Masao Kimata

Effects of Various Organic Acids on the Bacterial Decomposition of Extractive Matter of Meat of Scomber japonicus (HOUTTUYN)

As for the bactericidal action of various acids, studies hitherto reported show that the action is principally due to the hydrogen ion and is generally, with exceptions with certain acids, not very much due to the undissociated molecules or to the anions. So far as I am awere, however, little is known of the effects of acid on the physiological activity of bacteria, such as to decompose nitrogenous compounds. In the present paper are embodied the observations on the effects of Na-salts of several organic acids in varied concentration upon the bacterial decomposition of extractive matter of meat of Scomber japonicus (HOUTTUYN). The results may be summarized as follows: - 1. Na-salts of the organic acids have effect on the bacterial decomposition, They retard the beginning of decomposition, increase the velocity, but limit the final rate of decomposi-tion. 2. With mono-basic fatty acids, the greater the molecular weight, the effects are ob-served the lower consentration. But, as the degree of dissolution has certain limit for those acids, of which molecular weight is greater than lauric acid, the effects are indifferent for concentrations of such acids over this limit. 3. So far as the present study is concerned, the effects in question are the greatest for mono-basic fatty acids, next for di-basic fatty acids, and the least for hydroxy-fatty acids.

Bacterial Decomposition of Extractive Matter of Meat of Aquatic Animals

So far as I am aware, little is known of the relation of the age of an animal to the rate and the velocity of the bacterial decomposition of it. According to HUNTER, (1) in salmon, which are actively feeding when caught, enzymic decomposition as well as bacterial decomposition may take place. It has been found that immature salmon caught on the feeding ground, with alimentary tract full of feed, is decomposed in a manner different from the mature salmon caught on the spawning migration after the fish have ceased feeding, and when the alimentary tract is empty. In the immature feeding salmon enzymic decomposition accompanies bacterial spoilage, causing a softening of the belly wall and disintegration of the viscera of the fish. In mature salmon caught on the spawning migration, the alimentary tract is sterile since no food is present and invasion by bacteria in such fish is solely from the outside inward. After the salmon have reached the spawning grounds, the type of spoilage again differs from that of the mature migratory salmon even though the bacterial flora in both are almost identical.(2) The chemical composition of the meat may vary with time in the life-cycle of the animal. Therefore, it seems that the rate and the velocity of the bacterial decomposition depend also upon the time in the life-cycle of the animal at which it is caught. The present paper embodies my observations on the rates and the velocities of the bacterial decomposition of extractive matter of meat of aquatic animals of different ages, and on the growth rate of bacteria in the solution of the extractive matter.

The Relation between the Growth of Bacteria and the Agar-concentration of the Medium

It is well known that the chemical properties of the media play an important part on the bacterial growth. On the contrary, our knowledge of relationship between the bacterial growth and the physical properties of the media is rather scanty, only a few papers having been published on such items as surface tension, osmotic pressure, and what not. TAKJTA1) has found abundant growths of Vibrio cholerae, tubercle bacilli etc., when the viscosity of media is of certain degrees. Semi-solid media have been successfully used by several authors for some kinds of bacteria2, 3, 4, 5) In the present experiment, I have studied the effect of the density of the medium upon the growth of bacteria, using various concentrations of agar.

On the Bacteria from Decomposing “Kamaboko”

The Japanese food matter called “Kamaboko” is prepared by steaming fish paste which is seasoned with salt, sugar and “mirin.”(1) It produces some mucous matter on its surface when left in a non-antiseptic condition. Our study on the bacteria in this mucous matter resulted to obtain 14 species from 45 cultures. Of these, 15 cultures were found referable to Micrococcus, 2 to Rhodococcus, 9 to Leuconostoc, 11 to Bacillus, 4 to Achromobacter, 4 to Flavobacterium. Among them those of Bacillaceae group were found to be motile, but Bacteriaceae group to be non-motile, and the following 6 species appear to us to be new to science. 1. Micrococcus gilvus n. sp. It resembles Micrococcus ochraceus. However, it differs in the following characters. Nitrates: reduced to nitrites; Broth: slight turbid with sediment (viscid), no pellicle; In a broth medium containing sugar, growth was abundant. 2. Micrococcus solidus n. sp. It possesses all the characteristics of the genus Micrococcus as given by BERGEY (1930). But it is quite different from any species of this genus which does not undergo chromogenesis but liquefies gelatin. Characterisation runs as follows: - Morphology: -Spheres (1.0 ?? 1.3μ dia.), occurring in irregular masses. Gram-positive. Nonmotile. Cultural features: -Agar colonies: irregular, dull, flat, opaque, white, lobate or auriculate, granular; Gelatin colonies: concave, cloudy, white, lobate, crateriform liquefaction; Gelatin stab: stratiform liquefaction; Potato: graysh-white, dull, dry; Broth: turbid with pellicle and scanty sediment (viscid). Physiological characters: -Litmus milk: acid, coagulation, peptonization, litmus reduced; Indol: not formed; Nitrates: reduced to nitrites; Fermentations: acid was produced from dextrose, sucrose, lactose, mannit, glycerine, salicin, raffinose; Starch not hydrolyzed. Opt. temp. 27°C. 3. Bacillus canus n. sp. It resembles Bacillus clostroides, differs in the following particulars. Potato: graysh-white, rugose; Broth: slight turbid with slight pellicle; In a broth medium with sugar growth was abundant. 4. Bacillus catenatus n. sp. It possesses all the characteristics of the genus Bacillus as given by BERGEY (1930), but not referable to any species of it. Characterisation is as follows: - Morphology: -Rods (0.8 ?? 1.0×3.5 ?? 7.0μ), occurring in long chains, peritrichous flagella; Gram-positive; Spores (0.8×1.8μ), central. Cultural features: -Agar colonies: curled or irregular, rough, flat or raised, gray, auriculate, curled or granular (internal structure); Agar stroke: echinulate or spreading, dull, rugose, flat or raised, opaque, gray to brown; Gelatin stab: liquefaction was slowly (napiform ?); Potato: dark-orange-white to brown, dull, dry, medium darkened; Broth: slight turbid with slight pellicle. Physiological characters: -Litmus milk: alkaline, peptonized; Indol: not formed ; Nitrates: reduced to nitrites; Fermentation: acid was produced from dextrose and sucrose: Starch not hydrolyzed. Opt. temp. 35°C. 5. Achromobacter immotum n. sp. It possesses all the characteristics of the genus Achromobacter as given by BERGEY (1930), but differing from any one of those species genus Achromobacter which do not liquefy gelatin and are non-motile. Characterisation is as follows: - Morphology: -Rods (0.8 ?? 0.9×1.8 ?? 4.0μ), with rounded ends occurring singly, in pairs and in short chains. Non-motile.

The Effect of Protein on the Thermal Death Rates of Bacteria

LANGE(1), BEHRENS(2) and HUCKEL(3) heated several kinds of bacteria to death and added them to suspensions of the living bacteria. Then they heated the mixtures and found that the admixture of the dead bacteria prolongs the thermal death time of the bacteria tested. A hypothesis was put forth by them to the effect that the added dead bacteria perhaps prevents the living ones from dying when heated. It was shown by HUCKEL that this protective activity does not depend on the kinds of the bacteria used. It would not be without interest, therefore, to ascertain whether or not the protection in question is due to protein of any kind of organism. One mg. of the bacteria cultivated for 48 hours at 28°C on agar stroke were put into l0c.c of the sterilized buffer solutions (pH=6.9) which contain 50mg., 100mg. or 200mg. of gelatin or the cells of 2 strokes killed by heating, and were shaken carefully, then put into ampullae (capacity, ca. 0.2c.c.; dia., ca. 3mm.), and were sealed. After heating at various temperatures they were cooled in water. The ampullae were disinfected with alcohol and the ends of them were cut off. Their contents were pipetted in the agar media with sterilized capillary, and the agar plate culture by the shaking method was made. The results obtained are shown in the table 1. From the table it is clear that the addition of 1-2% of gelatin prolongs about 2 times the thermal death time of the bacteria tested.

Decomposition of Betain by Pscudomonas fluorescens

Trimethylamin has been known to be produced from cholin and lecithin in the putrefactive pocess. It was thought that betain is similar to these compounds in construction and might produce trimethylamin as a decomposed product. Therefore, I have studied the decomposition of betain, which is contained in the muscle of cuttle-fish in great quantity, by the action of Pseudomonas fluorescens. In the first place, I made a synthetic medium which is free from the nitrogen- and carboncompounds, consisting of 500c.c. of water, 1gr. of K2HPO4, 0.5gr. of MgSO4, 0.5gr. of NaCl and trace of FeSO4 and Ca3(PO4)2. This p-oved to be suited for the growth of Pseudomonas fluorescens. In the second place, to 500c.c. of this synthetic medium 1 gr. of betain hydrochloride was added and neutralized with N NaOH. In this medium was cultivated for 3 weeks at 28°C. Pseudomonas fluorescens. Then MgO was addel to the culture which was afterwards distilled by means of the steam distillation. The volatile alkali obtained in the distillate were determined by the WEBER and WILSONs method(2), and the WQODWARD and ALSBERGs method(3). From the results of the experiment, it is clear that trimethylamin was not produced from betain, but ammonia was formed from it by the action of Pseudomonas fluorescens. Moreover organic volatile acids, alcohol and amino acids were not found as the decomposition products.

A Nephelometric Method for the Determination of Small Amount of Chlorine

It is customary to determine chlorine by the titration with silver nitrate, but it is almost impossible to find out the element in the tissues of organisms or in culture media by this method. The present paper embodies our determination of small amount of chlorine by a nephelometric method, chlorine being measured as silver chloride hereby. Experiment 1 A varying amount of N/50000 NaCl solution was first acidified with HNO3, 0.2 c.c. of 10% AgNO3 solution was added, and finally distilled water was put to make the mixture 30 c.c. in volume. The chlorine content of this series of the mixtures was determined by a nephelometer in reference to a standard solution which consists of 20 c.c. of N/50000 NaCl solution, 0.2 c.c. of 10% AgNO3, solution, and 3 c.c. of HNO3. It may be mentioned that I c.c. of N/50000 NaCl solution contains 0.0007 mgr. of Cl. It is clear from the experiment that chlorine was almost exactly determined in the series within the range from 0.008 to 0.018mgr., the experimental error being ±0.001mgr. Experiment 2 Firstly, 1 c.c. of a broth diluted to ten times was made 100 c.c. in volume with distilled water. Secondly, 1 c.c. of the broth diluted to ten times was mixed with 5 c.c. of N/2000 NaCl solution and then distilled water was added until 100 c.c. Two c.c. and 3 c.c. of each of these two solutions were acidified with HNO3, mixed with 0.2 c.c. of 10% AgNO3 solution, and made 30 c.c. with distilled water. The nephelometric determination of the samples was done with the satisfactory results

On the Optimum pH for Mold-Lecithase

AKAMATSU found by the estimation of the increased acidity and of the inorganic phosphoric acid that Taka-phosphatase (which is contained in Taka-diastase from Aspergillus oryzae) decomposes not only the synthetical glycerin-phosphoric acid, but also the glycerin-phosphoric acid which is a decomposed product from lecithin. One of the authors measured cholin, a decomposition product of lecithin by lecithase, and inorganic phosphoric acid, a decomposition product by phosphatase, and found that the estimation of cholin is a very convenient method for the study of lecithase. He observed that the degree of injurious effect of ultraviolet ray on the formation of inorganic phosphoric acid is different from that on the formation of cholin from lecithin by Taka-diastase, the former being the highest at pH 6.6. After Pincussens reports, the injurious effect of ultraviolet ray upon the enzyme is the greatest at the optimum pH for the enzyme. Therefore, we suspected that the optimum pH for lecithase may be different from that of phosphatase. In the following experiments, we have determined the optimum pH for lecithase by the measurement of cholin. For the estimation of cholin freed, we have used the following method: We added a small quantity of magnesium chloride solution and that of ammonia to the reaction mixture, filtered off the precipitates of magnesium hydroxide which includes the nondecomposing lecithin, added a iod-iodkali solution to the filtrate, separated the precipitate of cholin-periodide and solved in chloroform, and finally titrated freed iodine with N/100 thiosulphate solution. 1c.c. of N/100 Na2S2O3=0.14mgr. of cholin. To the lecithin-methylalcohol solution-0.5gr. of lecithin was solved in 5 c.c. of methylalcohol-40 c.c. of M/15 phosphate buffer solution and 20 c.c. of 5% Taka-diastase solution were added, and then the flask was filled to 100 c.c. with distilled water. After standing for a definite time at 36°-37°C, 10 c.c. of the reaction mixture was taken out, was filled to 11 c.c. with magnesium chloride solution and ammonia, and was filtered. Two c.c. of its filtrate was neutralised with hydrochloric acid, was added 0.2 c.c. of iod-iodkali solution, then cholin-periodide was collected by centrifuging, washed with the saturated NaCl solution at 0°C and was solved in chloroform. The iodine freed was titrated with N/100 Na2S2O3 solution. From the result of the experiment we see that the optimum pH-value for lecithase is near 7.0 (Table I.). Next, from the data we calculated the velocity-constant by using monomolecular equation and showed that the reaction velocity obeys the monomolecular reaction within 3 hours (Table II).