To develop an isoelectric focusing apparatus using a cellulose acetate membrane (Separax EF), we have designed a thermoelectric cooling isoelectric apparatus. This apparatus has two characteristics. Firstly, the cooling system was switched to a thermoelectric cooling system from an ice-cooling system. Secondly, the chamber lid of the electrophoretic apparatus was also devised so that samples could be applied without opening the chamber lid. With this apparatus we could perform the isoelectric focusing without worrying about room temperature and humidity in the laboratory. Applying 2000 V for an extra 5 min with our module cooling system, we achieved a much higher degree of resolution with three sheets of cellulose acetate membrane (Separax EF) overlaid for simultaneous electrophoresis. Thus, three types of information could be obtained from only one electrophoretic procedure.
We examined the effect of α-linolenic acid (18 : 3 (n-3)) pretreatment on the metabolism of ω-3 and ω-6 polyunsaturated fatty acids and histamine content and release of RBL-2H3 cells. RBL-2H3 cells grew without reduction in number when incubated with subculture media for 3 d and then placed again in serum-free medium with bovine serum albumin (BSA) and epidermal growth factor (EGF). Cholesterol pullulan (10 μg/ml) emulsified α-linolenic acid (20 μg/ml) was recommended as an additional form serum free medium. We determined the fatty acid composition in all neutral lipids, free fatty acids and all phospholipids in α-linolenic acid-treated cells. In all cases the concentration of α-linolenic acid and docosahexenoic acid (DHA, 22 : 6 (n-3)) was increased, while linolenic acid (18 : 2 (n-6)) was slightly and arachidonic acid (20 : 4 (n-6)) was markedly decreased. Content of histamine in α-linolenic acid-treated cells was remarkably lower than that of untreated cells. Accordingly, net histamine release stimulated by antigen or A23187 was also markedly decreased in the α-linolenic acid-treated cells, as was the percent histamine release stimulated by antigen. Results from our in vitro experiment suggest that the anti-allergic effect of α-linolenic acid may be caused either by the decrease in histamine content or by inhibition of the release of chemical mediator resulting from changes in the fatty acid composition.
Arylsulfate sulfotransferase purified from Eubacterium A-44 has higher specific activity than the enzymes from Klebsiella K-36 and Haemophilus K-12. Propylparaben and butylparaben were good substrates among several parabens. The antibacterial activity of parabens was reduced by the sulfation of the phenolic hydroxy group. Tyrosine-containing peptides, kyotorphin, enkephalin and cholecystokinin non-sulfate, were effective as acceptor substrates by A-44, K-36 and K-12 sulfotransferases.
Helicobacter pylori (HP) produces strong urease [EC 126.96.36.199], which is considered to play a role in the pathogenesis of gastritis and peptic ulcers. Inhibitions against this enzyme have been studied with hydroxamic acid (HXA) derivatives of aliphatic or aromatic carboxylic acids, amino acids and dipeptides. A number of HXAs potently inhibited the urease (I50 values were near the order of 10-6M), and H-Ile-Gly-NHOH (I50=0.20×10-6M) was the most potent inhibitor among the derivatives. HP urease was inhibited more potently, in general, than Jack bean (JB) urease by HXAs, and a correlation between the chemical structures of HXA derivatives and their inhibitory effects on HP urease was observed, in comparison with JB urease.
Intrinsic factor (IF) is a vitamin B12 binding protein that is secreted from the gastric mucosa. We tested secretagogues which stimulate IF secretion in rat gastric perfusion and found that carbachol and cholecystokinin octapeptide (CCK-8) stimulated secretion, but histamine and tetragastrin did not. To confirm these results, we examined IF secretion from isolated rat chief cells. For this purpose, we established an enzyme immunoassay (EIA) using an avidin-biotin peroxidase complex to measure small amounts of IF. To prepare an anti-rat IF, IF was isolated from the stomach, and was injected into a rabbit for immunization. Rat gastric chief cells were isolated from the gastric mucosa with Dispase and a Percoll gradient centrifugation, and were cultured. We examined the effects of chemicals by adding them to culture dishes of chief cells in a CO2 incubator. Released IF in culture medium was determined by EIA. Carbachol, CCK-8 and secretin stimulated IF secretion from cultured chief cells, while histamine and tetragastrin did not ; Forskolin and A23187 also stimulated the secretion. We concluded that carbachol and CCK-8 stimulated IF secretion via an increase of intracellular Ca2+ concentration and that secretin did so via a cAMP accumulation.
DX-9386, a traditional Chinese medicinal prescription consisting of ginseng, polygala, acorus and hoelen in the ratio of 1 : 1 : 25 : 50 (dry weight), was studied regarding the formation of long-term potentiation (LTP) in the dentate gyrus of anesthetized rats. Single oral administration of DX-9386 did not affect LTP formation evoked by suprathreshold tetanic stimulation ; however, it significantly intensified the spike amplitude evoked by a subthreshold stimulation. LTP formation induced by suprathreshold tetanus was significantly inhibited by ethanol given either orally or intracerebroventricularly. DX-9386 significantly antagonized this inhibitory effect of ethanol. Basal spike amplitude was not influenced by DX-9386. These results indicate that DX-9386 potentiated LTP formation in the hippocampus and suggest that the ameliorative effect of this prescription on learning deficit model animals was, at least partly, due to its direct action on the hippocampus.
Effects of diazoxide on norepinephrine-induced vasocontraction in vitro and global ischemia-induced lactate accumulation in the myocardial tissue in vivo were studied in rats. Diazoxide produced relaxation of the isolated rat aorta contracted by norepinephrine in a dose dependent manner. The relaxation of the aorta was associated with reduction of intracellular Ca2+ concentration. This reduction may be due either to activation of KATP channels or Na+-K+ ATPase, or to both. Global ischemia induced by aorta constriction for 30 min in vivo increased the myocardial tissue level of lactate. Pretreatment with diazoxide (10 mg·kg-1, i.v.) significantly attenuated the accumulation of lactate due to global ischemia. The present study suggests that diazoxide reduces ischemic influence on the myocardium partly through its vasodilatory action.
Effects of crude alkaloids extracted from the stem bark of Hunteria zeylanica GARD. (H. zeylanica) on nociceptive responses, capillary permeability, yeast-induced hyperthermia, pentobarbital-induced sleep, and spontaneous motor activity were investigated. Oral administration of 50 mg/kg H. zeylanica alkaloid extract significantly decreased the number of writhings induced by intraperitoneal acetic acid. The extract at 100-200 mg/kg significantly increased nociceptive threshold of the inflamed but not the non-inflamed paw in the Randall-Selitto test. Moreover, in the formalin test, the extract (100 mg/kg) significantly decreased licking activity in the late phase without affecting the activity in the early phase. However, the extract did not produce antinociceptive effect in the hot plate test, while it inhibited increase of vascular permeability induced by acetic acid in the capillary permeability test. Moreover, the extract dose-dependently reduced yeast-induced hyperthermia in rats without affecting normothermia. It did not affect pentobarbital-induced sleep, but significantly increased locomotor activity at 100 mg/kg. These results suggest that H. zeylanica alkaloid extract possesses antinociceptive and antipyretic effects, and that the former effect may be mediated by its anti-inflammatory action.
1, 3-Dichloropropene induced time- and dose-dependent toxicity and lipid peroxidation were examined in isolated rat hepatocytes. HPLC method with chemiluminescence detection (CL-HPLC) was employed to determine phosphatidylcholine hydroperoxide (PCOOH) and phosphatidylethanolamine hydroperoxide (PEOOH) contents. The release of lactate dehydrogenase (LDH) as a toxicological parameter was significantly increased after 90 min incubation at 1 mM of 1, 3-dichloropropene and after 60 min incubation at 5 mM, respectively. The cellular PCOOH and PEOOH contents were increased after 90 min incubation at 1 mM of 1, 3-dichloropropene, and after 15 min for PCOOH and 30 min for PEOOH at 5 mM, respectively. The increase of cellular phospholipid hydroperoxide preceded the cytotoxicity. Hepatotoxicity was effectively prevented by preincubation with d, l-α-tocopherol (α-toc.) accompanied by prevention of the membrane phospholipid peroxidation. In conclusion, the peroxidation of phospholipid preceded cytotoxicity, and cytotoxicity was effectively prevented by α-toc. These results indicated that the peroxidative degradation of membrane phospholipid is one of the main causes of cytotoxicity by 1, 3-dichloropropene.
We fractionated normal urinary proteins obtained from 40 healthy subjects using cellulose acetate membrane electrophoresis and stained them with Acid Violet 17. The electrophoretic patterns were classified into four groups. Each of groups I, II, III, IV had an albumin peak and 1, 2, 3, and 4 additional globulin peaks, respectively. Within-day variation study showed that the pattern was fundamentally specific to the individual, although some intermediate cases were observed. We were unable to determine which type was standard for normal subjects. However, the concentration of Tamm-Horsfall protein was speculated to be an important factor in determining the patterns. Group III showed significantly higher values than group I in urine albumin, total protein, and β-N-acetyl-D-glucosaminidase and this group was believed to include subjects in the subclinical stage of a glomerular disease. All specimens belonging to group IV showed an obvious fraction of α1 globulin which is often found in urine specimens of patients with renal diseases of tubular origin or other pathological conditions.
A highly sensitive detection system for acid proteinase separated on polyacrylamide gel was established. This system consisted of two-dimensional electrophoresis, combined with isoelectric focusing and polyacrylamide gel electrophoresis, and casein clotting (caseogram). Human urine, serum and gastric tissues obtained from normal individuals and gastric cancer patients were analyzed using this system. The previous electrophoretic method was not sufficiently sensitive to detected small amounts of pepsinogen (PG) C in normal urine. However, the new rapid and sensitive method clearly revealed its presence. In gastric tissue containing cancer cells, an additional proteinase, which was not present in normal tissue, was detected and named medium moving proteinase (MMP). MMP resembled PGs in alkaline stability rather than the non-PG proteinase, slow moving proteinase (SMP).
This study was conducted to determine the beneficial effects of treating digestive disorders of (6E, 12E)-tetradecadiene-8, 10-diyne-1, 3-diol diacetate (TDEYA) detected in the plasma in hydrolyzed form : (6E, 12E)-tetradecadiene-8, 10-diyne-1, 3-diol (TDEY), following the oral administration of a decoction of Atractylodes rhizome to rats. Assessment was also made of the efficacy of TDEYA in experimental gastric disorder models. Oral administration of TDEYA at doses of 300 to 500 mg/kg suppressed the formation of gastric lesions induced by indometacin in a dose-dependent manner. TDEYA at a dose of 200 mg/kg suppressed gastric lesions induced by an ischemia-reperfusion injury model. TDEYA at doses of 100 to 300 mg/kg did not show suppressive effects on water immersion stress-induced gastric lesions. TDEYA showed no active oxygen species scavenging action, nor did it have any effect on superoxide dismutase activity in the stomach tissue. TDEYA at doses of 200 to 500 mg/kg significantly suppressed xanthine oxidase (XO) activity in the stomach tissue follwing its oral administration. The suppressive effects of TDEYA on lesion formation induced by indometacin and ischemia-reperfusion injury models would thus appear to be due in part to the inhibition of XO activity in the stomach tissue.
Examination was made of the urinary and biliary excretion of metabolites of daidzin and daidzein, the major components of roots of Pueraria lobata OHWI (Leguminosae) in rats. The urine of rats administered daidzin orally contained four major metabolites, daidzein 7, 4'-di-O-sulfate (M-1), daidzein 7-O-β-D-glucuronide (M-2), daidzein 4'-O-sulfate (M-3), daidzein (M-4), as determined from spectroscopic and chemical data. The urine of rats treated with daidzein contained M-2-M-4 in the above metabolites. Total cumulative amounts of the four metabolites excreted in the urine at 48 h following the oral administration of daidzin and daidzein were approximately 4.8% and 4.6% of the doses administered, respectively. The bile of rats administered daidzin orally contained M-1-M-4. Daidzein 7-O-β-D-glucuronide 4'-O-sulfate (M-5), a major biliary metabolite, was identified by the high-performance liquid chromatography (HPLC), liquid chromatography-mass spectrometery (LC-MS) and nulcear magnetic resonance (NMR) spectra. At least daidzin appeared to be hydrolyzed to aglycone after absorption in the body, and as a part of metabolites, M-1-M-4 having free hydroxyl, glucuronided or sulfated hydroxyls at the C-7 position, may then be excreted in the urine and bile.
The effects of various surfactants and protease inhibitors on the nasal absorption of recombinant human granulocyte colony-stimulating factor (rhG-CSF) were examined in rats. No effects of bile salts and acids such as sodium glycocholate or taurocholic acid, amphoteric surfactants such as lauryldimethyl betaine, or anionic surfactants such as sodium lauryl sulfate on the absorption were found at a concentration of 1%. But non-ionic surfactants with hydrophile/lipophile balance (HLB) of 13 to 18 increased the total leukocyte numbers maximally by about 250% as a relative increase ratio to the control without surfactants. The increase in the plasma rhG-CSF concentration was obviously observed only in the presence of non-ionic surfactants, and in particular, the effects of Laureth-9 on the increase in total leukocyte numbers and plasma rhG-CSF concentration were maximal. In the presence of various kinds of protease inhibitors, the increasing effect of rhG-CSF on the total leukocyte numbers was not changed. Consequently, it is considered that the permeation of rhG-CSF through the nasal epithelium can be improved by non-ionic surfactants, but the effect of a protease inhibitor is smaller than that of the surfactant.
Glutathion (GSH) was covalently attached to dextrans with various molecular weights of 2, 5, 10, 40, and 70 kDa by the cyanogen bromide activation method. The conjugates obtained synthetically were white or pale yellowish powders containing 6-10% (w/w) of GSH. The average molecular weights of the conjugates were estimated to be larger and the molecular weight distribution was a little broader than that of each original dextran. The conjugates significantly stabilized GSH and liberated it gradually under physiological conditions (t1/2=0.99-1.6h). Mice depleted of GSH by treatment with buthionine sulfoximine, a potent inhibitor of γ-glutamylcysteine synthetase, exhibited a significant increase in hepatic GSH level after intravenous injection of the conjugates. In mice given a hepatotoxic dose of acetaminophen, the survival rate increased progressively with coadministration of the conjugates, whereas a small improvement was found when free GSH was given. The conjugate of GSH attached to dextran with the molecular weight of 40 kDa exhibited the highest prophylactic effect on acetaminophen-induced hepatotoxicity in mice. The prolonged retention of the conjugates of larger molecular weight in the circulation would cause a higher hepatic accumulation. These results suggested that molecular size would be the most critical factor in the delivery of GSH, as a dextran conjugate, into the liver.
To examine how efficient primary cultured brain capillary endothelial cells are as an in vitro blood-brain barrier (BBB) system with regard to drug transport, we compared bovine brain capillary endothelial cells (BBCEC) with bovine aortic endothelial cells (BAEC). Paracellular and transcellular transport were examined. [14C] Sucrose and fluorescein isothiocyanate-dextran (FITC-dextran) (average molecular weight about 4400 and 71200) were used as indices of paracellular transport. The permeability coefficients of [14C] sucrose and FITC-dextran through BBCEC monolayers were about 3 and 30-40 fold lower than those through BAEC monolayers, respectively. Transcellular transport was examined by means of [3H] eucine and [3H] alanine uptake studies. The Km value (Michaelis constant) of the amino acid uptake was lower in BBCEC than in BAEC. BBCEC had higher alkaline phosphatase (ALP) and γ-glutamyl transpeptidase (γ-GTP) activity than BAEC when cultured in vitro. These results suggest that BBCEC are suitable for drug transport studies across the BBB in vitro.
The solubility of 6-mercaptopurine (6-MP) in water increased as the concentration of sodium benzoate or sodium hippurate in the solution increased. The solubility of 6-MP in 20% (w/v) sodium benzoate or sodium hippurate solution was about 6-fold larger than that of 6-MP alone. The stability constant of the soluble complex of 6-MP with sodium benzoate was estimated to be 2-8M-1 from (1) phase-solubility study and (2) analysis of chemical shifts observed in 1H-NMR. Partition of 6-MP from the saturated solution to n-octanol was also greatly increased by the addition of sodium benzoate or sodium hippurate, the degree being less in the latter. Administration of 6-MP with 20% (w/v) sodium benzoate to rat rectum resulted in enhanced absorption and the area under the plasma concentration-time curve was comparable to that obtained by intravenous administration (bioavailability=100%), while the bioavailability after intrarectal administration of 6-MP with 20% (w/v) sodium hippurate was only 9%. The reason for the difference was discussed.
The suitability of sampling via microdialysis for a lipophilic drug, valproate (VPA), was evaluated by the elimination rate constant of VPA solution in an in vitro experimental first-order elimination system. The elimination rate constant of VPA in dialysate was found to be 0.43±0.05 h-1, which was in good agreement with the real elimination rate constant (0.46±0.02 h-1). A change in VPA concentration in the solution surrounding a microdialysis probe was well maintained by the microdialysis method, suggesting no adsorption between the membrane of the microdialysis probe and VPA. On the basis of the in vitro experiment, the effect of a penetration enhancer, 1-[2-(decylthio) ethyl] azacyclopentan-2-one (HPE-101), on the transdermal absorption of VPA was examined in rats by the use of microdialysis in vivo. An intradermal microdialysis was performed at a flow rate of 1.0 μl/min for 7h after the dermal application of 50 mM VPA solution with or without 3% (w/v) HPE-101. HPE-101 increased the transdermal absorption rate of VPA by 80 times compared with the control. The microdialysis system was found to be quite useful for assessing the in vivo transdermal absorption of a lipophilic VPA.
The mechanism of gastrointestinal absorption of glycyrrhizin (GZ) was examined in rats. Using an in situ loop technique, it was found that the intestinal absorption of glycyrrhetic acid (GA), a major metabolite of GZ, was larger than that of GZ and that the absorption of GA was larger in the small intestine than in the large intestine. Although GZ was poorly absorbable, both GZ and GA were detected in rat plasma after oral administration of GZ, suggesting that GZ can be absorbed in both parent and metabolite forms, although their bioavailabilities were low. GZ was hydrolyzed to GA by rat gastric and large-intestinal contents, but not by the small-intestinal contents. This hydrolysis was diminished after boiling of the gastrointestinal contents and was not observed in the gastrointestinal contents of kanamycin-treated rats. These results suggest that GZ is hydrolyzed by bacteria in the stomach and large-intestinal contents and that most of the GA formed is absorbed from the large intestine. Since GZ was extensively excreted in bile after intravenous administration, the first-pass elimination might be the reason for its low bioavailability, in addition to the poor mucosal permeability.
We previously classified chloroflavone congeners into 3-methylcholanthrene (MC)-type, phenobarbital (PB)-type and mixed (MC plus PB)-type inducers of hepatic microsomal monooxygenases in rats. In the present study, we examined the structure-activity relationship involved in the capability of congeners to induce the described regioselective O-demethylation activity of scoparone. The steric and electronic parameters of congeners were calculated by the MM2 molecular mechanics and Extended Huckel MO methods, respectively. The molecular rectangle-area/depth ratios related well to the ratios of scoparone 6-/7-O-demethylation activities induced by the congeners. The molecular dimensions characterized the MC-type congeners as nearly planar molecules and the PB-type congeners as bulky and nonplanar. Moreover, the ratio of scoparone 6-/7-O-demethylation activities had significant correlation with both the LUMO energy (ELUMO) and the difference (ΔE) between ELUMO and HOMO energy for the congeners. The ELUMO and ΔE were less in the MC-type congeners than in the PB-type. These correlations suggest that the steric and electronic features of chloroflavone congeners are responsible for the induction of cytochrome P450 isozymes and associated monooxygenase activities.
The influence of aging on the reductase activity of acetohexamide, an oral antidiabetic drug with a ketone group, was examined in liver microsomes and cytosol of male rats. Acetohexamide reductase activities in liver microsomes of male rats at 26 and 31 months of age were much lower than that in liver microsomes of male rats at 9 weeks of age. Testectomy markedly decreased acetohexamide reductase activity in liver microsomes of the 9-week old rats and the decreased enzyme activity was significantly increased by testosterone administration. These results indicate, at least in part, that aging decreases the enzyme activity by decreasing the secretion of testosterone from the testes. On the other hand, aging (26 months of age) did not affect acetohexamide reductase activity in liver cytosol of male rats, although the enzyme activity at 31 months of age was slightly but significantly lower than that in liver cytosol of male rats at 9 weeks of age. Testectomy or testosterone administration had no effect on the enzyme activity in liver cytosol of 9-week old male rats.
The reactivity of malondialdehyde (MDA) with deoxyribonucleosides in the presence of acetaldehyde (AA) was investigated. Although MDA is known to be reactive towards deoxyguanosine (GdR), deoxyadenosine (AdR) and deoxycytidine (CdR) under acidic conditions, MDA had only slight reactivity towards GdR under physiological pH. However, when AA was present, MDA exhibited much higher reactivity towards GdR, and a reaction with AdR also took place.
Reticuloendothelial system (RES)-avoiding liposomes are known to accumulate in tumor tissues due to passive targeting. Dipalmitoylphosphatidylfluorouridine (DPPF), a potent antitumor agent readily incorporated into the lipid bilayer, was embedded in RES-avoiding liposomes modified with a uronic acid derivative, palmityl-D-glucuronide (PGlcUA). The therapeutic effect of DPPF in PGlcUA-liposomes was examined in tumor-bearing mice. Free or liposomal DPPF was injected intravenously into BALB/c mice bearing subcutaneously implanted Meth A sarcomas. The RES-avoiding liposomal formulation using PGlcUA was effective in reducing tumors, and prolonging survival time compared with free DPPF and also DPPF in conventional liposomes. Therefore, PGlcUA-liposomes might be of practical use as drug carriers for anticancer agents, especially their derivatives for embedding in liposomal membranes.
The inhibitory effects of 50% ethanolic extracts from dried leaves of 38 plants collected in the herbal garden of Kinki University were investigated in vitro on melanin biosynthesis which is closely related to hyperpigmentation. Of the 38 extracts, Prunus yedoensis, P. zippeliana, P. amygdalus, P. persica, P. armeniaca, Thea sinensis and Chaenomeles sinensis showed a potent inhibition of tyrosinase, the enzyme which converts 3-(3, 4-dihy-droxyphenyl) alanine (dopa) to dopachrome in the biosynthetic process. Furthermore, the extracts from the leaves of P. yedoensis and P. zippeliana among the Prunus plants used in this experiment inhibited the production of melanin from dopachrome by autoxidation. These inhibitory effects of P. zippeliana on melanin biosynthesis were observed in cultured B-16 mouse melanoma cells. These results suggest that the leaves of P. zippeliana inhibit melanin biosynthesis which is involved in hyperpigmentation and could be used as a whitening agent for the skin.
The in vitro skin permeability of 16 drugs with a wide range of lipophilicity (log P ; -0.95-4.40) was evaluated by the use of an ethanol/water (60/40) binary vehicle with of without lauric acid as a permeation enhancer. The enhancing effect by the addition of lauric acid to the ethanol/water (60/40) binary vehicle could be observed from the aspect of both permeation rate and lag time. The permeation rate increased with an increase in the hydrophilicity of the drugs. It was considered that lauric acid exerts an effect on both the polar and nonpolar regions of lipids of the stratum corneum, and the cooperative interaction of ethanol and lauric acid increases the participation of the polar pathway of drugs. The relationship between lipophilicity and skin permeability of the drugs from the ethanol/water (60/40) binary vehicle with lauric acid showed a parabolic shape, with its peak at a more hydrophilic range (log P ; 0.19) compared with other past references (log P ; 2-3). The ethanol/water (60/40) binary vehicle with lauric acid appears to be a good candidate as a vehicle for transdermal therapeutic systems for hydrophilic drugs.
To further clarify the pharmacokinetic characteristics of phenytoin (DPH) and its derivatives, DPH-1-methylnicotininate (MNDPH), valeroyl DPH (VADPH) and valproyl DPH (VPDPH), in plasma and brain, we have investigated their physicochemical properties and protein binding characteristics. Additionally, the hydrolytic conversion of these derivatives to DPH was also studied using small intestine, liver and brain tissues, as well as rat plasma. The log partition coefficient (PC) values of all derivatives were much higher than that of DPH. Judging from their pKa values (5.68 and 5.91 for VADPH and VPDPH, respectively) and pH-solubilities, VADPH and VPDPH were acidic compounds, while MNDPH was basic. These data indicated that most fractions of VADPH and VPDPH existed as an ionized form (these fractions existed in an ionized form, 0.98 and 0.97, respectively) at physiological pH, whereas MNDPH existed as a unionized form under the same conditions. Rosenthal or Scatchard plots of the binding data of DPH and its derivatives to both rat plasma protein and bovine serum albumin (BSA) exhibited straight lines over their concentration ranges used, indicating that DPH and its derivatives have a single binding site on the protein. The binding potencies (K or n·P1 value) of the derivatives to both proteins were much greater than that of DPH. No DPH produced from VADPH and VPDPH was found in the biological fluids over a period of 24 h. However, the hydrolysis of MNDPH to DPH was observed in plasma and the tissues used, with the most rapid hydrolysis in the small intestine, and the hydrolysis rate constant in plasma was ca. 20-fold greater than that in the brain. The present results lead us to propose that the low uptake of VADPH and VPDPH into the brain, as well as their rapid elimination from plasma is mainly ascribed to both the high protein binding and the large dissociation of derivatives in the plasma, compared with that of DPH.
Six herbimycin A (HBM) derivatives were examined for their anti-angiogenic effects in a bioassay system involving chorioallantoic membranes (CAMs) of growing chick embryos on the basis of our previous observation that HBM is a potent angiogenesis inhibitor. 17-Cyclopropylamino-HBM dose-dependently inhibited embryonic angiogenesis. The ID50 value was 0.1 μg (160 pmol) per egg and thereby lower than that of the parent compound HBM (ID50=0.15 μg (260 pmol) per egg). In contrast, 19-dimethylamino-, N-acetyl-, 2, 3, 4, 5-tetrahydro- and 7-decarbamoyl-HBM at doses of 0.01-10 μg/egg failed to affect angiogenesis in CAMs. These results strongly suggest as follows : (1) C-19 position, amino group between positions C-1 and C-20 and carbamoyl group in C-7 are essential for the anti-angiogenic action of HBM ; (2) HBM needs certain fixed conformation for expression of angiogenesis inhibition ; (3) it is expected that the modification of C-17 with a suitable functional group results in increased anti-angiogenic potency of HBM - - that is, a more potent angiogenesis inhibitor than the parent compound would be developed.
1-Acetamino-3-(1-naphthyloxy)-2-propanol which is an acetyl conjugate of N-desisopropylpropranolol (AcNDP) was detected as a new metabolite of propranolol (PL) from the incubation medium of isolated rat hepatocyte system and from the urinary extracts of a patient under PL therapy as well as a healthy volunteer given PL. In the case of the hepatocyte systems, the optical selectivity of PL elimination and the metabolite formation were discussed by HPLC determination, and the effect of pretreatment by phenobarbital (PB) or 3-metylcholanthrene (3-MC) on the metabolism was also clarified.