and the tendency for decomposition of imidacloprid and related compounds by oxygen, hydrolytic mediums, and simulated sunlight were studied to see whether these physicochemical factors have any relation to the biological activity of the compounds in vitro, in a greenhouse, or under field conditions. Lipophilicity indices based on HPLC bore no definite relationship with the binding affinity to the acetylcholine receptor. However, the compounds having high insecticidal potential in greenhouse tests were generally less hydrophilic. In neutral water or in an oxygen-saturated solution, the compounds tested were completely stable. An evident difference was observed in their behavior toward the sunlight wavelength, the nitromethylene compounds decomposing far more rapidly than nitroimines like imidacloprid. The photolability of the nitromethylenes is ascribed to their longer maximum absorption wavelength of over 320 nm.

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We isolated an alcohol-fermenting yeast, Torulaspora delbrueckii No.3110, which was more salt-tolerant than Saccharomyces cerevisiae IFO 0224 as measured by alcohol fermentation. Strain No.3110 produced 5.5 wt% of ethanol in a medium containing 1 M NaCl in 4 days, but strain IFO 0224 produced a mere 0.8%. Strain No.3110 constitutively accumulated intracellular trehalose (10-17% of dry cell weight). A mutant, T1, did not accumulate trehalose. T1 synthesized and assimilated less trehalose and was less salt-tolerant than No.3110. These results suggest that metabolism of trehalose is important for the expression of salt tolerance.

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The alcohol-fermenting yeast Torulaspora delbrueckii No. 3110 was less tolerant to high temperature than Saccharomyces cerevisiae IFO 0224 as measured by alcohol fermentation during mild agitation : at 40°C, ethanol production of the two yeasts was 0.8 and 5.2 wt% respectively. The No.3110 cells had much unsaturated fatty acid (C18 : 2) and little ergosterol, which suggests that the low tolerance might be caused by high membrane fluidity. Two types of miconazole-resistant mutants were isolated and characterized. Strain M47 had less unsaturated fatty acid and was found to be more temperature tolerant than No.3110. Strain M59 was defective in ergosterol synthesis and was less temperature tolerant than No.3110. These results indicate the importance of membrane rigidity in temperature tolerance. M59 aaccumulated much less trehalose than No.3110 did. Addition of trehalose to the permeabilized cell system of M59 restored the temperature sensitivity, but not when the trehalase inhibitor deoxynojirimycin was also added, which suggests that the accumulation and metabolism of trehalose is important for the expression of temperature tolerance.

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The taste responses to five alkali metal salts of warfarin, aqueous rodenticides, in laboratory mice were observed at a short measurement time of 20 min by the two-bottle preference test using a licking time monitoring system newly developed. It was made clear that the order of taste avoidance to the warfarin salts was as follows : cesium salt > rubidium salt > lithium salt > sodium salt > potassium salt. The relationship between the hydrophobicity of the warfarin salts and the effectiveness of the taste avoidance was analyzed. The effectiveness of the taste avoidance largely depended on the hydrophobicity.

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The taste responses to the optical isomers of potassium warfarin, dipotassium glutamyl glutamate, and a mixture of both the substances in laboratory mice were studied using licking activity as an indicator of taste effectiveness. The offensive effectiveness of the optical isomers of potassium warfarin was the same. Every optical isomer of dipotassium glutamyl glutamate was avoided, the taste avoidance of dipotassium D-glutamyl D-glutamate being the strongest. The taste avoidance of potassium warfarin was greatly increased by the addition of every isomers of dipotassium glutamyl glutamate, and its cause was related to synergistic effects.

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3, 4-Dimethoxyphenol (3, 4-DMP) was converted into the corresponding glucoside in suspension-cultured cells of Coffea arabica. The maximum efficiency of glucosylation was attained, more than 40%, within 96 h after the addition of 1mM 3, 4-DMP when cultured in a modified Murashige and Skoog's medium with 5μM 2, 4-dichlorophenoxyacetic acid and 0.5μM kinetin. The glucoside was identified as 3, 4-dimethoxyphenyl β-D-glucopyranoside (3, 4-DMP glucoside) by ¹H-NMR and hydrolysis with α- and β-glucosidases. As the application of the glycosylation by C. arabica cells, eleven glycosides were obtained from seven methoxyphenols including 3, 4-DMP.

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The reaction conditions for γ-CD production by a purified CGTase from Brevibacterium sp. No. 9605 were investigated. The optimum pH and temperature for γ-CD formation were 7.0 and 50°C, respectively. The addition of calcium ion increased heat stability of the CGTase and the CDs formation was affected by the concentration of calcium ion. In the presence of ethanol, the yield of γ-CD from soluble starch was increased.

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Membrane clarification of green tea extract was studied as a treatment to reduce sediments in packaged drinks and as a pretreatment for concentration processes. The flux and variation of components were examined in dead-end and crossflow filtration with several types of membranes. In dead-end ultrafiltration, the flux reduction rate was small, although the initial flux was similar to the final flux in microfiltration. Prefiltration was effective in decreasing the reduction rate of flux. As the pore size of microfiltration membranes became smaller, the dry weight decreased gradually and the optical transmission at 660nm increased. By ultrafiltration, 30-50% pectin, 3-11% catechins and, 7-20% caffeine were rejected. Crossflow filtration was effective in keeping the flux high. The ultrafiltration spiral membrane (pore size : 0.008μm) was selected for repeated batch clarification of prefiltered green tea crude extract and showed reproducible performance.

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Eleven oligosaccharides formed by a transglycosylation reaction during lactose hydrolysis with Bacillus circulans β-galactosidase were purified by gel permeation chromatography, charcoal chromatography, and HPLC. From the results of methylation analysis, and MS and NMR studies, it was concluded that these oligosaccharides were β-D-Galp-(1→3)-D-Glc, β-D-Galp-(1→6)-D-Glc, β-D-Galp-(1→2)-D-Glc, β-D-Galp-(1→4)-β-D-Galp-(1→4)-D-Glc, β-D-Galp-(1→6)-[β-D-Galp-(1→2)]-D-Glc, β-D-Galp-(1→6)-[β-D-Galp-(1→4)]-D-Glc, β-D-Galp-(1→4)-β-D-Galp-(1→3)-D-Glc, β-D-Galp-(1→4)-β-D-Galp-(1→2)-D-Glc, β-D-Galp-(1→4)-[β-D-Galp-(1→2)]-D-Glc, β-D-Galp-(1→4)-β-D-Galp-(1→6)-D-Glc, β-D-Galp-(1→6)[β-D-Galp-(1→3)]-D-Glc. The last five are newly observed oligosaccharides. The results of a use test (in vitro) by human intestinal bacteria showed that the oligosaccharides containing lactose units were predominantly used by human intestinal bifidobacteria.

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Glucoamylase, acid protease, and acid carboxypeptidase in namazake were inactivated with micro-bubble supercritical carbon dioxide (SC CO₂) at 25 MPa, 35°C for 30 min. After the treatment, their activities decreased from 330, 20. 1, and 115 U/ml sake to 51. 8, 1. 8, and 0 U/ml sake, respectively. During the storage at 20°C for 60 days, no increase in amino acid value was observed. On the other hand, glucose concentration in the sake treated with SC CO₂ increased as well as namazake. The formation of lactic acid, acetic acid, and ethanol were depressed, and acidity was constant during storage. On the contrary, the decrease of pyruvic and malic acid after 20 days of storage could be depressed. Analysis of volatile compounds suggests that the treatment with SC CO₂ did not cause the formation of volatile compounds in contrast to heat-treatment. As a result of sensory evaluation, the sake treated with SC CO₂ at 20 MPa, 35°C for 30 min was given sensory scores near to those of namazake.

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The syntheses of (S)-13-hydroxy-(2E, 4E, 8E)-tetradecatrienoic acid (1) and (2E, 4E, 8Z-tetradecatrienoic acid (2) were carried out by using the Wittig reaction as the key step. The asymmetric center at C-13 and the double bond between C-8 and C-9 for natural compound 1 were reconfirmed as being of (S) configuration and E, respectively. The relationship between the structure of the unsaturated hydroxy fatty acids and their inhibitory effect on the growth of lettuce was investigated.

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Eight terpenes, including four new compounds, were isolated from Boletinus cavipes collected from two different districts (strains A and B) in Japan. Their structures were determined by spectroscopic and chemical methods, these compounds being geranylgeraniol (diterpene) or farnesol (sesquiterpene) derivatives. The geranylgeraniol derivatives were isolated from strain A, while the farnesol derivatives were isolated from strain B. These results indicate the presence of two chemotypes of the fungus B. cavipes.

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A large (80kbp) plasmid designated as pSLL was isolated from Streptomyces laurentii ATCC 31255 (strain P0). Another covalently closed circular plasmid, pSLS (16kbp), was found in pSLL-cured P0 cells (named P1), which spontaneously formed pocks on a solid medium. The frequency of the generation of pSLL-loss P1 cells coordinated positively with the frequency of pock appearance. pSLS was also found as an integrated form in the chromosome of strains P0 and P1. The results indicated that strain P0 harbored two plasmids, pSLL and integrated form pSLS, and pSLS seemed to be generated as a free-form plasmid with the loss of pSLL. It was also suggested that pSLL may suppress the excision of the integrated pSLS from the chromosome and that the free form of pSLS may participate in pock formation.

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Photothermal signals from some biological materials exposed to modulated CO₂ laser radiation were detected using the concept of photothermal interferometry. The magnitude and a phase of the signals were found to contain qualitative information about subsurface structural changes and variations of moisture content.

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Three different carbohydrate-depleted enzymes were prepared from an endo-β-1, 4-glucanase of Aspergillus niger IFO31125 by treatment with endo-β-N-acetylglucosaminidase or α-mannosidase. They were purified by Concanavalin A-Sepharose affinity and DEAE ion-exchange column chromatographies. The molecular sizes of these enzymes had been decreased from 40 kDa containing 9.0% carbohydrate to 39, 38, and 37 kDa with carbohydrate at 4.5, 1.3, and 0.8% (wt/wt), respectively. The native and these carbohydrate-depleted enzymes were compared in their enzymatic properties, and it was found that they were identical in their catalytic activities and both thermal and pH stabilities. However, the 37-kDa enzyme was more susceptible to proteolysis by Savinase, proteinase K, and Pronase E. On the other hand, the specific protease trypsin showed no such effect on activity of all enzymes. These results suggested that the core structure of the asparagine-linked sugar chain, which consisted of three monosaccharide residues, contributed to the high stability of the endo-β-1, 4-glucanase against protease digestion.

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The presence of copper and carbohydrate was demonstrated in an acidic ascorbate oxidase purified from Acremonium sp. HI-25. The enzyme was inhibited by cyanide, azide, and sulfide. The reaction product from L-ascorbic acid was identified to be dehydro-L-ascorbic acid. This enzyme did not oxidize other electron donors than ascorbate derivatives. Stoichiometric investigations on the oxidation reaction found that this enzyme belongs to the EC 1. 10. 3. 3 group, using molecular oxygen as an electron acceptor. The K_m for L-ascorbic acid was estimated to be 0.29 mM. The activation energy for inactivation of A cremonium ascorbate oxidase, 129kcal/mol, was four and three times higher than those of ascorbate oxidases from cucumber and pumpkin, respectively.

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Two arylsulfatase-producing streptomycetes that desulfated etoposide 4'-sulfate were isolated from soil samples. Taxonomical study identified one soil isolate as Streptomyces griseorubiginosus S980-14 (Es-1 arylsulfatase producer), while the other was considered new and tentatively designated Streptomyces sp. T109-3 (Es-2 arylsulfatase producer). Both strains produced extracellular arylsulfatase activities, provided that cultivation media were prepared with distilled water. Unlike the two known types of arylsulfatases, which had significant activity on p-nitrophenyl sulfate but none on etoposide 4'-sulfate, the crude streptomycete arylsulfatases efficiently desulfated etoposide 4'-sulfate and p-nitrophenyl sulfate, which supports the establishment of a new type of arylsulfatases.

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A new arylsulfatase designated Es-1, which desulfated etoposide 4'-sulfate and p-nitrophenyl sulfate, was isolated from Streptomyces griseorubiginosus S980-14 and purified to protein homogeneity by ammonium sulfate fractionation, ion exchange column chromatography, and chromatofocusing. The enzyme was active in monomeric form with an approximate molecular weight of 45, 000, had a pI value of 4.95, and required calcium for full activity. At an optimum reacion pH of 8.5, iodoacetate, mercurous chloride, and EDTA severely inhibited the activity of Es-1 arylsulfatase.

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Streptomyces sp. T109-3 arylsulfatase (Es-2), which desulfated p-nitrophenyl sulfate as well as etoposide 4'-sulfate, was purified to protein homogeneity by sulfated cellulose affinity and DEAE-cellulose column chromatographies. Es-2 required calcium for enzyme activity and was severely inhibited by SH and chelating reagents. Comparative characterization showed that, although distinct in recognition of the binding moiety of substrate, Es-1 (Streptomyces griseorubiginosus S980-14 arylsulfatase) and Es-2 shared high desulfating activity on etoposide 4'-sulfate and many other common enzymological characteristics, which suggested they would be acceptable as the enzyme component of antitumor antibody-enzyme conjugates for target chemotherapy.

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Chemical modifications of rye seed chitinase-c (RSC-c) with various reagents suggested the involvements of tryptophan and glutamic/aspartic acid residues in the activity. Of these, the modification of tryptophan residues with N-bromosuccinimide (NBS) was investigated in detail. In the NBS-oxidation at pH 4.0, two of the six tryptophan residues in RSC-c were rapidly oxidized and the chitinase activity was almost completely lost. On the other hand, in the NBS-oxidation at pH 5.9, only one tryptophan residue was oxidized and the activity was greatly reduced. Analyses of the oxidized tryptophan-containing peptides from the tryptic and chymotryptic digests of the modified RSC-c showed that two tryptophan residues oxidized at pH 4.0 are Trp72 and Trp82, and that oxidized at pH 5.9 is Trp72. The NBS-oxidation of Trp72 at pH 5.9 was protected by a tetramer of N-acetylglucosamine (NAG₄), a very slowly reactive substrate for RSC-c, and the activity was almost fully retained. In the presence of NAG₄, RSC-c exhibited an UV-difference spectrum with maxima at 284 nm and 293 nm, attributed to the red shift of the tryptophan residue, as well as a small trough around 300 nm probably due to an alteration of the environment of the tryptophan residue. From these results, it was suggested that Trp72 is exposed on the surface of the RSC-c molecule and involved in the binding to substrate.

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Four novel oligosaccharide chains were isolated from the glycoproteins of Fusarium sp. M7-1. Their chemical structures were resolved mainly by 400 MHz NMR spectrometry and mass spectrometry. The following structures were identified. Man α1→2Man-ol-6-P, Manα1→2Manα1→2Man-ol-6-P, Rhaα1→2Manα1→2Manα1→2Man-ol-6-P, and GlcNAcα1→4GlcAα1→2(GlcNAcα1→4)GlcAα1→2Galfβ1→6(Rhaα1→2)Man-ol.

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We have purified the p-nitrophenyl α-D-glucuronide-hydrolyzing enzyme (PNP-GAase) from snail (Helix pomatia) acetone powder by chromatographies on DEAE-cellulose, CM-Toyopearl, and Toyopearl HW-55F. The molecular weight of the enzyme was 180, 000 by gel filtration chromatography with Superose 6, and 97, 000 by SDS-polyacrylamide gel electrophoresis, and the pI was 6.8 by isoelectric focusing. The enzyme showed the maximum activity at pH 3.0 and 50°C, and was stable at pH between 3.0-7.0 and up to 50°C. The enzyme activity was greatly inhibited by Hg²⁺, p-chloromercuribenzoic acid, sodium dodecylsulfate, and N-bromosuccinimide. The enzyme released D-glucuronic acid not only from PNP-GA but also from 2-, 3-, and 4-O-α-D-glucuronosyl-D-xyloses, O-α-D-glucuronosyl α-D-glucuronide, and benzyl 4-O-α-D-glucuronosyl-β-D-glucoside. The results suggest that the α-glucuronidase from snail acetone powder had a broad substrate specificity comparing with α-glucuronidases from Aspergillus niger and basidiomycetes.

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A sialyl Lewis X ganglioside analogue containing 5-glycolylamido-3, 5-dideoxy-D-glycero-D-galacto-2-nonulopyranosylonic acid in place of N-acetylneuraminic acid was synthesized, clarifying the structural requirement for an N-acetyl group in the sialic acid moiety for selectin recognition.

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We had constructed an L-threonine-hyperproducing strain of E. coli K-12 by recombinant DNA techniques. In this paper, culture conditions for the practical production of L-threonine were investigated using this strain. Cultivation temperature, concentration of required amino acids, and dissolved oxygen greatly influenced the yield of L-threonine. High production of L-threonine was obtained at a high level of dissolved oxygen for the recombinant strain, but not for the parent. This improved production was accompanied by a high copy number of recombinant plasmids and high activity of aspartokinase. Initial addition of L-threonine together with required amino acids greatly reduced the net production of L-threonine. To remove the reductive effect, mothods for the addition of the required amino acids were tested. Lowering the required amino acids at a later stage of cultivation seemed to be effective to avoid the reductive effect of the accumulated L-threonine. By using the optimal conditions, the highest level of L-threonine production, 65g/l, 48% yield, was achieved.

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Two cosmid clones containing distinct types of self-defense gene of a 6-demethylchlortetracycline producer, Streptomyces aureofaciens NRRL3203, were isolated. The gene responsible for chlorination of tetracycline (chl gene) was subcloned from one of the cosmid clones by complementation of a chlorination-deficient mutant, using a gene cloning system for strain NRRL3203 developed in this study. The nucleotide sequence analysis of a 4.4-kb SacI-BamHI fragment containing the chl gene showed that the predicted product of the chl gene is a polypeptide of 452 amino acids, and that the chl gene was preceded by two open reading frames, which could endode polypeptides of 50 kDa and 32 kDa, respectively. A search for sequences homologous to these ORFs found that the former product strongly resembles that of the 6-hydroxylation enzyme for oxytetracycline biosynthesis, and that the latter product has a weak but significant similarity to the hydroxyindole O-methyltransferase of bovine pineal gland. By Northern blot analysis, these three genes were suggested to be polycistronically transcribed.

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Two kinds of proteinases, type-I and type-II, were purified or partially purified from salted muscle of anchovy, Engraulis japonica. Mol. wts. of type-I and type-II proteinases were estimated to 25, 000 and 37, 000, respectively, on electrophoretic analysis. Both proteinases strongly hydrolyzed synthetic tri or tetrapeptide substrates specific to trypsin, α-thrombin, and an activated protein C, while they hardly hydrolyzed Arg-MCA and benzoyl Arg-MCA derivatives. The proteinases were inhibited by common trypsin inhibitors. Optimal pH for the proteinase activities were pH 6.8 (type-I) and pH 7.0 to 7.5 (type-II), and the proteinases showed the highest activities at 45°C (type-I) and 50°C (type-II). The N-terminaI amino acid sequence of type-I proteinase, ¹I-²V-³G-⁴G··· (29 residues were identified), was significantly similar to sequences of trypsins and tryptases. Based on these findings, both proteinases were presumed to be kinds of tryptases in E. japonica muscle.

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In this study, the metabolism of phosphatidylcholine (PC) in bivalves and their feed plankton was investigated in the summer and winter. Lysophosphatidylcholine (lysoPC) and free fatty acid (FFA) appeared to be the major metabolites of PC in both bivalves and feed plankton. Production rates of both lysoPC and FFA relative to protein in plankton were much higher than those in bivalves, oysters, and scallops. Seasonal change in these metabolites in bivalves coincided with that in the plankton. In oysters, the activity of phospholipase A₁ (PLase A₁) was higher than that of PLase A₂ at both pH 4 and 7. The involvement of lysophospholipase in FFA release in oysters seemed to be small. PLase A activity in the digestive glands of oysters was much higher than that in the white portion. The optimum pH and the effects of inorganic ions or chelate agents on the production of these metabolites were different between plankton and oysters, showing that they have their own PLase A characteristics.

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The fragmentary form of alanine racemase from Bacillus stearothermophilus is composed of two sets of two different polypeptides corresponding to the two domains of the subunit of wild-type enzyme. It was denatured with 6M guanidium hydrochloride to be separated into pieces, and renatured by dilution with about 50% recovery of the activity. The two kinds of polypeptides (i. e., large and a small fragments) were isolated by gel filtration in the presence of 4M guanidium hydrochloride. The CD spectra obtained by summation of the spectra of the refolded fragments were closely similar to that of the native fragmentary enzyme. The lysine residue to which PLP is bound in the wild-type enzyme occurs in the large peptide of the fragmentary enzyme containing the amino terminus of the wild-type enzyme. The visible spectrum of the large peptide refolded, however, indicated that PLP was not bound to it. The large peptide alone had no significant activity, but it was activated by incubation with the small peptide. Accordingly, co-existence of both peptide fragments is necessary for folding of a complex of the two kinds of peptide to form an active structure.

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Effects of cold treatment and day length on endogenous gibberellins (GAs) in the stems and leaves of Japanese radish (Raphanus sativus L. cv. Taibyo-sobutori) were investigated. The concentrations of putative active GAs (GA₁ and GA₄) and their precursors (GA₉ and GA₂₀) in both stems and leaves of R. sativus grown without cold treatment were higher in the long-day condition than the short-day condition. The concentrations of the above four GAs in the stems and leaves generally tended to decrease during the cold treatment, although GA₁ in the stems was at almost the same level before and after the cold treatment. These results suggest that the GA biosynthesis is promoted by the long-day condition rather than cold treatment. Considering that application of GAs after the cold treatment as well as cultivation in the long-day condition after cold treatment induces the bolting of R. sativus, both the cold treatment and activation of GA biosynthesis by cultivation in the long-day condition seem to be essential to the bolting, although it remains to be seen whether the cold treatment affects sensitivity or responsiveness of the stems to GAs. It was also shown that the endogenous levels of GAs in the upper part of the bolting stem were one or two orders of magnitude higher than those in the stem in the rosette plant.

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Koninginin C, a congener of koninginins A and B, was isolated from Trichoderma koningii fermented on a shredded wheat medium. The compound inhibited the growth of etiolated wheat coleoptiles by 100% at 10^-3 M. It was a fine, white cystalline substance with a molecular formula of C₁₆H₂₈O₄ and a melting point of 70-72°C.

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The tube growth of Pinus pollen was strongly inhibited by paraquat, while that of Typha pollen was not. Among superoxide dismutase, catalase, and ascorbate peroxidase, which scavenge superoxide and hydrogen peroxide, the activity of ascorbate peroxidase in Pinus pollen was completely inactivated by paraquat before it began to elongate the tube.

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The antifeedant activity toward larvae of Pieris rapae crucivora of aromatic carbonyl compounds was studied. When an H atom of aldehyde group of aromatic aldehydes was replaced by a CH₃ group, it was observed that arylmethylketones were more active toward larvae than aromatic aldehydes.

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An autonomously replicating plasmid is expected to increase the frequency of Aspergillus transformation. To construct this type of plasmid, we developed a rapid method of re-isolating autonomously replicating plasmids from Aspergillus transformants. Transformants grown in MM medium under selective pressure for 1-2 days were converted to protoplasts with a cell wall lytic enzyme (e. g. Yatalase). The protoplasts were lysed with phenol/chloroform followed by precipitation with ethanol. The total DNA was treated with RNaseA, re-precipitated with PEG, and then used to transform E. coli. These re-isolated plasmids were mainly the plasmid monomer.

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β-N-Acetylglucosaminidase (EC 3.2.1.30) was purified from the outer membrane of a marine bacterium, Alteromonas sp. strain O-7. The enzyme (GlcNAcase A) was purified by successive column chromatographies. The purified enzyme was found to be homogeneous on sodium dodecyl sulfate polyacrylamide gel electrophoresis. The molecular mass and pI of GlcNAcase A were 92 kDa and 4.9, respectively. The optimum pH and temperature were 6.0-7.0and 45°C, respectively. GlcNAcase A was stable up to 40°C at pH 7.0, and hydrolyzed N-acetylchitooligosaccharides from dimer to hexamer. The amino-terminal 16 amino acid residues of GlcNAcase A were sequenced.

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A mechanism of menaquinone (MK) production in a detergent supplemented culture, in which only MK-4 but not MK-6 was released into the culture medium, was studied. MK-4 as well as MK-6 was found in the cell membrane fraction. Migration of MK4 from the inside to the outside of cells in the extracellular production might be due to release from cells rather than active excretion. Selective release of MK-4 was found to require the envelope structure of the cell membrane.

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(+)-Eucamalol (1) and (-)-1-epi-eucamalol (2) were synthesized from (S)-(-)-perillaldehyde to determine the absolute configuration of 1, the structure of natural (+)-eucamalol being determined to be (1R, 6R)-(+)-3-formyl-6-isopropyl-2-cyclohexen-1-ol. (+)-Eucamalol (1) and its 1-epimer (2) exhibited significant repellent activity against Aedes albopictus, and inhibited its feeding as well as DEET.

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The transglycosylation reaction of partially purified β-glucosidase from sesame seeds with cellobiose is described. Sesame β-glucosidase was partially purified by ammonium sulfate fractionation and gel filtration. The molecular weight of the enzyme was 200, 000 by gel filtration. Sesame β-glucosidase showed strong transfer activity to synthesize the trisaccharide from cellobiose. The optimum pH and temperature of the transglycosylation reaction were pH 4.0 and 60°C.

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The effect of storing beer on the generation of stale flavor was investigated by a sensory evaluation and quantitative determination of the volatile components. The results of the sensory evaluation revealed 2-furfuryl ethyl ether (2-FEE) to be an important component responsible for the astringent stale flavor, and (E)-2-nonenal (E2N)for the cardboard-like stable flavor. However, the stale flavor could not be reproduced by adding 2-FEE or E2N alone to fresh beer, but was closely reproduced when 2-FEE and E2N were added together. The results of experiments on adding furfuryl acetate (FAC) and furfuryl alcohol (FAL) to a 5% ethanol solution or beer indicate that 2-FEE was generated by the reaction between ethanol and FAL, which was formed by the hydrolysis of FAC, during the storage of beer.

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Angiotensin I converting enzyme (ACE, E. C. 3. 4. 15. 1) inhibitory activity were measured with 11 kinds (31 items) of fermented foods. Strong inhibitory activity was detected in soy sauce, fish sauce, natto, nyufu, and cheese, but not in mirin, sake, or vinegar.

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Natural rubber serum (NRS), was found to have a remarkable growth-promoting effect on various kinds of microorganisms, in particular, anaerobes. NRS stimulated the growth of all the species of Bifidobacterium tried and synergism was noted between the effects of NRS and nutrients in media that contained yeast extract, meat extract, and the casein as nutrients.

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Random mutations were introduced into recombinant barley β-amylase by modified PCR to increase its thermostability. Two clones were obtained. One was found to have a change of Ser-351to Pro and another, a change of Ala-376 to Ser, and 2.3°C and 1.0°C increases, respectively, in the thermostabilities compared with that of native recombinant β-amylase.

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We isolated three organic solvent (OS)-tolerant bacterial strains DS-1051, DS-1902, and DS-313 from a depth of 1, 168m in Sagami Bay, Japan. These isolates were tolerant to various kinds of toxic OSs such as benzene, toluene, and p-xylene. They also could degrade polyaromatic hydrocarbons, naphthalene or biphenyl, in a medium-OS (9:1) two-liquid-phase system. Percentage degradation of polyaromatic hydrocarbons in OS by these strains were higher than those obtained from cultures in which substrates were in the medium without OS.

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Malonylated anthocyanin was isolated from the receptacle of 3 Japanese strawberries. The chemical structure was identified to be pelargonidin 3-O-(6-O-malonyl-β-D-glucoside) (1) mainly by ¹H-NMR and FABMS. The anthocyanin content was about 30, 25, and 12% of total anthocyanins in Fragaria × ananassa Duch. cv. Nyoho, F. ananassa Duch. cv. Himesodachi and F. ananassa Duch. cv. Reiko, respectively. On the other hand, the cultivars Toyonoka and Ai-berry did not contain 1 at all.

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The effect was investigated of dietary 0.5% sodium phytate on the hepatic and serum lipid status in rats fed on sucrose for 29 or 30 days. The increase in hepatic weight, levels of hepatic total lipids, triglyceride, cholesterol, serum triglyceride and phospholipid, and the activities of hepatic NADPH-generating enzymes due to sucrose feeding were generally depressed by dietary sodium phytate.

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A practical synthesis of (2S, 4S)4-hydroxyproline (1) based on DCC-induced inversion of the hydroxyl group of (2S, 4R)4-hydroxyproline (2) is described.

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The substrate-binding sites of a bifunctional enzyme, cyclic-ribonucleotide phosphomutase-5'-phosphodiesterase from Aspergillus niger, FS-44, were investigated by kinetic studies. A series of adenine nucleotides. inhibited the mutase and diesterase reactions, but adenosine and adenine did not at all. Adenosine monophosphate derivatives acted as competitive inhibitors with respect to both adenosine 3', 5'-cyclic monophosphate in the mutase reaction and adenosine 5'-P-nitrophenyl phosphate in the diesterase reaction with the same decreasing order of inhibition constant. However, the inhibition constants of these compounds in the mutase reaction were 1 order of magnitude lower than those in the diesterase reaction. These results suggest that the active site(s) on the enzyme catalyzing the mutase and diesterase reactions have similar properties, but the substrate-binding sites of both reactions are distinct at least.

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Bovine K-caseinoglycopeptide (residues 106-169, CGP) bound to mouse plastic-adherent cells. A culture supernatant prepared from mouse spleen cells and plastic-adherent cells cultured with CGP significantly inhibited the lipopolysaccharide-induced proliferative response of mouse spleen lymphocytes. The inhibition of proliferation by the culture supernatant was completely overcome by the addition of the anti-interleukin-1 receptor antagonist antibody.

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