Cell surface carbohydrates modulate a variety of cellular functions, including recognition and adhesion. The HNK-1 carbohydrate epitope, which is recognized by the monoclonal antibody HNK-1, is specifically expressed on a series of cell adhesion molecules and also on some glycolipids in the nervous system over a wide range of species from insect to mammal. The HNK-1 epitope is spatially and temporally regulated during the development of the nervous system and associated with neural crest cell migration, neuron to glial cell adhesion, outgrowth of astrocytic processes and migration of cell body, as well as the preferential outgrowth of neurites from motor neurons. These observations together with the unusual chemical nature of the HNK-1 epitope, namely a non-reducing terminal 3-sulfoglucuronic acid residue, prompted us to study the biosynthesis of the HNK-1 epitope, in which a unique glucuronyltransferase (s) plays a key role. We found that the respective glucuronyltransferases are involved in the biosynthesis of the HNK-1 epitope on glycoproteins (GlcAT-P) and on glycolipids (GlcAT-L). Then, we isolated a novel glucuronyltransferase (GlcAT-P) specific for glycoprotein substrates and its cDNA from the rat brain. The primary structure deduced from the cDNA sequence predicted a type II transmenbrane protein with 347 amino acids. Transfection of the GlcAT-P cDNA into COS-1 cells induced not only expression of the HNK-1 epitope on the cell surface but also marked morphological changes of the cells, suggesting that the HNK-1 epitope was associated with the cell-substratum interaction. The GlcAT-P cDNA obtained in this study will be a useful molecular tool to open the way for further steps in the elucidation of the biological function of the HNK-1 carbohydrate epitope in the development of the nervous system.
In this study, a very reliable HPLC method was developed for the determination of fenofibric acid and reduced fenofibric acid in the biological samples described as follows. After addition of the internal standard solution and 0.5M HCl to the biological sample, fenofibric acid, reduced fenofibric acid and the internal standard were extracted with a mixed solvent of n-hexane and ethyl acetate (90 : 10) from the mixture. The acids were back-extracted from the organic phase with 0.1M Na2HPO4 and then re-extracted from the aqueous phase with a mixed solution of n-hexane and ethyl acetate (95 : 5) after addition of 0.5M HCl. The organic phase was evaporated to dryness under the vacuum. The residue was dissolved in MeOH and diluted with distilled water. An aliquot of the resulting solution was injected on to the HPLC. High reproducibility was observed in this HPLC method (C. V.% less than 4%). Moreover it was confirmed that the conjugates in the urine could be hydrolyzed by incubation at 37°C for 18h after addition of 400IU of β-glucuronidase.
We applied capillary electrophoresis (CE) to quantify and examine the content uniformity of ingredients of pharmaceutical formulations (diltiazem hydrochloride tablets and trimetoquinol hydrochloride tablets), and also studied separation of enantiomers through the addition of dextrin or cyclodextrin (CD) as a chiral selector. We evaluated these methods in the light of testing procedures and system suitability described in the USP forum. The fast assay and content uniformity of both tablets, including optical purity testing, were performed within 300s. From the results of the system suitability testing, content uniformity and assay, these CE methods were found to be useful as an alternative to the conventional HPLC method.
In order to characterize the pharmaceutical utility of the root of Fukuchiyama-jio, which was produced as the hybrid of Rehmannia glutinosa LIBOSCH. var. purpurea MAKINO (Akaya-jio in Japanese) and Rehmannia glutinosa LIBOSCH. forma hueichingensis HSIAO (Kaikei-jio in Japanese) (Scrophulariaceae), we have investigated the chemical constituents of the root of Fukuchiyama-jio in detail in comparison with those of Akaya-jio and Kaikei-jio. Chemical analysis as well as quantitative one by means of gas liquid chromatography (GLC) have shown that the root of Fukuchiyama-jio contains a larger amount of iridoid glucosides as compared with Akaya-jio and Kaikei-jio. During these chemical analyses, a new ionone glucoside named oxyrehmaionoside B (7) along with melasmoside (6) were isolated as the characteristic constituents of Fukuchiyama-jio and the structure of 7 has been determined.
The in vivo effect of Hypsizigus marmoreus on the plasma antioxidant status of tumor-bearing mice was examined. Female DBA/2 mice treated with subcutaneous injection of 3-methylcholanthrene were fed on a basal diet (CE-2) or CE-2 containing 5% fruit bodies of the mushroom for 76 weeks. Antioxidant activities (AOA) of mice with tumor were significantly higher than those of mice without tumor. The high levels of AOA were attributable to the increase of high molecular weight AOA in the plasma. A similar increase in plasma AOA was also observed in sarcoma-180 solid tumor-bearing mice. The mushroom feeding exhibited a potent antitumor effect and caused a significant decrease in lipid peroxide levels (as thiobarbituric acid reactive substances ; TBARS). These results suggest that the increase of AOA in the plasma of tumorbearing mice is one of the mechanism of cancer preventive effects and also suggest that Hypsizigus marmoreus might play a role in decreasing TBARS by controling AOA induction.