The venom of “Mamushi” (Agkistrodon halys blomhoffi BOIE) was submitted to chromatography over diethylaminoethylcellulose and substances with phosphodiesterase activity were obtained as two large peaks. These were designated diesterase-I and -II. The diesterase-I was further submitted to chromatography over carboxymethylcellulose and a peak with phosphodiesterase activity was found besides the large peak. This was found to be different from either diesterase-I or -II and was named diesterase-III. The diesterases-I, -II, and -III hydrolyzed RNA and DNA besides calcium bis [bis(p-nitrophenyl)phosphate] and the ratio of activities to these three substrates were different between diesterase-I and -II, or -III.
Of the three kinds of phosphodiesterase fractionated from the venom of Agkistrodon halys blomhoffi BOIE by the combination of diethylaminoethyl- and carboxymethylcellulose, diesterase-I with the largest yield showed marked activity against deoxyoligonucleotides and hydrolyzed ca. 70% of RNA to form the corresponding 5′-mononucleotides and left non-dialyzable oligonucleotide. Activity of diesterase-III to deoxyoligonucleotides is weak but it hydrolyzes RNA completely. Characteristics of diesterase-II are still obscure at present. These three kinds of diesterase show similar behavior to optimal pH, stability, and effect of group reagents but behave differently to the effect of metal ions, Diesterase-I is not affected by magnesium ion, and diesterase-II and -III are activated by it. The nucleotide pyrophosphatase and phosphodiesterase of snake venom are considered to be the same enzyme.
It has been reported that diphosphopyridine nucleotide is decomposed by snake-venom enzyme to form nicotinamide but there has been no proof that this decomposition is effected by diphosphopyridinenucleotidase. This nucleotidase activity was examined in nine kinds of Formosan snake venom and the presence of this diphosphopyridinenucleotidase was found in the venom of Bungarus multicinctus and Trimeresurus gramineus. The nature of this nucleotidase of snake venom was also examined and this activity was found to be inhibited non-competitively by nicotinamide and isonicotinic hydrazide. This enzyme also possesses transglycosidase activity and produces isonicotinic hydrazide-diphosphopyridine nucleotide from these two components. The diphosphopyridinenucleotidase in the venom of Trimeresurus gramineus was purified by column chromatography using carboxymethyl- and diethylaminoethylcellulose, and a sample with six-fold increase in specific activity was obtained. This sample is totally free from phosphodiesterase.
In order to obtain pyridinecarboxylic acid derivatives with strong antibacterial action from alkylpicolinic acid, two kinds of compound (A and B) possessing a strong electron-releasing group substituted in 4-position of the pyridine ring were synthesized. (A) compounds, 4-X-picolinic acids (X=OMe, OEt, OPh), were prepared by substitution of the nitro group in 4-nitro-2-methylpyridine 1-oxide with X, followed by deoxygenation and permanganate oxidation. (B) compounds, 4-Y-5-ethylpicolinic acid (Y=OMe, OEt, Cl), were prepared also by substitution of the nitro group in 5-ethyl-2-methyl-4-nitro-pyridine 1-oxide, followed by change of 2-methyl with acetoxymethyl by the Boekelheide rearrangement, deacetylatian with hydrochloric acid, conversion to the formyl group by the Ball reaction, and mild oxidation with silver oxide or hydrogen peroxide to the objective carboxylic acid.
The structure of a new quaternary base, phellodendrine, isolated from Phellodendron amurense RUPR. (Japanese name “Kihada”) and Ph. amurense RUPR. var. sachalinense FR. SCHM. (Japanese name “Hiroha-kihada”), was found to be not the assumed benzyltetrahydroisoquinoline type (I) as reported in the preceding paper but a tetrahydroprotoberberine-type base. The O, O-dimethylphellodendrine methine derived from this base was found identical with norcoralydine methylmethine (VI) and O, O-dimethyl-N-demethylphellodendrine with L-(-)-norcoralydine (VIII). Therefore, the structure of phellodendrine was assumed to be represented by formula (IV).
The position of two phenolic hydroxyls in phellodendrine was examined. Oxidation of L-(-)-O, O-diethyl-N-demethylphellodendrine (IV) with potassium permanganate results in formation of 1-oxo-6-methoxy-7-ethoxytetrahydroisoquinoline (VI) and 4-methoxy-5-ethoxyphthalic anhydride (V). Since the infrared spectrum in chloroform solution is identical with that of dl-O.O-diethylcoreximine, phellodendrine was determined as L-(-)-N-methylcoreximine (VII).
Crystallization temperature of aqueous solutions of dl-, d-, and l-threo-1-p-nitrophenyl-2-amino-1, 3-propanediols was measured in various concentrations and respective supersolubility curves approximately parallel to ordinary solubility curves were obtained. It was thereby found that these substances tended to form a stable supersaturated solution and that this tendency was especially strong in the racemic compound. The direct resolution of optical isomers proposed by Amiard was elucidated from this supersolubility curves and the solubility curves reported in the preceding paper, and its reason was clarified. These results provided fundamental data for a more theoretically advantageous procedure.
The plant material used in the present series of experiments is different from the plant which is now known to be Angelica dahurica (FISCH.) BENTH., from whose fruits Noguchi and others had earlier extracted furo-coumarins like imperatorin, phellopterin, and byak-angelicin. This plant is an allied species of Angelica pubescens MAXIM. and some workers consider the former to be a synonym of the latter. Three kinds of coumarin derivative were isolated from the fruit of this plant by the method shown in Fig. 3; (I), C17H16O6, m.p. 86-87°, (II), C16H14O5, m.p. 140-141°, and (III), C9H6O3, m.p. 223-225°. (I) formed isobyak-angelicolic acid (V) by saponification and byak-angelicin (VII) on being boiled with 1% oxalic acid, so that the structure of (I) was considered to be the same as byak-angelicol. However, infrared spectrum and other physical properties of (I) do not agree with those of byak-angelicol and further examinations of the structure of (I) seemed necessary. From its melting point, composition, and chemical properties, (I) is assumed to be identical with ferulin. The melting point, composition, and derivatives of (II) were found to be identical with those of oxy-peucedanin, and (III) was found by mixed fusion to be umbelliferone. Besides these three substances, a small amount of a neutral substance melting at 223-225° but not identical with (III) was obtained.
In continuation of the preparation of dyes having 2, 5-dimethyloxazole as the fundamental ring, five kinds of styryl-type dyes and 12 kinds of aminovinyl-type dyes were prepared with 2, 5-dimethyl-4-isobutyloxazole ring as the fundamental skeleton, and nine kinds of styryl-type dyes with 2, 4, 5-trimethyloxazole as the fundamental ring and 9 kinds of aminovinyl-type were prepared. 2, 5-Dimethyl-4-isobutyloxazole was prepared by acylation of dl-leucine to 3-acetamido-5-methylhexan-2-one and its dehydrative cyclization. 2, 4, 5-Trimethyloxazole was prepared similaly from dl-alanine through 3-acetamidobutan-2-one. Antibacterial tests on these dyes are reported.
Following the discovery of few compounds useful as antipyretic-analgesic among dimethylamide derivatives of N-substituted glycine, dimethylamide (X to XV) of N-p-alkoxyphenylglycine and dimethylamides (XVIII to XXV) of N-acetyl-N-p-alkoxyphenylglycine were prepared in order to examine variation in pharmacological effect and toxicity by the increase or decrease in number of carbons of the alkyl group of ethoxyl in N, N-dimethyl-2-(p-ethoxyanilino) acetamide.
As a part of studies on the syntheses of new antipyretic-analgesics, a number of N-o-alkoxyphenylglycine dimethylamides (XVI to XXI) and their N-acetylated compounds (XXII to XXVII) were prepared in order to compare pharmacological activities and the change in pharmacological action according to the number of alkyl group in alkoxyl in para-substituted aryl derivatives of N-arylglycine dimethylamide reported in the preceding paper. N-p-Ethoxyphenylglycine amides were derived to respective piperidide (XXVIII), morpholide (XXIX), and pyrrolidide (XXX), which were acetylated to form N-acetyl-p-ethoxy-phenylglycine amides (XXXI to XXXIII). N-Acetyl-p-ethoxyphenylglycine diethylamide (XXXIV) was obtained directly from acetophenetidine.
Heating of thebaine with dil. hydrochloric acid, followed by addition of 4-amino-antipyrine and potassium ferricyanide, and basification of this mixture with ammonia results in red coloration, the color changing to reddish brown on heating to 100°. The limit of detection by spot reaction is 4γ/0.05cc. Presence of morphine, codeine, narcotine, and papaverine does not interfere in this reaction as these alkaloids do not undergo coloration. By utilization of this coloration reaction, colorimetric determination of thebaine can be made in the presence of morphine, codeine, narcotine, and papaverine. The procedure of the colorimetry is as follows: One cc. of the test solution (aqueous solution of thebaine salt containing 40-310γ/cc. of thebaine) is placed in a 10-cc. measuring flask, 1cc. of 7% hydrochloric acid is added, and the mixture is warmed in a boiling-water bath for 25 minutes. When cooled, 1.5cc. of 0.5% 4-aminoantipyrine solution and 0.6cc. of 1% potassium ferricyanide solution are added and the mixture is shaken. After exactly 1 minute, 2cc. of 10% ammonia water is added, shaken again, and warmed in a boiling-water bath for 5 minutes. The mixture is cooled with tap water to room temperature, diluted with 1cc. of ethanol and water to make 10cc., and absorbance of the solution is measured at 435mμ. Blank test is carried out with the same reagents and in a same manner using 1cc. of water in place of the test solution.
Pechmann reaction was carried out on α-acetyl-γ-butyrolactone with 73% sulfuric acid or phosphoryl chloride as the condensation agent. Corresponding 4-methyl-coumarin derivatives were obtained from resorcinol, pyrogallol, phloroglucinol, and 1-naphthol and ultraviolet absorption spectra of these derivatives were measured. 7-Hydroxy-3-(2-hydroxyethyl) derivative, m. p. 226-228°; 7-hydroxy-3-(2-chloroethyl) derivative, m. p. 186-188°; 7, 8-dihydroxy-3-(2-hydroxyethyl) derivative, m. p. 241-242°; 7, 8-dihydroxy-3-(2-chloroethyl) derivative, m. p. 206°; 5, 7-dihydroxy-3-(2-chloroethyl) derivative, m. p. above 300°; 7, 8-benzo-3-(2-hydroxyethyl) derivative, m. p. 164°.
The method of quantitative and qualitative analyses by the use of infrared absorption spectra was examined for dyes used for foodstuff (Ponceau 3R, Amaranth, Erythrosine, Ponceau SX, Oil Red XO, Ponceau R, New Coccine, Eosine, Phloxine, Rose Bengal, and Acid Red) and one not used for foodstuff (Rhodamine B). Each of these dyes was able to be identified in a limit of 40-300γ by the characteristic bands (Table I). Attempted quantitative determination of Rose Bengal (I) and Rhodamine B (II) showed that they can be determined by the use of a characteristic band at 961cm-1 for (I) with standard deviation of 0.519 and at 1078cm-1 for (II) with standard deviation of 0.633. Separately determination of a mixture was tested with a mixture of (I) and (II). The key bands used were that at 961cm-1 for (I) and at 1078cm-1 for (II), and standard deviation of analytical result of each component in the mixture was presumed from Youden's method, by which quantitative determination was found possible with standard deviation of 0.624 for (I) and 0.659 for (II).
Water-soluble derivatives of phenobarbital were prepared in order to test their pharmacological action. Derivatives were prepared from phenobarbital and sodium salt of its p-chloro derivatives by the route shown in the Chart and the compounds obtained are listed in Tables I and II. Results of pharmacological test on these derivatives have been reported elsewhere.
A new synthetic process for thioctic acid on industrial scale is described. The use of sodium borohydride was therefore avoided. Addition of benzyl alcohol to methyl 6-oxo-7-octenoate (III) gave methyl 6-oxo-8-benzyloxyoctanoate (IV) which was reduced over Raney nickel to methyl 6, 8-dihydroxyoctanoate (VI), and its O-tosylate was reacted with sodium disulfide to form the 1, 2-dithiolane ring and derived to thioctic acid.
As a process for synthesis of thioctic acid, a new way of forming 1, 2-dithiolane ring was utilized and the acid was prepared by decomposition of the “Bunte” salt obtained by the reaction of methyl 6, 8-dihaloöctanoate (II and III) and methyl 6-bromo-8-mercaptoöctanoate (VIII) with sodium thiosulfate.
As a new process for synthesis of selenoctio acid, a process is described for the reaction of methyl 6, 8-dichloroöctanoate (I) and the “Bunte” salt thereby obtained, same as reported in Part II of this series, submitted to decomposition to form the objective acid.
A new process for formation of a carbon chain of thioctic acid is described, by the condensation of adipic acid ester chloride and acetylene to form methyl 6-oxo-8-chloro-7-octenoate (II) and the formation of thioctic acid from methyl 6-oxo-8, 8-dimethoxyoctanoate (III), methyl 6-oxo-8-methoxy-7-octenoate (IV), and 6, 6, 8, 8-tetrakis-(benzylthio) octanoic acid (XVI), all derived from (II).
As a new process for synthesis of thioctic acid, formation of dihydrothioctic acid (X) by high-pressure reduction with cobalt polysulfide as a catalyst and formation of 1, 2-dithiolane ring from the ester of (IX) by the action of sulfonyl chloride are described. Materials used for this reaction were methyl 6-oxo-8-chloro-7-octenoate (I), obtained by condensation of adipic acid ester chloride and acetylene, and methyl 6-oxo-8, 8-dimethoxyoctanoate (III), 6-oxo-8, 8-diacetoxyoctanoate (IV), 6-oxo-8-thiocyano-7-octenoate (V), dimethyl 8, 8′-thio-bis (6-oxo-7-octenoate) (VI), methyl 6-oxo-8-acetyl-thio-7-octenoate (VII), and methyl 6-oxo-8, 8-bis (benzylthio) octanoate (VIII), all derived from (II).
Presence of a cofactor or cofactors necessary for increased production of streptolysin-S of Streptococcus by ribonucleic acid in peptone or meat infusion had already been assumed by Bernheimer and by Hosoya and Egami. In order to purify and separate this factor from meat infusion, various examinations were made and a non-dialyzable polypeptide was successfully separated which indicated a high degree of cofactor activity biologically and which was identified as chemically uniform. The route of this separation was as follows: Removal of a substance precipitating with alcohol; collection of a substance from its supernatant, precipitation with acetone, removal from this acetone precipitate a substance precipitating in ammonia alkalinity (pH 8.6), and dialysis of the centrifugation supernatant against distilled water. The dialysate was passed through a column of Amberlite IRA-411 (Cl form) and the initial effluent therefrom was dialyzed, followed by lyophilization.
Ethyl 6-hydroxycomenate (R) was obtained by bromination of comenic acid followed by hydrolysis and esterification but the shape of its ultraviolet spectrum, alkali titration curve, and formation of a metal chelate compound are different from those of other meconic acid derivatives. Moreover, its ultraviolet spectrum in aqueous solution is extremely labile and it was considered that the acid might have changed into 2-pyrone derivative during the course of this synthesis. However, the compounds formed by methylation of the two hydroxyls to O-methyl were negative both to coloration for β-diketone and to ferric chloride, and the coloration became positive only after ring cleavage by basification. Such evidences and the fact that methoxyacetone was formed by alkaline decomposition proved that the original compound was actually ethyl 5, 6-dihydroxy-4-pyrone-2-carboxylate. The extreme lability of ultraviolet spectrum of this compound in aqueous solution was considered to be due to its facile ring cleavage to form β-keto acid (cf. Chart 1). This compound (R) shows two kinds of coloration to ferric chloride, one of violet and the other of red, and this was found to be due to the strong reducing effect of the enediol, the mechanism of coloration to violet and red being respectively represented as shown in Charts 2 and 3.
The product of the condensation of 5-amino-1, 2, 4-triazole (I) and diethyl ethoxy-methylenemalonate (II) was proved to be 6-ethoxycarbonyl-7-hydroxy-1, 2, 4-triazolo-[2, 3-α]pyrimidine (XI). Reaction of ethyl 2-cyano-3-ethoxyacrylate afforded 6-ethoxycarbonyl-7-amino-1, 2, 4-triazolo[2, 3-α]pyrimidine (XTV), 6-cyano-7-hydroxy-1, 2, 4-triazolo [2, 3-α]pyrimidine (XV), and ethyl 2-cyano-3-(1, 2, 4-triazol-5-ylamino) acrylate (XXI). 5-Hydroxy-6-ethoxycarbonyl-1, 2, 4-triazolo[4, 3-α]pyrimidine (X) and 5-hydroxy-6-cyano-1, 2, 4-triazolo [4, 3-α] pyrimidine (XX) were prepared and these were found to undergo isomerization on being heated to fusion, forming (XI) from (X) and (XV) from (XX).
Application of benzyl chloride to 5-methyl-7-hydroxy-1, 2, 4-triazolo[4, 3-α]pyrimidine (II), m. p. 300°(decomp.), obtained by the reaction of 2-hydrazino-4-hydroxy-6-methyl-pyrimidine (I) and formic acid, gave three kinds of benzyl derivatives. One of these agreed with 5-methyl-8-benzyl-1, 2, 4-triazolo[4, 3-α]pyrimidin-7(8H)-one (VII) prepared from 2-hydrazino-3-benzyl-6-methyl-4(3H)-pyrimidone (VI) and formic acid. Heating of 3-phenyl-1, 2, 4-triazolo[4, 3-α]pyrimidines (XIV: R=H and CH3) in formic acid results in isomerizatlon to 2-phenyl-1, 2, 4-triazolo[2, 3-α]pyrimidine (XVI: R=H and CH3) which can be obtained by oxidation of N-(2-pyrlmldlnyl) benzamldlnes with lead tetraacetate.
The extract obtained by microbiological oxidation of Reichstein's substance S (I) by Corticium sasakii was submitted to paper partition chromatography with the solvent system of propyleneglycol/toluene·dioxane (7:3) and was separated into A, B, C, D, and E zones (Fig. 1). E zone was found to be the starting material (I), C zone, the hydrocortisone (II), and A zone was found to be a mixture of 11-epihydrocortisone (III) and a monohydroxylated substance S (IV), m. p. 233-236°, [α]D+143°(EtOH), +127°(dioxane), U.V.λmaxEtOH243.5mμ.
The monohydroxylated substance S (IV), m. p. 233-236°(decomp.), obtained by microbiological oxidation of Reichstein's substance S (I) with Corticium sasakii, was oxidized with sodium bismuthate and afforded the known 19-hydroxy-4-androstene-3, 17-dione (VI), m. p. 165-167°, whose further oxidation with chromiumn trioxide gave 3, 17-dioxo-4-androsten-19-al (VII), m. p. 129-133°. Reduction of (IV) with zinc and acetic acid afforded the known 19, 21-dihydroxy-4-pregnene-3, 20-dione (VIII), m. p. 153-156°. From these experimental results, (IV) was proved to be 17α, 19, 21-trihydroxy-4-pregnene-3, 20-dione which is also obtained by enzymatic oxidation of progesterone or deoxycorticosterone by adrenal homogenate. The oxidation of a methyl at the ring juncture (C-19) of a steroid by microörganism was effected for the first time by Corticium sasakii.
Adaptive formation of mandelic acid oxidase was attempted using the ghost of Pseudomonas fluorescens and incubated with l-mandelic acid, l-mandelic acid and supernatant fraction, l-mandelic acid and supernatant fraction added with ATP, and l-mandelic acid, casamino acids, ATP, and fructose 1, 6-diphosphate. In none of these cases, formation of the enzyme was noticed. Activity of succinic acid oxidase, α-ketoglutaric acid oxidase, and p-phenylenediamine oxidase was much higher in the ghost of Ps. fluorescens and insoluble fraction from disrupted cells than that of the soluble fraction.
Pseudomonas fluorescens was suspended in hypertonic phosphate buffer and sound waves of 10kc, 100w, were passed through the suspension to disrupt the cells. Even when the cells were highly disrupted, viable cells were found in the insoluble fraction. Therefore, formation of mandelic acid oxidase was examined using the insoluble fraction containing viable cells and incubated with addition of mandelic acid alone and mandelic acid, casamino acids, and ATP. The ability of enzyme formation in the insoluble fraction was approximately comparable to the ability of enzyme formation shown by living cells remaining in that fraction.
Infrared absorption spectra were measured of orotic acid, barbituric acid and its derivatives which have NH-CO-NH group in its molecule and water of crystallization, and assignment of absorption bands of νOD and νND produced by deuteration was made. The anhydrate of calcium 5-ethyl-5-pentylbarbiturate showed the shift of νC=O to the higher wave number and νC=N to lower wave number. This was considered to be due to the degree of ionic bonding between calcium and oxygen in the 2-position. On the other hand, the anhydrate of deuterated barbituric acid showed the shift of νND absorption band to the higher wave number and this was assumed to be due, not to the electronic state of the componental atoms in the molecule but to other causes such as the changes in interaction between molecules, from magnetochemical data.
Application of hydroxylamine to N-(p-acetamido- or p-amino-benzenesulfonyl)-2-acetylpropionamide (Ia or Ib) gives their oximes (IIa or IIb) and dehydrative cyclization of these oximes with chlorosulfonic acid as the dehydrative agent in glacial acetic acid affords 5-(p-acetamido- or p-amino-benzenesulfonamido)-3, 4-dimethylisoxazole (IIIa or IIIb), which were confirmed by admixture and ultraviolet spectral comparison with the products obtained by the reaction of N-(p-acetamido- or p-amino-benzenesulfonyl)-2-acetylpropionamidine (VI) and hydroxylamine, and also by preparation of p-aminomethylbenzenesulfonamide derivatives. Formation of oxime of 2-acetylpropionamide (VIII) and cyclization of this oxime were examined and it was found that although the oxime is formed easily, its reductive cyclization to 3, 4-dimethyl-5-aminoisoxazole was not effected and a different substance of m. p. 110-112° was obtained.
Examinations were made on the bovine testicular hyaluronidase, partially purified by zone electrophoresis for relationship between enzymic activity and intradermal diffusion of dye. Treatment of this hyaluronidase at 65° for 30 minutes results in disappearance of enzymic activity but the spreadig activity remains and this also occurs when enzymic activity is destroyed by tryptic digestion. The enzymic activity decreases to 25% by acetylation but the spreading activity was found to increase by this treatment. However, iodine treatment of hyaluronidase results in disappearance of both these activities.
Studies made to date suggest that bovine testicular hyaluronidase has a diffuing action due to mechanisms other than the enzymic action of hyaluronidase. However, this spreading activity is measured by intradermal diffusion of dye with a rabbit and for this reason there is a possibility of nonspecific diffusion due to foreign protein. The present series of work was undertaken in order to clarify this point. The hyaluronidase prepared from rabbit testis was found to have a spreading activity free from enzymic activity of a hyaluronidase and this spreading activity remained after disappearance of enzymic activity by heat treatment. These facts show that the diffusing action of bovine testicular hyaluronidase other than its enzymic action is not a nonspecific response by foreign protein.
Reaction of 2-methylacetoacetic ester (IV) and N4-acetylsulfanilamide (II a) or sulfanilamide (II b) in the presence of a basic substance like potassium carbonate results in dealcoholationcondensation at N1 in (II a) or (II b) to form p-aminobenzenesulfonyl-2-acetylpropionamide (Va or Vb). (IV) and (II a) do not react in the absence of a basic substance but (IV) and (II b) undergo condensation with dealcoholation at N4-position to form N-(p-sulfamoylphenyl)-2-acetylpropionamide (VI). These products, (Va), (Vb) and (VI), were confirmed by syntheses through another route, preparation of derivatives, and decomposition reactions.
As a process for synthesis of N1-acetyl-N1-(3, 4-dimethyl-5-isoxazolyl) sulfanilamide (I), there is the reaction of N1-(3, 4-dimethyl-5-isoxazolyl) sulfanilamide (II) and acetic anhydride, in the presence of an organic base, such as pyridine. According to reexamination of this reaction, yield of (I) is low due to formation of N1-(3, 4-dimethyl-5-isoxazolyl)-N4-acetylsulfanilamide (III) as a by-product. Use of excess acetic anhydride results in formation of N1, N4-diacetyl-N1-(3, 4-dimethyl-5-isoxazolyl)sulfanilamide (V) and the use of a great excess gives (V) quantitatively. In order to improve these points, the sodium salt (IV) of (I) was acetylated with acetic anhydride, without addition of an organic base and (I) was obtained in quantitative yield. Even the use of excess acetic anhydride failed to produce the by-product (V). This process was therefore found to be the selective acetylation of N1-position. The use of a calcium salt (VI) of (II), in place of its sodium salt (IV), also gives (I) in a good yield.
Acetylation of N1-(3, 4-dimethyl-5-pyrazolyl) sulfanilamide (III) with theoretical amount of acetic anhydride in the presence of pyridine results in the formation of N1-(3, 4-dimethyl-5-pyrazolyl)-N4-acetylsulfanilamide (IV) and N1, N4-diacetyl-N1-(3, 4-dimethyl-5-pyrazolyl) sulfanilamide (V), while acetylation of the sodium salt (VI) of (III) with theoretical amount of acetic anhydride in the absence of pyridine results in formation of (IV) and N1-acetyl-N1-(3, 4-dimethyl-5-pyrazolyl) sulfanilamide (VII). The structure of (VII) so formed was presumed by comparison of its ultraviolet absorption spectrum with those of N1-(3, 4-dimethyl-5-isoxazolyl) sulfanilamide (I), (III), and their derivatives.
Substrate specificity of L-amino acid oxidase in the venoms of Japanese “Mamushi” (Agkistrodon halys blomhoffi) and “Habu” (Trimeresurus okinavensis) was examined with 46 hinds of amino acid. The enzymes from these snake venom were stronger than that of cobra venom and were especially marked in oxidizing arginine and isoleucine. This specificity to these two amino acids has not been found in any literature, either in Japan or abroad, or in cobra venom.
Three kinds of 1-(x-bromobenzyl)-1, 2, 3, 4-tetrahydroisoquinoline derivative, with bromine in different position in each, were reduced with zinc dust and acetic acid. It was found that bromine atoms in 3- and 4-positions remain inert and only that in 2-position undergoes reductive debromination. Reduction with metallic sodium in liquid ammonia effects quantitative debromination in all these positions.
About 1mg. of phenylmercuric acetate in aqueous acid solution of pH 2-2.5, is titrated with 1.5×10-4M carbon tetrachloride solution of copper diethyldithiocarba-mate as the standard solution, the end point being slight tinting of the carbon tetrachloride layer to yellowish brown. Mean rate of determination error is less than -0.5% and assumed value of standard deviation is 0.5%.