Separatory microdetermination of binary system metal ion mixture by the oxinebromate azotometry was attempted. The optimal pH at which the metal-oxine complex precipitates quantitatively differs with each ion and this was utilized in the separatory determination of Zn2+ and Mg2+, and Cd2+ and Al3+, estimable in the range of 10-200γ/cc. Before precipitating the metal-oxine complex, sodium cyanide was added as a masking agent, and the determination was made of Zn2+ in the presence of Hg2+, Cd2+ in the presence of Cu2+, and Al3+ in the presence of Fe3+. Determination of Mg2+ in the presence of Ca2+ was made by the use of ammonium oxalate as the masking agent. The estimable range in this case was also 10-200γ/cc. The determination of cadmium, zinc, and nickel was effected by the separation of Cd2+ and Zn2+ with cation exchange resin (Amberlite IR-120) and of Ni2+ and Zn2+ with anion exchange resin (Amberlite IRA-400), and subsequent oxine-bromate azotometry, the estimable range in this case being 50-150γ/cc.
In an aqueous solution, streptomycilideneisonicotinylhydrazine (Streptohydrazid) exists in an equilibrium of streptomycin and isonicotinic acid hydrazide (INAH). This equilibrium constant differs according to the pH of the solution, the value decreasing with increasing value of pH. This indicates that the equilibrium exists between streptomycin, INAH, and Streptohydrazid and that the difference in their concentration results in different dissociation of each compound and the original and product systems according to pH.
Mephenesin monocarbamate is reported to show the same pharmacological activities as mephenesin itself and sometimes slightly prolongs the duration of the action. The fact was examined together with the syntheses and examination of pharmacological activities of mephenesin carbanilate and the carbamates and carbanilates of 3-(o-methoxyphenoxy) propane-1, 2-diol (I) and 3-(1, 2, 3, 4-tetrahydro-7-naphthyloxy) propane-1, 2-diol (II). The muscle relaxing action of these compounds is weaker than that of mephenesin but the monocarbamate of (II) showed stronger anticonvulsant action against pentetrazol than mephenesin, and about equal anticonvulsant action against electric shock as mephenesin, The safety margin, LD50/MED50 and LD50/EED50, of these compounds was greater than that of mephenesin. The monocarbamates of both (I) and (II) showed longer duration of action than mephenesin in paralysis and protection against electric shock seizure.
In order to elucidate the cause of significant differences existing between individual operators in elemental analytical procedures, standardization of analytical method and process of its control were devised. Examinations were made on the weighing and burning procedures, which had hitherto been left to the discretion of individual operators, and on portions likely to become the cause of variance. A standard was fixed which would prevent the introduction of individual error by the operators into analytical values.
In order to elucidate the cause of error in carbon and hydrogen determinations revealed by the previous work, studies were made based on the statistical design of experiment. The three factors, balance, analytical apparatus, and operator, were placed in a combination of 3-way layout of two levels each, the order of analysis in each combination was stratified into four portions, two operations were alloted to each, and a total of 64 experiments were designed. During the experiments, precaution was taken to collect the data at random and the experiments were carried out during continuous analyses of general assignment. Statistical analysis of the results revealed that, of the three factors, variation by the balance alone showed significant difference, and that the precision of analytical values was C 0.1844% and H 0.2034% at σ, much smaller than that of C=H 0.25% in the preceding report. These results indicate that the cause of error in elemental analyses is due almost entirely to the decreased precision of the balance.
Fresh leaves of Gaultheria Miqueliana TAKEDA were extracted with 50% methanol, the extract treated with lead acetate solution, and the portion not precipitating with the lead acetate solution afforded 1.1% of monotropitoside. The portion precipitating with the lead acetate solution afforded brownish yellow needles, which recrystallized from water to brownish yellow microneedles, m.p. 180-185°. Its anhydrate begins to color from around 220°, gradually turns black, and decomposes at around 285° with effervescence; [α]D10-22.93° (EtOH). Analytical values agree with the formula C21H18O13⋅5H2O. The anhydrate, when left in the air, absorbs 4 moles of water. From the result of its hydrolysis, this substance was found to be composed of 1 mole each of quercetin and glucuronic acid. The position of the bonding of glucuronic acid was found to be the 3-position by the formation of 5, 7, 3′, 4′-tetramethylquercetin by the methylation of this substance with diazomethane, followed by hydrolysis. The substance, therefore, is quercetin-3-glucuronide which is a new substance and is here designated as miquelianin.
By the utilization of the coloration reaction of dimethylglyoxime and that of dimethylglyoxime and thiosemicarbazide, a new detection method for some organic nitrogenous compounds, such as hydrazine, carboxylic acid hydrazides, hydrogen cyanide, barbitals, bromovalerylurea, guanidines, arginine, creative, and riboflavin, was examined.
In connection with the cleavage reaction of nitrodiphenyl ethers with sodium hydroxide-liquid ammonia, effect of carboxyl and methyl groups in the benzene ring not possessing a nitro group was examined. It was found that the cleavage reaction is not affected by carboxyl or methyl group but is caused by the effect of the nitro group in the ortho or para to the ethereal oxygen. This suggests that, although sodium hydroxide would be present as a salt in liquid ammonia, it may actually take part in the reaction as an acid or alkali.
The formation of 3-hydroxy-2, 4-lutidine by the application of acetic anhydride to 2, 4-lutidine-1-oxide was found to have been erroneous. The product was found to be 5-hydroxy-2, 4-lutidine by later syntheses of 3- and 5-hydroxy-2, 4-lutidine by the nitration of 2, 4-lutidine and reduction and diazotization of the 3- and 5-nitro-2, 4-lutidines obtained, after each had been isolated. Diazotization of 3- and 5-amino-2, 4-lutidine afforded, besides the phenolic bases, 3H-1, 2, 6-triaza-7-methylindene from the 3-nitro compound and 3H-1, 2, 6-triaza-5-methylindene from the 5-nitro compound.
Aqueous solution of chlorotetracycline hydrochloride, when warmed for 45 minutes at 30° with the addition of aminopyrine, sodium hydroxide, and potassium ferricyanide, colors reddish violet and this color transits to the amyl acetate layer when the aqueous solution is acidified with acetic acid and shaken with amyl acetate. The optical density of this colored acetate layer was measured at 564mμ to make colorimetric determination of chlorotetracycline. The presence of oxytetracycline and tetracycline in less than 20% amount makes it possible to determine chlorotetracycline with below 2% accuracy. Determination of chlorotetracycline hydrochloride, in an aqueous solution, allowed to stand for a long period of time and expected to have undergone a certain amount of decomposition, by the biological assay, hydrochloric acid method, and the present method gave well agreeing value. It was also found that the markedly decomposed solution determined by the present method gave values approximating those by biological assay than the values by the hydrochloric acid method.
Separatory determination of pyrazinamide and its in vitro decomposition product, pyrazinoic acid, in the urine and blood appearing after the oral administration of pyrazinamide, was carried out by the utilization of bright reddish orange coloration formed by the reaction of pyrazinamide and pyrazinoic acid with sodium nitroprusside, in sodium carbonate alkalinity. The urine sample or the sample blood added with sodium citrate, hemolysed, and deproteinized with methanol, is passed through a column of ion exchanger, Amberlite IRA-400, and pyrazinamide in the effluent is first determined. The pyrazinoic acid adsorbed on the resin is eluted with warm sodium chloride solution and the acid in this solution is determined. It was thereby found that the blood level of pyrazinamide is far higher than that of pyrazinoic acid at any time, the acid being hardly detected. On the contrary, the level of pyrazinoic acid in the urine is far higher than that of pyrazinamide.
p-(2-Benzoylacetamido) benzenesulfonyl chloride was prepared in a good yield by the chlorosulfonylation of benzoylacetanilide with chlorosulfonic acid and seven kinds of these sulfanilamide couplers, including sulfanilamide, sulfanilanilide, sulfanilamidobenzothiazole, and sulfanilamidobenzimidazole were obtained.
It was found that p-toluidine and crotonaldehyde reacted in glacial acetic acid, in the presence of 1 mole of sulfuric acid, without any tendency for resinification, to afford a crystalline precipitate corresponding to the formula of C13H17O2N⋅H2SO4. This substance formed 2, 6-dimethylquinoline in a good yield when heated with ferric chloride in glacial acetic acid. From these results, it was found that 2, 6-dimethylquinoline could be obtained in a good yield directly from p-toluidine and crotonaldehyde under mild conditions. In a similar manner, quinaldine is obtained from aniline and crotonaldehyde.
The reaction conditions for condensation of crotonaldehyde and p-toluidine or aniline reported in the previous paper were applied to the reaction of aniline and methyl vinyl ketone and 4-anilinobutan-2-one was isolated as the intermediate of lepidine synthesis. This ketone easily changes to lepidine when heated with ferric chloride in acid medium, but the addition of aniline to methyl vinyl ketone is reversible and there was found no marked difference between the yields of lepidine directly from aniline and methyl vinyl ketone and of lepidine from 4-anilinobutan-2-one. Similarly, aniline and methyl isopropenyl ketone afforded 3, 4-dimethylquinoline and aniline and tiglic aldehyde afforded 2, 3-dimethylquinoline.
Two kinds of new crystalline glycoside were isolated from the root of Paeonia suffruticosa ANDL. (P. Moutan SIMS.) and were named paeonoside and paeonoside. The isolation of the two glycosides was effected by the utilization of column chromatography through an ion exchange resin and starch-celite. The presence of paeonoside had been predicted by Péron and others in 1911, which was confirmed by the present series of experiments. Although Péron and others reported that it would not be a β-type since it does not submit to hydrolysis by emulsin, the present experiments revealed that paeonoside is hydrolyzed by emulsin and its structure was indicated as paeonol-β-D-glucoside. Paeonolide is hydrolyzed by emulsin to liberate a pentose and a hexose, so that its sugar portion is known to be an oligosaccharide. It was also clarified that the bonding of the sugar is a β-type glycoside, same as in paeonoside.
The hydrolyzate of paeonolide was submitted to paper chromatography and its sugars were clarified as L-arabinose and D-glucose. Partial hydrolysis of paeonolide with formic acid affords paeonolide and L-arabinose, so that its structure must be paeonol-β-D-glucose-α-L-arabinose. A modified synthesis of paeonol was devised and the use of nitrobenzene as a solvent to effect two-layer reaction resulted in increase of the yield from the usual 15% to 79%.
Ethyl pyrrole-2-carboxylate derivatives, 3-ethoxycarbonylmethyl-4-acetyl-5-methyl (Ia), 3-ethoxycarbonylmethyl-4-ethyl-5-methyl (Ib), 3, 5-dimethyl-4-acetyl (Ic), 3, 4-dimethyl (Id), 3, 5-dimethyl (Ie), and 3-ethoxycarbonylmethyl-4-ethyl-5-bromo (If) compounds were derived to N-ethoxycarbonyl compounds and submitted to catalytic reduction with platinum catalyst, at ordinary temperature and pressure. (Ia, b, c) afforded the corresponding ethyl pyrroline-1, 2-dicarboxylate derivatives (IIIa, b, c) and (Id, e, f) afforded the corresponding ethyl pyrrolidine-1, 2-dicarboxylate derivatives (III′d, e, f). In these cases, the 4-acetyl group is reduced at the same time to 4-ethyl group and 5-bromo group in (If) is liberated. Saponification with hydrobromic acid of (III) and (III′) afforded pyrroline-2-carboxylic acid derivatives (IV) and proline derivatives (IV′). It was concluded that the double bond in pyrroline is not conjugated but is in the 3-position because the imino group in (IVa) gives pK′ value of 10.25 and the infrared absorption of the carboxylic ester group at 2-position is identical with that of the carboxylic acid ester at 2-position of pyrrolidine.
Infrared absorption spectra of threo and erythro compounds of dl-chloramphenicol and its allied compounds were measured, and relationship between each isomer and its spectrum was pointed out and discussed. In dilute solutions, these compounds exhibited absorptions due to free and bonded hydroxyls and the spectra confirmed the presence of an intramolecular hydrogen bond. Further, the role of hydrogen atom in 1-OH group was pointed out. There was a regularity in the amide-I and -II bands of the N-acyl compounds and it was shown that, in a crystalline state, the erythro compounds invariably possessed intramolecular hydrogen bond between the N-acyl groups stronger (2-3 times) than that in the corresponding threo series. A single absorption always appeared in the region of 950-1000cm-1 in the threo compounds and 900-950cm-1 in the erythro compounds. It is proposed that this absorption is probably due to the C-H bond of the asymmetric carbon atom. dl-2, 2-Dimethyl-5-dichloracetamido-6-phenyl-1, 3-dioxane showed various phenomena entirely reverse of other compounds, indicating that its conformation is not parallel to that of other compounds.
Using anhydrous propionic acid as a solvent, diphenhydramine and its acid salts can be directly titrated potentiometrically with perchloric acid by glass-calomel electrodes with good accuracy. The effect of the combined acid on the direct titration of the salts was comparatively examined by the titration curves. It was thereby found that the order of the strength of the acids in anhydrous propionic acid is almost equal to their strength in aqueous solution and anhydrous acetic acid, but organic acids, especially picric acid, are relatively weaker. It would be because of the physical and chemical interactions of the solution and the solute, for example the affinity of solvation. It may be concluded that organic acids with pK below 2 in aqueous solution and inorganic acids have some effect on the titration and lower the accuracy of the measurement but weaker acids would have almost no effect.
In order to measure the metabolism of thyroid hormone and its blood and gland levels, a series of amino acids related to thyroid gland, tyrosine, diiodotyrosine, and thyroxine, were derived to their dinitrophenyl derivatives by reacting with 2, 4-dinitrofluorobenzene and their identification and separation were attempted. Fundamental experiments on their method of preparation, melting point, ultraviolet absorption spectra, paper chromatography, and stability to hydrolysis showed that these dinitrophenyl derivatives are fully utilizable for the foregoing purposes.
Drug preparations containing non-ionic surface active agents as a solubilizer is apt to become molded and the addition of an antifungal agents in such preparations at times do not exhibit the anticipated effect. This was assumed to be due to the fact that the solubilized antifungal agent is enveloped inside the micelle formed by the surface active agent. Using Tween 20 as the surface active agent and four kinds of p-hydroxybenzoic acid esters as the antifungal agent, solubility of the antifungal agent in the presence of surfactant and effect on Aspeyillus niger were tested and the relationship between the degree of hydrophilic or lipophilic properties of the antifungal agents and micelle formation of surfactants was considered. Addition of Tween 20 solution increases the solubility of the esters (Fig. 1) but the antifungal activity decreases (Fig. 2). Since the rate of decrease is different in each ester, antifungal activity of each ester differs in magnitude above a certain concentration of Tween 20. The ratio of solubility to antifungal concentration is approximately constant in each ester and this value is smaller in the esters with smaller number of carbon atoms (Fig. 3). It was thereby concluded that the antifungal agents easily soluble in aqueous phase outside the micelle possessed greater antifungal activity.
Synthesis of DL-dimethyl-, DL-dibenzyl-, DL-S8-methyl-S6-benzyl-, and DL-S8-benzyl-S6-methyl-dihydro-α-lipoic acids was carried out and they were all obtained as an oil, except the S-dibenzyl compound. In order to obtain crystalline derivatives, formation of various salts were attempted. Although S-benzyl- and S-(p-nitrobenzyl)-thiuronium salts of the four compounds crystallized, they showed no depression of the melting point when fused with each other. Their benzhydryl-amine salts, however, showed about 10° depression of the melting point on admixture with each other and were found to have good properties as salts for their identification.
In vitro transacetylation was attempted between 1 mole of ethyl DL-diacetyldihyd-ro-α-lipoate and 2 moles of aniline or benzylamine. In the case of aniline, 1 mole of aniline changed to acetanilide and only the acetyl group in S8-position easily underwent transition, while that in S6-position remained resistant. In the case of benzylamine, 2 moles of the amine changed to benzylacetamide and the acetyl groups in both S8-and S6-positions were found to undergo transition. Such difference occurs by the difference in the basicity of the two amines.
As a color reagent for santonin 2, 4-dinitrophenylhydrazone, 0.33% potassium hydroxide solution was used instead of l% sodium hydroxide or 1% dehyd. ethanolic solution of sodium hydroxide used in the previous methods. As a result, turbidity of the test solution was completely eliminated and stability of the color formed was also improved. Light absorbency (e) of this reddish purple color was measured by the Beckman spectrophotometer at 500mμ. Concentration (r) of the hydrazone in 100cc of the solution was calculated by the following equation: r=1573e+6.10 Test conditions were determined for assay of santonin in drugs or chocolate. The existence of bisatin, phenobarbital, kainic acid, phenylthiourethan, precipitated sulfur, quinophen, ethyl p-aminobenzoate, taurin, methionine, dextrose, and extracts of digenea or melia did not disturb the results. When curcumin is present in the sample, it must be eliminated by partition chromatography before this procedure.
In order to find hypotensive drugs, synthesis of cyclohexane derivatives was attempted and cis-1, 4-bis(trimethylammoniummethyl)cyclohexane dibromide, its di (hydrogen tartrate), cis-1, 4-bis(1′-methylpyrrolidiniummethyl)cyclohexane dibromide, and its di (hydrogen tartrate), and their trans compounds were obtained. During the course of this work, it was found that the reduction of diethyl terephthalate with Raney nickel catalyst afforded entirely different reduction products according to the degree of washing with water after alkali treatment of Raney nickel.
Treatment of the reaction product of ketene and crotonaldehyde with 35% sulfuric acid, by the method of Boese, afforded sorbic acid (Table I). Examinations were made on the formation of sorbic acid by the treatment of the foregoing reaction product with alkali hydroxide and showed that the acid can be obtained by treatment with 30% potassium hydroxide (Table II), and that sorbic acid changes into an unknown substance by the action of alkali. This sequential reaction was assumed to be one of the causes in reducing the yield of sorbic acid during treatment with alkali. Antifungal action of sorbic acid was examined with Trichophyton interdigitale (Table III).
Application of hydrogen peroxide to pyrrolidine yielded succinic acid. Oxidative cleavage of N-benzoylpyrrolidine and N-p-toluenesulfonylpyrrolidine with potassium permanganate respectively yielded benzamide and succinic acid, and p-sulfaminobenzoic acid and succinic acid. The foregoing results have provided experimental evidences that the cleavage of the pyrrolidine ring by the application of oxidation agents occurred on both sides of the ring nitrogen.
Eleven kinds of monobasic acids were prepared by the electrolysis of mixed salts, a kind of Kolbe reaction. As the half-ester of dibasic acid, ethyl hydrogen adipate was used and, by crossing with monobasic acids, crossed-coupling products were obtained. Platinum plate was used as the electrode, and methanol, ethanol, or water as the solvent. The addition of tetrahydrofuran as a solubilization agent during the synthesis of higher fatty acids was found to successfully effect progress of the electrolytic reactions. The purity of the acids obtained was fairly high.
Hydrochloric acid hydrolyzate of zinc-casein was submitted to paper chromatography and colored with ninhydrin. A bright scarlet spot appeared at Rf 0.3, which does not appear in the casein hydrolyzate and differs from the violet-colored spots of various amino acid peptides (Fig. 1). Dithizone coloration of the zinc-casein hydrolyzate chromatogram also gave violet red spot at Rf 0.3 and at Rf 0.1 with zinc sulfate solution (Fig. 2). Absorption spectrum of zinc-casein shows maximum absorption at 270mμ while those of casein and of casein mixed with zinc sulfate shows specific absorption curve for casein protein, with maximum absorption at 274mμ (Fig. 3). Zinc-casein hydrolysate exhibits the maximum absorption at 274mμ, in which tyrosine absorption appears with a strong intensity. The portion containing zinc does not contain tyrosine and shows the maximum absorption at 266mμ, while the portion not containing zinc contains tyrosine and shows maximum absorption at 274mμ (Fig. 4). If zinc is removed by dithizone treatment of the. portion containing zinc, the maximum absorption at 266mμ disappears (Fig. 5). These results indicate that the zinc-casein clearly contains chelated zinc.
In order to examine the structure and pharmacological action of reserpine, compounds in which the E-ring is an aromatic ring and D-ring has been cleaved at N4 and C20, i.e. 1, 2, 3, 4-tetrahydro-β-carboline and 15, 16, 17, 18, 19, 20-hexadehydroyohimbane derivatives, were prepared. General method for the syntheses of compounds possessing such fundamental skeleton has already been established but attempts were also made to cyclize the acid amide (III) to 3, 4-dihydro-β-carboline (IV) and its reduction to 1, 2, 3, 4-tetrahydro-β-carbolines (V).
The triterpene-carboxylic acid in Poria cocos WOLF. from Kumamoto Prefecture does not agree with pachymic acid and its properties were found to be extremely similar to those of a 3:1 mixture of eburicoic and dehydroeburicoic acid reported by Robertson et al. The water-soluble sugar component contained glucose, sucrose, and a minute amount of unknown sugar, and fructose was not detected.
The laboratory-cultivated mycelium of Aspergillus chevalieri (MANGIN) THOM ET CHURCH on Czapek-Dox medium with 10% sucrose was treated as indicated in Chart 1 and echinulin, flavoglaucin, auroglaucin, parietin, and white needle crystals of m. p. 181-182° were obtained.