Copper and zinc-containing superoxide dismutase (Cu, Zn-SOD), and manganese-containing superoxide dismutase (Mn-SOD) were purified from human placenta by heat treatment at 70° for 5 min, following precipitation with ammonium sulfate and column chromatographies. The molecular weights of Cu, Zn-SOD and Mn-SOD estimated by gel filtration were 32000 and 81000, respectively. From the result of SDS polyacrylamide gel electrophoresis, Cu, Zn-SOD consisted of two subunits with an equal molecular weight of 16000 and Mn-SOD consisted of four subunits with an equal molecular weight of 21000. One g-atom each of copper and zinc was contained in the subunit of Cu, Zn-SOD, and one g-atom of manganese was contained in the subunit of Mn-SOD. Immunological properties of superoxide dismutases showed that no organ specificity existed but that a species specificity obviously existed. The method of Laurell's rocket immunoelectrophoresis which can determine Cu, Zn-SOD or Mn-SOD on the basis of the immunological difference has been established. The contents of Cu, Zn-SOD and Mn-SOD in various human tissues were measured by this method.

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Copper and zinc containing-superoxide dismutase (Cu, Zn-SOD), and manganese containing-superoxide dismutase (Mn-SOD) were purified from human placenta with immunoadsorbent columns using anti-human SOD-antibody conjugated Sepharose 4B. The final preparation has a high purity with a good recovery. The Cu, Zn-SOD purified by an immunoadsorbent column had less zinc content and was more labile than the native Cu, Zn-SOD, but this difference disappeared completely by dialyzing the enzyme obtained by immunoadsorbent column against the buffer containing 1 mM ZnCl₂. Other enzymological properties of two SODs were identical with that of native SODs.

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In order to obtain evidences supporting the previous suggestion that restraint and water immersion stressed (RWIS) animals are possibly in the state of partial sympathicotonia, further investgations have been carried out. 1) In the experiment on the influence from the adrenal, the preadministration of dexamethasone, spironolactone or metopirone did not affect the enhancement of response to acetylcholine (ACh) of the duodenum isolated from RWIS mice, while adrenalectomy inhibited both the enhancement of response to ACh and the reduction of response to noradrenaline (NA) of isolated duodenum and vas deferens of RWIS mice. The potentiation of the response to ACh of the duodenum isolated from the adrenalectomized RWIS mouse, on the other hand, was little affected by the pretreatment of dexamethasone. These results suggest that in the RWIS animal the influence from the adrenal medulla is greater than that from the adrenal cortex. 2) In the experiment on the influence from the autonomic nervous system, the treatment of RWIS mice with an adrenergic neuron degenerating agent to cause chemical denervation was found to suppress the potentiation of the response to ACh of isolated duodenum and vas deferens, suggesting the participation of the adrenergic neuron. A vagotomy showed little effect on responses either to ACh or NA of isolated duodenum and vas deferens from RWIS mice. A sympathectomy, however, suppressed these changes in responses, particularly reduction of response to NA was markedly inhibited to the normal level. These findings indicate that the excessive sympathicotonic state in the RWIS animal was remitted by sympathectomy. It may be concluded that the results of present investigations utilizing adrenalectomy, vagotomy and sympathectomy may present evidences for the previous suggestion that the RWIS animal might be in the state of partial sympathicotonia.

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The transport of cephalothin (CET) to the exudate was investigated in croton oil-induced inflammatory rats following the intramuscular administration of CET-aqueous solution, CET-O/W emulsion, or CET-W/O emulsion. The lymphatic transport of CET was the highest following the administration of W/O emulsion. The exudate level of CET was in a decreasing order of W/O emulsion, O/W emulsion, and aqueous solution. In the rat whose blood vessels at femoral rectus, the site of administration, were ligated, the plasma level was decreased by the administration of aqueous solution, but not by W/O emulsion. The exudate level of CET was significantly higher in case of W/O emulsion. Possible mechanisms involved are discussed.

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2-Amino-5-methoxy-2'(or 3')-methylindamine and 2-amino-5-methoxy-2'(or 3')-methylindoaniline were isolated from a coloring matter formed by oxidative condensation of 2, 5-diaminotoluene with 2, 4-diaminoanisole, and their structures were elucidated on the basis of nuclear magnetic resonance and mass spectral data. These two main components of the oxidation product formed at the early stage exhibited a positive result in the mutagenicity test on the thin-layer chromatography plate. The mutagenic activities of purified aminoindamines and aminoindoanilines against Salmonella typhimurium TA 98 were as strong as that of 2-acetamidofluorene. It was also demonstrated that 5-methoxy-2'(or 3')-methylindamine was formed in 10% yield under the conditions used for hair dyeing and the yield was dependent upon the molar ratio of 2, 5-diaminotoluene to 2, 4-diaminoanisole.

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New hapten-carrier conjugates were prepared from 9, 10-dihydro-N-[1-(hydroxymethyl) propyl]-6-methylergoline-8-carboxamide 1-hemisuccinate (dihydromethylergometrine hemisuccinate) by coupling with bovine serum albumin employing the mixed anhydride technique. The specificity of anti-dihydromethylergometrine antiserum elicited in the rabbits by immunization with this antigen was tested by cross-reaction studies with methylergometrine, dihydroergotoxine, dihydroergocristine and dihydroergotamine employing [9, 10-³H]-dihydromethylergometrine and [9, 10-³H]-dihydroergocryptine in the radioimmunoassay procedure. These cross reactivities were observed 100% for methylergometrine, 48.3% for dihydroergotamine, 16.6% for dihydroergotoxine and 6.6% for dihydroergocristine using labeled dihydromethylergometrine. When labeled dihydroergocryptine was the results were 18.3%, 112.1%, 100% and 63.8%, respectively. Methylergometrine concentrations in the postpartal plasma and milk at 2 hr after oral administration of methylergometrine maleate (0.75 mg) to woman were 5.1 ng/ml and 1.3 ng/ml, respectively. Dihydroergotoxine concentrations in the rabbit sera after oral administration of dihydroergotoxine methanesulfonate (4 mg) were studied. The assay was satisfactory to permit the measurement of ergot alkaloid levels in the biological fluids.

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Saframycin S, structurally the decyano-saframycin A and biosynthetically the precursor of saframycin A, is a new antibiotic and is assumed to be the active principle for the formation of the antibiotic-DNA complex to manifest its activity. In the present study on the biological activity of saframycin S, the antibiotic showed a marked activity against Ehrlich ascites tumor with the doses of 0.5 to 0.75 mg/kg/day for 10 days, and the rate of 40-day survivors was 80 to 90%. However, this antibiotic was less effective against P388 leukemia than saframycin A at the doses tested. Saframycin S also showed the highest antimicrobial activity, in particular against gram-positive bacteria, among the saframycin group antibiotics identified to date. The LD₅₀ of saframycin S was 3.2 mg/kg (i.p.) for ddY mice. The biological activity and toxicity of saframycin S in relation to its structure were described.

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The hepatic transport of sulfobromophthalein (BSP) and its glutathione conjugate (BSP-GSH) were compared in rats by pharmacokinetic analysis in an attempt to determine the effect of conjugation on the over-all transfer of BSP from blood into bile. In the control rats, the plasma disappearance of BSP was faster than that of BSP-GSH, while the biliary excretion rate of BSP-GSH was significantly higher than that of BSP for the first 10 min. No significant difference in the total amount excreted in the bile for 4 hr was observed between two dyes. In the rats chronically intoxicated with carbon tetrachloride, typical delays were shown in both the plasma disappearance and the biliary excretion of BSP-GSH. In the control rats, the pharmacokinetic parameters, k₁₂ and k₂₃ of BSP-GSH were significantly smaller than those of BSP, while k₃₄ was significantly greater than those of BSP. In the intoxicated rats, significant decreases were observed in k₁₂, k₂₃ and k₃₄. The binding of BSP and BSP-GSH to the Y-fraction prepared from the control and intoxicated rats were studied by equilibrium dialysis. In the control rats, the number of high affinity bindings sites (n₁) of BSP-GSH was significantly smaller than that of BSP, while no significant difference was revealed in the binding constant of the high affinity binding site (K₁). In the intoxicated rats, both n₁ and K₁ of BSP-GSH were significantly smaller than those of the control rats. It was suggested that the glutathione conjugation might not play as the rate-determining step in the over-all hepatic transport of BSP.

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Hypotensive actions of single and repeated administrations of nifedipine were evaluated in comparison with hydralazine in normotensive, spontaneously hypertensive and renal hypertensive rats. Substantial and prolonged falls in blood pressure were observed following oral dosing with 3 mg/kg of nifedipine in renal hypertensive rats, with 10 mg/kg of nifedipine in normotensive and spontaneously hypertensive rats ans with 10 mg/kg of hydralazine in all three groups of animals. The hypotensive effect of nifedipine was greater in both forms of hypertensive rats than the normotensive rats, while there was no difference in the hypotensive effect of hydralazine in normotensive and hypertensive animals. In spontaneously hypertensive rats tolerance to nifedipine and hydralazine was not observed with oral administrations for 4 weeks at a daily dose of 10 mg/kg. An increase in heart rate always accompanied the hypotension induced by both drugs, but this tachycardia was seen to a lesser extent with nifedipine than with hydralazine. The mechanism of the selective action of nifedipine in hypertensive rats in contrast to normotensive rats is discussed.

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The present investigation was undertaken to study effects of methylxanthine derivatives on rotational behaviour produced by dopamine receptor stimulating drugs and properties of methylxanthine-induced rotation in mice with unilateral 6-hydroxydopamine (6-OHDA)-induced lesions of the striatum. L-Dopa 10 mg/kg, i.p., produced contralateral turning which lasted for about 40 min. When L-dopa 10 mg/kg was biven to mice in combination with theophylline 25 mg/kg, total turns for 2 hr were significantly higher than those of L-dopa and theophylline alone. Caffeine and theophylline in doses of 25 and 50 mg/kg, i.p., respectively, produced marked increase in contralateral rotation in a dose-dependent manner in mice with lesions. Theobromine 100 mg/kg, i.p., also produced a moderate contralateral rotation. Total turns of ipsilateral rotation produced by methylxanthine derivatives were less than 10% of those of contralateral circling. Theophylline-induced contralateral rotation was reduced to nearly 30% of the control by alpha-methyl-p-tyrosine (alpha-MPT). It was also suppressed by spiroperidol, at a dose higher than that to apomorphine or methamphetamine. These results suggest that methylxanthine derivatives produce contralateral rotational behaviour due to not only phosphodiesterase inhibition but also dopamine receptor stimulation.

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