The effects of oral administration of methyl (+) (3S)-1, 2, 3, 4-tetrahydro-3-hydroxymethyl-β-carboline-2-carbodithioate (THC) on acute liver injuries induced by carbon tetrachloride (CCl₄), bromobenzene (B.B.), D-galactosamine (GALN) and α-naphthylisothio-cyanate (ANIT) in rats and allyl alcohol (A.A.) in mice were studied. THC suppressed the elevation of plasma transaminase activities and hepatic lipid contents induced by CCl₄ in rats when the animals were treated with THC for 4 consecutive days simultaneously with CCl₄ administration. THC also suppressed the elevation of hepatic lipid contents induced by the 4 d-treatment of rats with CCl₄ when administered for 4 consecutive days from the next day after treatment with CCl₄. This effect of THC was histopathologically confirmed. In addition, pretreatment with THC protected rats against liver injuries induced by B.B., GALN, and ANIT, but not against A.A. in mice. This protection was evident from the suppression of elevated activities of plasma transaminase and/or elevated contents of hepatic lipid induced by these hepatotoxic substances. Furthermore, THC inhibited the formation of lipid peroxide in rat liver microsomes with an IC₅₀ of 3.2 μM. These results suggested that THC protected rats against liver injuries induced by CCl₄, B.B., GALN and ANIT and had a protective effect on the microsomal membrane against lipid peroxide in vitro.

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Peritoneal macrophages elicited in various ways exhibited marked wheat germ agglutinin (WGA)-dependent cytolytic activity against MM46 cells [nearly 100% lysis at effector to target ratio (E/T)=40], while macrophage cell lines J774.1 and WEHI-3 cells showed less activity (-25% at E/T=0.63-2.5) and no cytolysis (E/T=40), respectively. Studies on the kinetics of WGA-binding showed that peritoneal macrophages had an association constant (K_a) of 0.09-0.18×10⁶ M^-1 and the number of their binding sites (N) was 2-7.7×10⁷/cell, while the K_a and N values of J774.1 and WEHI-3 cells were 1.2×10⁶ M^-1 and 4.1×10⁷/cell and 0.6×10⁶ M^-1 and 5.4×10⁷/cell, respectively. On the other hand, cytolytic target MM46 cells showed values of 0.27×10⁶ M^-1 and 1.8×10⁸/cell. Lower affinities and moderate numbers of binding sites coincided with the efficient WGA-dependent cytolytic activities of the effector cells. The surface of peritoneal macrophages and J774.1 and MM46 cells was iodinated, the cells lysed, the lysates fractionated on a WGA-Sepharose column and the WGA-binding surface-glycoproteins in GlcNAc-specific fractions of the eluate were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Peritoneal macrophages gave three major bands of 153 kilo dalton (kDa), 140 and 90 kDa proteins, whereas J774.1 and MM46 cells both gave 6 bands of very similar molecular weights and intensities that were quite unlike those of peritoneal macrophages. These results suggest that the three major WGA-binding glycoproteins on peritoneal macrophages are involved in adherence and the lytic process in WGA-dependent macrophage-mediated cytolysis.

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The antitumor activity of a newly obtained cyclic hexapeptide, RA-700, from Rubiae Radix was studied using several murine experimental tumor systems. In P388 leukemia (inoculated intraperitoneally (i.p.), administered i.p. : i.p.-i.p.) the maximal increase in life span (ILS_max) resulting from an administration of RA-700 (4 mg/kg/d for 9 d) was 134% and the therapeutic ratio was 400. These values indicate that RA-700 has higher antitumor activity and broader active dose range than that of mitomycin C (MMC, 1 mg/kg/d for 9 d) which was used as a positive control. In the study of the treatment schedules on P388, RA-700 had the highest activity by consecutive injections. In MOPC-104E mouse plasmacytoma system (i.p.-i.p.), ILS_max of RA-700 (2.5 mg/kg/d for 9 d) was 84%. In a solid tumor, colon adenocarcinoma 38 (subcutaneously (s.c.)-intravenously (i.v.)), RA-700 (4 mg/kg/d for 11 d) showed complete cures (8/8) compared to MMC (0.5 mg/kg/d for 11 d) which showed one cure out of 8 animals. In the study of a model of the inhibition of lymph node metastasis using P388 leukemia, the administration of RA-700 at more than 2.5 mg/kg/d i.v. for 7 d resulted in the survival of all animals (5/5) for over 60 d. In the amputation system of the same metastasis model at a dose of 4 mg/kg/d i.v. for 7 d, 3 animals out of 5 survived over 60 d. With regard to spontaneous metastasis using a highly metastatic B16 melanoma, B16-BL6, RA-700 (4 mg/kg/d i.v. for 11 d) showed 97% inhibition of the number of pulmonary nodules compared to the control and this activity was similar to that shown by doxorubicin (2.5 mg/kg i.v. for 11 d). Thus, RA-700 was considered a good candidate of a new type of antitumor agent.

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We investigated the promoting effect of medium chain fatty acid salts such as sodium caprylate, sodium caprate (Na C10) and sodium laurate on rat nasal absorption of insulin. Further, we studied the possible mechanism of the absorption promoting effect by fatty acid salts in terms of hemolytic activity, osmotic pressure, chelating effect and inhibition of leucine aminopeptidase activity. Na C10 exhibited the strongest absorption promoting effect on insulin among these fatty acid salts. The promoting effect of Na C10 was very rapid reaching the maximum immunoreactive insulin level within 5 min and its optimum concentration was about 1%. The amount of insulin absorbed after a nasal administration with Na C10 was about 98% of one half subcutaneous dose. The absorption promoting effect of fatty acid salts on insulin was at least in part associated with the chelating ability for calcium ions and the inhibitory action on leucine aminopeptidase activity.

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By using isolated rat hepatocytes, the mechanism of uptake of a zwitterionic β-lactam antibiotic, cephalexin, was clarified. The uptake followed the combination of saturable carrier-mediated and nonsaturable first-order rate processes. The kinetic parameters were estimated as follows (mean±SD) : maximum uptake rate (V_max), 2.28±0.24 nmol/min/mg of protein ; Michaelis constant (K_t), 6.28±0.31 mM and first-order rate constant (k_d), 0.11±0.012 nmol/min/mg of protein/mM. There was no inhibitory effect by amino acids, dipeptides or organic cations, whereas an organic anion, probenecid, markedly inhibited the hepatic uptake of cephalexin. Several β-lactam antibiotics including zwitterionic and anionic derivatives inhibited cephalexin uptake significantly. The inhibition kinetics revealed that benzylpenicillin and the stereo-isomer l-cephalexin competitively inhibited cephalexin uptake. Furthermore, the efflux of cephalexin from the cells was stimulated by adding benzylpenicillin in the extracellular medium. These results demonstrated that all β-lactam antibiotics have a common transport system with an organic anion such as probenecid, irrespective of their ionic charges, though a cationic charge on the molecule is less advantageous for being recognized by the carrier system.

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In order to more objectively evaluate drug effects in the forced swimming test proposed by Porsolt et al. as a screening method for antidepressants, vibrations of the wall of a tank caused by rats trying to escape from water were recorded. Locomotor activity was also measured in an activity cage. Male Wistar rats were forced to swim once daily for 15 min in a tank of 25°C water filled to a depth of 20 cm. After 4 d sessions of swimming, drugs were administered i.p. 3 times (24, 5 and 1 h prior to the test session). On day 5, 45 min after the last injection of a test drug, locomotor activity was measured for 15 min and then the rat was subjected to the forced swimming test. All antidepressants tested, dose-dependently increased tank vibration. Nomifensine, atropine, methamphetamine, chlorpheniramine and diazepam increased locomotor activity. The remaining drugs had no effect or reduced locomotion. In addition, the pattern of the tank vibrations, caused by rats treated with most drugs, like atropine, showed a burst during the first 5 min followed by sporadic vibrations. Nomifensine and methamphetamine, on the other hand, caused vibrations throughout the 15 min test session. A specific effect of antidepressants was revealed by this forced swimming test in combination with the measurement of locomotor activity.

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The antitumor mechanism of grifolan NMF-5N, a β-1, 3-glucan obtained from mycelia of Grifola frondosa, was examined. Grifolan NMF-5N did not show direct cytocidal effect on cultured tumor cells. However, intraperitoncal injection of grifolan NMF-5N increased the number of peritoneal exudate cells and peritoneal adherent cells which showed cytostatic activity towards syngeneic tumor cells. In an in vivo assay, the administration of carrageenan, an inhibitor of macrophage function, reduced the antitumor activity of grifolan NMF-5N. The delayed-type hypersensitivity reaction was augmented in the grifolan NMF-5N-administered mice. The administration of NMF-5N augmented the induction of cytotoxic T cells but the antitumor activity of grifolan NMF-5N was reduced in athymic nu/nu mice. In addition, the treatment with anti-Thy 1, 2 antibody and complement C' of spleen cells taken from mice which showed regression of tumor due to grifolan NMF-5N, reduced the neutralizing effect in Winn assay. These results suggested that grifolan NMF-5N shows antitumor activity via hostmediated mechanisms and both macrophages and T cells play important roles in the mechanisms.

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A major metabolite of carbamazepine (CBZ), CBZ-10, 11-epoxide (EPO), has been reported to possess anticonvulsant properties. Therefore, the present study was undertaken in order to develop a pharmacokinetic model to predict the behavior of EPO in the body after administration of CBZ. The serum concentration-time curves after oral administration of solution of CBZ (200 mg) or EPO (150 mg) in six healthy subjects showed the characteristic "nose", suggesting that disposition of CBZ or EPO could be described by the two-compartment model. The kinetic parameters of disposition for CBZ and EPO were calculated by the method of Wagner, assuming the absolute bioavailabilities of CBZ and EPO to be 1.0 and 0.81, respectively. Total body clearance and elimination rate constant of EPO were very much larger than those of the parent drug but there was no statistically significant difference in the distribution volume between CBZ and EPO. The formation rate of EPO was calculated by a deconvolution method, and obeyed Michaelis-Menten kinetics. Based on these findings, a pharmacokinetic model of the fate of CBZ and EPO in humans was developed and the time courses of CBZ and EPO in serum after oral administration of three tablet preparations and a solution containing 200 mg of CBZ were simultaneouly fitted to this model by solving the differential equations by the Runge-Kutta-Gill method. There was good agreement between calculated and observed serum values, suggesting that the present model is appropriate to describe the formation and disposition of EPO from CBZ. The formation rate constant of EPO (V_max/K_m/V₁) was approximately one-fifteenth of the elimination rate constant of EPO. This suggested a flip-flop model in which the formation of EPO was rate-limiting in humans. The observation that the serum concentrations of EPO after administration of CBZ were one-tenth to one-twentieth of those of the parent drug was well explained by the flip-flop kinetics of EPO, together with the large differences in total body clearance and elimination rate constant between CBZ and EPO.

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Nifedipine was selected as a representative compound to investigate a method for improving transdermal bioavailability. The general strategy explored to improve the percutaneous transport of nifedipine was the manipulation of thermodynamics of the drug substance by the use of volatile/nonvolatile systems as vehicles. To investigate the potential of the strategy, diffusion studies were conducted using an ethylene-vinyl acetate copolymer (EVA) membrane and full-thickness excised abdominal skin of rats. Little penetration through EVA membrane or rat skin was found either from the volatile solvent ethanol (EtOH) or from the nonvolatile solvent diethyl sebacate (DES). When the vehicle was changed to a mixed solvent containing both EtOH and DES in a volume ratio of 75 : 25, penetration through EVA membrane or rat skin was increased up to 3 to 4 times, compared with those values for DES. The increase in the penetration was accounted for by the increase in the thermodynamic activity of the drug in the nonvolatile vehicle caused by the evaporation of the volatile component. The bioavailability of percutaneous nifedipine in rats was determined from the drug solutions containing different proportions of EtOH and DES. Once again, the highest bioavailability was achieved from the mixed solvent containing EtOH and DES in a volume ratio of 75 : 25. The area under the plasma nifedipine concentration-time curve for the mixed solvent was higher by about 4 times than that for DES.

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The histamine H₂-receptor antagonist, nizatidine (LY 139037), was tested for its effect on the intestinal smooth muscle. Isolated segments of guinea pig ileum were used in Tyrode solution at 37°C. Nizatidine (from 3.2×10^-6 to 3.2×10^-4M) caused a concentration-dependent contractile response by the guinea pig ileum. The average maximum response to nizatidine (3.2×10^-4M) was about 89% of the average maximum response to eserine (3.2×10^-6M). The contractile responses induced by nizatidine were not modified by pyrilamine (10^-8 and 10^-7M). On the other hand, atropine (10^-8 and 3.2×10^-8M) significantly prevented, while eserine (10^-8 and 3.2×10^-8M) significantly enhanced the nizatidine-induced responses in a concentration-dependent manner. These findings suggest that nizatidine exerts an excitatory effect on the guinea pig ileum which seems to be associated with the cholinergic system, probably through a direct and/or an indirect mechanism (inhibition of acetylcholinesterase and/or increased release of acetylcholine).

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A method for the preparation of calibration curves for acetaminophen glucuronide (NAPAG) and acetaminophen sulfate (NAPAS) in rabbit urine without use of authentic compounds in high-performance liquid chromatography was examined. Rabbits were dosed intravenously with acetaminophen (NAPA, 30 mg/kg). Urine was collected and diluted. A plot of the peak area ratio of NAPAG to internal standard against NAPA concentration after the hydrolysis of diluted urine with β-glucuronidase was linear and passed through the origin. A linear tendency was also observed in the plot of the peak area ratio of NAPAS to internal standard against NAPA concentration calculated by the difference between the peak area ratio of NAPA after the hydrolysis with β-glucuronidase and that with β-glucuronidase/arylsulfatase. Thus, once the calibration curve has been prepared following the enzyme hydrolysis of NAPAG and NAPAS, then the concentration of NAPAG and NAPAS in the sample solution can be calculated from the peak of NAPAG and NAPAS, respectively. The method is simple, and has the advantage that pure standards of the individual NAPA metabolites are not required.

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We studied the effect of loxistation (EST), an orally applicable cysteine proteinase inhibitor, on dystrophic hamsters, UM-X7.1.E-64-c, an active form of EST, was continuously found in plasma when the feed containing EST was given ad libitum. The drug concentration in plasma of dystrophic hamsters receiving EST feed showed distinct dose dependency. A diet containing EST from weaning to death significantly prolonged the survival period of the dystrophic hamsters.

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The alleviation of acute toxicity of paraquat dichloride (PQ) by sodium dextran sulfate (DS), which is clinically used for antihyperlipemia was studied in mice and rats. When mice were orally given DS (2000 mg/kg) or sodium polystylene sulfonate (SPS), (2000 mg/kg) within 1 h (0, 10, 20, 30, 60 min) after PQ ingestion (200 mg/kg), the effectiveness of DS in alleviating PQ toxicity was greater than that of SPS. In rats treated with DS (2000 mg/kg) or SPS (2000 mg/kg), within 4 h (0, 2, 4 h) after PQ administration (200 mg/kg), the effectiveness of DS was less than that of SPS. However, the effectiveness of DS in the alleviation of toxicity was similar to that of SPS, when given to mice and rats (2000 mg/kg) within a short time after PQ ingestion (200 mg/kg). These results suggested that DS might serve as an antidote for acute toxicity of PQ.

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