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The development of robust and reactive chiral catalysts is a fundamental aim in asymmetric catalysis, and crucial for providing efficient methods for synthesizing chiral molecules. Chiral paddle-wheel bimetallic complexes provide a highly tunable chiral environment in rhodium-catalysed asymmetric carbene/nitrene transfer reactions and Lewis acid-catalysed reactions. Chiral paddle-wheel complexes having other transition metals as the reactive metal centre, however, have not yet been identified in asymmetric catalysis. Here, we report the synthesis, structures and high catalytic performances of chiral paddle-wheel diruthenium complexes. The cationic chiral diruthenium complex [Ru2((S)-BPTPI)4]+ exhibited remarkable reactivity as a Lewis acid catalyst for asymmetric hetero-Diels–Alder reactions, achieving a turnover number of up to 1,880,000 with high enantioselectivity (>90% e.e.). The chiral diruthenium complexes also exhibited good reactivity and high enantioselectivity in C–H amination and cyclopropanation reactions under oxidizing conditions, indicating their high tolerance towards oxidation. Our results reveal the chiral paddle-wheel diruthenium scaffold as a promising platform for asymmetric catalysis. The development of chiral catalysts is of fundamental importance in asymmetric catalysis. Now, chiral paddle-wheel diruthenium complexes are reported that are stable under oxidizing conditions and effective in asymmetric C–C and C–N bond-forming reactions with turnover numbers of up to 1,880,000.

A novel indole-2,3-epoxide equivalent, 2-hydroxyindoline-3-triethylammonium bromide, was found to be a convenient reagent for formal C3-electrophilic reactions of indoles with various nucleophiles. By taking advantage of the nucleophilic character of the oxygen of the 2-hydroxyindoline, the interrupted retro-Claisen and interrupted Feist-Bénary reactions with 1,3-dicarbonyl compounds were efficiently achieved.

An experimental study on CH 4 /air propagating flames in a packed pebble bed was performed over a wide range of ambient pressures and flow velocities to clarify the combustion characteristics in a packed bed at high pressure. It focused on the turbulent flame propagation in cases that the flame thickness was sufficiently smaller than the void scale of the packed bed at high pressure. The flame speed was successfully measured at pressures of 0.1–1.0 MPa and flow velocities of 5–130 cm/s using a high-speed video camera, and then the relationships between the flame speed and pressure, flow velocity, pebble diameter and pebble Reynolds number, Re p , were examined. Turbulence measurements at high pressure were also conducted using a 2-D pseudo packed pebble bed to investigate the turbulence intensities in the flow channels and the estimated turbulent flame structure. Results of flame propagation experiments showed that the flame speed in a packed bed had a minimum value at Re p ≅ 150 and rapidly increased with the increase of the flow velocity and pressure. Results obtained in the turbulence measurements indicated that this change of the flame propagation mode was caused by the transition from laminar to turbulent flow when the pressure increased. A significant correlation was seen between the flame speed in a packed bed, S r , at high pressure and the turbulent burning velocity, S T , estimated from the results of turbulence measurements and previous studies on turbulent premixed Bunsen flames by Kobayashi et al., indicating the similarity between the turbulent premixed combustion with and without packed pebbles at high pressure. Flame extinction during propagation at high pressure was also seen as well as at ordinary pressure, the extinction mechanism being explained based on the competition between the heat release in turbulent flames and turbulent heat transfer to the pebbles.

Effects of nucleotides and cations on 2-[125I]iodomelatonin binding sites in the goldfish brain were examined. Nucleotides (10–6–10–3 M) dose-dependently inhibited the specific binding with the following order of potency: guanosine 5''-O-(3-thiotriphosphate) (GTPγS) >GTP = GDP >GMP = ATP >cyclic GMP. Cyclic AMP was ineffective. The treatment of membranes with GTPγS induced rapid dissociation of 2-[125I]iodomelatonin from membranes when added at the steady state, increased the Kd and decreased the Bmax values as revealed by saturation analysis, and increased the IC50 value of melatonin to inhibit the specific binding. The treatment decreased the specific binding to membrane preparations obtained from six brain regions as well. Inorganic salts (5–200 mM) dose-dependently inhibited the specific binding with the following order of potency: CaCl2 > MgCl2 > LiCl > NaCl > choline chloride > KCl, except for 5 mM MgCl2, which enhanced the specific binding. Saturation experiments demonstrated that 75 mM CaCl2,100 mM MgCl2 and 200 mM NaCl increased the Kd and decreased the Bmax while 5 mM MgCl2 increased the Bmax value. These results imply that G protein and physiological concentrations of cations are involved in the regulation of melatonin binding sites in the goldfish brain.

In an attempt to define the role of the pineal hormone melatonin and two analogues (5-methoxytryptamine, 5MT, and 6-hydroxymelatonin, 6HM) in limiting oxidative stress, the present study investigated the changes in glutathione, lipid peroxidation, and the activity of the antioxidant enzyme glutathione peroxidase after exercise (swimming for 60 min) with or without treatment with the indolamines mentioned. Lipid peroxidation was measured by estimating tissue levels of malondialdehyde and 4-hydroxyalkenals; the experimental animals in these studies were male Sprague-Dawley rats. In the liver, swimming exercise increased the levels of reduced glutathione (GSH) and also significantly increasing oxidized glutathione (GSSG), while decreasing the GSH/GSSG ratio, an index directly related to oxidative stress. When the animals were treated with melatonin, the concentrations of GSH and GSSG were also increased after swimming; however, no reduction in the GSH/GSSG ratio appeared. In the animals treated with 6HM the changes were the same as in those treated with melatonin. In muscle as well, the concentration of GSH and the GSH/GSSG ratio were decreased following 60 min of swimming. Pretreatment of the rats with melatonin prevented these effects. Pretreatment of the rats with both 5MT and 6HM also prevented the changes. Brain GSH/GSSG ratio was not affected by either exercise or indolamine administration. Swimming enhanced lipid peroxidation in the liver, muscle and brain; however, this was prevented in animals treated with melatonin or 6HM before swimming. Glutathione peroxidase was significantly elevated after exercise in the brain but not in the liver and muscle. It is concluded that swimming imposes a severe oxidative stress and suggests that melatonin and, to a lesser degree, 5MT and 6HM confer protection against the oxidative damage associated with swimming for 60 min. This mechanism may be reasonably attributed to their indole structure, which possibly allows these molecules to act as free-radical scavengers.

In order to develop a stain with increased sensitivity and selectivity for cadmium (Cd), we synthesized and characterized 2-(8-quinolylazo)-4,5-diphenylimidazole (QAI). This chelating agent was more than twice as sensitive for Cd than the best conventional staining agents, including benzothiazolylazo-beta-naphthol. Differentiation between Cd and zinc (Zn) was achieved by immersing tissue sections in TRIS(2-aminoethyl)amine before they were stained with QAI. This pretreatment made it possible to selectively stain for Cd by blocking Zn.

The present study used male Sprague-Dawley rats to investigate changes in glutathione [reduced (GSH) and oxidized GSH (GSSG)], lipid peroxidation (as indicated by tissue levels of malonaldehyde and 4-hydroxyalkenals), and the activity of the antioxidant enzyme glutathione peroxidase after a bout of swimming (30 min.) with or without melatonin (N-acetyl-5-methoxytryptamine) treatment. In muscle, the concentration of GSH and the GSH/GSSG ratio were decreased following 30 min. of swimming ; these changes are indicative of enhanced oxidative stress. Pretreatment with melatonin prevented these effects. In liver, swimming increased significantly both GSH and GSSG, and decreased the GSH/GSSG ratio. When animals were treated with melatonin, concentrations of GSH and GSSG were also increased after swimming ; however, the reduction in the GSH/GSSG ratio was prevented by melatonin. Brain GSH/GSSG ratio was not affected by exercise or by melatonin. Swimming enhanced the levels of lipid peroxidation products is muscle ; this was prevented in animals treated with melatonin. Glutathione peroxidase activity was significantly elevated after swimming in both liver and brain with the change not being influenced by concurrent melatonin treatment. It is concluded that swimming imposes an oxidative stress on liver and skeletal muscle and the results show that melatonin confers partial protection against oxidative toxicity, especially in muscle.

Immunocytochemical detection of ubiquitin in the nucleus of rat LH cells and the effects of castration and testosterone replacement on the occurrence of immunoreactive ubiquitin in the nucleus were investigated. Immunoreactive ubiquitin occurred in certain nuclei, mostly belonging to identified LH cells. The concentration of testosterone in blood was altered by castration and implantation of testosterone into castrated rats, and the occurrence of ubiquitin was examined weekly for the following 4 weeks. In castrated rats, the proportion of LH cells with ubiquitin-immunoreactive nuclei was high throughout the experiment. In castrated rats implanted with testosterone, on the contrary, the proportion remained significantly lower. Ubiquitin may be involved in the cellular activity of LH cells in the rat pituitary.

Distribution of fibronectin, laminin, and collagens type I, III, IV, and V in the lobular regions of regenerating rat liver was studied by indirect immunofluorescence. Little or no laminin was detected in sham-operated controls throughout the experimental period, while it was detected in sinusoids of regenerating liver as early as 6 h after partial hepatectomy (PH). After reaching a maximum at 24 h, it decreased and was barely detectable 6 days after PH. Changes in the other extracellular matrix (ECM) proteins were evident 3 days after PH, but not earlier than 24 h. Hepatocytes isolated from regenerating rat livers were tested in a short term assay for attachment to the substrates coated with the ECM proteins. The attachment of hepatocytes to laminin substrates increased 12 h after PH, reached a maximum at 24 h, and decreased to the control level 6 days after PH, while that of the control remained constant. The attachment to fibronectin substrates was not different between regenerating livers and controls at any time point. The attachment to collagen did not change earlier than 24 h after PH, but increased slightly 3 days after PH. Primary rat hepatocytes cultured on the substrates coated with the ECM proteins were determined for replicative DNA synthesis in response to epidermal growth factor. Both in normal liver and in regenerating liver 24 h after PH, laminin was one of the most effective substrates in supporting the responsiveness of hepatocytes to the growth stimulus. Taken together, these results suggest the importance of hepatocyte-laminin interaction during the early stage of liver regeneration possibly in growth stimulation of hepatocytes and/or maintenance of hepatocyte-specific functions.

Ocular melatonin rhythms in the goldfish were studied and compared to those in the pineal organ and plasma. Under light:dark (LD) of 12 h light:12 h dark, melatonin contents in the eye as well as the pineal organ and plasma exhibited clear day-night changes with higher levels at mid-dark than at mid-light. However, melatonin contents in the eye at mid-light and mid-dark were approximately 100 and 9 times greater than those in the pineal organ, respectively. Day-night changes of ocular melatonin persisted after pinealectomy, which abolished those in plasma melatonin under LD 12:12. Ocular melatonin contents in the pinealectomized fish at mid-light were significantly higher than those in the sham-operated control. Under constant darkness (DD), circadian melatonin rhythms were observed in the eye but damped on the 3rd day, whereas plasma melatonin rhythms generated by the pineal organ persisted for at least 3 days. Under constant light, ocular melatonin contents exhibited a significant fluctuation with a smaller amplitude than that under DD, whereas plasma melatonin remained at low levels. These results indicate the involvement of LD cycles, a circadian clock, and the pineal organ in the regulation of ocular melatonin rhythms in the goldfish.

We studied growth hormone (GH) and prolactin (PRL) messenger ribonucleic acids (mRNAs) in 14 cases with densely granulated somatotroph (DG) adenomas and 10 cases with sparsely granulated somatotroph (SG) adenomas using in situ hybridization with digoxigenin-labeled probes and correlated these data with their immunohistochemical results. A good correlation between in situ hybridization results and immunohistochemical data was found in most cases examined. The DG adenomas generally had a diffuse and intense GH immunoreactivity and GH hybridization signal, whereas in SG adenomas the number of cells exhibiting a GH mRNA signal and the strength of the GH mRNA signal in these cells were relatively lower than those of DG adenomas, indicating that lower expression of the GH mRNA signal is responsible for lower GH production in SG adenomas. In addition, PRL mRNA expression differed in the two types of adenoma; in DG adenomas, seven cases (50%) expressed PRL mRNA signal with a focal or scattered distribution despite normal serum PRL levels, but two showed no PRL immunoreactivity. In contrast, in SG adenomas, only one case contained a few cells possessing a PRL mRNA signal despite having no PRL immunoreactivity. It can be concluded that DG and SG adenomas, which have been considered variants of the same tumor, display definite differences as to GH or PRL gene expression in each type of adenoma.

An addition of methanol into the aqueous solution of Chrome azurol S (CAS) accelerates the reaction of CAS with Cr. Mechanism of the accelerative effect of methanol was discussed spectrometrically. CAS has four chemical species, and the third chemical species (in which three protons released) is most liable to chelate Cr. In a high pH range, the third chemical species increases in the solution, however, at the same time, an increase in hydroxy ion interrupts the reaction between CAS and Cr. Results of spectrometrical measurements indicate that the added methanol accelerates the dissociation of the second chemical species of CAS in a low pH range and the reduction of the fourth chemical species in a high pH range. Consequently, the third chemical species is highly concentrated in the solution, that leads to accelerating the Cr-CAS reaction without any interruption caused on an increase of hydroxy ion.

A highly sensitive and specific method for staining exogenous chromium and iron in tissues is described. This method is superior to conventional complex-forming methods with regard to its sensitivity and specificity for these metals. The staining reaction is based on the metal-catalysed oxidation of phenylamines. Tissue sections were incubated in a medium containing hydrogen peroxide and phenylamines, p-phenylenediamine or phenylhydrazine. Results obtained from test-tube experiments concerning the catalytic activities of metals indicated that the staining reactions depends on the activities of metals in tissues.

Thiazolylazocresol (TAC) was found to demonstrate cadmium and zinc in different colors in tissue sections, enabling differential staining by coloration without a masking agent. The metals in stained sections were microscopically identified by the aid of a laser microprobe.The absorption spectra obtained from TAC-metal chelates in a test tube show three absorption maxima: around 650 nm (λ2), 550 nm (λ1) and 400 nm. The ratio of the absorption of λ1 and λ2 with cadmium differs considerably from that of zinc. The difference in the ratios (λ2/λ1) is presumed to be the cause of the differential staining by coloration.

As part of the continuous developmental research for a highly sensitive chelating agent for cadmium, benzothiazolylazophenol derivatives (BTAP) were synthesized and their staining properties for cadmium were examined. The compounds synthesized for this study were identified as being BTAP by analysis of their melting points and C,H,N,S content. Benzothiazolylazo-p-methoxyphenol, -p-chlorophenol and -p-cresol stained cadmium with the highest optical density and the most precise localization amongst dyestuffs reported to date. Differential sequestration by pyrophosphate makes it possible to distinguish cadmium masked by the presence of zinc. With the modification of a chelating agent to increase sensitivity, the improvement in molar absorptivity was also considered theoretically.

We describe a new autoradiographic technique for differentially demonstrating the presence of trace amounts of cadmium that remains unbound to metallothionein in tissue sections. This procedure involves [14C-1,4,5,8--naphthol]-1-(2-thiazolylazo)-2-naphthol (TAN) instead of a metal radioisotope. Tissue sections were stained using labeled TAN, and were then subjected to routine autoradiographic procedures. The results obtained indicated that the distribution of photographic grains truly reflected the distribution of metallothionein-unbound Cd in the sections. This demonstrates the suitability of this procedure for the differential demonstration of cadmium present in different chemical forms in tissues.

1. The mayfly larvae of a heavy metal resistant species (Baetis thermicus) and two heavy metal susceptible species (B. yoshinensis and B. sahoensis) were exposed to cadmium at a concentration of 10 μg/1 and the comparative induction of cadmium-binding protein was correlated to their susceptibility to the metal. 2. Amounts of cadmium accumulated in the larvae were increased linearly depending on the duration of exposure. However, there was little difference in the cadmium concentration in each species. 3. Cadmium-binding protein was induced only in the larvae of resistant species (B. thermicus). 4. Copper and zinc concentrations in the two susceptible larvae were decreased by cadmium exposure, while zinc but not copper concentrations were increased in the resistant larvae.

An electron microscopic study was performed to clarify the effects of tunicamycin, a glycosylation inhibitor, on rat anterior pituitary cells. Tunicamycin (10, 50, and 100 micrograms/250 g B.W.) was intraperitoneally injected into rats, which were sacrificed 24 hrs later. Protein hormone producing GH and prolactin cells, and ACTH cells which are known to have a glycosylated precursor, showed no recognizable ultrastructural changes. TSH cells and gonadotrophs, both of which secrete glycoprotein hormones consisting of alpha and beta subunits, showed remarkable dilatation of the rough endoplasmic reticulum, and decreased numbers of secretory granules. These results suggest that the role of glycosylation in TSH cells and gonadotrophs may have a different biological significance from that in ACTH cells.

The compound 2-(5-bromo-2-pyridylazo)-5-diethylaminophenol (Br-PADAP) was found capable of staining very small amounts of metals in tissue. This finding stemmed from the examination of the staining properties of pyridylazophenol analogs; 2-(2-pyridylazo)-phenol, 2-(2-pyridylazo)-5-dimethylaminophenol, 1-(2-pyridylazo)-2-naphthol, 4-(2-pyridylazo)-resorcinol, and Br-PADAP. The unusually high sensitivity of Br-PADAP is attributed to the presence of an alkylamino group at the 5-position of the benzene ring. The presence of this alkylamino group forms a charged quinoid structure, and contributes resonance and optimum solubility to the hybrid by introducing a bromine atom into the pyridine ring. We report the staining results obtained from a variety of metals and the molar absorptivity of their metal chelates.