The product obtained by fission of 1-methoxydibenzo-p-dioxin (II) with sodium and liquid ammonia was examined and two kinds of compounds were found to be formed. One was 3-methoxy-2′-hydroxydiphenyl ether (III) and the other was found to be 2-methoxy-6, 2′-dihydroxybiphenyl (IVa). It was also reavealed that the yield of (III), considered to be the normal reaction product, increased when the reaction temperature was lower and on addition of ammonium chloride during the reaction.
Examinations were made on the reduction of azo compounds with hydrazine hydrate in the presence of copper powder. Azobenzene and 4-methylazobenzene were reduced to hydrazo compounds, while 4-aminoazobenzene, 4-dimethylaminoazobenzene, Methyl Orange, Methy Red, and Orange IV suffered reductive cleavage to the amines. 4-Hydroxyazobenzene and 4-hydroxyazobenzene-4-sulfonic acid were also cleaved but did not form aminophenol. Refluxing of azobenzene with alkali hydroxide and hydrazine hydrate in diethylene glycol for 30 minutes, with further heating for 2 hours at 200° with distillation column, afforded 2-amino- and 4-aminodiphenylamines. These aminodiphenylamines were also obtained when hydrazobenzene was reacted under the same conditions or when azobenzene was heated with hydrazine hydrate in diethylene glycol. Therefore, these aminodiphenylamines must be formed after azobenzene has been reduced to hydrazobenzene and the latter had undergone rearrangement.
Experiments were carried out in order to obtain basic knowledge on the change of filling behavior of a powder when a tablet is compressed. Kaolinite-rich aluminum silicate was prepared into five kinds of sample with different compessional force and adsorption of water vapor at 20° was carried out to obtain various adsorption and deso rption isotherms. Uncompressed sample shows the BET II-type adsorption isotherm while it changes to BET IV-type when compressed, indicating that capillary condensa tion had taken place. Moreover, a marked hysteresis loop appears and this clearly indicates formation of a bottle-neck pore, and a linear relationship was found to be established between the logarithm of the area of hysteresis loop and compressional force.
From the adsorption isotherm described in the preceding paper, specific surface area, and heat of adsorption, were calculated by the BET method. Decrease of void space by compression indicates increased strength of molded powder. Heat of adsorption of water is one of important mechanisms for disintegration and decrease of this heat by compression agrees with experimental fact that disintegration becomes poor if compressed too much. The mean particle diameter is 50-52Å and the most developed pore diameter is 124.8Å, and this shows that the pore is not formed on the particle, but a void is formed by aggregation of primary particles with mean diameter of 50-52Å, and distribution of pore diameter has a peak at 124.8Å. Decrease of specific surface area is small and differs greatly from that of sulfathiazole reported by Higuchi. When calculated according to his data, coordination number of particles, i.e. the number of particles coming in contact with one particle, is approximately 8, 000 and indicates that a plastic deformation is in force, but present experiments gave the value of 3.6 and this indicates that the system is porous and rigid.
The manner in which the void space, thought to represent flaws in compressed tablets, changes with increasing compressional force was examined by comparing the pore volume distribution of four samples from their adsorption isotherm by Shull-Wheeler's method. The peak of distribution in samples compressed at 0.76 ton/cm2 was at pore radius of 62.4Å, the pore volume at this point was 148×10-4cc/gÅ, with the distribution making a sharp upward curve, and showed Gauss distribution in the region with small pore radius and Maxwell distribution in that with large pore radius. The peak shifted to 42.4Å in samples with compression of 1.5 tons/cm2 and the pore volume decreased to 81×10-1cc/gÅ. When the compression was increased to 2.9tons/cm2, the peak of distribution became still lower, to radius of 21Å, and pore volume at that point became 60×10-4cc/gÅ, indicating that larger pores gave way to smaller pores.
A separatory determination of pyrazinamide and its biological decomposition products excreted in urine, after concurrent administration of pyrazinamide and INAH, had been devised and this was utilized in comparative examination of biological changes of pyrazinamide after its single administration and in conjunction with INAH. The sample urine was treated with anion exchange resin (Amberlite IRA-400) to separate pyrazinoic acid and urine components, the effluent solution was oxidized with bromine to change INAH and its metabolites to isonicotinic acid, and pyrazinamide was determined by the cyanogen bromide and sodium nitroprusside methods. It was thereby found that biological change of pyrazinamide was almost the same whether administered singly or with INAH and pyrazinoic acid was excreted in far larger amounts than pyrazinamide. Separatory determination of acetylated INAH and isonicotinic acid in urine was effected by the use of cyanogen bromide method, anion exchange resin, and acid-treated alumina, and excretion of these metabolites into urine after single administration of INAH and in conjunction with pyrazinamide was examined. As a result, acetylation rate of INAH was found to differ not greatly by its administration alone or with pyrazinamide, but excretion of isonicotinic acid was found to increase fairly greatly by the concurrent use of pyrazinamide.
Stability and rate of change of compounds related to pyrazinamide, 1-pyrazinoyl-2-isonicotinoylhydrazine (P-I) and p-pyrazinamidosalicyclic acid (P-P), which show better growth inhibitory action against tubercle bacilli than pyrazinamide, were compared in vitro and in vivo to those of pyrazinamide. Urinary level of P-P was determined by utilization of adsorption of P-P on alumina and the use of sodium nitroprusside, and this is not interfered by other biological decomposition products. Measurement of urinary level of P-I was made after separating it from other biological decomposition products by the use of acid-treated alumina and determined as isonicotinic acid by the cyanogen bromide method. The result of these determinations indicated that P-P and P-I are both far more stable than pyrazinamide, are absorbed better, and are excreted in far larger amounts in urine. Experiments using simulated gastric juice, simulated intestinal juice, enzyme solution from intestinal mucous membrane, intestinal bacteria, and liver homogenate all indicated that they are more stable than pyrazinamide, although the compounds were found to undergo slight decomposition with liver homogenate. P-I alone is decomposed by Escherichia coli communior but was found to be much more stable than pyrazinamide by measuring the rate of decomposition of pyrazinamide under the same conditions.
Of the four kinds of crystals reported as poisonous component of Japanese star anise (fruit of Illicium Anisatum) (Table I), Lane's anisatin alone seems to be a unity and others must be a mixture. Extraction of anisatin is best effected by using the fruits from which seeds have been removed. A good result was obtained by the countercurrent distribution method with water and ethyl acetate in isolating pure anisatin and pseudoanisatin from crude crystals (Fig. 1). Toxicity of various parts of this plant was compared by measuring LD50 with mice (Table II).
In order to prepare 2, 6-dimethyl-4-aminopyrimidine-5-carbonitrile (I), the method of Kenner, et al. was followed and it was found that their report contained some error. The compound obtained by their method was 1-(acetamidino) ethylidenemalononitrile (II) and does not contain a pyrimidine ring. The objective cyanopyrimidine compound (I) was obtained finally by warming (II) with acetamidine in ethanol or application of acetamidine in amount of 2 moles to 1 mole of malononitrile. The cyanopyrimidine compound (I) was derived through chloromethylpyrimidine compound (V) to the isothiuronium chloride (VI) and oxidized to the disulfide compound. (V) was derived to the methyl analog of thiamine, 3-(2, 6-dimethyl-4-amino-5-pyrimidinylmethyl)-4-methyl-5-(2-hydroxyethyl) thiazolium chloride hydrochloride (IX). This last compound was found to have about one-third the potency of thiamine in rats.
Purely synthetic preparation of biscoclaurine-type bases was planned by the Ullmann condensation of two kinds of benzyl-tetrahydroisoquinoline-type bases and to effect formation of diphenyl ether linkage in the final stage. One of the starting materials for this synthesis, 1-(4-bromobenzyl)-2-methyl-6, 7-dimethoxy-1, 2, 3, 4-tetrahydroisoquinoline (VII), was prepared.
dl-1-(3, 4-Methylenedioxybenzyl)-2-methyl-6, 7-dimethoxy-1, 2, 3, 4-tetrahydroisoquinoline (IX) was prepared from 3, 4-dimethoxyphenethylamine (V) and 3, 4-methylenedioxyphenylacetyl chloride (VI), and (IX) was decomposed with metallic sodium in liquid ammonia. The phenolic base obtained by this cleavage was proved to be dl-armepavine (X) from its Rf value, melting point, and infrared spectral data. The Ullmann condensation of (X) and 1-(4-bromobenzyl)-2-methyl-6, 7-dimethoxy-1, 2, 3, 4-tetrahydroisoquinoline (XI) to effect diphenyl ether bonding successfully afforded 4, 4′-bis (2-methyl-6, 7-dimethoxy-1, 2, 3, 4-tetrahydroisoquinolylmethyl) diphenyl ether (XII) by purely synthetic means, and the compound was identified through its oxalate and styphnate.
During the purification of whale insulin, a substance was obtained by fractional precipitation at pH 5.9 and acetone concentration of 40%. This substance effected a lasting hypoglycemia and this effect was found to be caused by mixing of insulin and a special protein in the pancreas. The protein itself had no physiological action and only conferred lasting properties on being mixed with insulin, but no such activity was found in other protein fractions of the pancreas. The protein fraction in question may have some physiological activities. Animal test with rabbits indicated that this fraction can be used clinically.
During studies on the relationship between stability of thiamine and diluent (bulking agent) or amount of moisture in powders, it was found that the decomposition of thiamine was markedly promoted in powders bulked with talc, in the presence of even a small amount of moisture in ordinary state of preservation, and it was also observed that this decomposition was chiefly towards hydrolysis of thiamine into the thiazole and pyrimidine moieties. The decomposition was also very much marked when aqueous solution of thiamine in which talc was suspended was heated. Examination of these phenomena revealed that this decomposition was not due to the buffer action of talc or the effect of impurities but to talc itself whose action was the strongest at pH 5-6. This was chiefly due to the lability of thiamine adsorbed on talc so that the talc that already has adsorbed other substances has less effect on thiamine by decreased adsorbability. This has shown that the decomposition of thiamine in thiamine-talc powders was not the effect of moisture but was due to adsorption phenomenon. This decomposition of thiamine was prevented by the addition of acid.
Since it was found that the marked decomposition of thiamine in thiamine-talc powders, in the presence of moisture, was due to adsorption of thiamine on talc, examinations were made on compounding of other adsorbents, such as silica gel, alumina, and aluminum silicate. It was thereby found that decomposition of thiamine occurred, both in suspension and compounded powders, in accordance with adsorptive power of adsorbents. Determination of thiamine in suspension samples was carried out by separatory determination of thiamine in solution and that adsorbed on the adsorbent to find the distribution and change of thiamine in the sample.
Some pharmacological properties of phosphorylcholine were exmined with its calcium salt. Even in a high dose no or slight activities of the componnd were observed so far tested. In comparison with calcium chloride, it was concluded that calcium ion of the salt might be responsible for these activities.
Oxidation of 3-phenylpropyne, 1, 3-diphenylpropyne, and 1-phenyl-2-heptyne with tert-butyl chromate or freshly sublimed selenium oxide results in the formation of corresponding acetylenic ketones, 3-phenylpropyn-3-one, 1, 3-diphenylpropyn-3-one, and 1-phenyl-2-heptyn-1-one. Oxidation of 1-hexyne and 1-phenyl-1-butyne with selenium dioxide gives the corresponding acetylenic alcohols, 1-hexyn-3-ol and 1-phenyl-1-butyn-3-ol,
Microbiological oxidation of progesterone was carried out with Stemphylium botryosum and monohydroxyprogesterone, m.p. 190-192°, [α]D+186° (acetone), extremely similar to 11β-hydroxyprogesterone, was obtained. This substance was identical with monohydroxyprogesterone obtained by microbiological oxidation of progesterone with Curvularia lunata or Cunninghamella blakesleeana. This was found to be not 11β-but 14α-hydroxyprogesterone. Microbiological oxidation of 17α-hydroxydesoxycorticosterone with St. botryosum afforded 14α, 17α-dihydroxydesoxycorticosterone.
Phenol system pharmaceutics generally have strong affinity to non-ionic surfactants of polyoxyethylene series such as Tweens and Brijs, and are known to form complexes by facile reaction. p-Hydroxybenzoates (Parabens) also have strong affinity to surfactant micelle and are absorbed well, and it has been pointed out that antifungal activity will not be sufficiently effected in a solution of such non-ionic surfactants. Therefore, effect of Paraben addition was examined in relation to solubilization system of non-ionic surfactants containing a solute, because the antifungal effect of Paraben will be lowered not only by its micelle affinity but also by the change in the system in the presence of a solute. Various amounts of Propylparaben crystals were added to the system containing caproic acid, dimethyl phthalate, or toluene as a solute, solubilized with 10%(w/w) solution of Emasol 4130 (Tween-80 type) and Nikkol BL-60 (Brij-35 type) and the mixture was shaken at 25°. Propylparaben is solubilized as usual in the surfactant solution not containing a solute but a marked turbidity occurred in the solution on addition of excess of Propylparaben in the presence of a solute. When this turbid solution was allowed to stand for a long period, creaming occurred with deposition of an oily subtstance at the bottom and leaving a clear supernatant. Residual solute and amount of Propylparaben in the supernatant solution were determined to presume change in the system. It was observed in the case of caproic acid and dimethyl phthalate as solutes that the effect of excess addition of Paraben differed according to the nature of the initial solute.
In order to examine pharmacological activity of allopregnane-3α, 20α-diol, a metabolite of progesterone excreted into the urine, a partial synthesis of this compound was undertaken. 20α-Acetoxy allopregnan-3β, -ol was derived to its 3-tosylate and subsequent Walden inversion gave the objective compound. It was found through pharmacological tests that allopregnane-3α, 20α-diol was effective for climacteric disturbances and amenorrhoeae, although to a lesser degree than pregnane-3α, 20α-diol.
Incubation of steroidal hormone (testosterone, progesterone, desoxycorticosterone, estrone) crystals in aqueous solution of bovine serum or lyophilized bovine serum at 36° results in appearance of characteristic absorption maximum in the range of 245-251mμ (276-278mμ in estrone) in their ultraviolet spectrum after 24-48hours. Electrophoretic pattern of the serum proteins indicated that the ratio of α-globulin fraction had decreased in the serum incubated with desoxycorticosterone, while that of β-globulin had decreased in that incubated with testosterone, with concurrent increase of the ratio of γ-globulin. The serum incubated with estrone showed decrease of albumin fraction and increased α-globulin fraction. The steroids became more hydrophilic by these incubation and intensity of this tendency was in the decreasing order of testosterone, desoxycorticosterone, estrone, and progesterone.
Fourteen kinds of steroidal hormone crystals were incubated with bovine serum albumin in a phosphate buffer of pH 8.0 at 36° and it was found that a part of the steroid became hydrophilic and lost solubility in organic solvents. The electrophoretic pattern of the bovine serum albumin incubated with steroids showed decreased height of the peak, thereby becoming asymmetric. In the serum albumin incubated with estrone, a small sub-peak appeared and δ-peak showed increased height. The amount of steroid in the buffer solution decreased with passage of time during incubation and reached a constant level after 24-48hours.
Fourteen kinds of steroidal hormone crystals were incubated with bovine serum globulin in phosphate buffer solution of pH 8.0 at 36°. The tendency of steroids to become hydrophilic was greater than that in bovine serum albumin. Estrogens like estrone and estriol showed about the same degree of affinity to both serum albumin and globulin, while testosterone and progesterone showed greater affinity to β-globulin, and corticosteroids, such as cortisone, hydrocortisone, and desoxycorticosterone, to α-globulin. Electrophoretic pattern of bovin serum globulin incubated with steroid crystals showed lower α-globulin peak when incubated with estrogen, corticosteroids, and 17-oxosteroids, and lower β-globulin peak in that incubated with testosterone and progesterone, with consequent formation of an asymmetric type and higher γ-globulin peak.
Fourteen kinds of steroidal hormones (crystals) were incubated with bovine serum proteins, serum albumin, or serum globulin, in a phosphate buffer of pH 8.0 and the product thereby obtained was lyophilized to examine its physicochemical properties. Paper chromatography using polar solvent (butanol·water) as a developing solution showed a spot for steroid, presumably separated from the adduct, while such separation was not effected by development with a non-polar solvent (ligroine·methanol·water). Combination with serum albumin was separated by paper electrophoresis but that with serum globulin was not separated and the complex compound migrated as such. Polarogram in ammonium chloride-ammonia buffer added with hexamine cobaltic chloride showed that the half-wave potentials of 2nd and 3rd waves of protein shifted to a lower potential by combination with a steroid. The isoelectric point of proteins combined with steroid showed somewhat higher values.
Combination of steroidal hormones with bovine serum albumin or globulin was examined. In the combination of steroids with bovine serum globulin, protein turbidity and the amount of combined steroid are in approximately linear relationship but in combination with serum albumin, this relationship is more like adsorption curve. The wave height of the 2nd wave of proteins in polarography in ammonium chlorideammonia buffer containing cobaltous chloride shows negative correlation to some extent with the amount of combined steroid. This combination is partially undertaken by -CO-NH-, -NH2, and -COOH in the protein molecule and the individual amino acid forming the serum albumin and globulin, and -S-S- or -SH in the protein molecule do not take part in this combination with steroids.
Effect of infrared or ultraviolet irradiation on the combination of steroidal hormones with bovine serum albumin or globulin was examined. Ultraviolet irradiation resulted in decreased amount of the combination, with decrease of 2nd wave and increase of 3rd wave in polarogram, both with serum albumin and globulin. Irradiation of infrared ray effected marked decrease of the 2nd wave and combination with steroid was almost nil. The effect of ultraviolet ray on the 3rd wave was greater that of infrared ray while the effect of the infrared ray was greater on the 2nd wave. Serum albumin was found to be affected more than serum globulin. The steroid combined with serum albumin was not separated by ultraviolet ray but was separated by infrared ray, but when it was combined with serum albumin free steroid combined with serum albumin was produced by both ultraviolet ray (especially that in a shorter wave-length region) and infrared ray. The amount of steroid generated by ultraviolet irradiation increaseswith length of irradiation time but no periodical increase was observed by infrared irradiation.
During the extraction of poisonous component from the bark of Illicium Anisatum L., a flavonoid glycoside was obtained as crystals and the substance was identified as quercitrin (quercetin 3-rhamnoside).
Determination of xanthine bases by amperometric titration was examined. Xanthine bases react quantitatively with silicotungstic acid in hydrochloric acid solution. A white precipitate is formed by bonding of 1 mole of silicotungstic acid with 4 moles of theobromine or theophylline. A trace of xanthine bases can be determined rapidly by titration under following conditions: Ec=-0.80V vs. S. C. E.; concentration of hydrochloric acid during titration, above 1.5N; mercury drop electrode.
3, 3-Dibromo-2-piperidone (II), obtained by bromination of 2-piperidone in the presence of phosphorus pentachloride, was submitted to reduction with Raney nickel at atmospheric pressure, in the presence of equimolar amount of triethylamine, and 3-bromo-2-piperidone (III) was obtained. Its hydrolysis with conc. hydrochloric acid and treatment of the crude product with conc. sodium hydroxide effected cyclization to (V). This was converted to (VI) with hydrochloric acid and subsequent treatment with silver carbonate followed by hydrogen sulfide gave DL-proline.
Antiviral activity against Japanese B encephalitis virus and rabies-fixed virus was examined with the following five kinds of diaryl sulfone compounds prepared in the present series of work, bis[4-(4-hydroxyphenylazo) phenyl] sulfone, bis[4-(2-butenylidene)-hydrazinophenyl] sulfone, bis[4-(1-carboxyethyliedne) hydrazinophenyl] sulfone, bis[4-(1-methyl-3-carboxypropylidene) hydrazinophenyl] sulfone, and bis(4-cyclohexylidenehydrazinophenyl) sulfone, together with 21 kinds of componds which were described in previous papers. None of these compounds showed inactivation of the viruses.