After oral administration of an antitussive, Eprazinone hydrochloride [1-(2-ethoxy-2-phenylethyl-4-(2-bezoylpropyl) piperazine dihydrochloride](I), two metabolites were identified from both urine and bile of rats, rabbits, and humans. Three Dragendorff reagentpositive spots were detected on thin-layer chromatograms of the benzene extracts from urine and bile. Methanol-eluted substance of each of the spots isolated from the thin-layer plate was submitted to gas-liquid chromatography, infrared spectroscopy, and rechromatography over thin-layer with the authentic Eprazinone [1-(2-ethoxy-2-phenylethyl)-piperazine](II), and 1-(2-hydroxy-2-phenylethyl) piperazine (III). These results suggested that II was a product formed by the cleavage of an N-C bond of the Mannich base moiety in Ia, and III was formed by its deethylation.
An examinations was made to determine whether the previously known in vivo N-C cleavage of the Mannich base moiety of an antitussive, Eprazinone [1-(2-ethoxy-2-phenylethyl)-4-(2-benzoylpropyl) piperazine] takes place enzymically or nonenzymically. When the drug was incubated with phosphate buffer (pH 7.4) alone, whole blood of a rat, or a 9000×g supernatant of boiled rat liver in vitro, only a small extent of N-C cleavage occurred. However, the drug was cleaved significantly by its incubation with a 9000×g supernatant of a rat liver homogenate. The N-C cleavage of Eprazinone decreased in an anaerobic atmosphere or by incubation without NADP+ and glucose 6-phosphate. Further, the enzyme with this activity was induced by treatment of rats with phenobarbital, and was inhibited by SKF 525-A in vitro. These results suggested that the N-C bond of Mannich moiety of Eprazinone was cleaved by an oxygenase in rat liver. It is of interest that Eprazinone was selectively cleaved at the N-C bond of its Mannich base moiety by a hepatic oxygenase.
An improved method for microassay of blood cholinesterase activity is described. The method is an application of fluoromotric titration for acetic acid. The reaction mixture, composed of 100 μl of substrate, 50 μl of Tris buffer (pH 7.6), and 50 μl of sample, is incubated for 10-20 min at 30°, and 1000 μl of umbelliferone (10-6M)-isopropanol is added to the mixture. After centrifugation, 1000 μl aliquot of isopropanol supernatant, containing 0.2-15 μmoles of acetic acid, is titrated by fluorometry with 0.02N isopropanolic NaOH. Linearity of the reaction rate is obtained within 40 min of incubation. This method is applicable to colored and insoluble materials such as erythrocyte, in the same way as in the case of serum.
The structure of lyofoligenic acid (II), obtained by acid hydrolysis of lyofolic acid2) (I), was investigated in comparison with protolyofoligenic acid (III), which is the genuine aglycone of I and has already been chemically proved to possess the same carbon skeleton as cycloartenol5).(i) Acid c1eavage of the cyclopropane ring in III has been elucidated to give a 9(10→19)-abeo-lanost-5(10)-ene system (II) by preparing a cyclohexadienone derivative (VII).(ii) The conformation of 3α-OH in II and III was discussed on the basis of 1Hnuclear magnetic resonance (1H-NMR).(iii) Chemical behavior of the tertiary carboxylic function in II was examined and the 14α-COOH in II Was examined by using the shift reagent in the 1H-NMR analysis of XX and by 13C-NMR analysis of I, II, and III.(iv) The absolute configuration of hydroxyl groups at C-3 and C-24 was examined by applying the modified Horeau's method an XI and XIX, and both proved to be R. The structure II assigned to lyofoligenic acid is consistent with the result obtained by X-ray crystallographic study of XXII.
Mass spectra of the isomeric indanones, 4, 5-dimethoxy-, 5, 6'-dimethoxy-, and 6, 7-dimethoxy-1-indanone, were studied. These indanones equally exhibited the fragment ions (M-15), (M-43), (M-57), and (M-71) in their mass spectra. On the other hand, the ions (M-28), (M-58), and (M-29) are characteristic fragment ions for 5, 6-dimethoxy-, 4, 5-dimethoxy-, and 6, 7-dimethoxy- indanone, respectively. Based on these findings, the fragmentation mechanism for each isomer was discussed, and it was confirmed that the ion (M-29) in the case of 6, 7-dimethoxyindanone was produced by the loss of CHO group by the migration of hydrogen from ortho-methoxyl group to the carbonyl-oxygen.
The effect of sodium polyacrylate (PANa) on the aggregation and sedimentation state of pyrimidine penicillin G (PYG) was examined by the measurement of the apparent degree of aggregation (γm/γ0), amount of adsorption (X), ζ-potential, porosity (ε), and tortuosity (q) of the sedimentation layer. PYG of various primary particle radius (γ0) and PANa of various molecular weight (Mη) were used as the sample. It was found that the adsorption isotherms of PANa on PYG were of Langmuir type. Amount of saturated adsorption (X∞ : mg/g) increased with the increase of Mη and the decrease of γ0. A little increase of X∞ was found in the presence of NaCl. X∞ in mg/cm2 showed a little higher value with the decrease of γ0. η-Potential increased with the increase of PANa concentration and decreased in the presence of NaCl. PYG forms secondary particles in water by aggregation and its γm/γ0 and ε increased with the increase of PANa concentration and Mη. This tendency was more pronounced in the case of larger γ0. The aggregation effect of PANa on PYG decreased in the presence of NaCl. q decreased and qv/qH increased with the increase of PANa concentration. It was concluded that the anisotropy of the sedimentation layer increased by the nonspherical nature of the aggregated secondary particle grown up by PANa.
Both the release of enzymes and the lipid peroxidation were induced by the addition of hydrogen peroxide to the lysosome suspension prepared from rat liver homogenate. In this experimental model with hydrogen peroxide, Tinoridine inhibited the lipid peroxidation more strongly than the release of enzymes, at concentrations from 1.0 to 100 μM. At a high concentration of 1000 μM, Tinoridine did increase the release of enzymes, but markedly inhibited the lipid peroxidation. Moreover, Tinoridine inhibited the spontaneous release of enzymes from lysosomes by incubation without hydrogen peroxide. Tinoridine was also found to inhibit the lipid peroxidation in the 9000×g supernatant fraction of rat liver homogenate by incubation with or without hydrogen peroxide at concentrations from 1.0 to 1000 μM. These properties of Tinoridine seem to be different from those of acetylsalicylic acid, prednisolone, or α-tocopherol. These evidences suggest that Tinoridine has not only a membrane-stabilizing activity on lysosomes but also an inhibitory activity against the lipid peroxidation.
A series of pyridoxine derivatives having a thiazolidine ring, thiazoline ring, or thiazolium ring at the 4-position were synthesized. Pyridoxal oxime (II), pyridoxal hydrazone (IV), pyridoxalazine (V), and pyridoxal phenylhydrazone (VI) were reduced to pyridoxamine (III) by zinc powder and acetic acid. 3-(3-Hydroxy-5-hydroxymethyl-2-methyl-4-pyridyl) methyl-4-hydroxy-4-methylthiazolidine-2-thione (VIII) was syntheized from chloropropanone, carbon disulfide, and III. Hydrolysis of VIII with 10% hydrochloric acid gave thiazoline-2-thione derivative (IX), whose treatment with hydrogen peroxide in 3% hydrochloric acid afforded thiazolin-2-one derivative (XIV). However, treatment of IX with hydrogen peroxide in 20-30% hydrochloric acid yielded a thiazolium derivative (XV).
In the Sandmeyer reaction of benzenediamnium ion with thiocyanate ion in the presence of cupric ion, the yield of phenyl thiocyanate (I) increased up to 30% with the increase in the molar ratio of thiocyanate ion to cupric ion, in the pH range of 8.0-8.5, whereas that of phenyl iso-thiocyanate (II) increased up to 40% with the decrease of the molar ratio of these reactsnts in the pH range of 1.0-1.5. For the explanation of the difference in the yield of these products, relationship between the reaction of Cu2+ to SCN- and the results of Sandmeyer reaction was examined. The amount of cuprous thiocyanate prpduced and the concentration of cupric thiocyanate in the solution were proportional to the yields of I and II, respectively. Accordingly, cuprous thiocyanate and cupric thiocyanate are considered to be responsible for the formation of I and II, respectively.
The effect of metals on the reduction of 4-nitroquinoline 1-oxide (4-NQO) was examined by using ascorbic acid as a reducing agent, and Fe(II), Cu(II), Zn(II), Mo(VI), Co(II), Mn(II), Cr(III), Al(III), and W(VI) as the metals. It was found that copper and molybdenum could reduce 4-NQO to 4-hydroxyaminoquino1ine 1-oxide (4-HAQO). It was further found that molybdenum was the only metal which was specific for the reduction of 4-HAQO to 4-aminoquinoline 1-oxide (4-AQO) among the metals examined. At a higher concentration of ascorbic acid, molybdenum reduced 4-HAQO to 4-aminoquinoline(4-AQ) via 4-AQO. The reduction mechanism of 4-HAQO by molybdenum was speculated as the facile reduction of 4-HAQO by the complex formation between 4-HAQO and molybdenum blue (reduced type of molybdenum). This result would support the role of molybdenum in the enzyme groups activated by molybdenum such as xanthine oxidase and nitrate reductase.
Acceleration of gelation reaction of limulus lysate was observed by the addition of a concentration higher than 10-3M of alkaline earth metals, such as Mg2+, Ca2+, B2+, or Sr2+. The limulus lysate containing Mg2+ or Ca2+ in a concentration of 10-1M as an additive showed the highest sensitivity which was 500 times that of original lysate. The sensitivity of the limulus lysates with Sr2+ or Ba2+ as an additive was lower than that of the limulus lysates with Mg2+ or Ca2+ as an additive. Acceleration of gelation reaction was interrupted by the use of any of alkaline earth metals in 1M concentration, and especially by the addition of Ca2+ in 1M concentration, approximately complete inhibition of gelation reaction being observed. Addition of a chelating agent such as ethylenediaminetetraacetic acid (EDTA) or diethylenetriamine-N, N, N', N", N"-pentaacetic acid (DTPA) reduced the sensitivity of limulus lysates to one-tenth of the sensitivity of the original limulus lysate in 10-2M concentration of such a chelating agent. The lowering of pH resulting from chelating streng-thens the lowering of the sensitivity of the limulus lysates. Complete inhibition of the gelation reaction was observed with marked lowering of the pH by the addition of EDTA to the limulus lysate containing a higher concentration of Mg2+. Correlation between the sensitivity of the limulus lysates and concentration of Mg2+ or Ca2+ was not observed. Reexamination of the optimum pH for the gelation readion necessitated the correction of pH to 6.75-7.25 from the previously reported pH range of 6, 50-6.75.
The structure-activity relationship between the hypotensive and some general pharmacological effects of aminoguanidine derivatives was investigated and the following results were obtained.1) A moderate length of the intermediate chain between the benzene ring and the aminoguanidino group of the test compounds was required to obtain a much longer hypotension, and the introduction of one methylene group gave the optimum activity.2) Among the test compounds, the benzyl- and benzylidene-hydrazino-3, 4, 5, 6-tetra-hydropyrimidines (BHTP) showed a relatively mild and slowly acting hypotensive effect with a long duration.3) Substitution of 2- and 6-positions in the benzene ring of BHTP with dichloro groups gave a marked hypotensive and some strong general pharmacological actions and, in particular, the anti-form isomer of 2-(2, 6-dichlorobenzylidenehydrazino)-3, 4, 5, 6-tetra-hydropyrimidines (VA) showed the most marked action.4) VA aid not show an α-sympathomimetic action as clonidine, etc., did.5) VA had an inhibitory action as potent as guanethidine on the contractions of the isolated guinea-pig hypogastric nerve-vas deferens induced by electrical stimulations.6) VA inhibited the contractile force of the isolated guinea-pig atria and decreased the heart rate of the isolated rat atria.7) VA had an inhibitory revolution mortor activity in mice, a taming effect in rats, and a sedative and hypnotic action in cats.8) A toxic dose of VA showed clonic convulsion, tremor, and hind leg ataxia, but did not show group toxicity and biting behavior as did clonidine in grouping mice.
The previously assigned strocture, which has a indanone moiety, for the Friedel-Crafts cyclization product of 3-(3, 4-dimethoxyphenyl)-1, 2, 3, 6-tetrahydrophthalic acid (1) and its anhydride (14) should be revised to the tetralone (12) based on its spectral data and its chemical reactions. The methylenedioxy analogs (7 and 8) also gave the tetralone (10) by the same reaction. Oxidative decarboxylation of the dihydroderivative (13) of 12 with lead tetraacetate in benzene gave the acetate (17) and the phenyl derivative (16). The former was hydrolyzed to the alcohol (18) which was converted to the tosylate (19). Lithium aluminium hydride reduction of the tosylate gave the dimethoxyphenyl-cyclohexene (20) as a fragmentation product. When the oxidative decarboxylation was carried out in the presence of lithium chloride, a mixture of two stereoisomeric chlorides (22a, b) was obtained. Raney nickel reduction of the mixture gave the tetralone (23), whose infrared spectrum exhibited a carbonyl band at 1663 cm-1. Furthermore, the synthetic sample of 6, 7-dimethoxy-1, 2, 3, 4, 4a.9a-hexahydrofluoren-9-one (15), derived from 3, 4-dimethoxybenzoyl chloride and cyclohexane, was not identical with the reduction product of the chlorides (22a. b), confirming the structures of the Friedel-Crafts cyclization products of the tetrahydrophthalic acid (1) and its analogs (2, 7, 8, and 14).
The reaction of 1-acetyl-3-indolinone with carbon disulfide in the presence of sodium hydride gave anhydro-1-mercapto-3-methyl-9-oxo (9H) thiazolo [3, 4-a] indolium hydroxide. Use of excess carbon disulfide and sodium hydroxide, followed by treatment with dimethyl sulfategave 3, 4-dihydro-1-methylthio-3-(methylthio) thiocarbonylmethyleneimidazo [1, 5-a]-indol-9(9H)-one (XI). Reaction of III with primary amines resulted in the formation of 2-substituted anhydro-1-mercapto-3-methyl-9-oxo (9H)-imidazo [1, 5-a] indolium hydroxides (IXa-e), which are a mesoionic compound having an imidazo [1, 5-a] indolium ring system. Reaction of III with hydrazines gave 10-hydroxy-3-methyl-2H-(1, 2, 4)-triazino [4, 5-a] indole-2-thiones (Xa, b).
A fluorometric method has been investigated for the determination of phenylephrine hydrochloride (I). I was converted with HCHO into a condensed substance (II) which was oxidized by K3Fe(CN)6 in 1N NaOH solution to a fluorescent substance (III). Based on these reactions, a fluorometric determination of I was established. Chemical structures of II and III were found to be respectively 4, 6-dihydroxy-2-methyl-1, 2, 3, 4-tetrahydro-isoquinoline and 6-hydroxy-2-methylisoquinolinium chloride which is in an amphoteric ion form in an alkaline solution. The anlytical procedure is as follows : To 1 ml of a solution of I (2.5 μg/ml), 1 ml of 20% HCHO solution is added, the mixed solution is allowed to stand for 15 min at room temperature, and 5 ml of 1N NaOH solution and 1ml of 0.25% K3Fe (CN)6 solution are added to this solution. After standing for 45 min at room temperature, 1 ml of 0.15% ascorbic acid solution is added, and the whole is diluted to 20 ml with methanol. The fluorescence intensity of the solution is measured at 450 mμ, exciting at 370 mμ.
From the essential oil of the flower of Wistaria floribunda DC.3-hexen-1-ol, 1-octen-3-ol, benzyl alcohol, phenetyl alcohol, γ-phenylpropyl alcohol, l-linalool, cinnamic alcohol, phenol, o-cresol, m-cresol, p-cresol, eugenol, anethole, methyl palmitate, ethyl palmitate, and 16 kinds of volatile acid were isolated and identified. Apigenin, luteolin, p-coumaric acid, and 13 kinds of paraffin were also obtained from the residue of steam distillation of the ether extract.
The electrode reaction and the adsorption characteristics of thiobarbituric acid (I) on a mercury electrode have been investigated in aqueous solution by the conventional polarographic technique. I shows four anodic waves on DC polarogram, which are attributed to the complex formation between mercury and I. The fact that the anodic polarogram of I depends on pH has been explained in relation to the acid-base equilibria of I. In the cathodic potential region, I gives a well-defined reduction wave in 0.1M NaClO4 solution due to the one-electron reduction of the proton produced through the dissociation of I. The presence of I causes a marked lowering of the electrocapillary curve. A peculiar peak is observed on AC polarogram of I at -0.75 V, but neither electrochemical oxidation nor reduction of I does not take place at this potential. Increasing concentration of I leads to an increase in the peak height with a tendency to approach a limiting value. These findings clearly indicate the adsorption of I on the electrode surface. The presence of I also affects the DC polarogram of Ni2+, i. e., I gives rise to a prewave of Ni2+ at a more positive potential than the main reduction wave of Ni2+. This fact has been discussed on the assumption of the formation and the subsequent reduction of a complex between Ni2+ and I adsorbed on the electrode surface. The experimental results obtained for barbituric acid have been used to confirm that the adsorption of I is to be attributed to its sulfur atom.
Absorption, distribution, and excretion of an antitussive piperazine derivative, Eprazinone hydrochloride (I), in mice, rats, rabbits, and men were studied by autoradiography, liquid scintillation counting, and colorimetry. In whole-body autoradiogram of mice, the radioactivity was found in a high level in bladder urine, already 15 min and was found only in the large intestine and gall bladder 12 hr after oral administration of 3H-I. Rapid absorption and excretion, and a little accumulation of this drug in mice were suggested by these findings. Estimation of the drug remaining in the gastrointestinal tract of rats after oral administration of I revealed that absorption of I was rapid. The radioactivity in blood of rats and rabbits reached a maximum 2 and 1 hr, respectively, after oral administration of 3H-I. The radioactivity in rat tissues was found in the carcase and in the liver in a high level, and in the brain in a trace. The excretion of Eprazinone (Ia) from rat feces was greater than that from urine after both oral and intravenous administration. Excretion of the drug from rat bile was great, and this fact suggested that the large excretion of Eprazinone (Ia) from rat bile played an important role. In the rabbit, excretion of the drug from urine was dominant, but the amount excreted from feces could not be neglected.
Silver is quantitatively extracted from an aqueous solution by shaking with thiothenoyltrifluoroacetone (STTA) solution in xylene in the presence of 1, 10-phenanthroline at pH 4.3-6.0, and the excess STTA in organic phase is removed by shaking with borate buffer (pH 11.5). Silver is determined by measuring the absorbance of the complex extracted at 360 nm against the reagent blank. The calibration curve follows Beer's law over the range of 2-45 μg of silver in 10 ml of the organic phase. The molar absorptivity is 16200 at 360 nm. The sensitivity is 0.0067 μg Ag/cm2 for the absorbance of 0.001. The coefficient of variation in precision is 1.2% for 30 μg of silver. The molar ratio of silver, STTA and 1, 10-phenanthroline in the extracted species is estimated to be 1 : 1 : 1 by the continuous variation method. The method described herein is more sensitive than the known method, 3) and the selectivity can be improved by the addition of ethylenediaminetetraacetic acid.
Two new flavonolignans, named 2, 3-dehydrosilymarin (I) and 2, 3-dehydrosilychristin (IIa), were isolated from the seeds of Silybum marianum (L.) GAERTN. (Compositae), together with silymarin and silychristin. Structures of these substances were determined from spectroscopic evidence.
Pharmacological activities of 3, 3-diphenyl-3-hydroxypropionic acid diethylaminoethyl ester hydrochloride (DPE-HCl) and its three quaternary ammonium salts (DPE-methyl iodide, DPE-ethyl iodide, and DPE-ethyl bromide) were investigated and following results were obtained. Intravenous LD50s in male mice were 25.5 mg/kg for DPE-HCl and 12.5 mg/kg for quarternary ammonium salts. The topical anesthesic action of DPE-HCl to rabbit cornea was assumed to be almost the same as lidocaine, while there was no such activity in the quaternary ammonium salts. The spontaneous movement of dog ileum was decreased by these compounds, and the intensity of this action in quaternary ammonium salts was stronger than DPE-HCl. These compounds did not affect the respiration of a dog markedly. DPE-HCl showed a transitory hypotensive action on the blood presesure of a dog, but the quaternary ammonium salts caused a marked and sustained fall in blood pressure.