An enzymatic method for determination of 5-phosphoribosylpyrophosphate (PRPP) was investigated. The method was based on a coupled reaction of orotidine-5'-monophosphate pyrophosphorylase and orotidine-5'-monophosphate decarboxylase obtained as cell-free extract from a mutant strain of Micrococcus glutamicus (Syn. Corynebacterium glutamicum). The enzymes catalyzed quantitative conversion of orotic acid to uridine-5'-monophosphate. In the method, the consumption of orotic acid was determined by measuring optical density at 295mμ and then PRPP was calculated from the amount of orotic acid consumed. The conditions of the enzyme reaction were examined in detail, and a modified method for determination of PRPP involving termination of the enzyme reaction by EDTA was est-ablished. According to the present method, 10^-1 μmol. of PRPP is determined with ac-curacy of 100±3% within 30 minutes. As an application of the method, chemical stability of PRPP was examined and the results were described.

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From mycelia of Asp. niger and Asp. awamori aurasperones A, B and C along with related two yellow pigments have been isolated.
Aurasperone A, C₃₂H₂₆0₁₀, is obtained in yellow prisms; m. p. 207°C; [α]D-136°; gives the diacetate and the dimethyl ether and is assumed to be a dimeric 2-methyl-5-hydroxy-6, 8-dimethoxy-4H-naphtho [2, 3-b] pyran-4-one (IV). Aurasperone B, [α]D+46.3°, is the main yellow metabolite, m. p. 186°C, and affords aurasperone A on hydrochloric acid-treatment. It has molecular formula C₃₂H₃₀0₁₂ and is supposed to have the structure (V). The other yellow pigments have been found to be also congeners of aurasperone A.

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Four fractions of glucoamylase, different chromatographically from one another, were isolated and purified from a culture of Aspergillus oryzae on steamed rice. All these fractions were found to be homogeneous in sedimentation analysis, and had almost the same enzymatic activities in the hydrolysis of soluble starch. The limit of hydrolysis of starch was about 60%, which was very similar to those of Taka-amylase B of Okazaki and Taka-amylase B₂ of Sawasaki.

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Benzyl 2, 3, 6-tri-O-acetyl-4-O-(2, 3-di-O-acetyl-4, 6-di-O-methylsulfonyl-β-D-glucopyrano-syl)-β-D-glucopyranoside (VI) was prepared from α-cellobiose octaacetate. Displacement of the sulfonyl esters of VI with acyloxy-groups in N, N-dimethyl formamide in the presence of sodium benzoate gave 4-0-β-D-galactopyranosyl-D-glucopyranose derivative (lactose derivative). Elimination of blocking groups of the derivative yielded lactose hydrate (IX), though the overall yield of lactose from cellobiose octaacetate was less than 2%.

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Acid hydrolysis of cytosinine gave each one mole of cytosine, levulinic acid, ammonia and carbon dioxide. Reduction of cytosinine with PtO₂ afforded a mixture of dihydro-cytosinine, 3-amino-tetrahydropyran-2-carboxylic acid and cytosine. Ozonolysis of N, N'-diacetylcytosinine methyl ester, followed by oxidation with hydrogen peroxide and acid hydrolysis gave erythro-D-β-hydroxyaspartic acid. These data permitted the assignment of structure (I) for cytosinine. Acid hydrolysis of uracinine gave uracil instead of cytosine, therefore, the structure (II) could be assigned to uracinine. Some stereochemical features and mechanism of levulinic acid formation were discussed.

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Blasticidin S was cleaved into cytosinine and blastidic acid by acid hydrolysis under selected condition. Chemical and NMR spectral evidences showed that blastidic acid has the structure VI. Alkaline hydrolysis of blasticidin S gave cytomycin which was further degradated into cytosinine and blastidone, whose structure was assigned as 4-ureido-N-methylpiperidone. Hydrogenolyses of blasticidin S and cytomycin afforded the hydro-genolytic compounds SC₁₃ and C₁₃ respectively. SC₁₃ was transformed into C₁₃ by alkali, which was further degradated via partial hydrolysis product PHC₁₃ into 3-aminotetra-hydropyran-2-carboxylic acid and blastidone. The sequence of these reactions established clearily the relationship of degradative compounds and provided evidences to assign struc-ture I and II to blasticidin S and cytomycin respectively.

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Conditions for the production of endo-polygalacturonase (endo-PG) with Aspergillus saitoi IAM 2217 in the submerged culture was examined. This strain was selected as the most potent producer of endo-PG. Endo-PG of this strain was produced in the absence of pectin, but the addition of pectin increased endo-PG activity when inoculated with proliferated mycelia.
As far as examined with a modified Czapek medium (ordinary constituents+pectin and ammonium tartrate), the addition of organic nitrogen sources, such as corn steep liquor, markedly reduced the enzyme producibility. As for the carbon and nitrogen amount in the medium, sucrose: 4%, pectin: 2%, NaNO₃: 1.15%, C/N=10, gave the best result among tested.

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Amino acid composition of crystalline Japanese-radish peroxidase c has been deter-mined by an automatic amino acid analyzer. Carbohydrate composition of this enzyme has also been investigated by means of the Tillmen and the Bial reactions. The analyses indicated that the peroxidase c had the following constituents in each molecule (mol. wt. 41, 500): Asp₃₆Glu₂₄Gly₃₄Ala₂₈Val₂₇Leu₂₇Ileu₁₆Ser₂₃Thr₁₆(GYs-)₁₀Met₃Pro₁₆Phe₁₄Tyr₂Try₂His₄ Lys₈Arg₃₀(-CONH₂)₂₇Hexose₇Pentose₂Hexosamine₃Protohematin₁. Since this composition was quite different from that of peroxidase a, two peroxidases could not be converted from each other, but they would be synthesized in vivo independently.

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Polyphenol oxidase in tea leaves was fractionated to three components, called A-I, A-II and B, by the combination of DEAE and CM-cellulose column chromatographies. They exhibited different migration on starch-gel electrophoregram at pH 5.0. The results of CM-cellulose chromatography and starch-gel electrophoresis indicated that A-I and A-II were basic proteins but B was not. Specific activities of purified components, A-I and A-II, were increased to about 200 and 140 times as much as that of tea leaf homogenate, respectively.
Optimum pH for A-I and A-II are 5.1 and 4.6, respectively. A-I was found to oxidize o-diphenol rather well, whereas, A-II oxidized well both vicinal-triphenol and o-diphenol. The activity of the above two enzymes was inhibited by a high concentration of substrate. Optimum temperature of enzymic reaction was about 35°C.

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A thin-layer chromatographic method is described for the separation, identification and determination of 2, 4-dinitrophenyl sulfides. The derivatives are easily obtained from mercaptan and dinitrofluorobenzene in the presence of sodium bicarbonate at room tem-perature. The sulfides are separated on silica gel plates using a solvent mixture of benzene-xylene-carbon tetrachloride (2:1:1, v/v). The individual sulfides are determined spectrophotometrically, at 330_??_335mμ in ethanol, εmax ca. 13, 000, after washing out the plate with hexane and extraction from the adsorbent with acetone.

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The enzyme system of NAD degradation was extracted from autolysate of Saccharomyces oviformis.
The enzymes were partially separated by ammonium sulfate fractionation and DEAE-cellulose column chromatography, and then the metabolic pathway of NAD in yeast was presented, in which four enzymes were contained. It has been found that, among them, the 5'nucleotidase has more affinity for AMP and the nucleosidase has strict affinity for nicotinamide riboside.
In the degradation of NAD in the yeast, nucleotide pyrophosphatase was main enzyme, but NADase, nucleotide pyrophosphorylase and adenosine deaminase seemed not to play an important role.

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The production of pimelic acid from azelaic acid by microorganisms was studied. About 100 strains of bacteria which were able to utilize azelaic acid as a sole carbon source were isolated from soil and other natural materials. Among these bacteria, several strains produced a large quantity of an organic acid (pimelic acid) from azelaic acid in their culture fluids during the cultivation. The acid was isolated from the culture fluid of strain A133 in crystalline form. The crystal was identified as pimelic acid by physico-chemical and biological methods.
From the results of investigations on the morphological and physiological characters, the bacterial strain A133 was assumed to be Micrococcus sp.

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A number of saligenin cyclic phosphonates and phosphonothionates were prepared and their insecticidal activity and stability were examined. They were less stable than corresponding phosphate esters. 2-Ethyl-4H-l, 3, 2-benzodioxaphosphorin-2-sulfide was the most effective against house flies.

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A peptidase was isolated from the cells of amylase-producing Bac. subtilis by means of cell lysis with egg white lysozyme, followed by freezing and thawing, salting out, dialysis and ion-exchanger column chromatography. The enzyme required manganese ion to show the enzyme activity. Also the enzyme was stable in the presence of magnesium ion. The enzyme hydrolyzed various synthetic peptides by stepwise removal of the amino terminal amino acid of peptides and thus the peptidase was found to be aminonentidase.

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