Bromocasein was prepared in accordance with that for iodocasein and free bromine was eliminated by electrodialysis. The bromine content of bromocasein was 4.0-4.2%. This bromocasein also showed acceleration of tadpole metamorphosis but in a degree far weaker than that of iodocasein.
The diazo reagent was prepared by the application of 1cc. of 2% sodium nitrite solution to 1cc. hydrochloric acid solution containing 0.5% 2-nitro-4-methoxyaniline, and after complete decomposition of excess nitrous acid with ammonium sulfaminate, the total volume was brought to 100cc. with 2 N sodium acetate. To 5cc. of 2% metaphosphoric acid solution of vitamin C (containing 5-22γ of vitamin C in each cc.), 3cc. of this diazo reagent was added, alkalized with addition of 2cc. of 10% sodium hydroxide, and the bluish violet coloration (λmax 570mμ) thereby formed was optically measured for the determination of vitamin C. The absorption spectral values are λmax 570mμ and E1%1cm 447.
Thiophene has recently become important as the raw material for the manufacture of analgesics and antihistamines. Attempts were made for the synthesis of thiophene from butyne-2-diol-1, 4 (I), obtained easily from acetylene and formaldehyde by the ethinylation. Hydrogenation of (I). gave butane-1, 4-diol (II) whose dehydrogenation with sulfuric acid catalyst at 150° and with alumina catalyst at 250° gave tetrahydrofuran (III). Reaction of (III) with hydrogen sulfide at 300°, in the presence of Al2O3, Al2O3-Cr2O3, or Al2O3-ThO2 as a catalyst, yielded thiolane (IV) whose dehydrogenation at 500° with Al2O3-Cr2O3 catalyst finally gave thiophene, the total yield being 27.8% calcutated from (I).
The previous paper described that the cause of the mass deaths of over 100 heads of milking cows in Kobe, Osaka, and Mie pref. was due to the toxic substance produced by the metabolism of a fungus, one of the Penicillium sp., which had adhered to the dried malt feed. It was observed that this toxic substance is produced only in a medium containing both sugar and proteins. This toxic substance is extracted and purified by ether, corresponds to formula C7H6O4, and its potency is 0.5mg./20g. mouse by intraperitoneal injection. Further investigations revealed that this toxic substance is identical with patulin, clavacin, claviformin, expansine, and penicidin, reported by Raistrick, Engel, Woodward, et al. The chemical structure proposed by Woodward, et al. seems to need further examination because of the infrared spectral values and the lack of optical activity.
Anthelmintic action of degenic acid, the active principle, sodium mannosidoglycerate, and volatile oil of Digenea simplex Ag., was examined pharmacologically. Digenic acid was found to accelerate the kinetic motion of Allolobophora foetida nerve muscles, in extremely smaller amount than that of 85% methanolic extract of Digenea, and also to inhibit the dehydrogenase in the muscle of hog ascaris. The volatile oil was also found to have similar action, although far weaker than that of digenic acid. Sodium mannosidoglycerate possessed no such action of suppressing the dehydrogenase but paralyzed automatic motion of A. foetida nerve muscles. Oral administration of these substances in puppy showed approximately parallel anthelmintic effect as that in in vitro tests. Parenteral administration of digenic acid in puppy was found to give a rather strong side-effects. Toxicity in mouse, LD50, was 0.024mg./g. by subcutaneous injection and 0.12mg./g. by oral administration.
Effects of various antibiotics, sulfa drugs, and tuberculosis chemotherapeutics against the growth of Streptococcus faecalis were examined. All the antibiotics tested possessed a marked inhibitive action, but the action of the other two kinds gave varied results, mostly showing accelerating action. On the other hand, it was found that penicillin and homosulfanilamide, which possessed a marked growth inhibitive action, developed a certain amount of resistance by repeated culture of these bacteria.
Effects of various anthelmintics, adsorbants, antiseptics, and numerous other pharmaceutics, that can be administered orally, on the growth of Streptococcus faecalis were examined. Tetrachloroethylene, an anthelmintic, was able to arrest the growth completely while a marked growth inhibition was observed in mercuric oxycyanide. None of the others showed any marked influence.
Titrations of antipyrine, aminopyrine, sulpyrine, antipyrine salicylate, migrenin, and pyrabital in glacial acetic acid with 0.1N perchloric acid were carried out. With the exception of sulpyrine, the determinations were found possible with an accuracy of 0.05-0.2%. Antipyrine was titrated one mole as one equivalent. Aminopyrine was titrated first one mole as one equivalent but could not be determined at the next neutralization point. Sulpyrine was titrated first one mole as two equivalents and the next neutralization point was obscure. The basicity of aminopyrine is stronger than that of antipyrine or sulpyrine. Antipyrine salicylate was titrated as in the case of antipyrine alone since salicylic acid does not show its acidity in the glacial acetic acid, as is the case with citric acid and barbital. Migrenin can be titrated at the same neutralization point as the mixture of antipyrine and caffeine it contains, as long as their ratio is known. Pyrabital can also be titrated as its componental aminopyrine and the determination values become varied when the mixture ratio of componental aminopyrine and barbital is varied. The titration was possible when glass-calomel electrodes were used but the use of an indicator sometimes necessitated the use of color comparators.
From the alkaline degradation products of Terramycin, Kuhn and Dury isolated a phenolic ketocarboxylic acid and confirmed its structure as 6-acetylsalicylic acid by its conversion to methyl 2-methoxy-6-acetylbenzoate. However, the synthesis of 6-acetylsalicylic acid has not been reported. The authors synthesized this compound through the route described in Table I. From the study of infrared absorption spectra of 6-acetylsalicylic acid and its derivatives, it was found that these compounds exhibit a ketolactol tautomerism represented by the general formulae: The results obtained are summarised in Tables II and III.
In the course of synthetic studies on degradation products of Terramycin, a series of amino- or hydroxy-substituted derivatives of 3-methylphthalide were prepared and their antibacterial effect was assayed against Staphylococcus aureus, Eberthella typhosa, Brucella melitensis, Vibrio cholerae, Shigella dysenteriae, B. C. G., and Mycobacterium tuberculosis typus hominus. The compounds prepared are presented in the accompanying table together with data on their antimicrobial activities.
1) Presence of digitalinum verum in the water-soluble glycosides obtained from the dried leaves of Digitalis purpurea was confirmed by a method different from that given in the previous paper. 2) For comparison, digitalinum verum was prepared from the seeds of Digitalis purpurea, and in this case the substance was obtained in crystalline form for the first time so far as is known.
1) Canned crab meat showed toxicity 12-18 hours after the can has been opened and left to stand in the air. 2) The toxicity is such that it causes internal hemorrhage in mouse by subcutaneous injection or causes death when rather strong (cf. Table I). 3) This toxin is not characteristic to crab meat but is caused by the increased growth of nonpathogenic, aerobic Bacillus subtilis and Bacillus cereus.
1) It has been proved that nonpathogenic putrefying bacteria, Bacillus subtilis and B. cereus, acquire transitory toxicity (cf. Tables I, II, and III). 2) Examination of the toxicity of these bacilli by parenteral administration of its shaking culture at every hour showed that its toxification occured in the latter logarithmic period of its growth curve (Table IV). 3) The toxified bacilli not only showed toxicity against mouse but also caused violent diarrhoae in kittens by oral administration (Table V). 4) This toxic bacillary component was composed of lethal and hemolytic toxins, the lethal component being quite unstable but the hemolytic component somewhat stable (Tables VI and VII).
Volatile biogenic amines, such as monomethylamine, dimethylamine, isopropylamine, ω-phenylethylamine, cadaverine, and piperidine, were submitted to paper electrophoresis and colored by ninhydrin by which these volatile amines were separately detected within a short time. The method was utilized for the examination of volatile amines excreted into urine and it was found that the majority of the amines were monomethylamine and dimethylamine, and only an extremely small amount of other amines. Piperidine was also detected from urine by this method.
Paper electrophoresis was utilized for the detection of substances giving positive Jaffé reaction, and creatine and creatinine were separately detected by simple procedures. This method was applied for the urine analysis of normal persons in pregnancy, and in patients suffering from Banti's disease, uterine carcinoma, pernecious anemia, cachexia, and chlio epithelioma. Urine from such persons contained a fair amount of substance giving positive Jaffé reaction, besides creatine and creatinine, and the tendency was especially marked in the case with pernecious anemia. The separation of creatine and glycothiamine, and of creatinine and glycothiamidine was found difficult.
Substances giving positive Pauly's diazo reaction in urine were examined by paper electrophoresis in combination with fractionation by ion exchangers. Urine contains various substances giving positive diazo reaction, other than histidine and tyrosine, and the migration patterns of such substances in the urine of pregnant women, and of patients from morning sickness, pernecious anemia, and Bunti's disease, are qualitatively different. The amount of urochromogen, tyrosine, and histidine was also different. The urine of pregnant woman used in the present experiment did not show as marked an increase histidine as has been reported. The urine from leucemia patient contained comparatively large amount of p-hydroxybenzoic acid and extremely small amount of p-hydroxyphenylacetic acid. Marked excretion of β-indoleacetic acid was also observed in such a case.
Auraptene, m.p. 68°, obtained as the crystalline component of orange oil and confirmed as 7-hydroxycoumarin geranyl ether, was catalytically reduced with platinum black at ordinary temperature and pressure from which the corresponding phenol, i.e. 7-hydroxycoumarin, was obtained. As a model experiment to confirm this fact, various ethers of hydroxycoumarins were prepared and their catalytic reduction was carried out, by which it was observed that the aromatic ethers possessing β, γ-unsaturated alkyl side chain easily underwent hydrogenolysis to give the corresponding phenols. 7-Hydroxycoumarin was also obtained by pyrolysis of auraptene.
By heating phenol and anisole with free carboxylic acids, in the presence of polyphosphoric acid, for 5-30 minutes in boiling water bath, 4-hydroxy- and 4-methoxyacylophenones were respectively obtained. The present method has made it possible to prepare 4-hydroxyacylophenones quite easily from phenol and carboxylic acids instead of the various indirect methods practiced to date.
Polyphosphoric acid reaction of phenol and dihydric phenols with benzoic, salicylic, p-hydroxybenzoic, or anisic acid resulted in the formation of pyrocatechol monobenzoate or monoanisoate, phloroglucinol tribenzoate, phenyl salicylate, hydroquinone disalicylate or dianisoate, or resorcinol dianisoate. The cold reaction of phenol with p-hydroxybenzoic, anisic, or benzoic acid gave 4-hydroxyacylophenones. Polyphosphoric acid reaction of phenyl anisoate resulted in its facile change to nuclear anisoylated compound. The condensation of phenol and benzoic acid and rearrangement of phenyl benzoate to 4-hydroxybenzophenone were practically impossible.
Leaves and bulb of Allium tuberosum Rottl. (A. odorum L.) and the bulb of leek (A. Porrum L.) were extracted with alcohol and the reaction products of these extracts with thiamine were examined by paper chromatography. The former plant was found to yield allithiamine (IV) and its methyl homolog (VIII), while the latter failed to yield these derivatives. In the case of A. tuberosum, (IV) and (VIII) were obtained as crystals and their formation confirmed.
The reaction products of the alcoholic extracts of following Allium species plants with thiamine were examined by paper chromatography: Onion (Allium Cepa L.), A. fistulosum L., A. schoenoprasum L., A. Bakeri Regel, A. victorialis L. var. platyphyllum Hultén, A. Grayi Regel, and A. Thunbergii Don. It was thereby found that the formation of allithiamine-like substances was practically nil by reaction with the extracts of A. Cepa, A. fistulosum, and A. schoenoprasum, while only the methyl homolog (II) of allithiamine was formed in reaction with A. Bakeri. Allithiamine (I) and (II) were formed in reaction with A. victorialis var. platyphyllum, the substances being isolated as crystals. Paper chromatograms assumed to be those of (I) and (II) were detected in the reaction products from A. Grayi and A. Thunbergii.
Application of equimolar amount of L-cysteine to allithiamine (V) results in its facile reduction to thiamine, and the allylmercapto group thereby liberated combines with L-cysteine to form S-allylmercapto-L-cysteine (VII). The mechanism of biological change of allithiamine (V) to thiamine is assumed to follow such a reaction by the SH-containing compounds, such as cysteine or glutathione, or by proteins present in the body.
The reaction of the quantitative reduction of allithiamine (I) by L-cysteine and concurrent formation of S-allylmercapto-L-cysteine also occur with other allithiamine homologs. Paper chromatography of S-alkylmercapto-L-cysteine thereby formed showed that chromatographic images with different Rf values are obtained by different alkyl groups present. This paper chromatography was utilized for the isolation and detection of allithiamine and its homologs in order to reëxamine the reaction products of the alcoholic extract of various Allium species plants with thiamine. It was thereby found that the allithiamine homologs formed were composed chiefly of allithiamine (I) and its methyl homolog (V), and a very minute amount of the formation of the propyl homolog (IX) was also detected.
Allithiamine (III) is formed by the reaction of thiamine (XIII) and alicin (II), which is said to be the secondary component formed by the enzymatic decomposition of alliin (I) by alliinase. If this relationship be true, the formation of allithiamine (III) by reaction with thiamine will confirm the presence of alliin (I) in plants as the sulfur-containing component and, at the same time, the presence of allicin (II). This relationship will also hold for the methyl (VI) and propyl (XII) homologs of allithiamine. The fact that the above relationship is correct was confirmed by using synthesized S-alkyl-L-cysteine sulfoxide and alliinase isolated from a plant. In other words, the presence of S-methyl-(IV) and S-propyl-L-cysteine sulfoxide (X) can be assumed in Allium sativum L., besides alliin. In other Allium species, a minute amount of (X) can also be assumed to be present besides a large amount of alliin (I) and (IV).
Various examinations were made on experimental method using the quantitative bridge. Figs. 1 and 4 show the rationality of the concurrent use of optical scanning in this method. In A, light is projected in an extremely small area (a and a′) or in too large a portion (with oblique lines) of the substance dispersed on a filter paper in optional form, while on the quantitative bridge (B), optical scannning is made over whole of the substance present. Fig. 2 (a) shows the examination of substances of the same concentration in the quantitative bridge of various width (D). It is interesting to see that the distribution of the substances differs by the width, that the product of its length (L) and the width (D) of the quantitative bridge, i.e. the area of a substance on the filter paper, is not necessarily definite, and that the maximum concentration (h) at each width is approximately definite so that the value of h did not necessarily became larger with decrease of L. Fig. 2 (b) shows that the concentration distribution of substances on the quantitative bridge is different, the substances easily soluble in developing solutions tending to move forward. It was found that when the length of the substance on the quantitative bridge is 2-30mm., the best results are obtained so that the widh should be determined in accordance with the amount of the substance. Fig. 3 shows the chromatograms of berberine developed at various temperatures and indicates that the developing is greatly affected by the temperature. It was found that, in general, the substances stayed on the quantitative bridge better and showed distinct border when chromatography was carried out at a higher temperature. When developed at 0°, the substances dispersed widely and became too indistinct to be practicable. When a sudden temperature variation is effected after a substance is on the quantitative bridge, the substance will be cut off and no quantitative relation will be obtained.
The two growth factors of Lactobacillus arabinosus obtained by the paper chromatography of biotin showed a different activity towards Streptococcus faecalis and Lactobacillus caseii at the same concentration. The one with smaller Rf value could not be the growth factor of these two bacteria. The antagonism of these two factors against acidomycin was examined and it was found that these factors showed approximately similar activity towards avian type tubercle bacilli and the bacilli, whose growth had been injured by acidomycin, recovered on addition of these two factors.
In the type of a high frequency oscillator (Fig. 1), where a sample is inserted into the coil, it was found that the placement of a copper plate (designated as S-plate, shown as a long line in Fig. 2) at one end of the coil generally resulted in the increased sensitivity and disappearance of the maximum and minimum ordinarily observed on the concentration-current curve and was replaced by the curves shown in Fig. 3 (b, c, d, e) and Fig. 7 (the descending phenomenon of the curve that occurs at around 5-6 N is due to the fall of conductivity) (cycle, ca. 20 megacycles). When the size ratio of the coil, S-plate, and beaker is varied, the curve becomes such as shown in Fig. 7. This effect of the S-plate was explained theoretically by the use of a simplified equivalence circuit shown in Fig. 5, the relationship shown in Table I, and equations (1) and (2) given in the main text. The inverted phenomena of titration curves and impossibility of titration, was often observed by changing the concentration, were distinctly eliminated by the use of the S-plate (see Fig. 11).
Lactobacillus fermenti can utilize ornithine or citrulline, the same as arginine but the synthesis of arginine from ornithine or citrulline shows the lag phase in the initial period of culture. Pyridoxal is effective in shortening this lag phase. Canavanine inhibits the utilization of arginine and strongly inhibits utilization of ornithine or citrulline, i.e. biosynthesis of arginine. Arginine can recover growth inhibition at any concentration of canavanine, but such activity is weak in citrulline. The presence of canavanine in high concentrations makes it impossible to completely recover growth inhibition even by the increase of the amount of citrulline or by the extension of culture period. The antagonism between canavanine and arginine, and between canavanine and citrulline were found to be not competitive. In other words, the amount of canavanine against the unit concentration of citrulline when complete growth inhibition is indicated decreases extremely with the increase in the amount of citrulline. Canaline failed to inhibit the utilization of arginine, citrulline, or ornithine. Reaction center of canavanine was discussed briefly.
Lactobacillus arabinosus 17-5 can synthesize arginine in a culture medium containing glutamic acid and pyridoxal but canavanine can completely inhibit growth under such cultural conditions in an extremely low concentration (1.76×10-5 moles). In this case, the addition of arginine makes it necessary to have higher concentration of canavanine for growth inhibition. The metabolic products which can recover growth inhibition of canavanine include arginine, which is the best, citrulline, ornithine, and glutamic acid, the strength becoming weaker in this order. This order seems to have some relation to the arginine metabolism of L. arabinosus. In the case of lower concentrations of canavanine (1-50 γ), simple extension of culture period can recover the growth. Canaline shows growth inhibitive action in culture medium in which arginine is synthesized from glutamic acid but does not show practically any inhibition on the utilization of ornithine, citrulline and arginine.
Arginine requirement and growth inhibition by canavanine were examined in the parent strains and arginine-requiring mutants of Streptococcus faecalis, Leuconostoc mesenteroides, Lactobacillus arabinosus, Lact. casei, Lact. fermenti, Escherichia coli No. 1, Staphylococcus aureus 209 P, and Salmonella enteritidis 1891. It was thereby found that Strept. faecalisand Leuc. mesenteroides required only arginine and did not receive inhibition by canavanine, while other strains were able to grow by synthesizing arginine from inorganic substances or from ornithine and citrulline in the medium and their growth was easily inhibited by canavanine. Neither canavanine, canalin, nor homoserine was able to substitute for arginine. The reaction center of the growth inhibition by canavanine was discussed from the experimental results of the writer and those of Snell and Walker.
Using a strain of Pseudomonas sp. isolated from the soil, decomposition of N-methylpyridinium hydroxide (I), trigonelline (II), and homarine (III), was examined. The strain that has been adapted to (I) decomposes (I) immediately but only effects the shortening of lag phase against the decomposition of (II) and (III). The strain adapted to (II) immediately decomposes (II) and its isomer, (III), but only effects shortening of the lag phase against (I). In other words, there is no successive adaptation and only a relative adaptation is found. Monomethylamine was detected as the decomposition product of (I). Decomposition ability of N-methylpyridinium homologs by 12 kinds of bacteria including Escherichia coli was examined and none was able to decompose these homologs.
Hog kidney homogenate was applied to cadaverine and the reaction product was examined by paper chromatography by which it was found that piperidine was formed from cadaverine. The same result was also obtained when partially purified diamine oxidase, obtained from hog kidney, was applied to cadaverine. It was, therefore, assumed that cadaverine was first oxidized to δ-aminovaleraldehyde by diamine oxidase, then underwent cyclization to piperidine. The intermediate of this reaction is still obscure.
Three alkanolamines, 3, 4-dihydroxy-, 3, 4-dimethoxy-, and 3, 4-methylenedioxy-N-methylephedrines, were prepared from piperonylpropiophenone. This latter easily underwent demethylenedioxidation by treatment in nitrobenzene with anhydrous aluminum chloride at 20-30° to give 3, 4-dihydroxypropiophenone in a good yield.
By heating phenylacetylcarbinol and methylbenzoylcarbinol, or their acetates, with secondary amines, such as dimethylamine, diethylamine, benzylmethylamine, piperidine, or N-methylpiperazine, in a sealed tube at 150-180°, α-sec-aminopropiophenones were obtained in 10-44% yield. The objective was also attained by treating a few other acetates with secondary amines. From the results obtained, the foregoing reaction may be termed as the Viogt reaction.
The isolation of thyroxine from iodocasein was confirmed to necessitate the treatment of the pH 2.4-2.6-precipitate by pH-fractionation after sodium hydroxide decomposition by the concurrent observations of tadpole metamorphosis.
Observations were made on the relationship between the sowing time and growth and yields chiefly of Ammi visnaga Lam. It was found that the autumn sowing should be carried out by the end of October at the latest. In the cultural experiments made during 1953, there was a large number of withered plants, the yield of fruits was extremely small, and there was no content of khellin in the fruit. These facts were assumed to be due to the large number and amount of rain during June and July, the extraordinary cool weather during the fruiting season, and the poor drainage of the soil in the experimental plot, as well as the generation of a kind of stem-borer and diseases of unknown origin. These indicate the necessity of a small amount of rain during the summer and a well-drained soil.