Shintaro Furusaki

Liquid membrane transport of amino acids by a calix[6]arene carboxylic acid derivative

Calix[6]arene hexacarboxylic acid was found to be a useful carrier for transporting amino acids through a liquid membrane. The calix[6]arene, which has a cyclic structure to include a guest molecule of the amino acid ester and bears six ionizable carboxylic acids to contribute electrostatic interaction, exhibited a high transport efficiency compared to its monomer analog and the other calix[n]arene derivatives. The novel carrier successfully transported hydrophobic amino acid esters from the feed phase to the receiving phase. The transport rate could be controlled by changing the pH gradient between the feed and receiving aqueous phases because the complexation proceeds by a proton-exchange mechanism. Furthermore, an optical resolution system was constructed by applying an enantioselective enzymatic reaction for a chiral separation of the amino acids. In the enzyme reaction, the l-form ester was selectively hydrolyzed to the free amino acid. The free amino acid hydrolyzed was not transported, while the unhydrolyzed d-form ester effectively moved to the receiving phase through the liquid membrane containing the calix[6]arene as a mobile carrier.

A molecularly imprinted polymer that shows enzymatic activity

Abstract An enzyme-mimic polymer was prepared by the surface molecular imprinting technique. In the active site of the polymer, a substrate analog was imprinted through the complex formation between a cobalt ion and alkyl imidazole that functions as the hydrolysis catalysis. The enzymatic performance of the imprinted polymer was evaluated by the hydrolysis reaction of an amino acid ester. Based on the Michaelis–Menten analysis, V max and K m were obtained. The imprinted polymer shows a high activity compared to that of the control unimprinted polymer. The result was supported by a low K m value of the imprinted polymer, indicating a high affinity to the target substrate. The enzyme-mimic polymer was found to possess a substrate specificity by using several substrates that have a different structure. A computational modeling also supports the structure of the imprinted sites formed on the polymer surface.

Enhanced inhibitory effects of extracts from Ginkgo biloba L. leaves encapsulated in hybrid liposomes on the growth of tumor cells in vitro

Abstract Inhibitory effects of extracts from Ginkgo biloba L. leaves with a variety of organic solvents and hot water on the growth of B-16 mouse melanoma (B-16 melanoma) and human lung adenocarcinoma (RERF-LC-OK) cells were examined. Although free extracts showed little inhibitory effects on the growth of tumor cells, extracts encapsulated in hybrid liposomes (HLs), which were composed of l -α-dimyristylphosphatidylcholine (DMPC) as a phospholipid and Tween 20 as a surfactant, exhibited inhibitory effects on the growth of B-16 melanoma cells. Significant inhibitory effects of petroleum ether extract (Extract “P”) encapsulated in HL on the growth of B-16 melanoma cells were observed. Further separation of Extract “P” using preparative layer chromatography (PLC) resulted in four fractions. Among them, one fraction had a remarkably high inhibitory effect (80%) on the growth of B-16 melanoma and RERF-LC-OK cells. These results suggest the presence of hydrophobic antitumor agents in G. biloba L. leaf.

Release of anthocyanin from strawberry cultured cells with heating treatment

Heating treatment as a permeabilization method for plant cells was examined on the release of anthocyanin pigments from strawberry cells. The release rate of anthocyanins followed an Arrhenius-type dependency on temperature. The death rate was observed to be peculiarly low at 45 °C, the temperature at which the heat shock protein was induced. Thus, the release of anthocyanin from viable cells depends on the functioning of the heat shock protein probably. The treatment with calcium after heating was observed to restore viability. An elevated rate of the release of anthocyanins, which were likely accumulated in vacuoles, was achieved after they had been gradually released for 45 min.

Chromatographic separation of amyloglucosidase from the mixtures of enzymes

Abstract An affinity chromatographic separation of amyloglucosidase (AMG) enzyme using β-cyclodextrin-chitosan matrix was investigated in this study. The adsorbent selectively adsorbed AMG from the mixtures of enzymes (lactase and α-amylase), and this selectively bound AMG was eluted from the column in two steps with a recovery yield of about 72%. A small fraction of AMG and other contaminating proteins did not bind to the column specifically and were eluted easily. Purified AMG was visualised on the sodiumdodecyl sulphate–polyacrylamide gel electrophoresis (SDS–PAGE) experiments.

Screening for transgenic plant cells that highly express a target gene from genetically mixed cells

To establish a strategy for stably and highly expressing a target gene in transformed plant cell suspensions, we developed the co-selection method that linked the hygromycin phosphotransferase gene (hph) to the β-glucuronidase (uidA, GUS) gene in the opposite direction under the same transcriptional regulation of the cauliflower mosaic virus (CaMV) 35S promoter. The linked genes were transferred into a tobacco BY-2 cell suspension, which was cultivated under various levels of the antibiotic pressure. Under the high pressure of hygromycin, GUS expression was increased and maintained over 1.5 years. We presented a successful example for selecting the plant cell suspension that highly expressed the target gene out of genetically heterogeneous cells.

Recovery of uranium from seawater by adsorption method

Uranium recovery from seawater was investigated in the sea by towing or mooring the adsorption bed. Amidoxime adsorbents showed promising results. The current flow and the wave motion were necessary to keep the effective adsorption. The cost of uranium recovered is estimated 50,000 yen/kg, which will cause the rise in the price of electricity by ca. 1 yen/kwh.