Various biological active substances were produced provided the appropriate stimulation was given to animal cells. Among them, human interferon is one of the substances most applicable to "drug." On the other hand, it is also important to develop interferon inducers (IFN-Ind.), a stimulater for producing interferon (IFN). This paper deals with a detailed description of the actual states of IFN and IFN-Ind. used as drugs, in which many pharmaceutical scientists are especially interested, and of a new effect obtained by the combined use of IFN and IFN-Ind. Moreover, our study to remove the hypreactivity which has been often detected in the inducer by the combined use of IFN was also described in detail. The clinical effect of IFN and its limitation at the present stage of the development was briefly discussed.
Twenty kinds of derivatives of 4-(2-dialkylaminoethyl)-3, 4-dihydro-2H-pyrido[3, 2-b]-1, 4-oxazine (13a-17d) were synthesized by two different methods, and obtained as hydrochloride. The following results were obtained by the use of the preliminary investigation on the local anesthetic and the analgesic activities (mouse writhing test) of these synthesized compounds. It was proved that the duration time of the local anesthetic action of almost all these compounds was inferior to that of cocaine or lidocaine, but, in quite few cases, superior to that of cocaine or lidocaine ; the analgesic action of these compounds was approximately equivalent to or slightly weaker than that of pyrabital.
Two glucosides were isolated from the whole plant of Cistanche salsa (C.A. MZY.) G. BECK and their structures were established as 8-hydroxygeraniol-1-β-D-glucopyranoside, a new monoterpene glucoside, and 8-epiloganic acid on the basis of chemical and spectral data.
A spin-labeled derivative of cortisol was prepared by coupling cortisol-3-(O-carboxylmethyl)oxime with 3-amino-2, 2, 5, 5-tetramethyl-1-oxypyrrolidine and used to develop a method for measuring free cortisol in the human urine. The samples were extracted by ethyl ether and measured by spin immunoassay (SIA) both with and without a preceding thin-layer chromatography (TLC). Although the values obtained from SIA without TLC purification were higher than those with TLC, the relationship between them was fairly good (r=0.980). Cortisol levels were determined in eleven normal urine samples simultaneously by SIA and radioimmunoassay without TLC purification. The coefficient of correlation was 0.988. The method was found to be speedy and simple, and to be useful for the routine use.
Conventionally, insulin was determined by using radioimmunoassay and radioisotope methods. We established the determination of insulin by high performance liquid chromatography, which is superior to another assay methods on simplicity, rapidity and accuracy, and identification limit of insulin is 0.5 ng. By using this method, we investigated the adsorption of insulin on glass surfaces. Adsorption of insulin was little independent upon pH and ionic strength. It was suggested that adsorption of insulin to glass surfaces was mainly hydrophobic interaction.
Reaction products of sugars with strongly basic anion-exchange resin (Dowex 1×2, OH- and BO2- forms) were analyzed by gas liquid chromatography-mass spectrometry (GC-MS) as trimethylsilyl derivatives and high-performance liquid chromatography (HPLC). When a suspension of Dowex 1×2 (OH- or BO2- form) in an aqueous solution of glucose was shaken mechanically, a part of glucopyranose was transformed into glucofuranose besides epimerizing to fructose and degrading to arabinose. The resin (BO2- form) was a more effective reagent for this transformation than the resin (OH- form). Although arabinonic, erythronic, glyceric, glycolic, and formic acids were detected in the reaction products of glucose or its epimers with the resin (OH- form) by HPLC and GC-MS, lactic acid was not detected. On the other hand, only a small amount of arabinonic and formic acids were found in the reaction products with the resin (BO2- form). A hypothetical mechanism of organic acids formation from glucose and its epimers was postulated.
The common tetracyclines (TCs) in fish tissues which are used in agriculture, tetracycline, oxytetracycline, and chlortetracycline, are analysed using an ODS-cartridge extraction device, and high performance thin layer chromatography (HPTLC) followed by densitometry. Fish tissue is blended in 0.1 M disodium ethylenediaminetetraacetate (di-Na EDTA)-McIlvaine buffer (pH 4.0) and centrifuged. The supernatant is adsorbed on the pretreated ODS-cartridge with 0.2 M di-Na EDTA aqueous solution, the cartrige is washed with water, and TCs are eluted with ethanol. The concentrated residue is spotted on a silica gel HPTLC plate predeveloped with saturated di-Na EDTA aqueous solution. The plate is then developed with chloroform-methanol-5% di-Na EDTA aqueous solution (65 : 20 : 5, lower layer). The amounts of TCs present on the developed plate are determined by ultraviolet absorption densitometry. Recoveries of TCs from fish samples fortified at level of 1 ppm are 60.2-78.6%.
The active site of an N-succinyl-L-trialanine p-nitroanilide-hydrolyzing protease purified from pronase E was investigated. When the enzyme was treated with [3H] diisopropyl phosphofluoridate, an active serine residue of the enzyme was modified, resulting in almost complete loss of the enzymatic activity. The amino acid sequence around the active serine residue (*) was Gly-Asp-Ser*-Gly-Gly. When a histidine residue in the enzyme molecule was destroyed by photooxidation in the presence of methylene blue, its activity decreased up to 15% of the original level. In this treatment, no change was observed either in tyrosine and methionine contents or in ultraviolet absorption spectra. The enzyme was also inactivated by glycine methyl ester in the presence of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide but not by p-bromophenacyl bromide. These findings suggest the possibility that serine, histidine, and aspartic or glutamic acid may be involved in the catalytic site of the enzyme to form the charge relay system.
1) Repeated administration of 5-hydroxy-L-tryptophan (5-HTP, 5, 10 or 50 mg/kg/d, 7d) did not produce any further increase in brain serotonin (5-HT) content in Wistar male rats as compared with that found in animals given a single dose of 5-HTP. On the other hand, 5-HT content in peripheral tissues (liver, kidney, spleen and small intestine) increased more after the repeated administration of 5-HTP than after a single dose of this compound. The accumulation of 5-HT in these tissues increased with an increase in the dosage and the number of administration times of 5-HTP. 2) Brain 5-HT content increased more and maintained a high level by the combined administration of DL-3-pyridylalanine (3-PA, 100 mg/kg) with 5-HTP (5 or 10 mg/kg) than by the single administration of these compounds ; the effect was additive. Liver, kidney, spleen and small intestine 5-HT contents after the combined administration of 3-PA and 5-HTP were not different from those after the single administration of 5-HTP. 3) The efficacy of combined administration of 3-PA and 5-HTP was further confirmed by the results of the determination of tryptophan (Trp)-5-HT metabolizing enzyme activities (Trp pyrrolase, Trp 5-hydroxylase, 5-HTP decarboxylase and monoamine oxidase) and pharmacological experiment (conditioned avoidance response) in rats given these compounds. 4) Data are discussed in relation to the side effect of 5-HTP for the treatment of depression.
To evaluate a possible utility of γ-cyclodextrin (γ-CyD) in an injecting agent, various pharmaceutical properties of inclusion complex such as dissolution, membrane permeation, chemical stability, protein binding, and biological fate were investigated using thiopental (TP) as a test drug. Inclusion complex formation of TP with γ-CyD in aqueous solution and in solid state was assesed by solubility method, spectroscopies, thermal analysis, and X-ray diffractometry. The 1 : 1 TP-γ-CyD complex exhibited a rapid dissolution in water, resulting in the net increase in the fraction of free TP available for membrane permeation. The TP in γ-CyD complex was significantly stable compared with sodium salt of TP during the accelerated storage conditions (40°C, RH 75%). The binding capacity of TP to rabbit serum albumin was found to be reduced by CyDs. Because of the small stability constant of TP-γ-CyD complex, no significant effect of γ-CyD was observed on the pharmacokinetic parameters of TP after the intravenous administration to rabbits. Many advantages obtained for TP-γ-CyD complex suggested great utility of γ-CyD in parenteral preparations without change in the biological fate of the drug.
The concentrations of o-ethoxybenzamide (EBA), N-(β-hydroxybutyryl)-p-phenetidine (HBP) and metabolites in the rabbit serum were investigated when EBA or HBP was administered orally with 2-bromo-isovalerylurea (BVU). 1) The concentration of EBA in the serum increased when 100 mg/kg of EBA was administered concurrently with 40 mg/kg of BVU as compared with the administration of 100 mg/kg of EBA alone. The con-administration of EBA and BVU also decreased the concentration of salicylamide (SAM) sulfate in the serum. 2) The concentration of HBP in the serum increased when 100 mg/kg of HBP and 40 mg/kg of BVU were administered together, while that of N-(β-hydroxybutyryl)-p-aminophenol (HBA) glucuronide decreased. 3) The concentrations of EBA and HBP in the serum increased when 100 mg/kg of EBA, 28.5 mg/kg of HBP and 45.5 mg/kg of BVU were administered together as compared with the co-administration of 100 mg/kg of EBA and 28.5 mg/kg of HBP, while those of SAM sulfate and HBA glucuronide decreased. 4) The deethylation of EBA and HBP was inhibited non-competitively by the addition of BVU in 9000×g supernatant fraction of the rabbit liver.
The metabolic rates of o-ethoxybenzamide (EBA) and N-(β-hydroxybutyryl)-p-phenetidine (HBP) were investigated when EBA or HBP was incubated with urea derivatives in 9000×g supernatant fraction of the rabbit liver. 1) The inhibitory effect of the deethylation on EBA and HBP was induced by urea as an agent capable of modifying the enzyme structure. Also urea derivatives increased the inhibitory effect as compared with urea. 2) Isovaleryl or isobutyryl substitution of urea increased inhibitory effect of deethylation, and the substituent of Br or allyl present in C2 of their molecules produced a large inhibitory effect. 3) The important factor of inhibition of deethylation was characterized in the presence of amide in their molecules, as shown by comparing 2-bromo-isovalerylurea and 2-bromo-isovalerylamide. 4) The cyclic ureides were recognized for the inhibition of deethylation, and secobarbital and phenobarbital produced an increase in inhibitory power. The substituent in C5 of barbiturates was an important factor for the inhibition of deethylation.
Absorption enhancing mechanism on propranolol by diltiazem was investigated in dog. No competition of plasma protein binding between two drugs was observed in equilibrium dialysis. When these drugs were intravenously administered in a combined injection, there was no alteration in the elimination behaviors. These results show that there occurs no substantial interaction between diltiazem and propranolol in the systemic circulation. In vitro metabolism study using liver homogenate revealed mutual inhibition of metabolism of propranolol and diltiazem, but the inhibitor constant (k1) was much higher than the plasma concentration. In vivo experiments showed that hepatic blood flow was increased substantially by intravenous and intraduodenal administration of diltiazem in anesthetized dogs. It seems that this pharmacological action causes reduction of the first pass metabolism of propranolol in the liver. Similar effects to enhance the absorption of propranolol observed by coadministration of vasodilators other than diltiazem, support the above idea. Relationship between absolute availability of propranolol in individual dogs and promoting effect of diltiazem on combined administration was significant. On the basis of these findings, it is considered that diltiazem enhanced the systemic availability of propranolol mainly by increasing hepatic blood flow.
Alterations in bioavailability of several drugs other than propranolol after the coadministration of diltiazem were investigated in dog and human. Diltiazem did not promote the bioavailability of dipyridamole and furosemide, but the plasma levels of diazepam and its metabolite, desmethyl-diazepam were remarkably increased by the coadministration of diltiazem in dog. On diltiazem-diazepam interaction in human, there was no significant observation. This species difference may be explained by the difference in metabolizing activity for diazepam in dog and human. Several other drugs were tested in dog and it was found that relationship between absolute availability of drugs and the promoting effect of diltiazem was significant. Namely, diltiazem enhanced the absorption of drugs with absolute availability less than 20%, indicating that underlying mechanisms were related to their first pass metabolism. It is considered that such a diltiazem interaction would be advantageous to reduce inter-individual variation in pharmacological effect and also to reduce doses of coadministrated drugs with high liver clearance.
Acute renal failure (R.F.) rabbits induced by the muscular injection (i.m.) of HgCl2 (10 mg/kg) produced a significant increase of blood urea nitrogen concentration (BUN) and plasma creatinine concentration (Cre.) after a few days. However, no significant change was observed for free fatty acid (FFA) level. The pharmacokinetics of furosemide in acute renal failure rabbit was also studied. The apparent volume of tissue compartment (V2) slightly increased. The elimination rate constant (k10) from central compartment significantly decreased, and mutually correlated with BUN or Cre. It was found from results of in vitro equilibrium dialysis measurement that the protein binding of furosemide (FR) significantly decreased in the plasma obtained from R.F. rabbits as well as that from patients with acute renal failure. Calculations made according to Sandberg-Rosenthal's formula revealed that the maximum binding concentrations (nP) was not changed but binding constant (K) decreased in the R.F. rabbit. The presence of endogenous substances that inhibit the plasma protein-FR binding competitively may be suggested from the above result.
The 4-substituted 5-amino-1-phenyl-1H-1, 2, 3-triazoles, in which 4-substituent is formyl (1a), hydroxyiminomethyl (1b), and acetoxyiminomethyl groups (1c), were thermally isomerized to the corresponding 4-substituted 5-anilino-1H-1, 2, 3-triazoles (2a-c) by heating at 90°C in dimethyl sulfoxide. On the basis of the results obtained from the reaction in several kinds of solvents, it appears that this isomerization depends on the nature of the solvent used and the reaction temperature. In acetylation of 1a-c with acetic anhydride a similar isomerization proceeded concurrently, and the corresponding 4-substituted 1-acetyl-5-anilino-1H-1, 2, 3-triazoles (2d-f) were formed. The isomerization mechanism is discussed.