1. The phenomenon of entangled fermentation previously found and expressed as follows: glucose+2 pyruvate→2 lactate+2 CO2+acetoin was reinvestigated in L. plantarum, and the above equation was confirmed. 2. Entangled fermentation was further studied by tracer techniques. All of the fermentation products, lactate, CO2, and acetoin were found to derive one half of their carbon atoms from glucose and the other half from pyruvate. 3. Periodate degradation of the acetoin molecule showed that the acetyl moiety of the molecule originates from pyruvate and the methyl carbinol moiety from glucose.
STUDIES ON LIPASE
II. HYDROLYTIC AND ESTERIFYING ACTIONS OF CRYSTALLINE LIPASE OF ASPERGILLUS NIGER
The hydrolytic and esterifying actions of lipase have been studied using the crystalline electrophoretically homogeneous enzyme of Aspergillus niger. The enzyme hydrolyzed vegetable oils, triglycerides of long or short chain fatty acids, and alkyl esters of long chain fatty acids, but not alkyl esters of short chain fatty acids. The hydrolytic action was found to depend on the chain length of either fatty acid or alcohol moiety of the substrate, and the maximal activities were found with fatty acids of eight and sixteen carbons using single esters as the substrate. The enzyme catalyzed the synthesis of the glycerides and the single esters of fatty acids, and the activity seemed to be influenced by the structure of the substrate. The esterification of oleic acid with glycerol was achieved up to 60% when the concentration of water in the reaction mixture was kept as low as possible. The reversibility of the reaction, fatty acid+alcohol (including glycerol) † ester (including glyceride)+water, was confirmed by the fact that the changes in the concentration of free oleic acid formed (or consumed) during the reaction was related to those of water and glycerol contents in the reaction mixture.
1. The utilization of various organic acids as sole source of carbon by M. glutamicus was studied. Organic acids examined are classified into three groups, namely (1) those which can be assimilated when the media contain biotin as sole growth factor, (2) those which can be assimilated when some auxiliary nutrients are added besides biotin, and (3) those which can not be utilized at all by the organism even with added auxiliary nutrients. 2. The culture conditions for the L-glutamic acid production from acetate were also studied. It was observed that a considerable amount of L-glutamic acid was produced in a medium having biotin as sole growth factor. 3. The culture conditions for the L-glutamic acid production from citrate were studied. An appreciable amount of L-glutamic acid was produced in a medium containing biotin and corn steep liquor. 4. Relationship between the concentration of biotin in the medium and the L-glutamic acid production was shown to be similar to the case of L-glutamic acid production from glucose. 5. Metabolic activities of the cells grown on glucose, citrate and acetate were studied. The yield of L-glutamic acid by these cells is discussed in correlation with oxidation of the substrates.
In order to clarify the mechanism of racemic lactate formation in Lactobacillus plantarum, the intracellular activities of NAD-linked D- and L-lactate dehydrogenases were estimated. Both enzymes were separated from each other and purified about 90-fold by ammonium sulfate fractionation and DEAE cellulose chromatography. Basal properties of these purified enzymes were examined. These enzymes were distinguishable by their substrate specificity, heat stability and oxamate inhibition. From the activities of both enzymes in a sonic extract, their respective intracellular activities were calculated by correcting these values for the intracellular conditions which have been reported previously. It was found that both of the enzymes operated with equal activity in glucose-metabolizing washed cells.
A method of the determination of L-lactic acid is described. A dried cells or an acetone powder from Lactobacillus saké GO1 in the presence of methylene blue oxidized L-lactic acid quantitatively and specifically. Oxygen consumption in a Warburg apparatus was used to determine the amount of L-lactic acid present in the sample.
The nature of the B12-binding principle existing in the wall of Lactobacillusdelbrueckii No. 1 was investigated. Pepsin destroys the B12-binding activity of the wall of this organism. On the other hand, when B12 has been bound to the wall, pepsin is ineffective. This situation is explained as follows: B12 binds to the same site where pepsin acts and that once bound B12 hinders sterically the action of the enzyme at the reactive center of the principle. Solutions of 0.2N HCl are suggested to cleave the principle at a closer position to the basal structure of the wall than the site upon which pepsin attacks.
(1) Magnetic susceptibility was measured of intact resting, metabolizing and heat-treated yeast cells in suspension. The accuracy of measurement was ±0.3%. (2) A susceptibility change of 4% as obtained by BAUER and RASKIN upon death of the cells was not observed. (3) The remarkable change in susceptibility given by BAUER and RASKIN seems to be erroneous. The authors are indebted to Miss K. NUNOMURA for her technical assistance.
DNA was extracted from dormant and germinating (1.5hr culture) conidia of Aspergillus oryzae. Cell disintegration was effected by sonic oscillation in the presence of glass beads, since the conidia were resistant to mild treatment with various lytic enzymes. The sonicate was subjected to extraction of DNA by the phenol method. The resultant crude DNA was treated with ribonucleases (RNase I and RNase T1) and further with isopropanol, and then subjected to zone-electrophoresis using Geon as supporting material: the latter procedure was effective for removing some contaminants contained in a large amounts. Purified DNA samples obtained from dormant and germinating conidia contained about 40% of polyphosphate, but gave only a single peak by sedimentation analysis. It was found that the base composition of the conidia DNA changed only slightly at early germination stages.
The photosynthetic activity as measured by the light fixation of 14CO2 by intact cells has been shown to be a measure of viability of microalgae. This method can be conveniently employed to examine the cell viability change in non-growing systems. Comparative studies on heat and UV sensitivities of various algae were carried out using this method.
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Edited and published by : Applied Microbiology, Molecular and Cellular Biosciences Research Foundation/Center for Academic Publications Japan Produced and listed by : TERRAPUB, Center for Academic Publications Japan/Shobi Printing Co., Ltd. (-Vol.60,No12), Center for Academic Publications Japan/InternationalAcademic Printing Co., Ltd.(-Vol.54,No1)