Fractionation of serum lipid by means of the droplet countercurrent chromatography (D.C.C.) by the ascending method was investigated. D.C.C. column system was built 50 of or 100 glass tubes of 2.4×1200 mm, which were joined with each other with Teflon tubes of 0.8×1500 mm. The solvents, heptane : butanol : chloroform : methanol : 60% acetic acid (3 : 2 : 2 : 3 : 5, v/v) and heptane : benzene : chloroform : ethanol : 60% acetic acid (2 : 3 : 1 : 4 : 3, v/v) were found to be suitable in consideration of their physical properties, such as density, distribution coefficient for lipid, and passing rate of droplet in columns of stationary phase. By this method, human serum lipid was able to be separated into three fractions of neutral fat, fatty acid, and phospholipid, and the separated phospholipid was found to be a mixture of cephalin, lecithin, and sphingomyelin by silica gel thin-layer chromatography.

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The inner structure of the bark of Cinnamomum burmanni BL. has been investigated. Some variation was observed in the structure and the bark was divided into three types ; Sinoa type, Padang type, and Tjibodas type. Sinoa type is characterized by the presence of small stone cell groups in the secondary cortex, many thick-walled and lignified parenchyma cells, and very few fibers. Padang type has an interrupted stone cell ring, a large amount of big stone cell groups rarely has thick-walled and lignified parenchyma cells, and not many fibers. Tjibodas type has a continuous stone cell ring, few small stone cell groups near the ring, many fibers in the secondary cortex, and many thick-walled and lignified parenchyma cells form a mechanical tissue.

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Histological examinations were made on cinnamon and allied barks in the market of Jawa, Indonesia. Barks marketed as"kayu manis"and"manis jangan"have a sweet taste and smell of cinnamaldehyde. Barks commercially designated as"sintok"or"kayu sintok"are pungent and/or astringent. Fourteen samples of such a bark were examined and classified into six types, with one of them further divided into two subtypes. Types A-α and A-β were identified respectively as the Tjibodas type and Padang type of Cinnamomum burmanni BL., and types B and C were identified as the bark of C. cassia PRESL. and C. sintoc BL., respectively. The remaining three types, D, E, and F were not identified.

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Reactions of carboxylic acids and esters with sodium borohydride were examined. With an equivalent mol of sodium borohydride, caproic acid (I) was reduced to 1-hexanol (II) in the absence of a solvent at 300°(bath temperature), and hexyl caproate (III) was obtained with 1/4 equivalent mol of sodium borohydride. Similarly, aliphatic and aromatic carboxylic acids were reduced to the corresponding alcohols with an equivalent mol of sodium borohydride. Reaction of esters with sodium borohydride gave the corresponding alcohols.

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Dendropanoxide, α-amyrin, β-amyrin, α-acetylamyrin, β-acetylamyrin, friedelin, epifriedelinol, uvaol, ursolic acid, and 18 kinds of paraffin were isolated from etherial extract of the leaves of Rhododendron degronianum CARR. by silica gel column chromatography. From the methanolic extract of the leaves, grayanotoxin I, rhododendrin, quercetin, and isoquercitrin were isolated, and from the aqueous extract of the leaves. 11 kinds of amino acid were identified. Ten kinds of inorganic cation were detected from the ashed leaves.

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Binding of six nonsteroidal anti-inflammatory agents, flutenamic acid, flurbiprofen, indomethacin, phenylbutazone, naproxen, and salicylic acid, to bovine serum albumin was examined using the equilibrium dialysis method at pH 7.4 and 37°. The nonlinear Scatch ard plots were obtained from the binding data except for indomethacin. On the assumption that there are two classes of binding sites on the protein molecule, the binding parameters, n₁, n₂, k₁, and k₂, were estimated. The partition coefficient, Kp, of these agents to several organic solvents was measured but there was no correlation between Kp and binding parameters. The theoretical percentage of drug free against total concentration in serum was calculated from the binding parameters assuming the concentration of human serum albumin as 4.11 g/dl. It was found that flufenamic acid and indomethacin strongly bind to the protein and the free fraction of these drugs exist within 1% over the therapeutic range. Phenylbutazone and naproxen also bind to the protein strongly, but the saturable nature of the binding sites with increasing drug concentration was observed.

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In order to explore the anticipated nitrone-like property of the N-oxide group of 3, 2'-diquinolyl 1-oxide (2), some usual nucleophilic reactions of 2 were examined and it was found that all attempted reactions proceeded normally. Thus, treatment with sulfuryl chloride, benzoyl chloride-potassium hydroxide solution, and benzoyl chloride-potassium cyanide gave 2- and 4-chloro (3 and 4), 2-hydroxy (6), and 2-cyano (7) derivatives, respectively. Reactions in the presence of acylating agents with ethanol-triethylamine, ethyl cyanoacetate, and 1-morpholinocyclohexene also afforded the respective 2-substituted products (10, 12, and 13) in the usual way.

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Carboxypeptidase S_a was composed of 716 amino acid residues, 90 hexose residues (as galactose), and 31 hexosamine residues (as glucosamine) ; these were Trp₁₄, Lys₄₂, His₁₅, Arg₂₁, Asp₇₇, Thr₄₂, Ser₆₀, Glu₅₀, Pro₃₄, Gly₄₈, Ala₅₇, Val₅₀, Met₆, Ile₅₆, Leu₉₃, Tyr₂₂, and Phe₃₄. Carboxypeptidase S_b was composed of 969 amino acid residues but was free from hexose and hexosamines ; these were Trp₂₁, Lys₆₂, His₁₉, Arg₃₅, Asp₁₀₉, Thr₄₉, Ser₇₃, Glu₈₄, Pro₄₉, Gly₈₈, Ala₇₄, Gys (half)₄, Val₆₂, Met₁₅, Ile₄₉, Leu₇₅, Tyr₃₄, and Phe₄₇. Carboxypeptidase S_a was stable in the pH range from 4.5 to 6.5 and in the temperature range from 0 to 50°, but carboxypeptidase S_b was quickly denatured at pH values below 5.0 or over 6.0. They were completely inhibited by the detergents, sodium dodecyl sulfate and cetyltrimethylammonium bromide, but not by Tween 80 or Triton X-100. The Km and V_max values of carboxypeptidase S_a for the hydrolysis of Z-Glu-Phe were 0.16 M and 0.29 μmol/min/·mg, respectively, and those of carboxypeptidase S_b were 0.053 M and 11.36 μmol/min·mg, respectively.

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An in situ fluorometric method was developed for the quantitation of 1-methyl-2-(2-naphthyl) aziridine (I) in biological materials. Three-phase thin-layer plate (Silica gel G, cellulose powder, and talc) was effective for the separation and fluorescence measurement of I. I in rat muscle or in rat blood was extracted with AcOEt n-hexane, respectively, and then 2-10 μl of the extract was spotted on the silica gel phase of the three-phase thinlayer plate. The plate was developed for 40 min with MeOH·ether (7 : 1, v/v) mixture, dried, and then isopropyl myristate was sprayed. Rf-value of I was about 0.5 in the talc phase of the three-phase plate. The plate was heated for 20 min on a hot plate (180°) and cooled in N₂ stream. Fluorescence intensity was determined by scanning with a spetrofluorodensitometer (excit, 340 nm, emission 480 nm) after the fluorescent spot was covered with a quartz plate. The linear dynamic range for the assay was found from 0.02 to 0.2 μg of I per spot. This method was applied for the determination of unchanged I in the rat blood and muscle, and 2-vinylnaphthalene was found as the metabolite of I in rat blood.

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Examinations on the effect of poly (vinylpyrrolidone) (PVP) on the gastrointestinal absorption of salicylic acid and related compounds in rats, and also on the intermolecular interaction between several compounds related to salicylic acid and PVP revealed that the absorption of small-molecule compounds from the stomach (absorption analysed by the loop method) and small intestine (by the circulation method) was inhibited by the presence of PVP and degree of inhibition decreased in the order of 2, 4-dihydroxybenzoic acid, p-aminosalicylic acid, salicylic acid, salicylamide, and o-anisic acid. This order of inhibition was independent of the easiness of absorption of these compounds in the absence of PVP. The inhibitory effect of PVP on the absorption was observed to be reflected on the blood level pattern of salicylic acid administered orally. Intermolecular interaction of 12 compounds related to salicylic acid with PVP was analysed by the equilibrium dialysis method. It was found that the degree of the interaction, expressed in terms of R-value, ran parallel with the degree of inhibition of the gastrointestinal absorption by PVP. From the relationship between the chemical structure and R, it was presumed that hydrogen bonding through the phenolic hydroxyl group of the small-molecule compounds played an important role in the interaction with PVP. Such a presumption was supported by the results of infrared spectroscopic examinations.

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1-Thioniabicyclo [4.4.0] decane bromide (I) derived from 1, 9-diphenoxy-5-mercaptononane was found to be composed of isomers, Ia and Ib, in the ratio of 2 : 3, each of which was purified by column chromatography on silica gel. On heating at 260-270°, Ia was partially isomerized to the equilibrium mixture of Ia and Ib in 3 : 2 ratio, as did Ib. The structures of Ia and Ib were established as cis and trans isomers, respectively, on the basis of the carbon-13 neuclear magnetic resonance spectral studies. The reaction of Ia and Ib with sodium amalgam gave 2-butylthiane and thiecane. Heating of Ia and Ib with aqueous potassium hydroxide gave 2-(3'-butenyl) thiane, trans-3, 4, 7, 8, 9, 10-hexahydro-2H-thiecine and 2-(4'-hydroxybutyl) thiane.

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The reaction of diphenides (IIa-f) with potassium cyanide in dimethyl sulfoxide was found to give 10-hydroxy-9-phenanthrenecarbonitriles (IIIa-f) in a reasonable yield via the intramolecular recyclization of a ring-opened intermediate (IV). However, the reaction of 1, 2, 10, 11-tetramethoxydiphenide (IIg) afforded 2'-cyanomethyl-5, 6, 5', 6'-tetramethoxy-2-biphenylcarboxylic acid (Ig), which could not be recyclized to IIIg because of steric hindrance of the methoxyl groups in Ig. The nuclear magnetic resonance spectra of III were discussed.

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Two new pyrimido-1, 4-diazepine derivatives, 4-phenyl-6, 8-dimethyl-1H-pyrimido-[4, 5-b] [1, 4] diazepine-2, 7, 9 (3H, 6H, 8H) trione (VIII) and 2-phenyl-6, 8-dimethyl-3H-pyrimido [4, 5-b] [1, 4] diazepine-4, 7, 9 (5H, 6H, 8H) trione (X), were synthesized. 5, 6-Diamino-1, 3-dimethyluracil (I) was condensed with ethyl acetoacetate under elimination of ethanol by heating in xylene, to give N-(6-amino-1, 3-dimethyl-2, 4-dioxo-1, 2, 3, 4-tetrahydro-5-pyrimidyl)-3-phenyl-3-oxopropionamide (VI). When the two components were heated in butanol, I gave ethyl 3-phenyl-3-(6-amino-1, 3-dimethyl-2, 4-dioxo-1, 2, 3, 4-tetrahydro-5-pyrimidylamino) acrylate (IX) under elimination of water. Heating of VI at 230-240°for a few minutes under a reduced pressure gave VIII in a poor yield (17%), while heating of IX in ethanol with sodium ethoxide on a water bath for a few minutes gave X in a good yield (74%).

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Chemical constituents of the wood and bark of Xanthoxylum cuspidatum CHAMP. (Fagara cuspidata ENGL.), a Formosan Rutaceous plant (Japanese name : Tsuru-Zansho), were examined. From the wood six alkaloids, nitridine (2) chloride, dictamine (6), γ-fagarine (7), skimmianine (8), robustine (12), and haplopine (13), β-sitosterol (9), and a new phenylpropanoid, cuspidio (11), mp 65-67°, were isolated. From the bark eight known alkaloids, 2 chloride, oxynitidine (4), des-N-methyl-chelerythrine (14), decarine (24), 7, 8, liriodenine (20), and 4-methoxy-1-methyl-2-quinolone (19), four new alkaloids, des-N-methylavicine (16), mp 290-295°, arnottianamide (18), mp 264-267°, isoarnottianamide (21), mp 254-257°, and base X-C-1 (23), mp 268-272°, 9, 11, and dihydro-p-coumaryl alcohol (22), were isolated.

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Examinations were made on the possibility of an in vitro test for bioinequivalence of chloramphenicol tablets, previously found by the measurement of their level in human serum, using the dissolution tests as a marker. Dissolution tests were made by the Japanese Pharmacopoeia VIII disintegration test, USP XVIII dissolution test, Levy's beaker method and Erweka's method. Erweka's method, using water as the solvent, was found to give a result that correlated well with the area under a serum level-time curve. Serum level of 10 products selected from 24 kinds of commercial chloramphenicol tablets was measured in a total of 135 healthy women in the first and 129 healthy women in the second experiment. Dissolution rate of these 10 products was also measured. In addition, serum level and dissolution rate of the original powder for these chloramphenicol tablets were measured and the difference between tablets and their original powder of chloramphenicol was confirmed. Correlation between the human serum level and dissolution rate of commercial chloramphenicol tablets manufactured in Japan was examined and a regression formula for them was calculated. It was found from these experiments that chloramphenicol tablets with 50% dissolution time of over 64 min have a high probability to be bioinequivalent, and 3 out of 24 products of commercially available chloramphenicol tablets on Japanese market in 1974 were found to be bioinequivalent.

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Oral administration of 4-hydroxypyrazolo [3, 4-d] pyrimidine (H.P.P.) to various species of animals was found to show sex difference in its blood level, excretion time, and serum uric acid value. Therefore, effect of H.P.P. on uricase activity was examined by using mice. Various concentrations of H.P.P. were applied to the homogenate and first and second fractions prepared from mouse liver, and their uricase activity was measured. It was thereby found that the sex difference in the decrease of serum uric acid and effective period after oral administration of H.P.P. were due, in part, to the difference in the degree of uricase activation by H.P.P. Discussions were also made on the difference in blood level and excretion time after administration of H.P.P.

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Anserine, carnosine, β-alanine, and methylhistidine contained in animal and vegetable tissues were measured with an amino acid autoanalyzer, and quantitative characteristic of their distribution was examined. Distribution of their content varied greatly with animal and vegetable species, but the variation was not related to taxonomy. Anserine and carnosine were not found in rice, wheat, soybean, and cow milk, and these constituents were recognized as not being a food nutrient and not affected by the intestinal bacteria. The striking feature in the animal kingdom was found in tuna (Thunnus thynnus (LINNE) in which a large quantity (89.43 μmol/g wet tissue) of anserine was detected but no carnosine. In the eel (Entosphenus japonicus (MARTENS)) 3.617μmol of carnosine was detected but not anserine. Some animal tissues contained one or both of these dipeptides. The human iliopsoas usually contained carnosine but anserine was detected in 3 of 58 cases examined, including a case that contained 8.984 μmol/g protein of anserine and 6.88 μmol/g protein of carnosine. In the case in which anserine was detected, an abnormally high (3.45 μmol/g protein) of free histidine was detected. β-Alanine was not detected in human hepatoma.

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Biochanin A, formononetin, ononin, sissotorin, genistin, β-sitosterol, and β-sitosterol-3-O-β-D-glucoside were isolated from the methanolic extract of the fresh flower of Pueraria thunbergiana BENTH.

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A rapid, simple, and accurate high-speed liquid chromatographic procedure was established for the quantitative determination of glycyrrhizin in pharmaceutical preparations including Glycyrrhiza radix. Glycyrrhizin, extracted from simple pharmaceutical preparations, was determined under the same conditions as previously. Glycyrrhizin in compound pharmaceutical preparations containing multiple components was separated on Permaphase AAX column (1 m×3 mm, internal diam.) with a mobile phase of 50% ethanol in phosphate buffer (pH 5.2)-0.005 M NaClO₄ solution at 45°. In most of preparations, good recovery was obtained, but in a few, sufficient recovery was not obtained because of incomplete extraction.

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From the fruits of Arctium lappa L. (Japanease name"goboshi, ""akujitsu, "or "dairikishi"), two new sesquilignan derivatives, AL-D and AL-F, were isolated along with arctin, arctigenin, and matairesinol. From spectroscopic and chemical investigations, the structure of AL-D was elucidated as 2-[2-(3-methoxy-4-hydroxy) phenyl-3-hydroxymethyl-7-methoxy-2, 3-dihydrobenzofuran-5-yl] methyl-3-(3-methoxy-4-hydroxy)-benzyl-buthyrolactone [I] or 2-(3-methoxy-4-hydroxy) benzyl-3-[2-(3-methoxy-4-hydroxy)-phenyl-3-hydroxymethyl-7-methoxy-2, 3-dihydrobenzofuran-5-yl] methyl-buthyrolactone [II]. AL-F was found to be a stereoisomer of AL-D.

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