An aqueous solution of lecithin, bile salts, and cholesterol (artificial bile) which resembles human bile was studied by means of polarizing microscopy, electron microscopy, spin-probe electron spin resonance (ESR) spectroscopy, and pancreatic lipase treatment to digest fatty esters. The relation between the structure of artificial bile and the digestion of fatty esters by pancreatic lipase was investigated, and the following results were obtained. Artificial bile consists of bile salt micelles and bile salt-lecithin-cholesterol mixed micelles. Fatty esters are solubilized in these micelles. Lipase hydrolyzes fatty esters solubilized in bile salt micelles but not in bile salt-lecithin-cholesterol mixed micelles. Ranging from 0.1 to 0.4μ in size, the macromolecular complex of bile salt-lecithin-cholesterol can be observed by electron microscopy.

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The intrinsic viscosity and the amount of adsorption of sodium and calcium chondroitin-6-sulfate (Na₂Chs and CaChs) on hydroxyapatite (HAP) were measured. The values depended on the degree of counter-ion dissociation of chondroitin-6-sulfate (Chs). Ion activity, α, and apparent degree of dissociation of counter-ions (Na⁺ and Ca²⁺), α, for Chs were determined by means of ion-selective electrodes in the presence or absence of HAP. In the case of salt-added systems (NaCl or CaCl₂ : 5-20mM), α, obtained from the slope of the relationship between α and [Chs], was almost independent of the concentration of HAP, [HAP] (0-5g/dl), although α decreased with increase in [HAP] due to counter-ion adsorption by HAP. In the case of salt-free systems, both α and α apparently decreased with increase in the amount of HAP added, because a portion of the counter-ions dissociated from Chs is consumed by adsorption at the surface of HAP. It was concluded that the apparent degree of ionization (i.e. apparent charge density along the polymer chain) of Chs adsorbed by HAP is almost the same as that for Chs free from HAP in the bulk solution, irrespective of the apparent value of α obtained from the relationship between α and [Chs].

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Electrochemical reduction of tertiary nitroalkanes, 2-substituted-1, 1-dimethyl-1-nitroethanes (1), was investigated in aqueous buffer solutions. In polarography, 1 with a phenyl group and/or a hydroxyl group at the 2-position showed an ill-defined wave in the weakly acidic and neutral pH region at relatively high negative potentials, in addition to a well-defined wave arising from the reduction of 1 to the corresponding hydroxylamine (2). Controlled potential electrolysis of 1 at the potential of the former wave gave the 1, 1-dimethylethylamines (3) as well as the hydroxylamines 2. The amines 3 are formed exclusively from 2, probably via the O-protonated form. It is suggested that a phenyl group at the 2-position facilitates the reduction of 2 to 3 by enhancing the adsorption of 2 at the mercury cathode and that a hydroxyl group, while it interferes with the adsorption, assists the reduction by intramolecular hydrogen-bonding to stabilize the O-protonated form of 2.

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2', 3', 4', 5, 5', 6, 7, 8-Octamethoxy-(Ia) and 2', 3', 4', 5, 5', 6, 7-heptamethoxyflavone (IIa) and their position isomers substituted with tetramethoxyl groups in ring B were synthesized to confirm the structures of agehoustin A and agehoustin B isolated from Ageratum houstonianum. The characteristics of the synthesized flavones were investigated by spectroscopic methods.

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Oxidation of 1H-pyrazolo [3, 4-b] quinoxalines (1a : R=H, 1b : R=CH₃) with m-chloroperbenzoic acid (MCPBA) gave the 4-oxides (2a, b). The structures of 2a, b were confirmed by synthesis, by condensing 2-chloroquinoxaline-3-carbaldehyde 4-oxide (6) with appropriate hydrazines. Further oxidation of 2a with MCPBA gave the 4, 9-dioxide (8). Treatment of 1, 2-dihydro-2-oxoquinoxaline-3-carboxamide (10) and 1, 2-dihydro-2-oxoquinoxaline-3-carbonitrile 4-oxide (13) with a mixture of POCl₃ and PCl₅ or POCl₃-dimethyl-formamide afforded 2-chloroquinoxaline-3-carbonitrile (11) and its 4-oxide (14), respectively. When 11 and 14 were reacted with hydrazines, the corresponding 3-amino-1H-pyrazolo [3, 4-b]-quinoxalines (12a, b) and their 4-oxides (15a, b) were obtained in high yields. The reaction of ethyl 2-chloroquinoxaline-3-carboxylate (16) with hydrazine hydrate afforded a mixture of uncyclized products, N, N'-bis (2-ethoxycarbonyl-3-quinoxalinyl) hydrazine (17), ethyl 2-hydrazinoquinoxaline-3-carboxylate (18) and 2-hydrazinoquinoxaline-3-carbohydrazide (19).

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The reaction of 3-(N, N-dimethylcarbamoyl) furo [2, 3-b] quinoxaline (2) with ethyl cyanoacetate in sodium ethoxide-ethanol occurred with ring transformation to give 4-cyano-2-ethoxy-carbonyl-3-hydroxy-3'-oxo-3', 4'-dihydrospiro [2-cyclobutene-1, 2'(1H)-quinoxaline] (5). When 5 was refluxed in acetic acid and in N, N-dimethylformamide, it afforded 2-cyano-3-hydroxy-3'-oxo-3', 4'-dihydrospiro [2-cyclobutene-1, 2'(1H)-quinoxaline] (6) and 7-cyano-5, 6-dihydro-6, 10-dioxo-8-hydroxy-10H-pyrido [1, 2-a] quinoxaline (9), respectively. Refluxing of 5 with hydrazine dihydrochloride and with o-phenylenediamine dihydrochloride in acetic acid also induced ring transformation to produce 6-(3-oxo-3, 4-dihydroquinoxalin-2-yl)-3-amino-5-hydroxy-7-oxo-1, 2-dihydro-1, 2-diazepine hydrochloride (12a) and 16-amino-1, 8-dioxo-1, 2, 8, 10-tetrahydro-quinoxalino [1', 2' : 1, 2] pyrido [4, 3-b] [1, 5] benzodiazepine hydrochloride (13a), respectively.

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5-Acetylspiro [benzofuran-2 (3H), 1'-cyclopropan]-3-one (3a, AG-629), previously found to be the most potent antiulcer compound in a series of spirocyclopropanes, was obtained by onestep synthesis starting from methyl 5-acetyl-2-[(tetrahydro-2-oxo-3-furanyl) oxy] benzoate (1a). In this reaction, 5, 10'-(diacetyl-5', 6'-dihydrospiro [benzofuran-2 (3H), 4'(3'H)-[2H] oxocino [3, 2-b]-benzofuran]-3-one (5a), which has a new ring system, and 1H, 3H-8-acetyl-4, 5-dihydronaphtho [1, 2-c : 4, 3-b'] difuran-1-one (6) were isolated as by-products. A possible reaction mechanism is presented.

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The main sapogenol and a prosapogenin from the fruits of Xanthoceras sorbifolia BUNGE (Sapindaceae) were examined. On the basis of chemical and spectral analyses, the structure of the new sapogenol obtained by acid hydrolysis of crude saponin fraction was established to be 21β, 22α-diangeloyloxy-3β, 15α, 16α, 28-tetrahydroxyolean-12-ene (=21, 22-di-O-angeloyl-R₁-bar-rigenol) (1). The structure of the prosapogenin was identified as 21β-(3, 4-di-O-angeloyl-β-D-fucopyranosyl) oxy-3β, 16α, 22α, 24, 28-pentahydroxyolean-12-ene (napoleogenin B) (6) on the basis of spectral and X-ray crystal diffraction analyses.

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The reaction of various types of acetyl compounds with a ketene dithioacetal, methyl 2-cyano-3, 3-bis (methylthio) acrylate, in the presence of potassium hydroxide gave the corresponding 6-aryl-and 6-styryl-3-cyano-4-methylthio-2H-pyran-2-one derivatives. The methylthio group on the pyrone ring reacted readily with nucleophiles such as amines, active methylene compounds and methoxy anion to yield the corresponding displacement products in good yields.

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A total synthesis of (±)-1β, 10β-epoxyfuranoeremophilane-6, 9-dione (8) and (±)-epoxydecompositin (25) is described. The key step in this synthesis is the epoxidation of the allylic alcohol (10). A model experiment was carried out using the readily available diketone (7) to give the desired 1β, 10β-epoxide (16) stereoselectively in good overall yields. The diketone (17) was reduced with NaBH₄ to give the 9β, 10α-diol (18a), which was treated with Ac₂O then dehydrated with SOCl₂ to give the unexpected rearranged product 19a. The 9-hydroxyl group of 18a was protected with a trimethylsilyl group to give 20. The desired allylic alcohol (10) was derived from 20 via 21a in excellent yield. Epoxidation followed by MnO₂ oxidation of 10 gave (±)-8. NaBH₄ reduction of (±)-8 followed by acetylation with Ac₂O gave (±)-25

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Two new germacranolides, ixerins H and I, and three new melampolides, ixerins J, K and L were isolated from the polar and less polar fractions of Ixeris tamagawaensis KITAM., respectively. The structures and stereochemistry were established partly by chemical transformations and mainly by the use of ¹H-and ¹³C-nuclear magnetic resonance spectroscopy.

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6, 5'-Cyclo-5'-deoxyuridine, a fixed anti form of uridine, was synthesized by a radical cyclization of 5'-bromo (or iodo)-5'-deoxy-2', 3'-O-isopropylidene-5-chloro (or bromo)-uridine with tri-■-butyltin hydride followed by dehydrohalogenation and deacetonation. The 5-bromo and 4-thio derivatives of the cyclouridine were also prepared and were converted to the 2', 3'-cyclic phosphates. These nucleotides were hydrolyzed by pancreatic ribonuclease. The result showed that the enzyme recognizes the pyrimidine nucleotides in the anti form. 6, 5'-Cyclo-5'-deoxycytidine was also synthesized by two routes.

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Two kinds of chiral segments, i.e. segment B-I (4) and segment B-II (5), which are potentially versatile building-blocks for construction of the right half of taxane-type diterpenoids, were synthesized from 3-methyl-2-cyclohexen-1-one (6) via optical resolution of the (2S, 3S)-2, 3-butanediol ketal derivatives (8, 15).

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A total synthesis of polyamines was developed using potassium phthalimide as a sole nitrogen source. ¹⁵N-Enriched phthalimide was prepared almost quantitatively from ¹⁵N-enriched ammonium sulfate by a modified method. Key compounds for the synthesis of ¹⁵N-enriched spermidine and spermine were ¹⁵N-enriched putrescine (I), N-(3-bromopropyl)-phthalimide (II), N-(4-bromobutyl) phthalimide (III), and benzylamine, which were easily prepared in high yields from potassium ¹⁵N-enriched phthalimide. Spermidine was synthesized by two alternative methods involving an alkylation of monobenzyloxycarbonyl putrescine with II and a stepwise alkylation of benzylamine with II and III in the presence of KF-Celite. The lattter method allowed the preparation of all seven kinds of various ¹⁵N-enriched spermidines with combinations of the three reagents containing ¹⁵N or ¹⁴N. Spermine was similarly synthesized by an alkylation of N, N'-dibenzylputrescine with II. In these methods, the alkylation using KF-Celite was extremely useful for the synthesis of spermidine and spermine. The present methods were also used to synthesize various other polyamines in high yields.

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Torosachrysone 8-β-D-gentiobioside (1) and physcion 8-β-D-gentiobioside (2) were isolated along with protocatechuic acid from the unripe and ripe seeds of Cassia torosa CAVANILLES, respectively. The structure of 1 was established on the basis of chemical and spectroscopic findings. In addition, the ¹³C nuclear magnetic resonance spectra of torosachrysone and related compounds are discussed.

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Several semisynthetic maytansinoids that differ in the structure of the acyl group at the C₃ position were prepared by acylation of maytansinol (3) using appropriate carboxylic acids or their active derivatives, and the effects of the compounds on the growth of Tetrahymena pyriformis and the survival of tumor-bearing mice were determined. Among these analogs, the C₃ esters having a straight chain aliphatic acyl (11, 12), cycloalkanecarbonyl (18-20) or phenylacetyl group (22), and those having a 2-(N-acetyl-N-methyl) aminohexanoyl (7) or (2-(N-acetyl-N-methyl) aminophenylpropionyl group (8), strongly inhibited the growth of T. pyriformis and exhibited potent activity against B16 melanoma in mice. The potencies were similar to those of maytansine and ansamitocin P-3. The most striking result was the finding that the phenylglycinate (31) was superior to maytansine in terms of its broader effective dose range against ip B16 melanoma and P388 leukemia in mice ; however, higher doses of the phenylglycinate were required.

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(+)-(1S, 5R, 7R)-5, 8, 8-Trimethylbicyclo [5. 1. 0] octan-3-one (2), which comprises the C/D-ring moiety of ingenol esters (1) and is an important chiral synthon for a total synthesis of ingenol esters from easily available (+)-3-carene (3), was synthesized. Intramolecular alkylation of 5b with a strong base under kinetic control gave only the undesired five-membered ketone (6). The intramolecular directed aldol reaction of a mixture of the silyl enol ethers, (9) and (10), easily derived from (+)-3 in good overall yield, was achieved in the presence of tin (IV) chloride to afford 11, 12, and 13 in 66, 9, and 6% yields, respectively. β-Elimination of the methoxy group of 11 gave an enone (14), which was methylated with lithium dimethylcuprate to afford the title compound (2) in good yield ; The configuration of the C-5 methyl group of 2 was deduced from the fact that catalytic hydrogenation of the enone (16), derived from 14 via 15, gave 2. Finally the stereochemistry of 2 was unambiguously determined by the chemical correlation of 18a, derived from 15, with 24b, derived from the known compound 19.

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A chemical examination of hydrolyzable tannins in a rhubarb of high quality (commercial name : ?? ?? ?? ?? ) has revealed the occurrence of four new acylated sugars, i.e., 2-O-cinnamoyl-β-D-glucose (I), 2-O-cinnamoyl-1, 6-di-O-galloyl-β-D-glucose (II), 2-O-p-coumaroyl-1-O-galloyl-β-D-glucose (III) and 1-O-galloylfructose (IV), as well as the known compounds 2-O-cinnamoyl-1-O-galloyl-β-D-glucose (VII), (-)-epicatechin 3-O-gallate (VIII), 1-O-galloyl-β-D-glucose (IX) and 1, 6-di-O-galloyl-β-D-glucose (X). A low-quality rhubarb (commercial name : ?? ?? ?? ) was found to contain three new gallates, 1-O-galloylfructose (IV), 2, 6-di-O-galloylglucose (V) and 3, 5-dihydroxyphenol 1-O-β-D-(6-O-galloyl)-glucopyranoside (VI), together with five known compounds, VIII, X, 6-O-galloylglucose (XI), 1, 2, 6-tri-O-galloyl-β-D-glucose (XII) and procyanidin B-1 3-O-gallate (XIII).

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The stereochemistry of 9-arylthioxanthene 10-oxides and 10, 10-dioxides was determined by detailed investigation of the ¹H-NMR spectra. These sulfoxides and sulfones were synthesized by oxidation of the corresponding 9-arylthioxanthenes with m-chloroperbenzoic acid in dichloromethane or with hydrogen peroxide in acetic acid.

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Products of the nitration of 2-thiazolylurea and 2-thiazolylthiourea derivatives with fuming nitric acid in concentrated sulfuric acid are described. The urea derivatives studied were 3-(4-methyl-2-thiazoly) urea and -thiourea (7), and their l-methyl (1 and 2, respectively) and 1, 1-dimethyl analogs. The products were the compounds nitrated at the 5-position of the thiazole moiety. With an excess of nitric acid, 1-methyl-3-(4-methyl-5-nitro-2-thiazolyl)-1-nitrourea was obtained from 1, while the corresponding 1-nitrosourea was formed from 2. A pale yellow compound was obtained from the nitration of 7. Unlike other nitrated thioureas, it did not form a colored Cu (II) chelate and was stable to acid, alkali, and heat. It was concluded to be 6-methyl-2-nitroiminothiazolo [3, 2-b] [1, 2, 4] thiadiazole from studies of its physicochemical properties, chemical reactions, and the results of X-ray crystallography. Corresponding compounds were obtained from other N-(4-alkyl-2-thiazolyl) thioureas.

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Six chromone and chromanone derivatives (I-VI) have been isolated from rhubarb (commercial name : ?? ?? ?? ?? ), together with torachrysone 8-O-β-D-glucopyranoside (VII), kaempferol (VIII) and kaempferol 3-O-α-L-rhamnoside (IX). The structures of I-VI were established on the basis of chemical and spectroscopic data to be 2, 5-dimethyl-7-hydroxychromone (I), 2-methyl-5-acetonyl-7-hydroxychromone (II), 2-methyl-5-carboxymethyl-7-hydroxychromone (III), 2-(2'-hydroxypropyl)-5-methyl-7-hydroxychromone (IV), 2-(2'-hydroxypropyl)-5-methyl-7-hydroxychromone 7-O-β-D-glucopyranoside (V) and 2-methyl-5-carboxymethyl-7-hydroxychromanone (VI).

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Nineteen stilbene derivatives (I-XIX), of which five are novel cis-stilbenes, have been obtained from a rhubarb of low quality (commercial name : ?? ?? ??, Imo-Daio). Based on chemical and spectroscopic evidence, these compounds have been characterized as rhapontigenin (I), rhaponticin (II), rhapontigenin 3'-O-β-D-glucopyranoside (III), rhaponticin 6"-O-gallate (IV), rhaponticin 2"-O-gallate (V), rhaponticin 2"-O-p-coumarate (VI), piceatannol (VII), piceatannol 3-O-β-D-glucopyranoside (VIII), piceatannol 3'-O-β-D-glucopyranoside (IX), piceatannol 3'-O-β-D-xylopyranoside (X), piceatannol 3-O-β-D-(6"-O-galloyl) glucopyranoside (XI), desoxyrhaponticin (XII), desoxyrhaponticin 6"-O-gallate (XIII), 3, 4', 5-trihydroxystilbene 4'-O-β-D-(6"-O-galloyl) glucopyranoside (XIV), cis-3, 3', 5-trihydroxy-4'-methoxystillbene 3-O-β-D-glucopyranoside (XV), cis-3, 3', 5-trihydroxy-4'-methoxystilbene 3-O-β-D-(6"-O-galloyl) glucopyranoside (XVI), cis-3, 5-dihydroxy-4'-methoxystibene 3-O-β-D-glucopyranoside (XVII), cis-3, 3', 5-trihydroxy-4'-methoxystilbene 3-O-β-D-(2"-O-galloyl) glucopyranoside (XVIII) and cis-3, 3', 5-trihydroxy-4'-methoxystibene (XIX).

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1α-Fluorovitamin D₃ has been synthesized from 6β-acetoxy-5α-cholest-1-en-3-one (2) via 1α-fluorocholesterol (11). The key reaction was the trans diaxial ring opening of the 1β, 2β-epoxide 5 with potassium hydrogen difluoride in ethyleneglycol by heating. The resulting 1α-fluoro-2β, 3β-diol was subjected to regeneration of the 5-ene function and reductive removal of the 2β-hydroxyl group by Barton's method to give 1α-fluorocholesterol (11). Transformation of 11 into 1α-fluorovitamin D₃ led to revision of the previously reported stereochemical assignment of 1α, 25-difluorovitamin D₃.

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trans Diaxial opening with potassium hydrogen difluoride of the 1β, 2β-epoxy-3β-ol derived from cholenic acid gave the 1α-fluoro-2β, 3β-diol derivative. Construction of the 25-hydroxy cholesterol side chain and deoxygenation of the 2β-hydroxy group afforded 1α-fluoro-3β, 25-dihydroxycholest-5-ene, which was transformed into 1α-fluoro-25-hydroxyvitamin D₃. A single dose of 1.3μg of 1α-fluoro-25-hydroxyvitamin D₃ produced neither an intestinal calcium transport response nor a bone calcium mobilization response in vitamin D-deficient rats. In the same rats, 50ng of 25-hydroxyvitamin D₃ produced a marked response. In contrast, 1α-fluoro-25-hydroxyvitamin D₃ was 30 times more active than 25-hydroxyvitamin D₃ in binding to the chick intestinal receptor for 1α, 25-dihydroxyvitamin D₃. Addition of 1α-fluoro-25-hydroxyvitamin D₃ to human promylocytic leukemia cells resulted in strong phagocytic activity.

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Spiro [benzofuran-2 (3H), 1'-cyclopropan]-3-one (VI-1) and its derivatives (VI-2-VI-75) were synthesized in the same manner as described previously for the synthesis of spiro [cyclopropane-1, 2'-[2H] indol]-3'(1'H)-ones (I). These spiro [benzofuran-2 (3H), 1'-cyclopropan]-3-ones were evaluated for gastric antisecretory activity and protective activity against lesions induced by water-immersion restraint stress in the rat. The most potent antiulcer compounds (VI-17 and VI-55) were obtained by the introduction of an acetyl or dimethylamino group at the 5-position on the benzene ring. The most interesting member of the series, 5-acetylspiro [benzofuran-2 (3H), 1'-cyclopropan]-3-one (VI-17, AG-629), was selected as a candidate for clinical studies.

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5-Acetylspiro [benzofuran-2 (3H), 1'-cyclopropan]-3-one (1a) shows potent gastric antisecretory and antiulcer activities in rats. In an attempt to improve the pharmacological profile of 1a, we synthesized positional isomers, as well as the prenyloxy and oxime derivatives. Evaluation of their antisecretory activities and protective activities against gastric lesions induced by water-immersion restraint stress in the rat indicated that the 7-acetyl derivative (1d) was equipotent to 1a.

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Various 1'-(o, m, and/or p-substituted benzyl) (5), 1'-(heterocyclic arylmethyl) (6), and 1'-acyl (7) analogs of spiro [isochroman-3, 4'-piperidin]-1-one were prepared and tested for inhibitory activity on the compound 48/80-induced release of histamine from mast cells. The biological results suggested that the activity is mainly affected by the lipophilicity rather than by the electrostatic character of the 1'-substituent. 4-Benzylspiro [cyclohexane-1, 3'-hexahydroisochroman]-1'-one (17) and 9-benzyl-1-oxaspiro [5.5] undecan-2-one (18) were prepared and found to be inactive, implying that the benzene moiety in the isochroman ring is essential for the activity.

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An 18-membered macrocyclic hexaamine, [18] aneN₆, interacts with histamine and its H₂-agonist dimaprit at physiological pH to yield stable 1 : 1 complexes with simultaneous liberation of H⁺, which mimics the histamine H₂-receptor-agonist interaction and the resulting gastric acid secretion. The polyamine H₂-receptor model does not interact with the histamine H₁-agonist 2-pyridylethylamine. Our model does interact with the H₂-antagonists cimetidine, metiamide, famotidine and ranitidine to form more stable 1 : 1 complexes than with the H₂-agonists, which offers a possible chemical model for the pharmacological ability of the H₂-antagonists to competitively block H₂-receptors and inhibit the gastric acid secretion induced by histamine. The known structural features distinguishing between histamie H₁-and H₂-agonist, and between histamine H₂-agonist and -antagonist are reevaluated in terms of our model.

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A new β-carboline alkaloid, named picrasidine E (I), was isolated from the wood of Picrasma quassioides BENNET (Simaroubaceae) together with seven known alkaloids, 1-methoxycarbonyl-β-carboline (II), 1-ethoxycarbonyl-β-carboline (III), 1-formyl-β-carboline (IV), 1-hydroxymethyl-β-carboline (V), β-carboline-1-propionic acid (VI), 4, 5-dimethoxy-canthin-6-one (VII), and 5-hydroxy-4-methoxycanthin-6-one (VIII). These structures were elucidated on the basis of spectroscopic and chemical evidence.

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A new dipeptide, lyciumamide (1), was isolated from the Chinese crude drug"Ti-ku-'pi, "root bark of Lycium chinense MILL. (Solanaceae), and was shown by spectroscopy, chemical degradation, and synthesis to be N-benzoyl-L-phenylalanyl-L-phenylalaninol O-acetate (1).

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Acetoacetate concentration in plasma was determined by head-space gas chromatography after decarboxylation by the use of acetoacetate decarboxylase, which was recently isolated from Bacillus polymyxa A-57 strain. Partial purification of acetoacetate decarboxylase solution was carried out by DEAE-cellulose column chromatography of the cell-free extract, which was prepared by ultrasonication. The properties of the enzyme were found to be particularly suitable for acetoacetate assay (optimum pH, 5.8 ; V_max, 230μmol/min/mg ; K_m, 5.9×10^-4M). Plasma was deproteinized by Somogyi's method. Acetone in the mildly basic supernatant was assayed by head-space gas chromatography. Acetoacetate was calculated by subtracting blank acetone from total acetone after enzymatic decarboxylation of acetoacetate. The minimal detectable concentration was 1μM. This method gave better reproducibility (CV=2.0-8.0%) and recovery (96.0-101.8%) than chemical decarboxylation with perchloric acid. Normal subjects (n=31) showed plasma acetone levels of 7.2±3.4μM and acetoacetate levels of 22.5±9.7μM. Diabetic patients (n=44), treated by diet control alone without drug therapy, gave plasma acetone levels of 8.1±3.5μM and acetoacetate levels of 25.0±8.0μM. There was no significant difference between the two groups.

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A sensitive and relatively specific enzyme immunoassay of 11-deoxycortisol in human plasma has been developed. Enzyme labeling of 11-deoxycortisol was accomplished by the active ester method. The use of 4-carboxymethylthio-11-deoxycortisol N-succinimidyl ester and β-galactosidase in an appropriate molar ratio provided an enzyme-labeled antigen suitable for enzyme immunoassay. The anti-11-deoxycortisol antiserum used was that elicited in the rabbit by immunization with the conjugate of the haptenic derivative with bovine serum albumin. In order to improve the assay specificity, cortisol was used as an agent for the blocking of less specific antibodies present in the antiserum. The specificity of the assay system was assessed by comparing the results of measurement of 11-deoxycortisol in human plasma with those obtained by a radioimmunoassay. The quantitation limit of 11-deoxycortisol was 800 fg per tube. The intra-and inter-assay coefficients of variation for 11-deoxycortisol in human plasma were 3.0-15.8% and 7.6-12.5%, respectively. The present enzyme immunoassay of 11-deoxycortisol was found to be useful in the metyrapone tests for evaluation of pituitary-adrenal function and for differential diagnosis of Cushing's syndrome. The assay can be done on methylene chloride extracts of plasma.

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The degradation of 2'-deoxycytidine 5'-monophosphate (dCMP), 2'-deoxythymidine 5'-monophosphate (dTMP), 2'-deoxyadenosine 5'-monophosphate (dAMP) and 2'-deoxyguanosine 5'-monophosphate (dGMP) with ozone in aqueous solution was investigated. The patterns of degradation were essentially the same as those of the corresponding ribonucleotides. The second-order rate constants of the reactions as determined by stopped-flow spectrophotometry at pH 6.9 and 15°C were 1.6×10⁴M^-1 s^-1 and 1.4×10³M^-1 s^-1 for dTMP and dCMP, respectively. The rate constants for dGMP and dAMP as estimated by a competitive technique under the same conditions were about 5×10⁴M^-1 s^-1 and 2×10²M^-1 s^-1, respectively. The pH dependencies of the relative rate constants, k_rel, of nucleotide pairs as well as the effect of a radical scavenger on k_rel suggested that the predominant reaction pathway is the direct attack of ozone on the base moiety in the cases of dCMP, dTMP and dGMP. On the other hand, radical reactions with the sugar moiety seemed to be predominant in the case of dAMP.

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The hydrolytic activity of lysozyme towards 4-methylumbelliferyl tetra-N-acetyl-β-chitotetraoside (4-MU-(GlcNAc)₄) was little affected by ionic strength, though the activity of lysozyme towards cell suspension of Micrococcus lysodeikticus varied markedly with ionic strength. About 40-60% of lysozyme activity with 4-MU-(GlcNAc)₄ as a substrate was inhibited by 0.1mM N, N', N"-triacetylchitotriose ((GlcNAc)₃), but the lytic activity of lysozyme towards M. lysodeikticus was little affected. The kinetics of hydrolysis of 4-MU-(GlcNAc)₄ by hen egg-white (HEW) lysozyme and human placental (HP) lysozyme and the inhibition of this hydrolysis by (GlcNAc)₃ were investigated. The K₅ values for 4-MU-(GlcNAc)₄ of HEW-and HP-lysozymes were 19.7 and 27.9μM, respectively, and the V_max values were 0.124 and 0.0833 nmol/min/mg, respectively. The k values of both enzymes were very low, implying a poor orientation of the scissile bond in the substrate molecule with respect to the active site of lysozyme. (GlcNAc)₃ inhibited lysozyme with hyperbolic mixed-type inhibition. The inhibition reduced the V_max values of both lysozymes. The K₅ value of HEW-lysozyme was increased by the addition of the inhibitor, whereas the K₅ value of HP-lysozyme was decreased. The K_i value was 29.6μM for HEW-lysozyme and 106μM for HP-lysozyme.

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Hot 50% MeOH extracts of crude drugs considered to have a hypotensive effect were tested for inhibitory effects on hog kidney angiotensin converting enzyme. Of 65 samples tested, 14 showed reproducible inhibition in the preliminary screening. The extracts of 8 potent samples were fractionated by MCI gel and Sephadex LH-20 gel chromatographies to concentrate the inhibitors. The most potent fractions of Arecae Semen, Ephedrae Herba, Epimedii Herba, Polygoni avicularis Herba, Potentillae Herba and Rhei Rhizoma showed more than 90% inhibition at the concentration of 20μg/ml. Those of Moutan Cortex and Cinnamomi Cortex showed 86 and 78% inhibitions, respectively, at the same concentration. All of these fractions consisted of tannin-type compounds.

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The administration of DL-3-pyridylalanine (DL-3-PA, DL-α-amino-3-pyridinepropanoic acid) or L-3-PA significantly increased brain serotonin (5-HT) concentration without affecting the other tissue 5-HT concentrations in male Wistar rats. The increase in brain 5-HT upon the administration of DL or L-3-PA was maintained for a long time (at least 96h). DL and L-3-PA decreased tryptophan pyrrolase activity in the liver and increased free tryptophan concentration in the serum. These effects found in rats given a 100mg/kg dose of DL-3-PA were not statistically different from those in rats given a 50mg/kg dose of L-3-PA. DL and L-3-PA hardly affected the activities of L-tryptophan 5-hydroxylase, 5-hydroxy-L-tryptophan decarboxylase and monoamine oxidase in the brain. D-3-PA had no effect on 5-HT concentration or tryptophan-5-HT metabolizing enzymes in rats. These results suggest that the action of DL-3-PA is due to L-3-PA, not D-3-PA.

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The mutagenic and cytotoxic effects of 18 kinds of methyl, ethyl, and isopropyl esters of straight-chain alkanesulfonic acids including isethionic acid were investigated on Salmonella typhimurium TA100. In order to correlate these biological activities with the chemical and/or physicochemical properties, the hydrolytic rates (as a measure of alkylating ability) and the capacity factors (as a measure of hydrophobicity) were determined. Some structural correlations were apparent, although quantitative structure-activity relationships were not obtained.

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The degradation of calf thymus deoxyribonucleic acid (DNA) (0.54mg/ml) with ozonecontaining oxygen gas (ozone content, 0.1mg/l ; flow rate, 70ml/min) was examined. The degradation rates of the four nucleotides in native and heat-denatured DNAs were in the following order : dGMP≃dTMP>dCMP≃dAMP and dTMP>dGMP>dCMP≃dAMP, respectively. In the case of heat-denatured DNA, the rapid degradation of the thymine moiety seems to be due to its relatively weak stacking ability, so that it is more exposed to attack, In intact DNA, the double-helical structure of DNA tends to protect the base moieties from attack by ozone. At the early stage of ozonization of native DNA, strand scission did not occur but the degradation of several guanine and/or thymine moieties was detected. As the ozonization proceeded, strand scissions of DNA and susceptibility to nuclease S₁ digestion were observed. Therefore a specific"ozone-denaturation"or"labilization"occurred, causing the double-helical structure to become increasingly loose due to the destruction of guanine and/or thymine moieties, and making the structure more accessible to nuclease S₁.

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The mutagenicity of 3'-methyl-N, N-dimethyl-4-aminoazobenzene (3'-Me-DAB) and 3'-CH₂OH-DAB, potent hepatocarcinogens, was examined. Microsomal and cytosolic fractions from rat, mouse, hamster, rabbit, and human livers were used for metabolic activation, with Salmonella typhimurium TA100 or TA98 as a tester strain. The mutagenicity of both aminoazo dyes mediated by liver microsomes from polychlorinated biphenyls (PCB)-induced rats was markedly enhanced by addition of the hepatic cytosol ; it was also enhanced by addition of the hepatic cytosol from uninduced rats and phenobarbital-or 3-methylcholanthrene (3-MC)-induced rats. Similar mutagenesis enhancement was produced by cytosols from PCB-induced mice, hamsters, or female rats. The cytosols from rabbit and human liver enhanced the mutagenicity of 3'-CH₂OH-DAB but not that of 3'-Me-DAB. Heating of the cytosol from uninduced rat liver to 55°C for 15 min caused a loss of most of the enhancement activity ; at 40°C, it was stable. About 30% of the enhancement activity for both aminoazo dyes was lost upon dialysis for 24h at 4°C. Protein fractions with the aminoazo dye mutagenicity-enhancing activity were obtained from the dialyzed cytosol by gel filtration through Sephadex G-100 ; the protein fraction with the greatest activity, representing an approximately 3-fold increase in specific activity, was assigned a molecular weight of 43000-47500. This protein preparation enhanced microsome-mediated mutagenesis by 3'-hydroxymethyl-N-methyl-4-aminoazobenzene and o-aminoazotoluene but not that by 3-MC, benzo [a] pyrene, or dimethylnitrosamine.

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The kinetics of the degradation and epimerization of cefsulodin in various buffer solutions were studied at 25°C and 0.6 ionic strength. The overall degradation was a pseudo-first-order reaction in the pH region studied. The rate law of the degradation could be approximated in terms of specific acid-base and water catalyzed hydrolysis, that is k₁ (total degradation rate constant)=k_H⁺×α_H⁺+k₀+k_OH^-×α_OH^-. The apparent activation energies of the degradation reaction were 20.7, 22.3, 23.0 and 27.7 kcal mol^-1 at pH values of 2, 4, 6, and 9, respectively. The epimerization of cefsulodin was proved to be catalyzed by hydroxide ion from the epimerization rate constant-pH profile, solvent effects using ethanol, and the apparent activation energies (which were 27.0 and 26.1 kcal mol^-1 for the apparent forward and reverse epimerization reactions at pH 9.0, respectively). The mechanism of epimerization of cefsulodin is proposed to involve removal of the α-proton of the benzyl side chain by hydroxide ion to form an anioic intermediate. Interactions of cefsulodin with amines and aminoglycosides were also examined. The reaction was pseudo-second-order and the second-order rate constants for various amines and aminoglycosides were compared. It was found that intramolecular catalysis is the predominant factor for amines. An equation is proposed for the second-order rate constants of aminoglycosides. Peaks of unknown products in the alkaline reaction solutions could be separated under the high performance liquid chromatography conditions of the current study.

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Porcine pancreatic kallikreins generated from their precursors"prokallikreins A and B"in three different ways, i.e., (1) kallikreins A and B obtained from autolyzed pancreas, (2) kallikreins A'and B'spontaneously generated during the purification of prokallikreins A and B and (3) kallikreins A"and B"generated from purified prokallikreins A and B by treatment with trypsin, were each highly purified and their properties were compared. The K_m values for BzArgOEt hydrolysis, the mobilities on immunoelectrophoresis and the elution profiles from a DEAE-Sepharose CL-6B column of kallikreins A'and B'and kallikreins A"and B"closely resembled each other but apparently differed from those of kallikreins A and B. Kallikreins A and B were confirmed to consist of two polypeptide chains (three chains to some extent), while kallikreins A", B"and B'were each determined to consist of a single chain. The amino acid compositions of the two-chain kallikrein and the single-chain kallikrein were very similar, though somewhat higher values of Leu, Glu and Lys residues were observed in the single-chain kallikrein as compared with the two-chain kallikrein. Thus, kallikreins A"and B"generated from prokallikreins A and B by the action of trypsin were considered to be very similar to, or identical with, kallikreins A'and B'spontaneously generated during the purification of prokallikreins A and B. It is speculated that two-chain kallikreins A and B are generated from single-chain kallikreins A"and B"by the action of some protease (s) other than trypsin during the autolysis process.

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The critical micelle concentrations and the micellar weights of polyoxyethylene cholesteryl ether, C₂₇H₄₆ (OCH₂CH₂)_nOH (Chol EO_n, n=25 and 30) and polyoxyethylene dihydrocholesteryl ether, C₂₇H₄₈ (OCH₂CH₂)_nOH (Dichol EO_n, n=30) were determined by surface tension measurement and membrane osometry. The critical micelle concentration (cmc) values and the micellar weights of these surfactants are similar in magnitude to those of nonionic surfactants of ethylene oxide adduct type with an alkyl or aralkyl group as a lipophilic group. On the basis of these results, it was concluded that these nonionic surfactants form the usual type of large micelles in water in contrast to the cases of cholic acids and their conjugates which form small aggregates in the relatively high concentration range.

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A triterpene not previously obtained from natural sources has been isolated from Vellozia compacta and its structure assigned on the basis of spectral data. For confirmation of this assignment, a partial synthesis was carried out. β-Amyrenyl acetate was oxidized and the resulting ketone reduced to a diol, which, after selective dehydration and reoxidation, furnished the natural product.

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The Vilsmeier-Haack reaction of 2-ethoxycarbonyl-1-hydroxyindole (2) did not give 3-formylindoles but gave 3-chloro-2-ethoxycarbonylindole (3). The electrophilic reaction of 2 with quinoline 1-oxide (4) in the presence of benzoyl chloride or tosyl chloride also failed, 1-benzoyloxy-2-ethoxycarbonylindole (6) or 3 being formed, respectively. The reaction of 2 with tosyl chloride, benzoyl chloride or acetic anhydride gave the corresponding 3-acyloxy-2-ethoxycarbonylindoles (7, 8 and 11) in unsatisfactory yields. Treatment of 2 with tosyl chloride and 1-morpholinocyclohexene in pyridine-dimethylformamide afforded 2-ethoxycarbonyl-3-(2-oxocyclohexyl) indole (13) after hydrolysis of the reaction mixture.

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Condensation of carboxylic acids with tert-butyl hydroperoxide has been smoothly achieved by the use of diethyl phosphorocyanidate and triethylamine under mild reaction conditions, giving tert-butyl peroxycarboxylates in good yields.

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In contrast to the thermal stability of the oxido anions 7, facile eliminative rearrangement has been shown to occur in the case of the oxido anions 2 which possess an aryl substituent on the carbon atom bearing the trimethylsilyl group. Thus, the one-pot synthesis of the allyloxysilanes 5, starting from the (E)-β-(trimethylsilyl) vinyl sulfone 1, was achieved via the facile 1, 4-silyl rearrangement of the oxido anions 2 with concomitant loss of the benzenesulfonyl group.

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Under the Reissert reaction conditions, quinazoline (1a) afforded 2'-formylbenzanilide (2a), o-aminobenzaldehyde (3a), and N-formylbenzamide (4). Similarly, 4-methyl-(1b) and 4-ethoxyquinazoline (1c) gave the corresponding benzanilides 2b and 2c, respectively, o-aminoacetophenone (3b, from 1b), and benzamide (5). It was confirmed that the same results were obtained in the absence of the cyanide ion. A substituent at the 2-position prevented the ring fission, and only the corresponding N³-benzoyl pseudo-base (14) was obtained. The generality of the ring fission was shown by the reactions of 1-phenyl-1H-pyrazolo [3, 4-d] pyrimidine (6) and 3-phenyl-3H-1, 2, 3-triazolo [4, 5-d] pyrimidine (8), giving 5-amino-1-phenyl-1H-pyrazole-4-carbaldehyde (7) and 5-amino-1-phenyl-1H-1, 2, 3-triazole-4-carbaldehyde (9), respectively.

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The reaction of 6-chloro-1, 3-oxazine-2, 4-diones (1) with o-phenylenediamines (2) in the presence of acetic acid afforded pyrimido [1, 6-α] benzimidazole-1, 3 (2H, 5H)-dione derivatives (3) via a 1, 3-oxazine-to-pyrimidine ring transformation.

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The amido linkage of various glycosylamine derivatives, N-(L-γ-glutamyl)-α-, and-β-D-glucopyranosylamine, N-(L-β-aspartyl)-α-and-β-D-glucopyranosylamine and N-glycyl-α-D-glucopyranosylamine, can be cleaved by Amberlite IRA-410 (OH^-), as evidenced by the appearance of the aglycone amino acids and the disappearance of the starting amide.

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A flow-injection analysis system is described for the determination of halide (chloride, bromide and iodide) anions. This method could be used as a post column detection method for these anions in water samples. The system is based on the specific and selective reaction between halide anion and fluorescein with an oxidant in an acidic flow. The product is detected at 530 nm in an alkaline flow. The detection limits of the three anions under the conditions tentatively established for the determination of bromide anion were as follows ; Cl^-, 1×10^-8mol ; Br^-, 1×10^-10mol ; I^-, 1×10^-10mol.

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