The binding of methyl orange with bovine lens protein, α-crystallin and β-crystallin, was studied in various pH values by equilibrium dialysis method in 0.1 M phosphate buffer of pH 4.7∼8.9. At equilibrium, the relationship between the amount of bound methyl orange and the concentration of methyl orange followed the Langmuir-type equation for each protein. The maximum possible number of methyl orange bound to each protein remained constant independent of the pH values examined and the number with α-crystallin was 10-4 (mole methyl orange/g. α-crystallin) and that with β-crystallin was 10-5. Binding constant (k) with each protein decreased markedly with increasing pH. The binding affinity (Δμ) of methyl orange with each protein was calculated, variation of Δμ with pH values was examined, and the effect of protein charge at the binding was discussed. It was concluded that in the binding of methyl orange with these proteins, the electrostatic force was the most important factor and that the binding with α-crystallin was greatly affected by electrostatic force than that with β-crystallin.
Thin-layer chromatography of 83 p-aminoazobenzene derivatives, which include carcinogenic azo dyes and their metabolites, was carried out on Silica Gel H plates. Simple method to determine the structure of a dye was devised ; an azo dye was reduced with sodium hydrosulfite to aniline derivatives which were identified with the authentic samples on the thin-layer plates. Rf values of 46 aniline derivatives related to the azo dyes are also listed.
Alkaloids in Formosan Stephania sasakii HAYATA isolated to date include tertiary bases cepharanthine (I), crebanine (II), berbamine (III), and phanostenine (IV). Examinations for water-soluble quaternary bases in this plant afforded a new base as a chloride of m.p. 159.5∼160°, [α]D ±0°(MeOH), C21H22O5NCl=C15H7(OCH3)4 (N+CH3)·(C=O) Cl-. Its reduction with sodium borohydride gave β-hydroxylaudanosine (IV). Comparison of ultraviolet, infrared, and NMR spectra of the original base with those of N-methylpapaveraldinium (VII) chloride proved their identity. This is the first time that N-methylpapaveraldinium (N-methylxanthalinium) (VII) was detected from a plant.
From the root tuber of Stephania rotunda LOUREIRO, (-)-tetrahydropalmatine (I) and stepharine (IIa) were isolated together with a new phenolic base of tetrahydroprotoberberine type, stepharotine (hydrobromide, m.p. 227∼229°, [α]34D-203°(MeOH), C31H25O5N·HBr). The structure of O-methylstepharotine (m.p. 133∼135°, [α]35D -270°(CHCl3), C22H27-O5N) was identified by comparison with the synthesized 2, 3, 9, 10, 11-pentamethoxytetra-hydroprotoberberine (VI a). The mass spectrum of stepharotine trideutero-O-methyl ether (VI b), revealed that the phenolic hydroxyl in stepharotine is present in either 9-, 10-, or 11-position. Cleavage reaction of stepharotine O-phenyl ether (VI c) with metallic sodium in liquid ammonia gave (-)-tetrahydropalmatine (I) and (-)-tetrahydropalmatrubine (V c). These experiments revealed that stepharotine is represented by formula III.
Anti-inflammatory action of several proteases and α-amylases, especially protease from Streptomyces griseus and α-amylase from Bacillus amylosolvens, was examined and their anti-edematous action in rat hind paw was observed. Anti-inflammatory mechanism was examined, and following results were obtained : Anti-inflammatory action of protease was marked in egg-white and dextran edema, and that of α-amylase in croton oil edema. Enzyme activity and anti-inflammatory action were not always in parallel, So were also increase in capillary permeability and anti-edematous action. Extract of inflammatory fluid activated spontaneous contraction of isolated rat uterus and this activation was not antagonized by atropine and diphenhydramine. Some components in intraperitoneal fluid of enzyme pretreatment inhibited spontaneous contraction of isolated rat uterus, but these components were not identified.
The dose-response relationships of phenethylamine derivatives and pentylamine as substrate or inhibitor of rat liver monoamine oxidase were investigated. Increased concentration of tyramine increased the rate of oxidative deamination which later decreased. This bell-shaped dose-response curve of tyramine agreed fairly well with the theoretical curve calculated from eq. (3). Dissociation constants of tyramine-monoamine oxidase complex were estimated to be Km=4.5×10-4M as a substrate and KI'=1.0×10-2M as an inhibitor. Phenethylamine, 4-methoxyphenethylamine, and pentylamine were more or less active as substrates of monoamine oxidase, but also inhibited monoamine oxidase at higher concentration partially non-competitively. From these results, it is concluded that these amines act both as a substrate and an inhibitor, and the dose-response curves of these amines can be represented by eq. (3).
Activities of aliphatic primary alcohols and amines on rat liver monoamine oxidase were investigated. Aliphatic alcohols and amines inhibited monoamine oxidase and, as the number of carbon atoms increased, 50% inhibitory concentration decreased. Comparison of the inhibitory activities of alcohols and amines showed that amines inhibited monoamine oxidase at lower concentration than alcohols having the same number of carbon atoms. Moreover, while the inhibitory action of alcohols was reversible, that of amines was partially irreversible. These results suggest some specific affinity of amines on monoamine oxidase. Some of aliphatic amines were oxidized by monoamine oxidase and a part of their inhibitory action on monoamine oxidase was proved to be the attack of active center of the enzyme, but the inhibitory action of aliphatic alcohols and amines was estimated to be mainly non-competitive.
Inhibitory action of several compounds on rat liver monoamine oxidase was investigated according to Ferguson's rule and Mullins' hypothesis. Aliphatic primary alcohols, their acetates, aralkyl alcohols, phenethylamines, and piperidinoacetophenones decreased monoamine oxidase activity to about one-half of non-treated preparation at the concentration giving thermodynamic activities of 10-1 to 10-2. These results suggest that these compounds might be classified as drugs having a "physical toxicitiy" and affected monoamine oxidase by their physicochemical properties. Volume fractions of alcohols were also calculated and a theoretical curve calculated from δblo=11.5 and XV=6.7 fltted the experimental values.
Attempts were made to examine the effect of substituents at 1-position of 6 (1H)-pyridazinone ring on ring-contraction reaction into pyrazole ring. It was concluded that methyl and non-substituted derivatives do not undergo ring-contraction, but para-substituted phenyl derivatives do regardless of the electronic character of substituents in the para position.
Treatment of ethyl 1-phenyl-2, 3-dioxo-4-pyrrolidinecarboxylate (I) with excess urea afforded the corresponding 2, 4-dihydroxy-6-phenylpyrrolo[3, 4-d]pyrimidin-7-ones (II). Ethyl 3-amino-1-phenyl-2-oxo-4-pyrrolinecarboxylate (III), prepared by the Leucalt reaction of I with ammonium acetate, gave 2-amino(and 2-methyl)-4-hydroxy-6-phenylpyrrolo[3, 4-d]pyrimidin-7-ones (IV) and (V) by reaction with guanidine hydrochloride (and acetamidine hydrochloride) by the use of a base catalizer. Heating of IIIa in formamide gave a small amount of VI.
Syntheses of 2-[2-(5-nitro-2-furyl) vinyl]-5-ethoxy (or chloro)-1, 3, 4-oxadiazole (III or V) and 4-substituted 2-[2-(5-nitro-2-furyl) vinyl]-1, 3, 4-oxadiazolin-5-ones are described. When 1-[3-(5-nitro-2-furyl) acryloyl]-2-ethoxycarbonylhydrazine (II) was refluxed with excess phosphoryl chloride, it gave III and 2-[2-(5-nitro-2-furyl) vinyl]-1, 3, 4-oxadiazolin-5-one (IV). However, when equivalent amount of phosphorus pentachloride was employed instead of phosphoryl chloride in this reaction, the main product was V. IV and 2-(5-nitro-2-furyl)-1, 3, 4-oxadiazolin-5-one (VI), when refluxed with equivalent amount phosphorus pentachloride, gave V and 2-(5-nitro-2-furyl)-5-chloro-1, 3, 4-oxadiazole (VII), respectively. 4-Substituted 2-[2-(5-nitro-2-furyl) vinyl]-1, 3, 4-oxadiazolin-5-ones (VIIIa∼i, IXa∼c) were prepared by acylation and disubstituted aminomethylation of IV and the products are shown in Tables II and III. Other oxadiazoline derivatives, 4-pyrimidinyl-2-[2-(5-nitro-2-furyl) vinyl]-1, 3, 4-oxadiazolin-5-ones (XIa, b) were obtained by the action of phosgene on the dioxane solutions of 1-[3-(5-nitro-2-furyl) acryloyl]-2-pyrimidinylhydrazines (Xa, b). Most of these nitrofuran compounds were found to be active against Escherichia coli NIHJ, Staphylococcus aureus FDA 209 P, and Trichomonas vaginalis 4 FM.
Photodecomposition of propericiazine (I) and its sulfoxide (II) was carried out by irradiation of fluorescent light (twelve 20 W tubes of cool white light, ca. 5000 luces) on their acid solution sealed in ampules, under various conditions. I is stable to light in nitrogen atmosphere but is photooxidized in thepresence of air, forming II in the main and partly 7-oxo-2-phenothiazinecarbonitrile (III), which was found to be responsible for the red coloration of the solution. II was found to be photoreduced to I under various conditions. Quantitative studies were made on the photodecomposition of I and II in buffer solution, by colorimetric determination using a palladium chloride reagent. At the same time, effect of various additives on the photodecomposition was also examined. It was thereby found that a certain equilibrium is reached between I and II during the process of photodecomposition and this equilibrium is dependent on pH of the solution and content of oxygen in the ampules. Rate of photooxidation increased with increasing amount of oxygen and lowering of pH, while the rate of photoreduction decreased with increasing pH and amount of oxygen. Photooxidation is not affected by the addition of maleic or picolinic acid but photoreduction is inhibited by these additives.
Photodecomposition of thioproperazine (III), acetylpromazine (V), and its sulfoxide (VI) was carried out by irradiation of fluorescent light on their acid solution and in the lighting cabinet described in the preceding paper. III rapidly underwent photooxidation in air, in the presence of maleic acid, to its sulfoxide (IV). This maleic acid is known to prevent photohydrolysis of the sulfamoyl group in the 2-position of phenothiazine ring. Stability of V and VI to light was compared with that of propericiazine and its sulfoxide, and that of III and IV, and it was found that V and VI are more stable than the other compounds.
Photodecomposition of phenothiazine drugs having a methoxyl group in the 2-position, such as methopromazine (I) and levomepromazine (V), was carried out. Irradiation of I in aqueous acid solution produced methopromazine sulfoxide (II) and bimethopromazine (3, 3'-bi[2-methoxy-10-(3-dimethylaminopropyl) phenothiazinyl]) (III). Results of elemental analyses, molecular weight determination, and NMR spectral measurements indicated that III is a dimer of I in which two phenothiazine rings are bonded in the 3-position. Further irradiation of III resulted in its oxidation to bimethopromazine monosulfoxide (IV). Similar results were obtained by the photooxidation of V. Levomepromazine sulfoxide (VI) was submitted to photoreduction in nitrogen atmosphere. In a simple aqueous solution, V was first formed and bilevomepromazine (VII) then appeared as the main product. V was the main product in the presence of ascorbic acid. This photoreduction was inhibited by the addition of maleic acid.
Excess dietary zinc resulted in the lowered growth, hemoglobin concentration, and ceruloplasmin activity, with significant decrease in the concentration of serum copper. The addition of 0.2% copper to the diet containing 1.0% zinc was effective in preventing anemia and decrease in ceruloplasmin activity. Administration of a large amount of copper alone caused an increase in serum copper and a marked increase in ceruloplasmin activity (Table I and II). The animals receiving excess dietary zinc developed a mild anemia and showed a significant decrease in ceruloplasmin activity, When copper-histidine chelate was administered intraperitoneally to the animals, the decrease in ceruloplasmin activity recovered, but the concentration of hemoglobin was little affected (Fig. 1). These results suggested that the excess dietary zinc induced a copper-deficient state, and an antagonistic correlation existed between zinc and copper.
2-[2-(5-Nitro-2-furyl)vinyl]-2-imidazoline (VI) was prepared by the following route. Reaction of ethyl 3-(5-nitro-2-furyl) acrylimidate (IV) with ethylene diammonium bis-p-toluenesulfonate gave 2-[2-(5-nitro-2-furyl)vinyl]-2-imidazoline p-toluenesulfonate (V) in good yield, which was treated with cold sodium hydroxide solution to separate the free amine (VI). Antimicrobial activities of these nitrofuran compounds were tested.
Ground part of Euphorbia adenochlora MORR. et DECNE. was extracted with petroleum benzine and the extract was treated by chromatography. A wax-like nonacosane (?), m.p. 54∼55°, and β-sitosterol were isolated, besides taraxerone and taraxerol, a five-membered ring triterpene, and cycloartenol and euphol, a four-membered ring triterpene.
The terrestrial portion of Euphorbia watanabei MAKINO was found to contain, besides nonacosane (?), tetracosanol, a wax-like, m.p. 59∼60°, and β-sitosterol, five-membered ring triterpenes taraxerol acetate, lupeol acetate, alunusenone (?), taraxerone, luperone, and taraxasterol. It is interesting from botanical aspects that euphane series triterpenes, which would be anticipated from Euphorbia spp., were not obtained.
The structure of neoxyberberine acetone which was obtained from acetone-berberine (I) by potassium permanganate oxidation in aqueous acetone at room temperature is discussed. From a consideration of the data of nuclear magnetic resonance spectra and some chemical reactions, the structure (III), C23H25O6N, previously reported was revised, and an alternative bridged structure (IV), C23H23O6N, was proposed. Some related compounds were also synthesized.