Cultural conditions suitable for bacteriochlorophyll a formation by Protaminobacter ruber classified as a non-photosynthetic bacterium were investigated. Optimal values for temperature, light intensity in the light followed by dark conditions, which was most suitable for bacteriochlorophyll formation, and apparent k_La were 20-25°C, ca. 2, 000 lux, and ca. 1.0 min^-1, respectively. Although aeration was indispensable for the growth of P. ruber, less aeration was favorable for bacteriochlorophyll synthesis. In this microorganism, bacteriochlorophyll was inducibly formed under special conditions, while vitamin B₁₂ and cytochrome were constitutively produced. In contrast, a photosynthetic bacterium, Rhodopseudomonas spheroides produced the highest amount of bacteriochlorophyll under continuous illumination conditions, which completely prevented bacteriochlorophyll formation in P. ruber, and also low oxygen tension conditions, under which P. ruber could not grow at all.

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Fermentative production of L-serine using glycine as a precursor and Corynebacterium glycinophilum as a microbial producer was improved by decreasing the enzymatic degradation of L-serine. The cellular activity of L-serine dehydratase (SD) [EC 4.2.1.13], which was responsible for the L-serine degradation, was decreased in mutants that could not assimilate L-serine as a nitrogen, source or that required amino acids for their growth. Typical examples of the mutants were an L-leucine and L-isoleucine auxotroph (AJ-3414) which completely lacked SD and an L-leucine and L-methionine auxotroph (AJ-3413) whose SD activity was 32% of that of the parent. They accumulated 13.8 and 13.9mg/ml of L-serine, respectively, from 30mg/ml of glycine with a production yield of 46% as compared to the parental productivity of 15.3%.

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The ability of bifidobacteria to synthesize six water-soluble vitamins (thiamine, folic acid, nicotinic acid, pyridoxine, vitamin B₁₂ and riboflavin) was systematically investigated with twenty-four strains of five species derived from human feces. The vitamins synthesized were determined as those accumulated in cultures grown in a semi-synthetic medium. For the vitamins other than vitamin B₁₂ and riboflavin, extracellular liberation was also examined with the supernatant fluids obtained after removing cells from the cultures. Many strains of the bifidobacteria investigated could indeed synthesize five of the vitamins, the exception being riboflavin. A large portion of each of the vitamins synthesized was excreted into the medium. The concentrations of the vitamins, especially thiamine, nicotinic acid and folic acid, accumulated varied widely among different species or strains. On the basis of the results, these bifidobacteria could be divided into three general types according to the abilities to accumulate thiamine, nicotinic acid and folic acid. These three vitamins were accumulated in all the strains of B. bifidum and B. infantis as well as in many strains of B. breve and B. longum but the vitamin concentrations were significantly higher in the former species (higher-accumulators) than in the latter species (lower-accumulators). On the other hand, none of these three vitamins were detected in most strains of B. adolescentis or in some strains of B. breve and B. longum (non-accumulators). Non-accumulator strains of B. adolescentis required thiamine and nicotinic acid for maximal growth in a complete synthetic medium. Further studies on thiamine synthesis revealed that the addition of exogenous thiamine to the medium significantly reduced the level of thiamine accumulated in a culture of a lower-accumulator strain of B. longum but did not affect the vitamin level in a higher-accumulator strain of B. bifidum. These findings were discussed in relation to the vitamin biosynthesis and its regulation in bacteria.

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Analysis of trace elements using a polarized Zeeman atomic absorption spectrophotometer was performed on breast milk, cow's milk, and a modified milk powder commonly consumed in Japan. The elements examined in the samples were cobalt, nickel, chromium, manganese, molybdenum, and iron.
Average values of contents (per g wet weight) of these elements in breast milk obtained from mothers 19-384 days post partum were 1.4ng, 3.6ng, 6.5ng, 9.5ng, 24ng and 0.32 μg, respectively and those in cow's milk were 2.4 ng, 9.5 ng, 14 ng, 27 ng, 50 ng and 0.23 μg, respectively. The ratio of contents of cobalt, nickel, chromium, manganese, and molybdenum in breast milk was similar to that in cow's milk. There was no infant formula deficient in these elements as compared to those in breast milk when the levels of trace elements were expressed per ml of 14% (w/v) milk reconstituted from each milk powder. In addition, higher contents of iron and manganese in milk powder were noted.

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The endogenous gibberellins in rather matured seeds of Pharbitis purpurea were qualitatively analyzed by combined gas chromatography/mass spectrometry, and gibberellins A_{3, 5, 8, 17, 19, 20, 26, 27, 29, 34, 44 and 53} were identified. A semiquantitative analysis of these gibberellins was performed on a small quantity of very young, immature seeds by gas chromatography/selected ion monitoring (GC/SIM) after purification by high performance liquid chromatography. The mass spectra of two types of these gibberellin derivatives, methyl ester-trimethylsilyl ethers and trimethylsilyl ester-trimethylsilyl ethers, were measured and their advantages and disadvantages in GC/SIM analysis are discussed.

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The effects of light intensity and wavelength on hydrogen evolution by Rhodopseudomonas rutila, a nonsulfur purple photosynthetic bacteria newly isolated from paddy soil, were investigated. The maximal specific rate of hydrogen evolution was achieved at a light intensity of about 12 klux with an incandescent light. As to wavelength dependence, four major peaks of hydrogen evolution were observed at 900, 860, 810 and 590 nm. Light of longer wavelengths above 590 nm that is absorbed by bacteriochlorophyll a was more effective for hydrogen evolution than light of shorter wavelengths below 540 nm that is absorbed by carotenoids.

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Five enzymes after 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase (DS) in the common pathway for aromatic amino acid biosynthesis in Brevibacterium flavum were examined for general enzymatic and regulatory properties. These enzyme activities were not feedback-inhibited by the end products, phenylalanine, tyrosine, and tryptophan. On the other hand, syntheses of dehydroquinate synthase (DQS), shikimate dehydrogenase (SD), shikimate kinase (SK) and 5-enolpyruvylshikimate-3-phosphate synthase (ESPS) were derepressed 2- to 8-fold by tyrosine limitation as is DS. The azaserine-resistant tryptophan-producing mutant A-100 had 1.6- to 2.9-fold higher levels of DS, DQS, SD, SK, and chorismate synthase than those of the parent. Moreover, further increases of DS and DQS activities were found in sulfaguanidine-resistant mutants derived from strain A-100. These enzymes in the azaserine- and sulfaguanidine-resistant mutants were still repressed by tyrosine but showed higher activities than did the respective parents at any tyrosine concentration or any growth phase.

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In order to investigate the substrate-specificity of α-L-rhamnosidase from Aspergillus niger, the following disaccharides were synthesized: 2-O-α-L-rhamnopyranosyl-α-D-fucopyranose (1), methyl 4-O-α-L-rhamnopyranosyl-β-L-arabinopyranoside (2), methyl 2-0-α-L-rhamnopyranosyl-α-Lrhamnopyranoside (3) and 6-O-α-L-mannopyranosyl-D-glucopyranose (4).
The action of α-L-rhamnosidase on compounds 1-4 and another fifteen disaccharides containing α- or β-L-rhamnopyranosidic bonds or an α-L-mannopyranosidic bond was examined. As the result, all the disaccharides having an α-L-rhamnopyranosidic linkage were hydrolyzed well, while the ones having β-L-rhamnopyranosidic or α-L-mannopyranosidic linkage could not be hydrolyzed at all. Accordingly, this enzyme might be used for the determination of anomeric configurations of the L-rhamnopyranosidic bond, although further studies on the specificity of the enzyme are required.

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D-Glucosamine selectively inactivated free PL-1 phage under the conditions in which the growth of host cells, Lactobacillus casei ATCC 27092, was not affected. Both the N-acetylated and non-aminated forms little affected the phage infectivity. The rate of phage inactivation by D-glucosamine was proportional to the time and temperature of the incubation, and about 0.1 M D-glucosamine showed the maximum inactivation effect. The phage inactivation was stimulated by Cu²⁺, and inhibited by catalase and several oxygen radical scavengers. The inactivated phage was identical with the intact one with respect to the electron microscopic appearance, the density and the ability of combining with host cells. The DNA in the virion was fragmented by D-glucosamine and the DNA fragmentation was also stimulated by Cu²⁺. With plasmid pBR322, D-glucosamine caused single-strand scission of ccc-DNA to give oc- and in succession linear-DNA. The inactivation of PL-1 phage by D-glucosamine was considered to be due to single-strand scission of DNA in the virion by active oxygens such as ¹O₂ formed during autoxidation of D-glucosamine in the aqueous solution.

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The unit peptides, cadystin A and B, in the cadmium-containing peptides occurring in a fission yeast were synthesized for confirmation of the structure. Four possible stereoisomers of the previously proposed structure for cadystin were also synthesized.

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We investigated the subunit structures of the three agglutinins (BRA-1, BRA-2, and BRA-3) isolated from the coelomic fluid of the acorn barnacle, Megabalanus rosa, by high-speed gel filtration, gel electrophoresis, CD spectra, and peptide mapping. BRA-2 (Mr 140, 000) and BRA-3 (Mr 64, 000) were glycoproteins composed of subunits having two identical basic units cross-linked by disulfide bonds. The basic units of BRA-2 and BRA-3 had molecular weights of 22, 000 and 18, 000, respectively. They did not have a common structure. BRA-1 (Mr 330, 000) was composed of subunits homologous to that of BRA-2. The binding properties of these agglutinins against rabbit erythrocyte ghosts were also investigated after labeling the agglutinins with fluorescein isothiocyanate.

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Niacin formation from tryptophan was compared in two groups of rats, one fed a niacin- and protein-free diet (group A) and the other, niacin-free 30% casein diet (group B) for one day after preliminary feeding of a niacin- and protein-free diet for three days (experiment I). In the other experiment, L-tryptophan was added to the last day's diets (experiment II). NAD and the total niacin contents in liver as well as urinary excretion of pyridine compounds were significantly higher in group I-B than in group I-A. L-Tryptophan supplementation reversed the total niacin content in liver, while total tryptophan intake was about two times higher in group II-B. Since the activities of kynurenine metabolizing enzymes had not changed, we considered that it was not due to a difference in kynurenine metabolism. Urinary excretion of quinolinic acid tended to be higher in group II-A than in group II-B. However, quinolinate phosphoribosyltransferase activity did not differ significantly. On the other hand, the activities of α-amino-β-carboxymuconate-ε-semialdehyde decarboxylase (ACMSDase) in both liver and kidneys were found to be significantly higher in group II-B than in group II-A. This might be responsible for the lower content of liver total niacin in group II-B.

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A screening test was carried out to obtain microbes which produce fruiting-body inducing substances and an inducer was found in the culture broth of an actinomycete, strain TO 447. This strain was identified as Streptomyces albulus from its taxonomical characteristics.
The fruiting-body inducing substance produced by this strain was isolated and identified as Cyclooctasulfur by spectroscopic evidence. Cyclooctasulfur induced the fruiting-body formation of Favolus arcularius in a dosage range of 0.01-1000ng/disc.

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A screening test was carried out to obtain microbes which produced pig pancreatic α-amylase inhibitor, and a new inhibitor was found in the culture broth of an actinomycete, strain No. 196. This inhibitor was designated as Paim, an abbreviation for pig pancreatic α-amylase inhibitor from a microbe. Strain No. 196 was identified as Streptomyces corchorushii from its taxonomic characteristics. When strain No. 196 was cultured aerobically in a medium consisting of 5% glucose, 2% soy bean flake extract and 0.3% NaCl, pH 7.0, at 35°C for 35 hr, maximum production of Paim was obtained. Paim inhibited animal α-amylases such as those of pig, dog, cow or horse, but not the human one.

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A 1-hydroxy-2-naphthoate-resistant mutant, strain HNA 12-D, which was derived from a menaquinone (MK)-6 producer, Flavobacterium meningosepticum IFO 12535, produced an increased amount of MK with the production of another type of MK in addition to MK-6. The newly formed MK was isolated and identified as MK-5. The total amount of MK reached 55.6mg/liter of culture broth and 9.19mg/g dry cells in the ratio of MK-6 and MK-5 of 1: 1.7.

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The distribution of amino acid dehydrogenases in lactic acid bacteria was investigated. Several lactic acid bacteria were found to have NADP⁺-specific glutamate dehydrogenase, but NAD⁺- specific glutamate dehydrogenase activity was not detected in the bacteria tested. No alanine, leucine or lysine dehydrogenase activity was detected either, though serine and proline dehydrogenases occur in some of the bacteria. NADP⁺-specific glutamate dehydrogenase was purified to homogeneity and crystallized from Lactobacillus fermentum IFO 3071. The enzyme had a molecular weight of about 300, 000 and consisted of six subunits identical in molecular weight (50, 000). The enzyme is highly specific for L-glutamate and α-ketoglutarate, and the activity of the reductive amination is much higher than that of the oxidative deamination. The pH optima for the deamination and the amination were about 9.0 and 8.0, respectively. The apparent Km values were determined to be 79 mM for L-glutamate, 44 μM for NADP⁺, 5.6mM for α-ketoglutarate, 6.76mM for NH₃, and 77.5 μM for NADPH.

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A fugus which utilizes diaminomaleonitrile (DAMN) as a source of nitrogen was isolated from soil, and identified as a strain belonging to the genus Trichoderma. The strain degraded DAMN and accumulated several fluorescent metabolites. The intact mycelia and a cell-free extract of the strain formed hydrogen cyanide and ammonia from DAMN, besides the same fluorescent compounds, as metabolites. The enzyme catalyzing the liberation of hydrogen cyanide from DAMN was partially purified and its properties were investigated. From the results, the enzyme was found to catalyze the liberation of hydrogen cyanide from DAMN but not the formation of ammonia from DAMN, and its substrate specificity was very narrow; it reacts only with DAMN of the compounds so far examined.

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Spheroplasts of Zymomonas mobilis (Z-6) were prepared by cultivating it in a hypertonic medium containing penicillin G or glycine at 30°C for more than 6 hr. The reversion of Spheroplasts to a bacillary form was observed at frequencies of 10^-2 to 10^-3 when Spheroplasts were incubated on hypertonic solid agar plates overlaid with soft agar. Spheroplast fusion was attempted with polyethylene glycol 6000 by crossing two mutants lacking the ability to ferment sugars. Fusants were selected, and obtained at high frequencies. The properties of a stable fusant are discussed.

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Four geometrical isomers of 5, 7-, 6, 8-, 7, 9- and 8, 10-dodecadien-1-ols were systematically synthesized by two routes. One involved the Wittig reaction between (E)-2-alkenal and a phosphorane with an appropriate carbon chain, and yielded a mixture of (E, Z)- and (E, E)-isomers in a ratio of ca. 3:2. The other route comprised of the Wittig reaction of a 2-alkynal and the stereoselective reduction of the triple bond to a (Z)-double bond via hydroboration with dicyclohexylborane to give a mixture of (Z, Z)- and (Z, E)-isomers in a ratio of 6:1-10:1. Both the mixtures were separately chromatographed on a silica gel column impregnated with silver nitrate to give four geometrically pure isomers. With 9, 11-diene they were analysed by GC on a capillary column, HPLC, silver nitrate impregnated TLC, and ¹³C NMR.

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A Stereoisomeric mixture of (±)-PM-toxin B [6, 14, 22, 30, 32-pentahydroxy-8, 16, 24-trioxotritriacontane] has been synthesized via three steps of Grignard addition and shows virtually the same biological activities as the natural PM-toxin B.

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The decarboxylation of L-tryptophan catalyzed by the aromatic L-amino acid decarboxylase from Micrococcus percitreus was competitively inhibited by α-methyl-L-phenylalanine. During prolonged incubation with the α-methyl amino acid, the enzyme lost its activity and its absorption spectrum changed drastically. The activity was restored by the addition of a cofactor, pyridoxal phosphate (PLP). The reaction products from α-methyl-L-phenylalanine were isolated and identified as methylbenzylketone and α-methyl-β-phenylethyl amine. PLP added to the reaction mixture was found to change to pyridoxamine phosphate (PMP) after the reaction. These results indicate that the enzyme catalyzes decarboxylation-dependent transamination besides the normal decarboxylation of α-methyl-L-amino acids.

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A new shikimic acid derivative was isolated from the culture filtrate of Penicillium sp. and characterized as methyl 5-lactyl shikimate lactone.

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Conditions suitable for the release of mycelial protoplasts of Coprinus macrorhizus and for colony formation by the isolated protoplasts were examined. The study culminated in the development of a method using commercially available preparations of chitinase, cellulase and β-glucanase, by which over 2×10⁸/ml protoplasts could be obtained within 2hr. The protoplasts from the wild type and mutant Fis^c of C. macrorhizus regenerated into hyphae in a complex medium containing malt and yeast extract, and subsequently formed colonies with frequencies up to 50%, which paralleled the percentage of nucleated protoplasts. The plating efficiency in a minimal medium was about 20% on average. Sucrose was found to be superior to mannitol as an osmoticum for colony formation. Protoplasts from auxotrophic mutants formed colonies at much lower frequencies even in the complex medium, but addition of N-acetyl glucosamine significantly improved their plating efficiency. Colonies derived from the protoplasts of C. macrorhizus Fis^c consistently developed fruiting bodies, when grown on a hypotonic medium.

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Bacteria capable of producing N-acetylneuraminate lyase (N-acetylneuraminic acid aldolase, EC 4.1.3.3) were sought by using a culture medium containing N-acetylneuraminic acid as a sole source of carbon. The enzyme was found to be distributed in a veriety of bacteria belonging to such genera as Pseudomonas, Escherichia, Aerobacter, Proteus, Micrococcus, Sarcina, Brevibacterium, Corynebacterium, Arthrobacter, Bacillus and Bacterium. No bacteria secreted the enzyme into the medium. Substantially N-acetylneuraminate lyase of bacteria seems to be an intracellular enzyme.
Potent N-acetylneuraminate lyase activities were detected in all of the strains belonging to Escherichia such as E. coli, E. freundii and E. intermedia. There was no difference in the enzyme production between a strain of E. coli which had the ability to produce colominic acid and one which had lost the ability.

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The emulsifying properties of β-lactoglobulin (β-Lg) in relation to its molecular structure were investigated at pH 3-9. In the acidic pH region, β-Lg showed relatively low emulsifying and surface activity, while its surface hydrophobicity was greater than at neutral pHs. The conformational stability of β-Lg varied depending on pH, i.e. its conformation was more rigid and resistant to denaturation at pH 3 than at pH 7. The low emulsifying and surface activity of β-Lg at acidic pHs was assumed to be due to such low denaturability (flexibility) of the molecule. Cleavage of the intramolecular disulfide bonds increased the denaturability, as well as the emulsifying activity, of β-Lg at pH 3.

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The β-anomer of N⁶-benzyladenosine has been long known as an artificial cytokinin.¹⁾ Recently, however, this compound was isolated from a cytokinin-autotrophic cell culture of anise, Pimpinella anisum L.²⁾ Thus, this compound now constitutes the third member of the naturallyoccurring cytokinins possessing an N⁶-benzyladenine structure, together with N⁶-(o-hydroxybenzyl)adenosine isolated from leaves of Poplus robusta³⁾ and N⁶-(o-hydroxybenzyl)-2-methylthio-9-β-D-glucofuranosyladenine isolated from fruits of Zantedeschia aethiopica.⁴⁾ No information has been available concerning whether or not the α-anomer of N⁶-benzyladenosine possesses cytokinin activity.
We report here the synthesis and cytokinin activity of N⁶-benzyl-α-adenosine (N⁶-benzyl-9-α-D-ribofuranosyl adenine).

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(R)-[2-¹⁴C]-Mevalonic acid (MVA) lactone was incorporated into (-)-4'-hydroxy-γ-ionylideneacetic acid (4'-hydroxy-γ-acid), which was first isolated from the culture broth of Cercospora cruenta. 4'-Hydroxy-γ-was then metabolized to (+)-(2Z, 4E)-4'-oxo-α-ionylideneacetic acid and (+)-(2Z, 4E)-1', 4'-dihydroxy-γ-ionylideneacetic acid. The latter was converted to (+)-abscisic acid (ABA) with a high incorporation ratio by the fungus.

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¹⁴C-labelled methionine, xanthosine, and 7-methylxanthosine were given to excised tea shoots. The methyl group of methionine was incorporated into 7-methylxanthosine (ca. 10%) in the earlier period of incubation after the uptake. About 50% of the radioactivity of xanthosine was rapidly incorporated into caffeine via 7-methylxanthosine, 7-methylxanthine, and theobromine within 24 hr. 7-Methylxanthosine was also converted into caffeine at a high rate. The results suggest that the pathway for caffeine biosynthesis is as follows: xanthosine → 7-methylxanthosine → 7-methylxanthine → theobromine → caffeine.

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GA₉ methyl ester (GA₉-Me) fed to a culture of prothallia of Lygodium japonicum was metabolized into three monohydroxy-GA₉-Me after three weeks. One of these was identified as GA₂₀-Me and the structures of the other two were defined to be 12α- and 12β-hydroxy-GA₉-Me by spectroscopic analyses.

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