In the previous paper an increase of lactic acid formation was shown when lactic acid bacteria were grown on a milk culture medium containing the aqueous extract of KUKO leaves.
In this paper the stimulation of bacterial growth were attributed to amphoteric sub-stances by the following observations. Active principles could not be extracted by organic solvents such as n-butanol or ethyl acetate at pH values of 9 and 2, and were adsorbed on the columns of Amberlite IR-120 and IR-410. The eluates from the column of Amberlite IR-120 were further fractionated to three fractions-acidic, basic and neutral-using Amberlite IR-4 B and IRC 50. The neutral fraction (containing neutral amino acids) and the basic fraction (containing bases and basic amino acids) showed stimulating effect on acid formation of Lact. acidophilus, Lact. bulgaricus, Lact. casei and Sc. faecalis.

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Non protein nitrogenous substances in the aqueous extract of KUKO leaves were fractionated by means of column chromatography on ion exchange resin, zone electro-phoresis and cellulose column chromatography. The presence of active principles responsi-ble for acid formation of lactic acid bacteria under a certain condition was examined in each fraction.
Twenty two amino acids including a-aminoadipic acid, α-aminopimelic acid, ethanol-amine, and β-alanine were detected. A mixture of these amino acids showed marked influence on acid formation of lactic acid bacteria, but any single amino acid was less effective in the stimulation of growth as compared with the mixture of amino acids. In a homologous series of monoamino dicarboxylic acids the acid formation was stimulated with the increase in the length of carbon chain.
Adenine, guanine, xanthine, adenosine and an unidentified nucleotide were detected and these nucleic acid derivatives stimulated the acid formation by lactic acid bacteria.

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The carbonyl compounds are trace constituents which generally relate to the food flavor and roast flavor. The direct gas chromatography of the 2, 4-DNPH derivatives of carbonyl compounds was investigated. Linear relationship between the logarithmic reten-tion time and the carbon number was found and the structures of unidentified components were inferred from this relation.
The separation of the 2, 4-dinitrophenylhydrazones of oxo-fatty acids methylated with diazomethane was attained by gas chromatography. This method appears to be a good supplementary means for the identification of oxo-fatty acids.
The 2, 4-DNPH's purified by sublimation under reduced pressure were separated by gas chromatography without decomposition. During the process of gas chromatography, the syn-and anti-forms of 2, 4-DNPH's appeared to isomerize into the thermal equilibrium state.

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Volatile carbonyl components of the MUGI-CHA flavor were converted to mono 2, 4-dinitrophenylhydrazones (2, 4-DNPHs) and his 2, 4-DNPHs. The volatile mono 2, 4-DNPHs were subjected to gas chromatography and fourteen peaks were detected. Among those, twelve 2, 4-DNPHs were isolated in crystalline form by using column and thin-layer chromatographies. These compounds were identified by their IR, UV-spectra and mixed melting point test. Compounds identified are methanal, ethanal, propanone, 2-methyl propanal, 2-methylbutanal, 2-butanone, 3-pentanone, 2-methyl-3-pentanone, furfural (syn, anti), and 5-methylfurfural (syn, anti).
2-Methyl-3-pentanone had a characteristic menthol-like flavor, but the dilute solution had sweet flavor which is one of characteristic flavors of roast barley.

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Enzymatic degradation of gentiobiose, a bitter component in the residual mother liquor in the production of D-glucose by acid hydrolysis of starch (hydrol), was studied.
Gentiobiase-producing mould, identified as Aspergillus japonicus SAITO, has been isolated from many wild moulds by using the synthetic medium containing gentiobiose as a sole carbon source. This mould has also been able to grow well in solid bran medium and induce strong gentiobiase activity. Enzyme was purified about 100-fold with a yield of 20% from the extract of bran culture by tannic acid precipitation, ammonium sulfate fractionation, DEAE-Sephadex chromatography, Sephadex G-75 gel filtration and starch block electrophoresis.
Analyses by free boundary electrophoresis and sedimentation showed the presence of almost single component with the mobility of -1.26×10^-5cm².V^-1. sec^-1 (pH 6.0, μ 0.1) and s_{20, w} of 5.5 (pH 6.0, μ 0.1, _??_ 0.75).
Optimal pH and temperature of the enzyme reaction were 4.0 to 4.5 and 55 to 60°C, respectively. The enzyme was stable at pH 2 to 6 (40°C, 24 hr) and inactivated when heated for 15 min over 50 to 55°C at pH 4.5.
The enzyme was found to be able to hydrolyze gentiobiose, cellobiose, laminaribiose, salicin and p-nitrophenyl β-glucoside with K_m, of 1.4, 1.5, 2.3, 5.4 and 1.35×10^-3M, respectively.

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Kinetics of gentiobiase activity as a function of temperature, the effects of inhibitor and the mode of action were investigated using the purified β-glucosidase preparation from the bran cultures of Aspergillus japonicus SAITO.
The temperature coefficient of gentiobiase reaction between 2 to 40°C was about 2.2 with the apparent activation energy of 14, 300 cal/mole.
The rate of heat inactivation is characterized by the apparent activation energies of 20, 400 cal/mole below, and 53, 300 cal/mole above the critical temperature of about 68°C.
The enzyme reaction was remarkably inhibited by MnO₄^- and heavy metals, such as Cu²⁺, and Hg²⁺ and Ag⁺; but Mn²⁺, Zn²⁺, Co⁺, Fe²⁺ and Fe³⁺ were less inhibitory. Molybdate and o-phenanthroline were to some degree activatory. Sulfhydryl reagents were not effective.
The enzyme action on gentiobiose of various concentrations ranging from 0.25 to 10% were examined by the chromatographic technique. At higher concentrations of the sugar, transfer action in addition to hydrolytic action, was recognized and oligosaccharides of gentiobiose series, mainly gentiotriose and small amounts of 6²-β-glucosyl-cellobiose, cellobiose, and laminaribiose were detected. On the other hand, the increases of enzyme concentration or reaction time, even in the case that the gentiobiose concentrations were higher, resulted in reduction or disappearance of these reversion products. With a mixture of gentiobiose and glucose, no reversion products were detected, even in higher concentrations of gentiobiose.
The action on glucose of various concentrations ranging from 2.5 to 50% resulted in the formation of reversion products with the increase in concentration of glucose. At 5% and over, gentiobiose was mainly recognized, while after a long incubation, gentiotriose was also detected in addition to gentiobiose, but no other oligosaccharides were observed.
The equilibrium between glucose and gentiobiose was investigated using glucose as a substrate. The equilibrium constant was 2.19×10^-2 at 10% of glucose and 4.3×10^-2 at 20%.

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Sake yeast (Saccharomyces cerevisiae) and Micrococcus sp., both of which were isolated from the mash of Sake brewing, were cultured together in media containing various concentrations of vitamins (niacin, pantothenic acid, biotin, thiamine and inositol).
It was shown that, at the lower concentrations of the vitamins, growth of both organisms increased in proportion to vitamin concentrations, and at moderate concentrations, Micrococcus sp. repressed the growth of the yeast. When the concentrations of vitamins was high, yeast grew better than Micrococcus sp.

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Growth of Saccharomyces cerevisiae and Micrococcus sp. in mixed cultures was examined with special reference to inositol concentrations added to the media. At higher concentrations of inositol (1μg/ml), growth of the yeast was more abundant than that of Micrococcus sp. The reverse was true at lower concentrations of inositol (0.5μg/ml). When these organisms were grown individually, yeast growth was stimulated by inositol, but alonger lag time was observed in Micrococcus. Utilization of inositol occurred more rapidly in pure culture of the yeast than in mixed cultures. The result indicates that yeast cells in mixed culture incorporates inositol more slowly than the cells in pure cultures.
The antagonistic relationship during the growth of both organisms in mixed cultures was observed at definite cell concentrations. They grew independently until the product of cell concentrations of both organisms became about 10¹³ (cells/ml)², and when the product reaches to this value Micrococcus sp. began to repress growth of the yeast.

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Streptonsyces caespitosus excreted not only protease but also α-amino acylase in its submerged culture broth. Several production conditions and enzymatic properties of the acylase were investigated. The acylase was purified 50-fold by ammonium-sulfate precipitation method and chromatography on columns of Amberlite CG-50 and Duolite A-6. The maximum activity of the acylase was obtained at pH 7.0_??_8.0 and 60°C, and no metal ion activated the enzyme activity. The enzyme was stable over the range of pH 4.0 to 8.5 at 20°C, and the activity was not inhibited by the addition of metal ions and EDTA. The acylase was stable below 60°C, but lost the activity at 70°C in 10min. The enzyme had lower substrate specificity than that of molds, bacteria and animal tissues. It was effective toward the α-N-acyl-L-amino acids with aromatic and aliphatic side chains.

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As one of the fundamental experiments on the sunlight off-flavor of foods, photo-sensitized degradation of amino acids with riboflavine in the atmosphere of air, O₂, and N₂ under visible light was investigated. Amino acids were oxidatively deaminized and decarboxylated to the corresponding aldehydes. Methionine was the most photolabile and gave almost exclusively methional. Degradation of methionine depended on the concentration of riboflavine and O₂, and was maximum at pH 7. One molecule of riboflavine converted 42 molecules of methionine into methional by the irradiation with 500W visible light for an hour. Among amino acids, valine, leucine, isoleucine, phenylalanine, methionine, tryptophan, and cysteine produced strong special odors. It is suggested that these amino acids may be responsible for the sunlight off-flavors of foods.

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The present paper reports on the effective application of the ternary synergism. Results were as follows.
(1) When two or more kinds of α-amino acids are added to the medium containing monosodium L-glutamate and paratable 5'-nucleotides, a distinctive improvement of the palatable taste is recognized. In case of two or more kinds of α-amino acids, the ternary synergism occurs as algebraic sum of the activation.
(2) Glutamic and aspartic acids in the seasoning or food can be employed as the substitute of monosodium L-glutamate and monosodium L-aspartate.
(3) As the substitute of the palatable 5'-nucleotides, the compounds obtained by the hydrolysis of ribonucleic acid with 5'-phosphodiesterase or by the sinthesis, or the natural substance such as dried sardine, dried bonito and the like, are usable.
(4) By the application of the ternary synergism, a lot of new application of α-amino acids, such as the development of new product, quality improvement of food (seasoning, beverage), and cost reduction, became possible.

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Body fat of the two scale insects, C. rubens and C. japonicus, was studied. After extraction, the triglycerides were isolated by preparative thinlayer chromatography. Composition of the triglycerides was determined by a pancreatic lipase hydrolysis and a gasliquid chromatographic analysis. A similar distribution pattern of triglycerides was found in these two insects. The completely saturated type (S₃) consisted of the most part of total triglycerides; 75.5 mole % in the rubens fat and 83.5 mole % in the japonicus fat. It was noted that capric and lauric acids were the two major components in these triglycerides. In the unsaturated triglycerides, only S₂U type was significant in amount.
The data may suggest that the body fats of Ceroplastes genus consisted mainly of medium-chain triglycerides containing a large amounts of capric and lauric acids.

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