The sake brewing is distinguished from other wine brewings by its formation of high concentration alcohol, usually forming about 20 vol.% of alcohol in final concentration. I. Yamasaki indicated that the presence of the solid part of koji in the medium was essential for the formation of high concentration alcohol. In the present paper the effective component of rice was investigated. The experimental method was simplified by a gradual addition of sucrose instead of the saccharification of rice starch by koji. Inoculum size of Saccharomyces sake was 3×10⁷/ml. The addition of a protein of rice, oryzenin as well as koji and rice, resulted in the formation of such high concentration alcohol as 18_??_20%.
The UV-absorption of tryptophol at 280 mμ which had proved to be most toxic to the yeast among higher alcohols, decreased by adding oryzenin. Consequently oryzenin was shown to have an ability to adsorb tryptophol.

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Influences of aerobic and anaerobic conditions of cultures on the formation of high concentration alcohol were investigated. Estimation of oxidation-reduction potentials (ORP) was carried out by the electrometric method. In a koji-containing medium ORP was comparatively high (rH above 20) in the earlier stage of the fermentation, and kept low (rH 6_??_11) in the later stage. In a koji-containing medium the concentration of alcohol formed was 21.6% while in the basal medium 17.6%. This result suggested that the condition which was aerobic in the earlier stage and anaerobic in the later stage might result in the formation of high concentration alcohol. Under the condition, with the basal synthetic medium, which was aerobic with a cotton plug for the initial 3 days and anaerobic after the seventh day of incubation by the paraffin-covering over the medium, 19.6% of alcohol was formed. E₀' of egg albumin which was effective as reported in the previous paper was determined by the titration method and it was 50 mV at pH 7.28 (rH 16). Egg albumin might contribute to maintain the ORP in the later stage of the cultures.

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The present paper deals with component of a koji mold which stimulates the yeast growth and with its stimulating mechanism. The washed mycelium of a black koji mold which was cultured in a synthetic medium was used as the material. Saccharomyces sake was inoculated in a medium to the extent of 3×10⁷/ml in a flask with a fermentation bung, and incubated at 20°C for 5 days. Yeast population in the basal synthetic medium alone was 1.8×10⁸/ml, while in the medium with the black koji mold added, 4.7×10⁸/ml. Various kinds of extracts of the black koji mold (by water, 10% NaCl, 0.1 N HCl, 0.1 N NaOH, and 70% EtOH) proved not to stimulate the yeast growth. This fact led to a conclusion that the effective component existed in an insoluble part of the mold. Fractionation of the insoluble component by centrifugation of the homogenate of the black koji mold gave the result that the mitochondrial fraction was shown to stimulate the yeast growth. The stimulating effect was not influenced under an anaerobic condition by N₂ gas replacement method. Hemoglobin also stimulated the yeast growth (3.2×10⁸/ml). In view of these fact, the effective component of the black koji mold was speculated as an electron acceptor to the yeast respiration.

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The distribution of free sugars in the seeds, leaf buds and roots of cereals (including six genera and two families) and legumes (including three genera) has now been investigated by the application of the cutting method of paper chromatograms and, more over, the method of fractional quantitative analysis of these free sugars in cereals and legumes was developed and their quantitative distribution was studied. The seed of cereals examined in this investigation contained 4.1_??_8.3% of sucrose and traces of stachyose, raffinose, maltose and glucose. The seeds of legumes examined in this investigation contained 4.9_??_6.9% of sucrose, 4.3_??_7.5% of stachyose and small quantities of glucose, raffinose and verbascose (?). The leaf buds and roots (after the germination of cereals seeds) contained 6.7_??_12.0% of sucrose and 4.5_??_11.1% of glucose. Stachyose and raffinose were not existent in the leaf buds, roots of brown rice and glutinous rice. The leaf buds (after the germination of legumes seeds) contained 6.8_??_9.3% of glucose and small quantities of sucrose, maltose and raffinose. The roots (after the germination of legumes seeds) contained 3.3_??_5.4% of glucose and small quantities of sucrose, stachyose, maltose and raffinose.

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The flavor components of MUGI-CHA were separated into sodium carbonate-soluble fraction, sodium hydroxide-soluble fraction and neutral fraction. The sodium carbonate and sodium hydroxide fractions were studied by paper chromatography, column chromato-graphy and gas chromatography. The following twenty-nine compounds were identified; anisole, phenetole, phenol, 2-furoic acid, m-cresol, guaiacol, benzoic acid, isomaltol, maltol, 4-methylguaiacol, catechol, m-toluic acid, p-toluic acid, resorcinol, 2, 6-dimethoxyphenol, o-hydroxybenzyl alcohol, eugenol (or 4-ethylguaiacol), salicylic acid, vanillin, cinnamic acid, m-hydroxybenzoic acid, p-hydroxybenzoic acid, gentisic acid, vanillic acid, protocatechuic acid, syringic acid, p-coumaric acid, ferulic acid and caffeic acid.
2-Furoic acid was isolated from the sodium carbonate fraction, as a main component, and this compound possessed no odor in the free state but the ester had characteristic flavor, which seemed to contribute to the roast flavor.
By gas chromatography of the trimethylsilyl derivatives, maltol and isomaltol were detected.

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By extraction of rice vinegar with a mixture of ether and n-pentane (2:1), an aromaconcentrate was obtained in which at least fifty-one components could be detected by means of gas chromatography by comparing their retention times with those of known compounds and thin-layer chromatography.
Alchols positively identified were ethanol, isobutyl alcohol, n-butyl alcohol, isoamyl alcohol, active-amyl alcohol, tetrahydrofurfuryl alcohol, 2, 3-butyleneglycol, phenol, o-cresol, β-phenethyl alcohol and o-nitrophenol. Carbonyl compounds positively identified were acetaldehyde, acetone, diacetyl, acetoin, furfural, phenylacetaldehyde, p-oxybenzaldehyde and vanillin. Esters positively identified were ethyl acetate, isobutyl acetate, isoamyl acetate, ethyl lactate, β-phenethyl acetate, ethyl caprate and p-hydroxybenzoic butylester. Other twenty-five components detected were not yet identified.
The rice vinegar has comparatively large number of volatile components comparing with other brewing materials. This can be attributed to the nature of the fermentable carbohydrate materials in rice used for production of the alcohol, subsequently used for vinegar manufacture. It appears that diacetyl and acetoin are the most important odor in rice vinegar.

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A chromatographic procedure was established to fractionate systematically the constituents of rice vinegar, without any loss of its components, into six main fractions, i.e., volatile basic, acidic and neutral fractions and non-volatile basic, acidic and neutral fractions by flush evaporation and ion-exchange resins. 17% of the total nitrogen of rice vinegar was found in the amine fraction and this suggested a possible role of some organic acid-base complexes in the taste of rice vinegar. Using gas chromatography, the amine fraction could not be separated further to its components because of low ionization. But, using thin-layer chromatography, the fraction was separated into nine spots which comprised such components as putrescine, cadaverine, histamine, methylamine, ethylamine, tyramine, phenylethylamine and two unknown compounds. Putrescine and cadaverine were crystallized.

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By making irradiation of ultraviolet ray on Asp. niger, 57 mutants were obtained and the proteolytic systems of these mutants were studied in comparison with that of the parent strain. The molds were grown on wheat bran and the culture extracts were made to act on milk casein for determining protease activities according to the modified Anson's method. The productivity of acid protease tended to decrease with mutation, and mutants were generally inferior to the parent in the acid protease activity. Most of the mutants were similar to the parent in that they had proteolytic systems that were principally composed of acid protease. But two of the mutants had considerably different proteolytic systems and consisted predominantly of alkaline protease. Both of these two mutants were belonging to Albino type.

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Effect of heat upon bovine skimmilk protein, whole casein and whey protein was studied by means of polyacrylamide-gel electrophoresis. Electrophoresis of casein was carried out in 7% acrylamide and 4.5M urea at pH 9.1 (tris-EDTA buffer) and that of whey protein in 8% acrylamide at pH 8.6 (borate buffer). Casein and whey protein solution, dialyzed against pH 7.0, 0.02M phosphate buffer, and skimmilk were heated for 20 minutes at 70, 80, 90, 100 and 120°C.
Heating casein at 120°C for 20 minutes caused definite changes in gel-electrophoretogram. Along with spreading zone, smaller and diffused α_s- and β-casein zone were observed. The gel-electrophoretograms of heated whey protein showed the increase of unmigrating protein and long diffused zone instead of clear β-lactoglobulin and α-lactalbumin zone. Marked increase of unmigrating protein was observed with heated skimmilk compared with heated casein or whey protein. This is probably due to the effect of salts and sugars existing in milk or due to the interaction between casein and whey protein during heating. Heat-aggregated whey protein, which remained in the starting slot, could be migrated into the gel and showed a new zone between α_s- and β-casein with the presence of urea and 2-mercaptoethanol.

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A threonine-auxotroph of Escherichia coli was found to accumulate a significant amount of an amino acid in the culture broth. The amino acid was isolated in crystalline form from the culture broth and was identified as L-homoserine. The cultural conditions of L-homoserine production by the organism was investigated. The concentration of L-threonine in the medium influenced remarkably on the accumulation of L-homoserine by the organism. L-Homoserine accumulation was maximum at a suboptimum concentration, about 300 micrograms of L-threonine per milliliter, for the growth of the organism. The addtion of L-isoleucine or α-aminobutyric acid to the medium stimulated L-homoserine accumulation. About 4 milligrams of L-homoserine per milliliter accumulated with the medium containing glycerol (5%).

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Riboflavin glucoside-like substance was found to be easily formed from maltose and riboflavin by the action of enzymes prepared from the grains of glutinous indian corn, glutinous millet, glutinous rice and potato. It was also revealed that various kinds of riboflavin glycoside-like substances were synthesized from riboflavin and sugars other than maltose.

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