“Molecular science of fluctuations toward biological functions ” should be important for understanding the origin of life. Membrane-fluctuations of tumor cells are very different from those of normal cells. In general, the membranes of tumor cells are more fluid as compared with normal cells. We have produced hybrid liposomes (HL) which can be prepared by sonication of vesicular and micellar molecules in a buffer solution. The physical properties of HL such as size, shape, membrane fluidity, and the temperature of phase separation can be controlled by changing the constituents and compositional ratio. The uniform and stable structure of HL-n composed of dimyristoylphosphatidylcholine (DMPC) and polyoxyethylenedodecyl ether (C₁₂(EO)_n : n = 21~25) with a diameter of 80 nm was revealed. The remarkable inhibitory effects of HL on the growth of various tumor cells were attained in vitro. The induction of apoptosis by HL was obtained and the pathway was elucidated. Hybirid liposomes fused and accumulated in tumor cell membranes, and the apoptotic signal firstly pathed through mitochondria, caspase-9, and caspase-3, secondly through Fas, caspase-8, caspase-3 and then reached the nucleus. A good correlation between the membrane fluidity of HL and inhibitory effects of HL for tumor cells was obtained. Significantly chemotherapeutic effects were obtained using mice model of carcinoma after the treatment with HL without any drug in vivo. There were no abnormal findings on normal rats after administering HL on the basis of chronic toxicity tests. In clinical applications, prolonged survival was attained in patients with lymphoma after the intravenous injection of HL without any side effect after the approval of the bioethics committee. The membrane targeted chemotherapy with drug-free HL was established without any side effects for the first time.

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Molecular dynamics of the drug delivery to fluid lipid bilayer membranes is crucial as a primary stage of the bioactivity in the cell. Lipid bilayer membrane, as a platform of vital functions, is a dynamic structure where molecules are always fluctuating under physiological conditions. The mechanism of drug deliveries is related to the molecular dynamics in such soft, fluid membrane interface.
To gain insight into molecular mechanisms of drug deliveries in a noninvasive manner, we develop the method to monitor dynamic properties of drugs and lipid components in membranes without labeled nuclei, by applying multinuclear high-resolution solution NMR in combination with the pulsed-field-gradient (PFG) technique. PFG NMR spectroscopy is a versatile method to elucidate the molecular motion in a natural manner. We have quantified the diffusivity, the kinetics of membrane binding, and the bound fraction of the drug in situ by using large unilamellar vesicles of egg phosphatidylcholine as model cell membranes. The PFG method unveils the bound component after the preferential decay of the free component at the high field gradient, where the chemical shift difference between these components is not enough to distinguish from each other. The rate constants of the binding and dissociation of the drug are determined by using the solution of the Bloch equation with diffusion and exchange terms. The combination of 1D and PFG NMR serves to quantify the kinetics of membrane binding where the bound and the free components are unable to distinguish because of the rapid exchange on the NMR timescale. A small-sized 5-fluorouracil and fluorinated bisphenol A are used as a model drug.

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Dynamics in lipid bilayers such as two dimensional domain formation and lateral molecular transportation are crucial factors in cell membrane reactions. In this review several recent studies about the domain formations and molecular diffusion using supported planar lipid bilayers (SLBs) are introduced. SLBs exist at the interface between solid substrates and aqueous solutions, 1 ∼ 2 nm away from the substrate. The fluidity of the membrane remains while the membrane is still affected by the structure and the physical properties of the substrate surface. Two dimensional domain structures on the lateral order of several micrometers in SLB are affected by the atomic scale structures of the substrate. At the latter half of this review, the method of single molecule tracking free from the restriction of substrate materials is described. Lipid diffusions in SLBs on opaque silicon wafer and high refractive TiO₂ substrate were observed in situ and the diffusion coefficients were successfully obtained.

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Lipid dynamics plays important roles in the maintenance of homeostasis, and understanding and control of these lipid dynamics is a key challenge in biophysics and cell biology. Time-resolved small-angle neutron scattering (TR-SANS) is a powerful technique to determine the rates of intervesicular exchange and flip-flop of lipids in situ and real-time. Time-resolved fluorescence anisotropy determines the order parameter of membrane lipids. Combination of these time-resolved methods will clarify the relationship between static structure and dynamic property of lipid membranes. In this study, we applied these techniques to large (LUVs) and small unilamellar vesicles (SUVs) consisting of 1-palmitoyl-2-oleoylphosphatidylcholine (POPC) or 1,2-dimyristoylphosphatidylcholine (DMPC) to elucidate the effect of curvature on the structure and dynamics of lipid membranes.
Time-resolved fluorescence anisotropy revealed that the order parameter (S) of DPH-PC is larger in SUVs than in LUVs. Moreover, TMAP-DPH represented significantly large S value when the probe was located at outer leaflet of SUVs, suggesting the acyl chain ordering.
TR-SANS showed that the rates of the intervesicular exchange and flip-flop of DMPC are higher in SUVs than in LUVs. In addition, both the activation enthalpy and entropy of the intervesicular exchange and flip-flop of DMPC increased in SUVs. These results suggest that the increase in the curvature induces tighter packing and reduces the enthalpy and entropy of lipids in membranes, which facilitates the intervesicular exchange and flip-flop of lipids by making these processes entropically less unfavorable.

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We developed a coarse-grained (CG) molecular model, which is useful to investigate a self-assembling process of lipids and surfactants in an aqueous solution or a phase transition process at the molecular resolution. The CG model is designed to reproduce the interfacial properties, solvation free energy, as well as molecular distribution functions obtained from simulations with the all-atomic detail. The systematic CG modeling enabled us to investigate large molecular assemblies such as liposomes with chemical characterization of molecule, which is not accessible by previous phenomenological models. Several application studies given in this paper will elucidate the power of CG molecular dynamics simulation to shed light on molecular processes in lipid membranes happening on the scale of ‹ 100 nm.

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In recent years, the use of membrane for water processing has been increasing rapidly in the fields of potable water purification and water recovery from sewage.
In these applications, the amount of water processed at some plants exceeds 100,000 m³/day, where highly-durable PVDF membrane modules are widely used to minimize operation costs. Application of membrane started with the treatment of relatively clean waters. However, as the reliability of membrane increases, the demand for the ability to treat highly turbid water is increasing. Under these circumstances, Asahi Kasei Chemicals developed pressurized and submerged types of membrane modules and evaluated their filtration performance. This evaluation revealed that the submerged type is more suitable for treatment of highly turbid waters than the pressurized type. Submersed-type membrane modules are now actually used for water recovery from backwash wastewaters from pressurized-type membrane modules.

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