Masao Sugawara

Channel mimetic sensing membranes for alkali metal cations based on oriented monolayers of calixarene esters

Abstract As a fundamental study of the development of channel mimetic sensing membranes, guest-induced permeability changes were examined for oriented monolayers composed of 4-tert-butylcalix[6]arene- O , O ′, O ″, O ‴, O ″″, O ″‴-hexaacetic acid hexaethyl ester and the cone conformers of 4-tert-butyl- and 4-(1,1,4,4-tetramethylpentyl)calix[4]arene- O , O ′, O ″, O ‴-tetraacetic acid tetraethyl esters. The permeabilities through these monolayers were evaluated by cyclic voltammetry using cationic, anionic and neutral electroactive compounds as the permeability markers. The permeability was evaluated on the basis of both voltammogram area and limiting semi-integral current. The voltammetric measurements were carried out for either a monolayer formed at the air|water interface under an applied surface pressure, or a monolayer transferred to an electrode by the Langmuir-Blodgett method. Alkali metal cations induced an increase in the permeability for an anionic marker ion such as [Fe(CN) 6 ] 4− and a decrease in the permeability for cationic marker ions such as [Co(1,10-phenanthroline) 3 ] 2+ and [1,1′-dimethyl-4,4′-bipyridinium] 2+ (methylviologen 2+ ). These permeability changes can be ascribed to an attractive or repulsive electrostatic interaction between the marker ions and the membrane calixarene hosts bearing positive charges by complexation with alkali metal ions. A cationic guest-induced increase in the membrane permeability was also observed when a neutral marker (1,4-benzoquinone) was used, possibly because of an increase in the intermolecular voids (or microscopic defects) owing to conformational contraction upon host-guest complexation. Selectivity was observed for the calix[6]arene monolayer, with the magnitude of the response being in the order Cs + > Rb + > K + > Na + , Li + , which is consistent with the selectivities in solvent extraction or transport and in potentiometric response.

Biomembrane mimetic sensing chemistry

Abstract A comprehensive approach to ‘biomembrane mimetic sensors’ based on both artificial and biological systems is described. Of a number of biomembrane functions that are indicative of novel principles of chemical sensing, special focus is placed on (i) membrane potential change, (ii) membrane permeability change, and (iii) active transport of target compounds. The advantages of these biomembrane phenomena have been exploited in the development of several series of novel classes of chemical sensors and their signal transduction/amplification functions evaluated.

Ionophore incorporated bilayer lipid membranes that selectively respond to metal ions and induce membrane permeability changes

Abstract Bilayer lipid membranes (BLMs) embedded with ionophores are described that selectively induce membrane permeability changes upon binding analyte ions at the membrane/aqueous solution interfaces. A variety of ionophores for various metal ions, such as valinomycin, a lipophilic derivative of 2,2′-bipyridyl, ETH 4120 and dihexadecyl phosphate, were directly incorporated into planar BLMs formed by two techniques, i.e. the folding and tip-dip methods. The selective changes in transmembrane ion permeability were generally observed upon complexation of respective primary ions with membrane ionophores, which was monitored as a membrane conductance change. It was found that the primary ion-induced membrane conductance changes were due to permeation across the ionophore-incorporated BLMs of not only the primary ions but also their counter anions such as Cl - and ClO − 4 . The membrane conductance thus observed was discussed in terms of selectivity and the extent of permeability change per unit ionophore concentrations in the BLM.

Electrochemical evaluation of chemical selectivity of glutamate receptor ion channel proteins with a multi-channel sensor

A new method for evaluating chemical selectivity of agonists towards receptor ion channel proteins is proposed by using glutamate receptor (GluR) ion channel proteins and their agonists N-methyl-D-aspartic acid (NMDA), L-glutamate, and (2S, 3R, 4S) isomer of 2-(carboxycyclopropyl)glycine (L-CCG-IV). Integrated multi-channel currents, corresponding to the sum of total amount of ions passed through the multiple open channels, were used as a measure of agonists selectivity to recognize ion channel proteins and induce channel currents. GluRs isolated from rat synaptic plasma membranes were incorporated into planar bilayer lipid membranes (BLMs) formed by the folding method. The empirical factors that affect the selectivity were demonstrated: (i) the number of GluRs incorporated into BLMs varied from one membrane to another; (ii) each BLM contained different subtypes of GluRs (NMDA and/or non-NMDA subtypes); and (iii) the magnitude of multi-channel responses induced by L-glutamate at negative applied potentials was larger than at positive potentials, while those by NMDA and L-CCG-IV were linearly related to applied potentials. The chemical selectivity among NMDA, L-glutamate and L-CCG-IV for NMDA subtype of GluRs was determined with each single BLM in which only NMDA subtype of GluRs was designed to be active by inhibiting the non-NMDA subtypes using a specific antagonist DNQX. The order of selectivity among the relevant agonists for the NMDA receptor subtype was found to be L-CCG-IV > L-glutamate > NMDA, which is consistent with the order of binding affinity of these agonists towards the same NMDA subtypes. The potential use of this approach for evaluating chemical selectivity towards non-NMDA receptor subtypes of GluRs and other receptor ion channel proteins is discussed.

Cation permselectivity in the phase boundary of ionophore-incorporated solvent polymeric membranes as studied by Fourier transform infrared attenuated total reflection spectrometry

Abstract The behaviour of permselective membrane transport was studied by Fourier transform IR attenuated total refelction spectrometry and potential measurements for the solvent polymeric membranes incorporating several different ionophores. The stoichiometric ratios between the complexed cations and their corresponding counter anions in the phase boundary, 0.31–1.8 υm in depth, of the membrane were determined for each membrane made in contact with various primary ion solutions. When hydrophilic counter anions were used for the primary ion solutions, the IR spectra of the membrane were exclusively those of the complexed cations, and no IR-active anionic peak appeared. The intensity of the former peaks was found to be dependent on an added anionic site, carboxylated poly(vinyl chloride) and a derivative of tetraphenylborate, in the membranes, and also on the penetration depths of the IR beams used. In contrast, when a hydrophobic counter anion SCN − , was used for the primary ion solutions, the spectra from both the complexed cation and corresponding counter anion were seen, of which the stoichiometric ratios changed depending on the concetrations of primary ion salts, and on the coexistence of a derivative of tetraphenylborate in the membrane. The results are discussed in terms of the extent of cation permselectivity and its relevance to the potential of ionophore-incorporated liquid membrane ion-selective electrodes.

Receptor based chemical sensing

This mini-review describes our recent approach of mimicking transmembrane and intracellular signalings displayed by various receptors in biomembranes for the development of new sensing membranes. Several important modes of receptor signaling have been utilized exploiting bio and synthetic receptors; (i) Ca2+ signaling by calmodulin, (ii) active transport of target and relevant compounds displayed by Na+/D-glucose cotransporter and Na+,K(+)-ATPase, (iii) membrane permeability changes induced by glutamate receptor ion channel proteins and (iv) membrane potential changes induced by synthetic receptors. The newly designed sensing systems are demonstrated and discussed in terms of their novel mode of signal transduction, sensitivity and selectivity.

An enzyme-modified capillary as a platform for simultaneous fluorometric detection of d-glucose and l- lactate

&NA; The preparation of a glass capillary pattered with lipid layers on which lactate dehydrogenase (LDH) and glucose dehydrogenase (GDH) were regionally adsorbed and its application for simultaneous detection of D‐glucose and L‐lactate in human serum is described. A lipid layer was formed on the surface of BSA‐unabsorbed octadecyltrichlorosilane (OTS) inner wall of a glass capillary. The electrostatic charge of the lipid layer was a key factor for adsorbing the enzymes on the lipid layer. The fluorescence intensities were observed at each enzyme site in the presence of diaphorase (DIA), &bgr;‐nicotinamide‐adenine dinucleotide oxidized (NAD), resazurin, D‐glucose and L‐lactate. The fluorescence intensities at each enzyme site increased with an increase in the concentration of D‐glucose and L‐lactate=with the detection limits of 32 &mgr;M and 4.9 &mgr;M, respectively.

A Planar Bilayer Lipid Membrane Sensor Using a Miniaturized Auto-patch System

A bilayer lipid membrane sensor was constructed with a miniaturized auto-patch system. The performance of the patch system was optimized to obtain an analytically relevant signal. A biosensor based on an anti-BSA-antibody as a receptor and gramicidin as a signal transduction element was demonstrated. The sensor for BSA exhibited a detection limit of pg/mL level for BSA.

Fundamental studies on chemical sensing based on active transport.

能動及び上り坂輸送に基づく化学センシングの基礎研究を総説の形で著した.まず,能動輸送の基礎的特性について考察した.次に,能動輸送により物質の濃度を増幅するための原理である供給槽/受容槽体積効果の理論的考察を行い,輸送パラメーターと体積効果の関係について検討した.又,上り坂輸送現象を物質移動の新しいモードとして組み込んだセンサー,すなわち上り坂輸送膜センサーの特性を考察し,その増幅能と応答時間について評価した.更に,輸送効率を高めるためのエネルギー循環システムについて考察した.最後に,能動輸送バイオ素子としてNa+/D-グルコース共輸送タンパクを用いる新しいグルコースセンサーの可能性について考察した.