1) The melting point of 2-methyl-6-bromobenzothiazole had been rather irregular but was determined as melting at 86-87° by the utilization of ferricyanide oxidation-cyclization of p-bromothioacetanilide and bromine substitution of 2-methyl-6-aminobenzothiazole by the Sandmeyer reaction. 2) Application of phosphorus pentasulfide and potassium sulfide to the acetanilide compound was found to be accompanied by the formation of a small amount of aniline as a by-product. 3) Dozens of 2-methlybenzothiazole derivatives having ether bonding at 6-position were obtained by the Ullmann reaction of 2-methyl-6-bromobenzothiazole and potassium salt of phenols. 4) Some discrepancies were found in the data of 2-methyl-6-phenoxybenzothiazole obtained in the present experiments and those appearing in past literature. In order to find correct data, the compound was submitted to synthetic examinations by the (i) Ullmann reaction of 2-methyl-6-bromobenzothiazole and phenoxides and (ii) oxidative cyclization of 4-phenoxythioacetanilide (m.p. 103-104.5°) with potassium ferricyanide. It was thereby found that the compound melting at 85-87° obtained in the present experiment was the correct one. 5) Antifungal tests (chiefly test with Candida) of the intermediates and new compounds failed to reveal any marked substances.
1) 2-Methyl-6-(4-nitrophenylthio)benzothiazole (V) was synthesized by the following three routes: (i) Oxidative cyclization of 4-(4-nitrophenylthio)thioacetanilide (III) with alkaline ferricyanide; (ii) Ullmann reaction of 2-methyl-6-bromobenzothiazole and p-nitrothiophenol; and (iii) application of sulfuryl chloride to the hydrochloride of 4-(4-nitrophenylthio)aniline (IV) and the reaction of the intermediate so obtained with sodium hydroxide and acetic anhydride. 2) Several kinds of 4-acetamidophenyl aryl sulfides, obtained by the condensation of p-acetamidothiophenol and aromatic or pyridine compounds possessing active halogen atom, was heated with potassium sulfide and phosphorus pentasulfide and corresponding 4-thioacetamidophenyl aryl sulfides were obtained. The oxidative cyclization of these sulfides afforded several new derivatives of 2-methylbenzothiazole with thioether-type linkage at 6-position; i.e. 2-methyl-6-(2, 4-dinitrophenylthio)-, 2-methyl-6-(2-carboxy-5-chlorophenylthio)-, 2-methyl-6-(3-nitro-2-pyridylthio)-, and 2-methyl-6-(5-nitro-2-pyridylthio)-benzothiazoles. 3) Anti-candida tests of these new compounds revealed that their growth inhibitory concentration was all below 10, 000 dilutions.
1) In order to find the effect on the bridging sulfur atom in diphenyl sulfide-type compounds by the Sandmeyer reaction, experiments were carried out on 4-(4-nitrophenylthio)aniline. It was thereby found that the use of hydrobromic acid in diazotization, reacted at 0°, chiefly afforded 4-(4-nitrophenylthio)bromobenzene (IV) while the reaction temperature of 0° to 40° chiefly afforded 4-(4-nitrophenylsulfinyl)-bromobenzene (V). The use of conc. nitric acid for diazotization afforded (IV) when the reaction temperature was -20°, but (V) if carried out at 0°. 2) By the electrolytic reduction of 2-methyl-6-(4-nitrophenylthio)benzothiazole (I), its 4-amino compound (II) was obtained in a good yield. 3) The amino compound was submitted to the Sandmeyer reaction by diazotization with hydrobromic acid at 0° and substituted with bromine, chlorine, iodine, and hydroxyl group. 4) Anti-candida tests of these new compounds showed that (II) alone was effective in inhibiting the growth of C. albicans at 10, 000 dilution.
In order to obtain effective anti-cancer agents, O-(1-aziridinyl)acetyl-DL-serine, O-(1-aziridinyl)acetyl-N-benzyloxycarbonyl-DL-serine, N-(1-aziridinyl)acetyl-DL-serine, ethyl 1-aziridinylacetate, DL-2-(1-aziridinyl)-3-phenylpropionic acid and its ethyl ester, and DL-2-(1-aziridinyl)succinic acid were synthesized but all showed only a weak action against Ehrlich ascites tumor. During the course of these syntheses, it was found that the use of acetic acid as a solvent in the O-chloro-acetylation of serine with chloroacetyc anhydride only afforded the O-acetyl compound while the use of ethyl acetate or ethyl or methyl chloroacetate as a solvent gave O-chloroacetyl-DL-serine in a good yield.
In order to obtain effective anti-cancer agents, phenyl, 1-naphthyl, and 2-naphthyl bis(1-aziridinyl)phosphinate, 2-naphthyl bis(2-methyl-1-aziridinyl)phosphinate, 2-naphthyl bis(1-aziridinyl)phosphinothionate, 2-naphthyl bis(1-aziridinyl)phosphinite, p-phenylene bis[bis(1-aziridinyl)phosphinate] and -phosphinthionate], m-phenylene bis [bis(1-aziridinyl)phophinate], o-phenylene 1, 2-[(1-aziridinyl)phosphonate], and p-phenylene bis[bis(1-aziridinyl)phosphinite] were synthesized. Of these compounds synthesized, p-phenylene bis[bis(1-aziridinyl)phosphinate] was found to be effective against sarcoma 180 (ascitic), Ehrlich carcinoma (ascitic), and Ridgway osteogenic sarcoma in mice.
Separatory estimation of putrefaction amines by paper chromatography was examined with 17 compounds. Approximately successful separatory estimation was effected by the one-dimensional ascending method for 20 hours at 25°, using a mixture of butanol, water, and glacial acetic acid (4:2:1) for development. The revelation indicators used were ninhydrin, Dragendorff, diazo, Sakaguchi, and nitric acid, potassium nitrite reagents. In the aliphatic amine series, a relationship was found to hold between the number of carbon atoms in the alkyl chain and the Rf values. Separatory estimation of amines by paper chromatographic method was carried out with the culture solution of squid homogenate inoculated with Escherichia coli or Achromobacter sp., and incubated at 37° for 48 hours. The amines detected were trimethylamine, butylamine, arcaine, tyramine, and histamine.
Serum cholinesterase inhibition was examined with 18 kinds of putrefaction amine, 4 kinds of acetylcholine-like substance, and 2 kinds of amino acid in 10-3 to 10-1 molar concentrations. It was thereby found that in the methylamine and ethylamine series, the inhibition increased with increasing number of methyl and ethyl groups, while in the butylamine series, the inhibition increased in the order of mono-, iso-, and di-butyl groups. In the monoalkyl- and dialkylamine series, cholinesterase inhibition also increased with increasing number of carbon atoms in the alkyl chain. Some examinations were also made on the mechanism of cholinesterase inhibition by trimethylamine. In putrefaction amines, piperidine, tyramine, tryptamine, arcaine, and agmatine showed a strong inhibitory action, while histamine, betaine, choline, putrescine, and trimethylamine N-oxide only showed a weak action. It was suggested that there is some relation between the cholinesterase inhibitory action of these amines and food poisoning caused by them.
Action of 22 kinds of putrefaction amine and related compound on excised intestine of a rabbit was examined by the modified Magnus method. Ten kinds of aliphatic amine were found to show correlation between the minimum effective concentration for accelerating the excised intestine contraction and serum cholinesterase inhibition, but such correlation was not clearly detected in other amines. It was found that some of the acetylcholine-like substances and amino acids had strong action of accelerating the intestinal contraction. With trimethyl- and dibutylamine, the mechanism of their action on accelerating intestinal contraction is parasympathomimetic and the same mechanism was found to be applicable to the metabolic product of squid by bacteria (Escherichia coil and Achromobacter sp.).
Biological distribution of trimethylamine and its change were examined using a cat, and variation of trimethylamine and its metabolic product, trimethylamine N-oxide, was examined with cat liver homogenate. After intraperitoneal injection of 0.5-1.0g./kg. of trimethylamine into a cat, the organs were taken out from the dead animal or the cat sacrificed after 1 hour, and the amount of the amine was determined by Dyer's method. Trimethylamine was found to transit and be distributed in various organs but it accumulates in the largest amount in the liver, and gradually moves to the kidney. Trimethylamine N-oxide was not found in the case of death from acute poisoning but was found in a small amount in the kidneys, spleen, and blood when sacrificed 1 hour after the injection. When trimethylamine was added to the liver homogenate, the amount decreased rapidly by 1 hour later but there was little change thereafter for 24 hours. Trimethylamine N-oxide appeared in 3 hours and about one-half of the amount of trimeth ylamine added was found to be oxidized into N-oxide during 24 hours.
It is seriously necessary to remove the impurities such as oxygen and water vapor contained in the nitrogen stream used in the direct determination of organic oxygen by the method of Schütz-Unterzaucher. The variation in blank values resulting from these impurities was investigated with the various temperatures of reduced copper provided for removal of oxygen and simultaneously with the different materials of connecting tube which should prevent the diffusion of atmospheric oxygen and moisture. It was concluded that the reduced copper maintained at 550° may eliminate the oxygen completely and further, the polyethylene connecting tube shows sufficient prevention of the permeation of atmospheric oxygen and moisture.
d-trans-8-Oxocamphor is used widely as a cardiotonic but its crystal is extremely unstable and is oxidized in the air, almost completely loosing its pharmacological effect in about 24 hours. A methanolic solution of this substance and thiourea separates some needle crystals on being cooled and the newly formed substance is composed of 8-oxocamphor and thiourea in 1:3 molar ratio. From X-ray diffraction and infrared absorption spectrum, the crystal is a rhombohedral and the substance was proved to be the so-called inclusion compound by Schlenk and others. Such 8-oxocamphor included in thiourea is extremely stable and residual ratio after 8 months at room temperature was 97.5%, that after 6 months at 40° being 93%. Extraction of this adduct in a Soxhlet extractor with warm ether was found to give 100% of 8-oxocamphor and the residual thiourea formed the original rhombic crystals. When this adduct is dissolved in water, it dissociated into the original two components, the solution showing the chemical properties of 8-oxocamphor, as well as its toxicity and pharmacological effect. There was no difference in its effect as a medicinal from the corresponding 8-oxocamphor.
Solubilization of vitamins A and D has been effected by the use of surface active agents such as Tween, Myrj, and Brij, but recently it has been found that the solubilization can be effected by the use of hydrogenated castor oil polyethylene glycol ether (HCO). In this case the best active agent differs with the kind of vitamin A and D. HCO of low polymerization degree was found to be suitable for vitamin A palmitate, while that of high polymerization degree was better for vitamin A alcohol and D. Some of HCO were found to have greater range of solubilization than Tween, Myrj, or Brij. HCO derived from hydrogenated castor oil was less toxic than that derived from non-hydrogenated castor oil. Especially, HCO of high polymerization degree was less toxic than Tween. Although hemolytic action of HCO was slightly stronger than Tween, Atlas G 1295 is low toxic and therefore suitable for injection. The irritation or pain caused by injection of HCO was not different from those caused by injection of Tween, but the taste of the former was better. From these results HCO was found to be a good solubillizing agent.
Anatomical studies were made on the fruits of Gleditsia japonica MIQ, and G. sinensis LAMARCK (Figs. 1-14). Saponin, the chief constituent of these fruits was found to be present only in the mesocarp and not in any other parts of the fruit, seeds, bark, or thorns, as confirmed by the blood-gelatin method.
Isolation of two kinds of flavonoid, fisetin (I) and fustin (II), and a new substance of the catechol series, gleditsin (III), from the extract of the heartwood of Gleditsia japonica MIQUEL. was reported in earlier paper. Later studies revealed that (III) C15H14O6⋅2H2O, belonged to the leuco-anthocyanidin series and was assumed to be a 3, 4, 7, 3′, 4′-pentahydroxyflavan.
Production of three kinds of basic antibiotic substance by mutant strain of Streptomyces racemochromogenus nov. sp. was confirmed by paper chromatography and counter-current extraction. These antibiotics were named racemomycin A, B, and C. Of these, racemomycin B was isolated and purified as its picrate, and was also obtained as the free base, various acid salts, and N-benzoyl derivative. Racemomycin B agrees well with the minimum molecular formula of C15H32O8N5, it had no free carboxyl group, and came as a very hygroscopic substance with a strong and a weakly basic groups. Racemomycin C was isolated in a small amount as its salt and was assumed to be a substance closely related to racemomycin B from its analytical values and infrared spectrum. Hydrolysis of racemomycin B with mineral acid was found to give two kinds of a basic substance and one kind of reducing sugar as a result of paper chromatographic examination.
In order to clarify the structure of racemomycin B, a basic antibiotic obtained from a strain of Streptomyces racemochromogenus, it was hydrolyzed with hydrochloric acid and two kinds of basic amino acid were obtained. One of these agreed with β-lysine obtained by the hydrolysis of viomycin, streptothricin, geomycin, streptolin, or roseothricin. The other was proved to be identical with roseonine obtained by Hirata and Nakanishi or geamin obtained by Brockmann, et al., by mixed fusion, infrared spectrum, physical constants, and chemical properties. From these facts, it was considered that racemomycin B would be one of a group of antibiotics of the streptothricin series.
Aspartic acid mono- and dihydrazides, and 3-hydroxyaspartic acid were synthesized and their action in inhibiting the growth of Leuconostoc mesenteroides P-60 and Escherichia coli No. 1 was examined. In the presence of 50γ/cc, of aspartic acid, the growth of Leuc. mesenteroides was completely inhibited by 100γ/cc. of monohydrazide, 500γ/cc. of 3-hydroxyaspartic acid, and by 2000γ/cc. of dihydrazide. So far as Leuc. mesentesoides is concerned, therefore, the growth-inhibitory action of monohydrazide is stronger than that of dihydrazide. On the contrary, the growth of E. coli No. 1, which requires no aspartic acid for growth, was more strongly inhibited by the dihydrazide, the effective concentration for 50% inhibition being 55γ/cc. of the monohydrazide and 33γ/cc. of the dihydrazide. It was also observed that the dihydrazide had stronger inhibitory action than the monohydrazide upon glutamic acid decarboxylase of E. coli. The antagonism of hydrazides to aspartic acid was non-competitive in all the cases but growth inhibition of E. coli was more easily recovered by asparagine rather than by aspartic acid. Considerations were made on the biological activity of amino acid hydrazides from foregoing experimental results.
Presence of saponin and sapogenin in the rhizome of Dioscorea septemloba THUMB. was examined and following results were obtained. 1) As a sapogenin, diosgenin was chiefly present, while the presence of a minute amount of three kinds of isteroidal substance was found. Δ-Desoxytigogenin was obtained and it was assumed to have been formed secondarily during hydrolysis of diosgenin glycosides. 2) Gracillin and dioscin were obtained as the water-insoluble saponins and the presence of a substance which may be one of the prosapogenin A was detected. Gracillin, which was found for the first time in the rhizome of D. gracillima with dioscin, was found to be present in the plant in larger amount than dioscin. 3) At least one kind of water-soluble saponin was found to be present and this saponin is diosgenin glycoside with D-glucose and L-rhamnose as the sugar, with larger number of sugars than that of dioscin and gracillin. This was tentatively named kikuba-saponin.
Presence of steroidal Saponins in the rhizome of Dioscorea Tokoro MAKINO, other than dioscin, was examined. Diosgenin was chiefly obtained as the sapogenin and a small amount of tokorogenin was isolated, indicating the presence of a minute amount of several steroidal substances. Δ-Desoxytigogenin was obtained and it was assumed to have been formed secondarily during the hydolysis of diosgenin glycoside. Apart from dioscin, the saponins obtained were minute amounts of gracillin, a new diosgenin glycoside (A) with glucose and rhamnose as the sugars, and a substance (B) which was assumed to be a saponin with a new polar steroid and glucose and rhamnose as the sugars. It was considered that dioscorea-sapotoxin obtained by Tsukamoto and Ueno is a mixture of dioscin, gracillin, (A), and (B), from the manner of its isolation and purification and its apparent solubility in water is thought to be due to the formation of a colloidal solution by the presence of the mixture. (A) and (B) were tentatively named dioscorea-sapotoxin A and B.
The structure of the substance (I), m. p. 113°, obtained on the application of benzenesulfonyl chloride to thiamine in alkaline medium, was examined and it was named thiamine anhydride by presumption of its structure. Its vitamin B1 action using rats was tested and its effect was about 1/100 of thiamine hydrochloride. In vitro treatment with liver slices made the compound show slight thiamine reaction but only around 1/200. It did not return to thiamine with cysteine or Takadiastase.
1) Phenols can be determined by the addition of a definite excess of diazonium salt solution of sulfanilic acid (diazobenzenesulfonic acid) to the sample to effect diazo coupling and measuring residual diazobenzenesulfonic acid by Iwasaki's azotometry. 2) Iwasaki's azotometry of diazobenzenesulfonic acid followed the usual known process which utilizes generation of nitrogen gas by its reaction with sodium azide in acid solution. 3) Optimal determinable range was 1-2.5×10-6M, and the error was less than ±1% in the determination of phenol, α-naphthol, β-naphthol, β-naphthylamine, and barbituric acid.
5-Alkylidene or 5-carboxyalkylidene-4-thiazolidinones were synthesized by the condensation of aliphatic ketones and ketocarboxylic acids with 4-thiazolidinones possessing thioxo, oxo, or imino group in 2-position. Since the synthesized compounds possess α, β-unsaturated carbonyl group in their structure, they were expected to react with SH-system enzyme groups, necessary for the bacterial metabolism, and thereby inhibit the growth of bacteria. It was found through their antibactrial tests with tubercle bacilli in vitro that the compounds having thioxo or oxo group in 2-position had the strongest growth inhibitory action, while antibacterial activity increased with the increase of the carbon chain at 5-position, the maximum effect being found in (IV) and (XVII) with seven carbon atoms. The antibacterial activity decreased in carbocyclic compounds and those with carboxyl at the terminal end of the alkylidene group. Effect of the presence or absence of methyl or phenyl group at 3-positiou was not clear.
In the titration of cyanate with silver, a soluble complex salt forms first and silver cyanide precipitates out later. In applying potentiometric titration in this case, the dead-stop and derivative polarographic titration curves were found to show specific shapes and indicated two end points. Titration curves with the use of platinum and silver electrodes, with addition of hydrogen peroxide, are shown and theoretical proof of these has been attempted by the procedure of potentiometric polarography.
6-Ethoxyquinoline was derived to its 1-oxide (III) and the Reissert reaction of (III) and 6(or 7)-chloroquinoline 1-oxide afforded 6-ethoxy- (IV) and 6(or 7)-chloroquin-aldonitrile (I or II). The Reissert reaction of 6-ethoxy- and 7-chloro-quinoline afforded 1-benzoyl-6-ethoxy- (V) and 1-benzoyl-7-chloro-1, 2-dihydroquinaldonitrile (VI). The Reissert reaction of halogenated compounds or 3-bromo- and 4(5 or 8)-chloroquinolines results in dehalogenation and 1-benzoyl-1, 2-dihydroquinaldonitrile is formed.
6-Chloro- (I) and 6-ethoxy-quinaldonitrile (II) were derived to their respective 1-oxides (III and IV) and the nitriles (I to IV) were all derived to the corresponding acid amides (V to VIII) by the Radziszewsky method. The nitriles (I and II) were hydrolyzed with sodium alkoxide to form 6-chloro- (IX) and 6-ethoxy-quinaldic acid (X), then derived to their respective methyl esters (XI and XII), and finally condensed with hydrazine hydrate to form 6-chloro- (XIII) and 6-ethoxy-quinaldic acid hydrazide (XIV). The Reissert reaction product, 1-benzoyl-6-ethoxy-1, 2-dihydroquinaldonitrile, was treated with hydrochloric acid and afforded benzaldehyde, 6-ethoxyquinaldamide (VI), 6-ethoxyquinaldic acid (X), and the benzoin ester (XV) of (X).
The dried leaves of Eupatorium japonica THUNB. (Compositae) have been used as home remedies as diuretic, menogogic, and for jaundice. The fresh leaves have no odor but an aroma begins to evolve with passage of time. Based on this fact, it was assumed that an aromatic substance is present as a glycoside in the living plant. Coumarin, o-coumaric acid, and thymohydroquinone were isolated from the leaves and coumarin was considered to be responsible for the aroma.
Condensation of 1, 3-di(2-pyridyl)thiourea, in the presence of desulfuration agent and alcoholic solvent, with aniline, p-aminophenol, p-alkoxyphenylamines (alkyl=CH3 to C12H25), sodium p-aminosalicylate, and isonicotinic acid hydrazide afforded the corresponding dipyridylguanidine derivatives. Antibacterial tests of these compounds in vitro against tubercle bacilli showed that there is no antibacterial action in compounds not possessing alkoxyl radical and that the action increased with increasing number of carbon atoms in the alkyl chain, attaining the maximum (in minimum inhibitory concentration) in the butoxyl group, becoming weaker with further increase, and the action disappearing entirely with more than eight carbon atoms.
Antibacterial tests on Mycobacterium tuberculosis H37Rv, Staphylococcus aureus 209P, Bacterium coli communior type, and Bacillus subtilis were carried out with 95 kinds of cyanoacetylhydrazone, benzoylhydrazone, p-aminobenzoylhydrazone, and o- and p-hydroxybenzoylhydrazone of aldehydes and ketones. Growth inhibition on tubercle bacilli was shown by cyanoacetylhydrazones of isovaleraldehyde, β-orcilaldehyde, 2-hydroxynaphthaldehyde, p-methoxybenzalacetone, and p-nitrobenzalacetone, and p-aminobenzoylhydrazone of usnic acid and 2-butanone in 12.5γ/cc. concentration, and by cyanoacetylhydrazones of m-hydroxybenzaldehyde, orcilaldehyde, orcilaldehyde dimethyl ether, cinnamaldehyde, ethyl acetoacetate, cyclohexanone, usnic acid, and salicylalacetone, by o-hydroxybenzoylhydrazones of perillaldehyde and cyclohexanone, and by p-hydroxybenzoylhydrazone of vanillin methyl ether in 25γ/cc. concentration. Growth inhibition of the staphylococci was shown by cyanoacetylhydrazone of cinnamaldehyde in 12.5γ/cc. and by cyanoacetylhydrazone of 3-chloro-2, 4-dihydroxy-5-iaoamylbenzaldehyde, o-hydroxybenzoylhydrazone of divaraldehyde, and by benzoylhydrazone of usnic acid in 25γ/cc, concentration. Growth inhibition of Bac. subtillis was shown by cyanoacetylhydrazone of usnic acid in 12.5γ/cc. and by benzoylhydrazone of usnic acid in 25γ/cc. concentrations. All of these compounds showed only a weak growth inhibitory action against coli bacilli type.
Growth curve of Zygosaccharomyces japonicus, a kind of yeast for soy sauce, in fluid medium was followed by electrophotometer. It was thereby found that when 0.1cc. of the bacterial suspension (1, 500, 000-2, 000, 000 cells) in 100cc. of the medium is incubated at 30°, approximately up to 18 hours is the lag phase, 24th hour is the increased logarithmic growth phase, and the growth become slow after 60 hours, coming into the stationary phase. The effect of the amount of inoculum on the growth curve of the yeast was found to be similar in the case of 0.15 and 0.2cc. as in 0.1cc., while the addition of methanol to the medium in 1% ratio was found to have no significant effect. Addition of propyl and butyl p-hydroxybenzoate in 3.33×10-4M concentration resulted in complete inhibition of the lag and logarithmic growth phases, and the same phenomena were observed with 2.5×10-4M of propyl and butyl orsellate. Butyl orsellate completely inhibited the growth at concentrations above 1.25×10-4M, while its addition at the initial stage of the lag phase or during the logarithmic growth phase caused complete inhibition of growth in 1.25×10-4M concentration. Cultural test revealed that its action is bactericidal