A facile ring transformation of 1, 2, 3, 5, 6, 7-hexahydropyrrolizinium perchlorates (1 and 2) to 1-substituted 5-oxoperhydroazocines (5) is described. The spectroscopic and chemical properties of these products are also presented.

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Addition of chlorine to the double bond of α-santonin derivatives such as santonin chlorohydrin (4) santonin α-epoxide (5), and 5-chloro-4-methoxysatonin (6) produced new chloro-santonins (13, 14, 16, respectively). So far, only trans addition of chlorine to the 1, 2-double bond of santonin has been known, but from the configuration of these products, it wa clear that cis addition had occuured. 'Santonin dichloride' (2), and 2-chlorosantonin (3), were chloromethoxylated and chloro-hydroxylated to produce tetrasubstituted compounds (9 and 10), and trisubstituted compounds (11 and 12), respectively. Their structure and configuration were determined by interrelating them to known santonin derivatives (2, 3, 4, 5, and 6) and by X-ray analysis of 13.

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The 4-methoxy-5-methylpyrano[3, 4, -b]indole-1, 3(4H, 5H)-dione (9), prepared from methyl 3-methoxycarbonyl-1-methylindol-2-yl acetate (6), underwent a strong base-induced cycloaddition reaction with 2-chloro-6, 6-ehylenedioxy-5, 6, 7, 8-tetrahydro-1, 4-naphtoquinone (11) to give the tetrahydronaphtho[2, 3-b]carbazole-7, 12-dione (10), regioselectively. The cycloadduct (10) was successfully converted to a D-ring indole analogue of daunomycin (1a).

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In acid hydrolysis of N¹-acyl-N¹, N²-diarylamidines, a water molecule attacked exclusively the amidine central carbon, and the reaction proceeded in parallel through two patheways. One of them leads to the formation of two N-acylarylamines, and the other leads to the formation of N, N-diacylarylamine and srylamine. Acid hydrolysis of N¹-tosyl-N¹, N²-diarylamidine was also examined. The results were similar to those of the hydrolysis of N¹-acyl-N¹, N²-diarylamidines.Alkaline hyrdolysis and alcoholysis of N¹-acyl-N¹, N²-diarylamidines occurred almost exclusively at the amide carbonyl group to give N¹, N²-diarylamidines and carboxylic acids or their esters.The effects of structural change and the aryl substituents on the rate and direction of the reaction were examined.

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The Fischer indolization of ethyl pyruvate 2-(2-chlorophenyl)methylhydrazone (17) with HCl/EtOH proceeded rapidly to give six 1-methylindoles, ethyl 7-chloro (19), 6-chloro (22), and unsubstituted (24)-1-methylindole-2-car-boxylates, and dichloro derivatives (21, 23, and 26), whereas no indolization had occurred with the correspinding NH-hydrazone, as described in a previous paper. This result shows that the abnormal Fischer indolization of 2-chloro-phenylhydrazones with HCl/EtOH occurred in an ortho-C₆ fashion, in the same manner as that of the corresponding 2-methoxyphenylhydrazones (1 and 2).The Fischer indolization of ethyl pyruvate 2-(2, 6-dichlorophenyl)methylhydrazone (18) with HCl/EtOH did not occur at all, whereas the reaction with ZnCl₂/AcOH gave a low yield of ethyl 5, 7-dichloro-3-methylindole-2-carboxylate (36), which was formed by migration of the N-methyl group during Fischer indolization.

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In connection with the stereoselective exo cycloaddition in the 1, 3-dipolar reaction of 3, 5-lutidine N-oxide and N-arylmaleimides, the inverse-type cycloaddition of some pryidine N-oxides with 1, 4-epoxy-1, 4-dihydronaphthalene was investigated. In these reactions, the armatized furopyridine-type compounds formed from the 1, 5-sigmatropic rearrangement products of the primary cycloadducts were isolated. During the course of the reaction, coloration due to charge-transfer complex formation was observed. The reaction obeyed a second-order rate law and showed little solvent effect. The observed reactivity and stereochemistry are discussed in terms of frontier molecular orbital (FMO) considerations.

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A novel, one-step synthesis of 5-deazaflavin developed, in which a [4+2]cycloaddition is presumably involved, to give the 5-deazaflavin derivative in moderate yield. This methodology was successfully applied to the stepwise preparation of its thio analogue, 5-deaza-10-thiaflavin, whose 1, 5-dihydro derivative was transformed into the corresponding sulfone in good yield.

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A nonenzymatic asymmetric reduction of ethyl benzoylformate with an achiral 1, 5-dihydro-5-deazaflavin derivative in chiral media was investigated as a model system for reruced nicotinamide adenine dinucleotide (phosphate)-dependent dihydrogenase. The chiral media include chiral nuclear magnetic resonance shift reagent, chiral Lewis acid, and a combination of metal ion an chiral ligand (additive). Of these reductions, a substantial asymmetric induction was observed in the presence of tris-[3-(heptafluoropropylhydroxymethylene)(+)-camphorato]europium to give ethyl mandelate possessing prediominantly S-configuration, in a optical yield of 24 to 36%. These values are among the highest so far reported in nonenzymatic reduction of ethyl benzoylformate with a 5-deazaflavin model. The discrimination of the prochiral face of the carbonyl compound was effectvie even when a catalytic amount of the chiral shift reagent was employed.

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O-(2-Alkynyl) S-alkyl dithiocarbonates (allylic xanthates) were pyrolyzed to give 2-alkenyl alkyl sulfides (allylic sulfides) via the corresponding allylically isomeric S-(2-alkenyl) S-alkyl dithiocarbonates. The reaction follows the first-order rate law with relatively low sensitivity to the ionizing power of the medium and sizeably negative entropies of activation. When a mixture of two dithiol esters having different S-(2-alkenyl) groups and different S-alkyl groups was pyrolyzed, a "cross product" was not observed. The reaction was found to be facilitated by the presence of phenolic compounds or Lewis acids.Based on these findings together with modified neglect of diatomic overlap (MNDO) calculation data, the mechanism for the formation reaction of 2-alkenyl alkyl sulfides is discussed.

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Four new azaphilones of angular type, named chaetociridins A, B, C and D, were isolated from the culture of Chaetomium globosum var. flavo-viridae. The absolute configuration of chaetoviridin A was established on the basis of the spectral data and chemical reactions. The structures of chaetoviridins B, C and D were suggested by the spectral data. The result of a biosynthetic study of chaetoviridin B from sodium [1, 2-¹³C₂]acetate gave support to the angular structure.

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Four new dihydrophenanthrene derivatives, spiranthesol, spiranthoquinone, spiranthol-C, and spirasineol-B, were isolated from the roots of Spiranthes sinensis (PERS.) AMES var. amoena (M. BIEBERSON) HARA (Japanese name "nezibana"). The structures 1, 2, 3, and 4 were proposed for these compounds, respectively, on the basis of spectroscopic data obtained by various methods including ¹H-detected heteronuclear multiple-bond multiple-quantum coherence (HMBC) spectroscopy. Chemical conversions of spiranthol-A to spiranthol-C and spiranthoquinone and of spirasineol-A-to spirasineol-B are also described.

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The synthesis of S-glycosyl donors from 2, 3, 4, 6-tetra-O-benzyl-α-glucopyranose (1) using S, S'-bis(1-phenyl-1H-tetrazol-5-yl) dithiocarbonate (2) was carried out by a one-step reaction. One of the S-glycosyl donors, S-1-(1'-phenyl-1H-tetrazolyl)2, 3, 4, 6-tetra-O-benzyl-β-D-glucopyranose (3a), was utilized for glycosylation of glycosyl acceptors such as methanol or cholesterol using silver triflate as a promoter.

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Eighteen new tetranortriterpenoids were isolated from the cotyledons of Swietenia mahagoni (Meliaceae) along with the known tetranortriterpenoids. Among them, the structures of wight new compounds, swietenins B, C, D, E, and F, 3-O-acetylswietenolide, 6-O-acetylswietenolide, and 3-O-tigloyl-6-O-acetylswietenolide, were determined by the use of two-dimensional nuclear magnetic resonance (2D NMR) techniques. Detailed analyses of the ¹H- and ¹³C-NMR spectra of the ten known compounds were also performed.

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A detailed account is given of the general synthetic route to 7, 9-dialkyladeninium salts (16-18) from N⁶-alkoxy-9-alkyladenines (type 5 or 11-13), readily obtainable from 9-alkyladenines in three steps involving N(1)-oxidation, O-alkylation, and Dimroth rearrangement. Alkylations of N⁶-methoxy-9-methyladenine (5) and 9-alkyl-N⁶-benzyloxy-adenines (11-13) with MeI, EtI, and PhCH₂Br in AcNMe₂ produced the corresponding 7-alkylated derivatives (15 and 19-21), together with small amounts of the N⁶-alkylated isomers (6, 8, and 9). Catalytic hydrogenolysis of the former compounds with hydrogen and Raney Ni yielded 7, 9-dialkyladeninium salts (16-18).

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The deoxy-cyanation of 2-substituted quinoxaline 4-oxides (Ia-k) with trimethylsilyl cyanide in the presence of 1, 8-diazabicyclo[5.4.0]undec-7-ene gave the corresponding 3-substituted 2-quinoxalinecarbonitriles (IIa-k). However, in the case of 2-(p-tolylsulfonyl)quinoxaline 4-oxide (II), the substitution with cyanide ion proceeded together with deoxy-cyanation to give 2, 3-quinoxalinedicarbonitrile (III).

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Acid-catalyzed transannular cyclization of 11, 12-epoxycembranolide (1), isolated from the soft coral Sinularia mayi, was studied. The major product was the trans-fused bicyclo[8.4.0]tetradecene derivative 2, and the maximum yield was 89% with boron trifluoride in benzane at 0°C. When treated with p-toluenesulfonic acid in benzene at 55°C, 1 gave a bicyclo[9.3.0]tetradecene derivative 11 in 36% yield. Its immediate precursor was shown to be the allylic alcohol 4, which was converted to 11 on treatment with p-toluenesulfonic acid. The stereochemistries of the cyclization products indicated that the reaction takes place throught a conformation in which the dispositions of 11, 12- and 12, 13-bonds are crossed, with respect to the 7, 8- and 3, 4-double bonds, respectively.

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Cardiac glycosides and pregnanes from the roots and the stems of Adenium obesum ROEM. et SCHULT. were investigated. Among 30 cardiac glycosides including 15 known glycosides and 15 new combinations of the known aglycones and sugars, the structures of 11 glycosides were elucidated. Oleandrigenin β-gentiobiosyl-β-thevetoside was the main glycoside. Neridienone A and 16, 17-dihydroneridienone A, common pregnanes in Apocynacear, were also isolated.

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S-Trimethylacetamidomethyl-L-cysteine {Cys(Tacm)] was easily prepared from N-hydroxymethyltrimethyl-acetamide and L-cysteine in trifluoroacetic acid. The S-Tacm group is stable to HF, but cleavable with mercury(II) acetate in trifluoroacetic acid or iodine in aqueous acetic acid. Cys(Tacm) is less susceptible to sulfoxide formation than three related groups, i.e., acetamidomethyl (Acm), benzamidomethyl (Bam) and (2-oxo-1-pyrrolidinyl)methyl (Pym).

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Various asymmetric or cyclic ester derivatives of the phosphonic acid moiety in the calcium antagonist fostedil (KB-944) were synthesized. The coronary vasodilator activity of these compounds was assessed by Langendorff's method. Among them, the ethyl isopropyl ester 12 showed the most potent activity, which was comparable to that of fostedil.

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A series of 4-phenoxy-2-(1-piperazinyl)quinazolines was synthesized and examined for anticonvulsive and antihypoxic activities. Many of the compounds exhibited potent anticonvulsive activity comparable to that of carbamazepine or phenytoin. Among them, 4-phenoxy-2-(4-propyl-1-piperazinyl)quinazoline (5w) was selected as the most promising candidate antiepileptic drug with few side effects. It seemed that potent anticonvulsive activity was a prerequisite for potent antihypoxic activity.

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Tetrahydronapthoquinones and tetrahydroanthraquinones bearing an amido group have been prepared by Diels-Alder reactions between (E)-l-(N-carbobenxyloxyamino)-1, 3-butadiene (2) or (E)-l-(N-benzoyl-N-benzylamino)-1, 3-butadiene (5) and venzoquinone or 5-substituted naphthoquinones. The stereochemistry of the cycloadditions was investigated. A high regioselectivity was observed in the reaction of the diene carbamate 2 with 5-methoxy and 5-acetoxy naphthoquinones. This latter gave the unezpected 1, 8-regioisomer 3d. The cycloadditions of the dienamide 5 with naphthoquinones 1 (R=OH, OMe, OAc) are regiospecific. Assignment of the structure of the tetrahydroanthraquinone 6b is in good agreement with the known directing effect of the 5-hydroxy group of juglone 1b in analogous Diels-Alder reactions. With 5-methoxy and 5-acetoxy naphthoquinones, the opposite regiochemistry observed is consistent with the electron-donating influence of the methoxy or acetoxy group, making the C-3 carbon atom more electron deficient. Aromatization of the adducts 6b and 7c was accompanied by an unusual elimination of the amido moiety. Thus, 1-hydroxy and 1-methoxy anthraquinones were obtained. Reactions of the dienes 2 and 5 with benzoquinone gave the tetrahydronaphthoquinones 9 and 10 with an endo stereospecificity. Oxidation of 9 by activated manganese dioxide gave the naphthoquinone 11. These compounds were submitted to in vitro cytotoxic assays towards murine L 1210 leukemia cells and clonogenic human tumor cell line MDA-MB 231. The naphthoquinone derivatives 9, 10 and 11 had significant activities with IC₅₀⩽0.4 μg/ml towards these two tumor cells systems.

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As part of our search for new topical antiinflammatory agents, a series of corticosteroid 17-(alkylthio)- and methoxyalkanoate derivatives was prepared and tested for vasoconstrictive activities. Several compounds were proved to have activity superior or comparable to that of 9α-fluoro-11β, 21-dihydroxy-16β-methyl-17α-valeryloxy-1, 4-pregnadiene-3, 20-dione (betamethasone 17-valerate, BV). Among these compounds, 21-chloro-11β-hydroxy-17α-(methylthio)acetoxy-4-pregnene-3, 20-dione (5Aa) was found to have the most potent activity, being more active than BV. The structure-activity relationships of the series revealed that introduction of a (methylthio)acetate function into the 17-position as well as the 21-position of corticosteroids was effective for enhancing the topical antiinflammatory activity.

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Two new phloroglucinol derivatives, isomallotolerin (1) and isomallotochromanol (2), were isolated from the cytotoxic fraction of the pericarps of Mallotus japonicus. The new derivatives were identified as 3-(3-methyl-2-hydroxybut-3-enyl)-5-(3-acetyl-2, 4-dihydroxy-5-methyl-6-methoxybenxyl)-phlorisobutyrophenone (1) and 6-acetyl-5, 7-dihydroxy-8-(3-acetyl-2, 4-dihydroxy-5-methyl-6-methoxybenxyl)-2, 2-dimethyl-3-hydroxychroman (2) from chemical and spectral data. Isomallotolerin and its acetate were found to be cytotoxic to KB cell line.

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Two new clerodane diterpenes, cajucarins A (1) and B (2), were isolated from the cortices of Croton cajucara. The conformations of 1 and 2 were studied by means of spectroscopic methods and molecular orbital calculations.

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Anti-allergic activities of lyophilized pig bile ([PB]) were examined in mice with picryl chloride-induced contact dermatitis (PC-CD), an experimental model of delayer-type hepersensitivity (DTH; type-IV allergy). PC-CD was markedly inhibited by an oral administration of [PB] within 4h after but not during 8 to 16h after challenge with picryl chloride.Anti-inflammatory activities of [PB] were also examined in acetic acid-induced mouse increased vascular permeability, hypetonic-hyperthermic lysis of rat erythrocytes and carrageenin-induced rat hind paw edema. [PB] had no effect on these models. The present study suggests that[PB] inhibits PC-CD through its immuno-modulation in the inductive phase of DTH rather than by an anti-inflammatory action.

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Inorganic constituents of many Cinnamomi Cortices (64 samples; almost all obtained commercially on the Osaka market) were investigated using energy-dispersive X-ray fluorescence spectrometry.The results can be summarized as follows : (1) The Cinnamomi Cortex contains K, Ca, and Fe (except for Japanese cinnamon) at lower levels than those of orchard leaves, while Mn and Sr are present at high levels. (2) A feature of the metals profile of Cinnamomi Cortex is high Mn-content. Especially, Chinese cinnamon (originated from Cinnamomum cassia) contains at extremely high levels, 300-900 ppm, whereas Mn concentrations of Java and Japanese cinnamons range from 100 to 300 ppm, and those of Ceylon cinnamon ranged from 50 to 150 ppm. (3) The contents of Mn and Rb depend on the kind of Cinnamomi Cortex, making identification possible.

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Two new triterpenoid sulfates were isolated from the fresh leaf blades of Schefflera octophylla (Araliaceae). On the basis of results of spectral and chemical investigations, their structures were characterized as free forms of 3-epi-betulinic acid 3-O-sulfate and betulinic acid 3-O-sulfate.

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In the course of our studies on the development of anti-plaque agents for prevention of dental caries, we investigated effects of some of tea preparations and their individual components on the glucan synthesis catalyzed by glucosyltransferase (GTF) from Streptococcus mutans. Extracts of green tea and black tea, and polyphenol mixtures showed appreciable inhibition in the synthesis of insoluble glucan. Among the components isolated from tea infusions. theaflavin and its mono- and digallates had potent inhibitory activities at concentrations of 1-10mM against GTF. (+)-Catechin, (-)-epicatechin and their enantiomers had moderate inhibitory activities at these concentrations, while galloyl esters of (-)-epicatechin, (-)-epigallocatechin and (-)-gallocatechin had increased inhibitory activities.

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The values of log γ for monosubstituted benzene derivatives can be expressed after regression analyses by a linear combination of the quantitative structure-activity relationship (QSAR) descriptors σ_s°, μ²/α, and σ^±_π. The inclusion of aniline series derivatives demands and additional descriptor pK_a, ascribed to the CH/N interaction between the stationary liquid and the nitrogen lone pair. Furthermore, for all cases of polyalkylvenzene derivatives, the descriptor σ_s° is effective, and the number of vicinal type CH₃-pairs represented at first by the descriptor ortho numver N_o can be transformed to the corrected σ⁺_π, representing the CH/π type charge-transfer interaction between the substrate and the stationary liquid.

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Methods employing colorimetry and high-performance liquid chromatography (HPLC) were developed for determination of D-glucosone. Colorimetry with phenazine methosulfate and nitro blue tetrazolium as electron acceptors made it possible to determine D-glucosone (sensitivity : 50 nmol) in the presence of excess amounts of D-glucose and D-fructose. Further, by using a high-performance liquid chromatograph equipped with a ligand-exchange mode column, we could separate D-glucosone in human and rat sera from D-glucose. The blood level of D-glucosone 5 min after intraperitoneal administration (1.68 mmol/kg) of the sugar to rats was determined by the HPLC method to be 1.3 mM in portal blood and 0.5 mM in postcaval blood. The maximum increase in the blood glucose level was observed 2 h after administration of the D-glucosone. A similar increase of blood glucose was observed after the administration of 2-deoxy-D-glucose, and seemed to continue until 5 h. The assay methods described are useful for biochemical studies on D-glucosone as an analogue of D-glucose.

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Inclusion complexes of clobazam with α-, β-, γ-cyclodextrins (CyDs) and heptakis(2, 6-di-O-methyl)-β-cyclodextrin (DM-β-CyD) in aqueous solution and in the solid phase were studied by the solubility method, infrared (IR) spectroscopy, differential scanning calorimetry (DSC), and X-ray diffractometry. In addition. inclusion complex of clobazam with heptakis(2, 3, 6-tri-O-methyl)-β-cyclodextrin and the solid dispersion of clobazam with methyl cellulose (MC) in a ground mixture were investigated by IR, DSC and X-ray diffractometry. It was observed that DM-β-CyD had the highest stability constant among the four CyDs in solution. Thermal and X-ray diffraction analyses showed that clobazam molecules existed in a molecularly dispersed state in the ground mixture of CyDs. Infrared spectra showed lower frequency shifts in the case of the ground mixtures of clobazam with natural CyDs, which can be attributed to the formation of hydrogen bonds between the two carbonyl groups of clobazam and hydroxyl groups of natural CyDs. In contrast, higher frequency shifts were observed in the case of the ground mixtures of clobazam with methylated CyDs and MC and these were considered to be due to the monomolecular dispersion of clobazam in a hydrophobic environment.The mode of interaction of clobazam with DM-β-CyD was different from that with natural CyDs in the ground mixtures. Furthermore, the crystalline inclusion complex of clobazam with DM-β-CyD was obtained by heating of the coprecipitate in vacuo at 120°C for 1 h.

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With the objective of modifying the micromeritic properties of tolbutamide (i.e., to manufacture a highly functional powder from), particle design was attempted using a quasi-emulsion solvent diffusion (QESD) method, and the micromeritic properties and dissolution rate of the obtained spherical agglomerates were evaluated by comparison with agglomerates prepared by the solvent change (SC) method. For the production of tolbutamide agglomerates by the QESD method, a necessary condition was the addition of a sucrose fatty acid ester to the system as an emulsifying agent. The particle diameter of the agglomerates obtained by the QESD method depended on the size of the initially formed quasi-emulsion droplets, which in turn depended on the viscosity of the solution. In addition, the agglomerates were nearly perfectly spherical in shape. In the QESD method, the quasi-emulsion droplets crystallized instantaneously from the droplet surface inward. The resultant agglomerates were dense, had great mechanical strength and showed excellent flowability due to their perfect spherical shape. On the other hand, the agglomerates produced by the SC method were conglomerates of primary crystals, and fine, needle-like crystals formed on their surface. As a result, these agglomerates had a large specific surface area, and they therefore showed greater solubility than the agglomerates prepared by the QESD method.

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A membrane permeation-controlled transfermal delivery system (MC-TDS) of isosorbide dinitrate (ISDN), a model drug, was prepared from polyvinyl alcohol aqueous gel containing the drug, a membrane consisting of ethylenevinyl acetate copolymer membrane and acrylic adhesive (EV-a). The permeability of ISDN through the EV-a membrane was 2.5 times higher than that through excised hairless rat skin. The ratio of plasma concentration of ISDN after application of MC-TDS on stripped (damaged) skin relative to intack skin was lower than that after application of Frandol tape-S, a marketed ISDN TDS, which suggests that the EV-a membrane might work as a control membrane for overall delivery rate of ISDN to the body. When MC-TDS stored at 30°C for 13.5-48h was applied to the damaged skin, however, the initial plasma concentration of ISDN was very much higher than the expected therapeutic level and was not controlled by the EV-a membrane. The initial high plasma concentration of ISDN after application of the stored MC-TDS on the damaged skin was due to migration of ISDN from the reservoir to the adhesive during storage at 30°C. The migration of drugs into the adhesive might be an important problem in developing efficient MC-TDS.

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In anionic-nonionic and cationic-nonionic mixed surfactant systems, when two surfactants below their individual critical micelle concentrations (cmc) were mixed, the formation of mixed micelles was confirmed and the cmc values were measured by the first derivative absorption spectrum (FDAS) method, the solubilization method (with Sudan Red dye) and the surface tension method (Wilhelmy plate method). From plots of the mole fraction normalized with respect to cmc (=[[sodium dodecyl sulface (SDS)]/cmc_SDS]/[[SDS]/cmc_SDS+[hyptaethylene glycol dodecyl ether (HED)]/cmc_HED] where [SDS] (or [HED]) is the concentration of SDS (or HED), and cmc_SDS (or cmc_HED) is the cmc of SDS (or HED)) on the abscissa against the sum of concentraions corresponding to the cmc (=[SDS]_*HED/cmc_SDS+[HED]_*SDS/cmc_HED where [SDS]_*HED (or [HED]_*SDS) is the concentration of SDS (or HED) in the cmc of mixed micelles of SDS-HED binary systems) on the ordinate, obtained by the FDAS method and the surface tension method, the boundary line between the regions forming mixed micelles and the regions of monomer was obtained. The results obtained are compared and the usefulness of the methods is discussed.

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pH-induced destabilization, aggregation and fusion of liposomes composed of phosphatidylethanolamine (PE) and various fatty acid were studied. Destabilization was examined as a fluorescent change caused by leakage of coencapsulated aminonaphthalene-3, 6, 8-trisulfonic acid (ANTS) and N, N-p-xylylenebispyridinium bromide (DPX). Fusion was monitored by two different methods, that is, intermixing assay of internal aquenous contents of liposomes, and lipid dilution assay of liposomes labeled with fluorescent phospholipids.Contents leakage from liposomes was observed by lowering the pH, and pH where the leakage began depended on fatty acid used. Fifty percent leakage of contents from PE liposomes containing α-hydroxypalmitic acid or α-hydroxystearic acid was observed at pH 5.5, that from liposomes containing stearic acid or palmitic acid was observed at pH 6.5-6.7, and that from ricinoleic acid at pH 7.2. Aggregation and fusion of the respective liposomes also occurred at a similar pH region. Those results were interpreted by the notion that the protonation of the fatty acid triggers a series of pH-sensitive events. The liposomes developed in this study may be useful as a drug carrier which could release the contents in response to pH changes in their environment.

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The aim of this study was to establish the best manufacturing conditions for preparation by the direct compresison method of tablets which contain microcapsules having a minimal destruction rate of the coating wall, show the same dissolution pattern as microcapsules, and have enough mechanical strength for practical use, ant to elucidate the internal structure of the tablets under the best manufacturing conditions. Degree of destruction of the microcapsule wall was evaluated by the dissolution rate of the medicine in the microcapsules. To learn the mechanical strength of tablets, the crushing strength and friability were measured; their internal structure was analyzed by the porosity and pore size distribution.The best manufacturing conditions for the tablets were thus determined, and it was clarified by analysis of the internal structure that these conditions are markedly affected by the flowability of prescribed powders and the packing state at compression.

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The effects of various chelating agents, sodium N-benzyl-D-glucamine dithiocarbamate (BGD), sodium N-p-methylbenzyl-D-glucamine dithiocarbamate (MBGD), sodium N-p-hydroxymethylbenzyl-D-glucamine dithiocarbamate (HBGD), and N-p-carboxybenzyl-D-glucamine dithiocarbamate (CBGD), which were newly synthesized, and sodium N-methyl-D-glucamine dithiocarbamate (MGD), on the distribution and excretion of inorganic mercury were compared in rats exposed to HgCl₂. Rats were injected i.p. with ²⁰³HgCl₂ (300μg of Hg and 74kBq of ²⁰³Hg/kg) and 30 min or 24h later, they were injected with a dithiocarbamate (1200 μmol/kg). At 30 min after mercury administration, BGD and MBGD significantly enhanced the biliary excretion of mercury, while CBGD, MGD, and HBGD enhanced the urinary excretion of mercury to a small extent. At 24h after mercury injection, BGD was the most effective on the biliary excretion of the metal, while MGD and HBGD significantly enhanced the urinary excretion of the metal. All of these dithiocarbamates were effective in mobilizing mercury from the kidney at 30 min after mercury treatment. At 24h after mercury treatment. HBGD and BGD effectively depressed mercury content in the kidney. These results show that the injection of BGD and HBGD at both 30 min and 24h after mercury treatment can much more effectively mobilize mercury from the kidney without redistribution of mercury to other tissues, such as brain, heart, and lung, when compared with injection of other chelating agents. The pattern of mobilization and excretion of mercury following treatment with each chelating agent was related to the organic/aqueous partition coefficient of each dithiocarbamate-mercury complex.

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In order ot gain further insight into the protective action of Na₂MoO₄ pretreatment (1.24 mmol/kg, once a day, i.p.) against the acute toxicity of HgCl₂ (30 μmmol/mmol/kg, once, s.c.), changes of renal function, tissue accumulation of mercury, and urinary excretions of mercury and phenolsulfonphthalein after exposure to HgCl₂ were investigated. Lactate content in the kidney and serum calcium were also measured.Na₂MnO₄ pretreatment enhanced urinary excretion of mercury. Renal function of Na₂MoO₄-pretreated rats was better maintained as compared to that of the rats given HgCl₂ alone at either dose (30 or 15 μmol/kg) although the metal content in the kidney of this group was almost the same as that of the latter HgCl₂-alone rats. This pretreatment prevented the rise in lactate content in the kidney and the reduction of urinary excretion of phenolsulfonphthalein caused by HgCl₂. Na₂MoO₄ reduced serum calcium. These results suggest that Na₂MoO₄ prevented mercury-induced acute renal failure by decreasing tissue accumulation of the metal through urinary excretion of mercury. Better renal hemodynamics attributable to hypocalcalcemia may be a causative factor in the enhancement of urinary excretion of mercury.

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A natural antibody with binding specificity for recombinant tissue-type plasminogen activator (t-PA) expressed in mouse C127 cells was present in almost all disease-free humans and patients with thrombotic disease examined. This antibody was specific for a carbohydrate, α1-3-linked galactose residue, and was isolated by affinity chromatography using Synsorb 90 coupled with the glycosidic epitope Galα1-3Galβ1-4Glc-R as an immunoadsorbent. The evaluation of various glycoproteins for ability to bind the purified antibody in ELISA demonstrated that not only recombinant t-PA from C127 cells but also recombinant erythropoietinc (EPO) and recombinant protein C produced in C127 cells have α1-3-linked galactose residues on their sugar side chains. This anti-α-galactosyl antibody also interacted with natural t-PA from human vascular trees (vascular t-PA) and placenta (placent t-PA), but not to melanoma t-PA, recombinant t-PA, EPO or protein C expressed in Chinese hamster ovary (CHO) cells.

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A high-performence liquid chromatographic (HPLC) method for quanifying unsaturated hexasaccharide and tetrasaccharide from Streptomyces hyaluronidase enzyme digestion products of hyaluronic acid was developed using a gel-permeation column packed with a sulphated polystyrene-divinylbenzene gel. For the oligosaccharides, the separation was accomplished in less than 7 min with a detection limit of 65 ng. An unsaturated non-sulphated disaccharide prepared from hyaluronic acid (ΔDi-HA) and an unsaturated sulphated disaccharide (ΔDi-4S) were analyzed by a HPLC method using a combination of two different gel-permeation columns. The separation of the disaccharides required less than 17 min at a flow rate of 0.7 ml/min with detection limits of as little as 4 ng for ΔDi-HA and 5 ng for ΔDi-4S. Both chromatographic methods were used for assay of a major component of hyaluronic acid and trace amounts of chondroitin sulphates in rabbit synovial fluid. The resulting contents of hyaluronic acid were compared to the values of polymeric hyaluronic acid directly measured by a HPLC method using two gel-permeation columns packed with a poly(hydroxyalkyl methacrylate) gel and the amounts of hyaluronic acid converted from uronic acid content determined by a colorimetric method.

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The two-stage carcinogenesis by 7, 12-dimethylbenz[a]anthracene and 12-O-tetradecanoylphorbol-13-acetate (TPA) in mice was inhibited by kaempferol and flavonol glycosides, whereas naringenin, a flavanone, had no effect. The induction of epidermal ornithine decarboxylase activity by TPA was also inhibited by kaempferol, whereas mauritianin, a kaempferol glycoside, failed to inhibit it. In addition, the effect of the flavonol glycosides on cell-mediated immunosuppression in the two-stage carcinogenesis, observed in term of initiation after 14 weeks, was antagonized by mauritianin and myricitrin. Cell-mediated immunosuppression in the two-stage carcinogenesis was unaffected by kaempferol and naringenin. These results suggest that the inhibitory effects of flavonol glycosides may have been at least partly due to activation of immune responses against tumors.

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The formation of complexes between cycotiamine (CCT) and fatty acids (FA) (tetradecanoic acid, hexadecanoic acid, and octadecanoic acid) in 1, 2-dichloroethane was studied by phase solubility analysis and ultraviolet absorption spectroscopy. The apparent stability constants of the equimolar complexes between CCT and FA were determined at 288, 298, and 310 k, and thermodynamically discussed. The complex formation ability of FA depended on the number of carbon atoms in FA, but the differences were small. The ability of 1-octadecanol was very small, and octadecanoic acid methyl ester had none. From these results CCT-FA were concluded to be hydrogen bonded complexes in which FA act as hydrogen donors. The complex formation reactions were exothermic anc enthalpically controlled.

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Some 2-methylisoselenochromanium salts were prepared. Proton nuclear magnetic resonance spectra in CDCl₃ and electron impact mass spectra showed that the selenonium tosylate (4) and mesylate (5) are selenuranes and that the tetrafluoroborate (2) and triflate (3) are selenonium salts. Their structures at the selenium atom are influenced by their counter anions. Selenonium salts (2-5) reacted with some nucleophiles to give a styrene derivative (6) and methyl phenethyl selenides (7, 9).

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Two new oleanane-type triterpene glycosides, aster saponins E and F, were isolated from the less polar saponin fraction of the root of Aster tataricus L. f. (Compositae), and their structures were elucidated on the basis of spectral evidence and by correlation to the other aster saponins reported earlier. Aster saponin E is 3-O-[O-α-L-arabinopyranosyl-(1→6)-β-D-glucopyranosyl]-2β, 3β, 16α-trihydroxyolean-12-en-28-oic acid (asterogenic acid) 28-[O-β-D-xylopyranosyl-(1→3)-O-α-L-arabinopyranosyl-(1→4)-O-α-L-rhamnopyranosyl-(1→2)-β-D-xylopyranosyl] ester. Aster saponin F is a 3β, 16α-dihydroxyolean-12-en-28-oic acid (echinocystic acid) glycoside, the sugar moiety of which is the same as that of aster saponin E.

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A series of 21-(alkylthio)acetates and 21-(methylthio)methoxides of corticosteroids were synthesized and examined for vasoconstrictive activities. The activities of seven compounds were equal to or greater than that of 9α-fluoro-11β, 21-dihydroxy-16β-methyl-17α-valeryloxy-1, 4-pregnadiene-3, 20-dione (betamethasone 17-valerate, BV). Among them, betamethasone 21-(methylthio)acetate 17-propanoate (2Ca) was found to have the most potent activity, which is superior to that of BV. A structure-activity relationship study revealed that substitution of the 21-hydroxy group of corticosteroids with the (methylthio)acetate function is a useful approach for obtaining potent activity.

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The effect of hot water extract of green tea on the collagen-induced aggregation of washed rabbit platelets was examined. The extract lowered submaximal aggregation and prolonged the lag time in a dose-dependent manner. After fractionation of the extract, it was revealed that the tea catechins (tannins) are active principles for inhibition and that ester-type catechins are more effective than free-type catechins. One of the ester type catechins, epigallocatechin gallate (EGCG), suppressed the collagen-induced platelet aggregation completely at the concentration of 0.2 mg/ml (=0.45 mM). Compargin CI₅₀ values of EGCG and aspirin it was found that the potency of EGCG is comparable to that of aspirin. Thrombin- and platelet activating factor (PAF)-induced aggregation was also inhibited by EGCG. The elevation of cyclic adenosine 3', 5'-monophosphate (cAMP) level was not observed in EGCG treated platelets.

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A novel type of arylsulfotransferase was partially purified from human intestinal bacteria and its enzymatic properties were examined. Polyphenols such as chalcone, xanthone and flavonoid were found to be sulfated by the bacterial arylsulfotransferase though the sulfation activity varied depending upon the positions of the hydroxyl groups. Quercetin, as an example of a flavonol, was rapidly sulfated when p-nitrophenyl sulfate (PNS) was taken as a donor substrate. At a ten-fold molar excess of PNS over quercetin, two products, the 3, 3'-disulfate and 3, 3', 7-trisulfate derivatives, were formed, but the 4'-and 5-hydroxyl groups were not sulfated. In the case of equimolar or two-fold molar excess of PNS to quercetin, only the 3, 3'-disulfate was produced and no monosulfate was formed. The enzymatic procedure is useful as a specific anc concenient method for the preparation of polyphenol sulfate esters.

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We investigated the enhancing effect of three alkyo-2-pyrrolidones on transdermal penetration of phenolsulfon-phthalein (phenol red) and indomethacin from an aqueous vehicle by using an in vitro technique with excised rat skin. The enhancers included 1-methyl- (I), 1-hexyl- (II) and 1-lauryl-2-pyrrolidone (III). These derivatives effectively enhanced the penetration and skin accumulation of phenol red and indomethacin. Lipophilic enhancers such as II and III showed particularly high enhancing effects. The penetration profiles of phenol red and indomethacin showed a lag phase followed by a linear increase. Compounds II and II showed long lag times. The enhancer penetration was also determined. Compounds I and II showed a slight penetration. Compound III showed little penetration but high skin accumulation.

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The stability of β-glactosidase dosage forms was studied by differential scanning calorimetry (DSC). It was found that the observed enthalpy of thermal denaturation was approximately in proportion to remaining enzyme activity, and denaturation temperature was related to protein stability. These results suggest that DSC can be used to determine native proteins in dosage forms and to clarify the factors affecting protein stability. The DSC method seems to be more convenient than conventional activity assay methods, and useful to follow protein denaturation during the manufacturing process and storage of dosage forms.

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We investigated whether interleukin-1 (IL-1) acts as an endogenous pyrogen (EP) on the fever caused by the cell wall skeleton of Nocardia rubra (N-CWS) in guinea pigs. IL-1 activity was expressed as potency of lymphocyte activating factor (LAF). When guinea pig peritoneal macrophages were pulse-stimulated with N-CWS(1-100 μg/ml), dose-dependent LAF activity was detected in the supernatants after culture for 4h. Gel filtration of the culture supernatants on Sephadex G-200 showed that the fractions with LAF activity were not the same as those with cytotoxic activity for L-929 cells, which was measured as an index of tumor necrosis factor (TNF) in parallel with LAF activity. Pyretic activity was detected both in the fractions with LAF activity and in those with cytotoxic activity for L-929 cells. Furthermore, when these macrophages were pulse-stimulated again, this time with the supernatant obtained from macrophages previously pulse-stimulated with N-CWS, LAF and cytotoxic activity for L-929 cells continued to be released from the macrophages.We suggest that IL-1 might be a possible EP in the process of fever elicited by N-CWS, and that such an EP stimulates the macrophages to release further IL-1 or TNF. The resultant long-lasting fever would thus be caused by the continous release of an EP.

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