A molecular mechanism for how pressure induces interdigitation of phospholipid bilayer membranes

The phase transition from the bilayer gel (ripple gel) phase to the nonbilayer gel (interdigitated gel) phase of bilayers of phosphatidylcholines (PCs) with two saturated long-chain fatty acids under high pressure was investigated by pressure-scanning microscopy, fluorometry, and dynamic light scattering (DLS) measurements. Microscopic observations for giant vesicles (GVs) of distearoyl-PC (DSPC) under high pressure showed that spherical GVs transforms into warped and distorted spherical ones instantaneously at the interdigitation. The fluorescence intensities of amphiphilic probe Prodan and hydrophobic probe Laurdan in the dipalmitoyl-PC (DPPC) bilayer steeply decreased and increased, respectively, at the interdigitation, suggesting that the conformational change of the polar head group of DPPC in the bilayer transiently occurred at the interdigitation. Further, the high-pressure DLS measurements revealed that the size of the vesicle particles of DPPC and DSPC transiently increased near the interdigitation pressure, whereas the ethanol-induced interdigitation of the DSPC bilayer under atmospheric pressure produced no such change in the particle size. Taking account of the critical packing parameter of the PC molecule, we could conclude that the pressure-induced interdigitation is attributable to the increase in repulsive interaction between the polar head groups of the PC molecules resulting from the orientational change of the head group from a parallel alignment to a perpendicular one with respect to the bilayer surface with pressure, namely the transient state: it occurs when the repulsive interaction exceeds a threshold value for the balance between the repulsive interaction and the attractive interaction among the hydrophobic acyl chains.

Biochimica et Biophysica Acta (BBA) - Biomembranes


Biocompatibility of zwitterionic polymer-modified surface under acidic condition

In this study, properties such as hydrophilicity and biocompatibility of surfaces prepared with 2-methacryloyloxyethyl phosphorylcholine (MPC) were evaluated under neutral to acidic conditions. The MPC-modified surface exhibit high hydrophilicity in neutral condition due to the ionization effect, while under acidic conditions below pH 3, the hydrophilicity gradually decreased due to the decrease in the negative charge of the PC groups. Biocompatibility property was evaluated by measuring the adsorption prevention effect against fibrinogen. The adsorption of fibrinogen was hardly observed on scanning electronic microscope (SEM) pictures under pH 7.6, which was significantly lower than that of the bare glass. However, the adsorption ratio increased under acidic conditions of pH 2.8 indicating a decrease in biocompatibility.


Phase Imaging of Phosphatidylcholine Bilayer Membranes by Prodan Fluorescence

Prodan (6-propiponyl-2-(N,N-dimethylamino)naphthalene) is well known as a polarity-sensitive fluorescent probe and has a high capability of detecting structural changes occurring within phospholipid bilayer membranes. In this study, we carried out the fluorescence spectroscopic observation of bilayer phase behavior for a series of symmetric saturated diacylphosphatidylcholines (CnPCs) with different acyl-chain length n (n = 12 15 and 19 22) using Prodan as a membrane probe to confirm the availability of Prodan along with the previous results for the CnPC bilayer membranes (n = 16 18). The results were discussed by constructing spectral three-dimensional (3D) imaging plots for visualizing the change in bilayer phase states with temperature or pressure to verify the functionality of this 3D imaging plot. It was found that the Prodan fluorescence technique is applicable to the detection of the changes in the bilayer phase states of all CnPCs with a few exceptions. One of the most crucial exceptions was that Prodan cannot be used for the detection of the bilayer-gel state of the C21PC bilayer membrane. It was also found that it is only to the CnPC bilayer membranes with n = 15 18 that the 3D imaging plot is adequately and accurately applicable as a useful graphical tool for visually detecting the bilayer phase states. This is a disadvantageous feature of this technique brought about by the high sensitivity of Prodan as a membrane probe. Therefore, further detailed studies on the molecular behavior of Prodan will enable us to find a more useful way of utilizing this membrane probe.


Temperature- and Pressure-Induced Bilayer Phase Transitions of an Amide-Linked Phosphatidylcholine: A Contrasting Effect of Chain Linkage Type

An unnatural phospholipid with two amide linkages, dipalmitoylamidodeoxyphosphatidylcholine (DPADPC), was synthesized and its bilayer phase transitions were observed by differential scanning calorimetry under atmospheric pressure and light-transmittance measurements under high pressure. The DPADPC bilayer membrane showed different phase transitions depending on the thermal history of the lipid sample under atmospheric pressure. The thermal-history dependent transitions were also observed under high pressure. The temperature pressure phase diagram and thermodynamic quantities of the phase transitions of the DPADPC bilayer membrane were compared with those of bilayer membranes of hydrophobic chain-linkage isomers, ester-liked dipalmitoylphosphatidylcholine (DPPC) and ether-linked dihexadecylphosphatidylcholine (DHPC). The thermodynamic quantities of the main transition for these PC bilayer membranes were similar to each other, whereas the suppression of the interdigitation and great stabilization of the hydrated crystal phase were observed for the DPADPC bilayer membrane. Furthermore, the results of nuclear magnetic resonance measurements indicated that the motility of PC molecules in both gel and liquid crystal phases of the DPADPC bilayer membranes were much lower than those of the DPPC and DHPC bilayer membranes. From the above results, we concluded that the molecular interaction in the PC bilayer membranes increases in the order of ether-, ester- and amide-linkage isomers.

Bulletin of the Chemical Society of Japan


Membrane fusion of phospholipid bilayers under high pressure: Spherical and irreversible growth of giant vesicles

Membrane fusion of giant vesicles (GVs) for binary bilayers of unsaturated phospholipids, dioleoylphosphatidylethanolamine (DOPE) having an ability to promote membrane fusion and its homolog dioleoylphosphatidylcholine (DOPC) having an ability to form GV, was investigated under atmospheric and high pressure. While DOPC formed GVs in the presence of inorganic salts with a multivalent metal ion under atmospheric pressure, an equimolar mixture of DOPE and DOPC formed GVs both in the absence and the presence of LaCl3. We examined the change in size and shape of the GVs of this binary mixture in the absence and presence of LaCl3 as a function of time under atmospheric and high pressure. The size and shape of the GVs in the absence of LaCl3 under atmospheric and high pressure and those in the presence of LaCl3 under atmospheric pressure hardly changed with time. By contrast, the GV in the presence of LaCl3 under high pressure gradually changed in the size and shape with time on a time scale of several hours. Namely, the GV became larger than the original GV due to accelerated membrane fusion and its shape became more spherical. This pressure-induced membrane fusion was completely irreversible, and the growth rate was correlated with the applied pressure. The reason for the GV growth by applying pressure was considered on the basis of thermodynamic phase diagrams. We concluded that the growth is attributable to a closer packing of lipid molecules in the bilayer resulting from their preference of smaller volumes under high pressure. Furthermore, the molecular mechanism of the pressure-induced membrane fusion was explored by observing the fusion of two GVs with almost the same size. From their morphological changes, we revealed that the fusion is caused by the actions of Laplace and osmotic pressure.


Subgel-phase formation of membranes of ether-linked phosphatidylcholines

The formation of subgel (so-called hydrated crystal) phase of membranes of ether-linked phospholipids, dialkylphosphatidylcholines containing linear saturated alkyl chain (Cn = 14, 16 and 18), was examined under atmospheric and high pressure. The results of differential scanning calorimetry in 50 wt% aqueous ethylene glycol solution and water showed that these PC membranes undergo the subtransition from the subgel phase to the gel phase at a low temperature with or without the thermal pretreatment of lipid samples called annealing. The subtransition in water could be clearly observed by light-transmittance measurements under high pressure and the transition temperature increased by applying pressure. The temperature-pressure phase diagrams and the thermodynamic quantities of subtransition were obtained from the phase-transition data and compared with those of membranes of ester-linked phospholipids, diacylphosphatidylcholines. The phase diagrams indicated that all gel phases of the ether-linked PC membranes exist as stable phases while parts of the gel phases of the ester-linked PC membranes are metastable. The subtransition temperatures of the ether-linked PC membranes were lower than those of the ester-linked PC membranes by more than 10 °C and the corresponding thermodynamic quantities were extremely small. Further, it was revealed by high-pressure fluorometry that the difference in subgel phase between ether- and ester-linked PC membranes results from their phase structures: the nonbilayer interdigitated structure is maintained after the conversion from the gel phase to the subgel phase in the ether-linked PC membranes whereas the ester-linked PC membranes form bilayer subgel phase with staggered structure.


Formation of intermediate gel-liquid crystalline phase on medium-chain phosphatidylcholine bilayers: Phase transitions depending on the bilayer packing

The bilayer phase transitions of medium-chain phosphatidylcholines with linear saturated acyl chains (Cn = 12, 13 and 14) were measured by high-pressure light-transmittance measurements and differential scanning calorimetry to investigate the formation of intermediate gel-liquid crystalline phase called Lx phase. The constructed phase diagrams showed that there existed a distinct region of the Lx phase between ripple gel (Pβ') and liquid crystalline (Lα) phases for multilamellar vesicle bilayers of C12PC and C13PC. The Lx phase of the C12PC bilayer was metastable at all pressures and disappeared at a higher pressure. In the C13PC bilayer, the Lx phase was stable and also disappeared at a higher pressure but its region markedly shrunk. By contrast, the Lx phase was not detected for the C14PC bilayer. Effects of other factors such as vesicle size and solvent substitution on the Lx phase of the C13PC bilayer were also examined. A decrease in vesicle size and solvent substitution from water to 50 wt% ethylene glycol solution promoted the Lx-phase formation as opposed to the effects of acyl-chain elongation and pressurization. The fluorescence data of the C13PC bilayer with different vesicle sizes showed that the Lx phase is caused by the difference of local packing in the bilayer. Considering these facts, we concluded that the Lx phase is an intermediate gel-Lα phase that has gel-phase monolayers with negative curvature and Lα-phase monolayers with positive curvature. The formation mechanism of the Lx-phase in stacked bilayers and dispersed vesicles is also explainable by this difference in packing state.


Salt effect on bilayer phase transitions of dipalmitoylphosphatidylglycerol in saline water under high pressure

The phase transitions of dipalmitoylphosphatidylglycerol (DPPG) bilayer membrane at high NaCl concentrations under high pressure were investigated to construct the temperature pressure phase diagram and determine phase-transition properties. The constructed phase diagrams exhibited the qualitative resemblance to that of the dipalmitoylphosphatidylcholine (DPPC) bilayer membrane: they underwent the gel-phase polymorphism including the pressure-induced bilayer interdigitation. The phase-transition properties of the DPPG bilayer membrane changed in a salt concentration-dependent manner, whereas qualitatively the phase behavior is not affected appreciably by the salt concentration measured. We discussed the salt effect on the DPPG bilayer membrane from the variation in interactions between the polar head group of the PG molecule.


Phase Behavior of Binary Bilayer Membrane of Dipalmitoylphosphatidylcholine and Stigmasterol

We investigated the phase behavior of a binary bilayer membrane of dipalmitoylphosphatidylcholine (DPPC) and stigmasterol (Stig) as a function of the Stig concentration (XSt) by means of differential scanning calorimetry (DSC) and fluorescence spectroscopy using 6-propionyl-2-(N,N-dimethylamino)naphthalene (Prodan) as a membrane probe. The DSC results revealed that the pretransition and the main transition are abolished at XSt = ca. 0.08 and ca. 0.50, respectively. The main transition enthalpy DeltaHm of the DPPC Stig binary bilayer decreased with XSt in a non-linear manner, which is contrast to the almost linear decrease in DeltaHm for the DPPC cholesterol (Chol) binary bilayer. This suggests that Stig has a stronger effect of disturbing the chain packing in a gel state and a weaker effect of suppressing the chain melting as compared to Chol. The XSt dependence of the peak width at half height (DeltaT1/2) suggests that a drastic change in the phase behavior occurs at XSt = ca. 0.14, which is similar to the DPPC Chol binary bilayer. The Prodan fluorescence spectra changed depending on the phase state, which was characterized by the wavelength at the emission maximum and allowed us to determine the transition temperatures of the binary bilayers. The compositional phase diagram of the DPPC Stig binary bilayer was found to be similar to that of the DPPC Chol binary bilayer. The phase behavior of the DPPC Stig binary bilayer could be well explained by applying the so-called superlattice view to interpret the phase diagram.

リン脂質ジパルミトイルホスファチジルコリン-スティグマステロール混合二分子膜の熱的相挙動を示差走査熱量測定と蛍光プローブ測定から調査した．本系に対して得られた温度-濃度相図はジパルミトイルホスファチジルコリン-コレステロール混合二分子膜のものと類似していたが，スティグマステロールはコレステロールに比べて，ゲル相では脂質アシル鎖に大きな撹乱効果を示す一方で，鎖融解の抑制効果は弱いことが明らかとなった．

Journal of Thermal Analysis and Calorimetry


Phase Behavior of Cholesterol-Containing Binary Membrane of an Ether-Linked Phospholipid, Dihexadecylphosphatidylcholine

The thermotropic phase behavior of the cholesterol (Chol)-containing binary membrane of dihexadecylphosphatidylcholine (DHPC) was investigated as a function of the mole fraction of Chol Xch by DSC and fluoresce spectroscopy using 6-propionyl-2-(N,N-dimethylamino)naphthalene (Prodan). The main-transition enthalpy Hm was almost constant at Xch < ca. 0.05, but it decreased almost linearly with increasing Xch above 0.05 until it reached 0 at Xch = ca. 0.50, suggesting that the interaction between DHPC and Chol is relatively weaker than that between dipalmitoylphosphatidylcholine (DPPC) and Chol. The Prodan fluorescence spectra changed depending on the phase state, which was characterized by the wavelength at the emission maximum. Particularly, the two-phase state of the interdigitated (LβI) and non-interdigitated gel (Lβ) was sensitively detected as the superposition of the two elementary spectra corresponding to each phase state, allowing us to explain the Xch dependent non-linear behavior of the Chol-induced Lβ phase by introducing a proportionality factor ξ(Xch) and by assuming a simple probabilistic process. By comparing the phase diagram constructed for the DHPC Chol binary membrane with that for the DPPC Chol binary bilayer, a difference was observed in the minimum Xch value for the entire formation of the liquid ordered (Lo) phase. This difference could be explained in terms of the hydrophobic mismatch at the bilayer midplane by taking into account the difference in the capability of hydrogen bonding with the 3β-OH group of Chol due to the presence or absence of the carbonyl oxygens at the glycerol backbone.

エーテル結合型リン脂質ジヘキサデシルホスファチジルコリン-コレステロール混合二分子膜の熱的相挙動を示差走査熱量測定と蛍光プローブ測定から調査した．コレステロール添加は指組み構造ゲル相の形成を抑制した．エステル結合型脂質ジパルミトイルホスファチジルコリン-コレステロール混合二分子膜系の相図との比較から，脂質頭部のカルボニル炭素の有無が二分子膜中における両分子の混和性に大きな影響を与えることが明確となった．


Ligand partitioning into lipid bilayer membranes under high pressure: implication of variation in phase-transition temperatures

The variation in phase-transition temperatures of dipalmitoylphosphatidylcholine (DPPC) bilayer membrane by adding two membrane-active ligands, a long-chain fatty acid (palmitic acid (PA)) and an inhalation anesthetic (halothane (HAL)), was investigated by light-transmittance measurements and fluorometry. By assuming the thermodynamic colligative property for the bilayer membrane at low ligand concentrations, the partitioning behavior of these ligands into the DPPC bilayer membrane was considered. It was proved from the differential partition coefficients between two phases that PA has strong affinity with the gel (lamellar gel) phase in a micro-molal concentration range and makes the bilayer membrane more ordered while HAL has strong affinity with the liquid crystalline phase in a milli-molal concentration range and does the bilayer membrane more disordered. The transfer volumes of both ligands from the aqueous solution to each phase of the DPPC bilayer membrane showed that the preferential partitioning of the PA molecule into the gel (lamellar gel) produces about 20% decrease in transfer volume as compared with the liquid crystalline phase, whereas that of the HAL molecule into the liquid crystalline phase does about twice increase in transfer volume as compared with the gel (ripple gel) phase. Furthermore, changes in thermotropic and barotropic phase behavior of the DPPC bilayer membrane by adding the ligand was discussed from the viewpoint of the ligand partitioning. Reflecting the contrastive partitioning of PA and HAL into the pressure-induced interdigitated gel phase among the gel phases, it was revealed that PA suppresses the formation of the interdigitated gel phase under high pressure while HAL promotes it. These results clearly indicate that each phase of the DPPC bilayer membrane has a potential to recognize various ligand molecules.

ジパルミトイルホスファチジルコリン(DPPC)二分子膜の相転移温度への2種類の膜活性リガンド(麻酔薬ハロセン，脂肪酸パルミチン酸)の効果を調べた．DPPC二分子膜の前転移および主転移は，ハロセン添加により降下，パルミチン酸添加により逆に上昇し，加圧は相転移を温度を共に上昇させた．熱力学的解析からハロセンは液晶相，パルミチン酸はラメラゲル相に親和性を有し，ハロセン添加により液晶相が膨張し，パルミチン酸添加によりゲル相が収縮することがわかった．また，ハロセンは高圧力下における指組みゲル相形成を促進し，パルミチン酸は逆に抑制した．指組みゲル相を含む相転移挙動へのリガンド効果についての議論が熱力学観点からなされた．


Effect of pressure on bilayer phase behavior of N-methylated di-O-hexadecylphosphatidylethanolamines: relevance of head-group modification on the bilayer interdigitation

The phase transitions of N-methylated di-O-hexadecylphosphatidylethanolamines (DHPE, DH-N-methyl-PE (DHMePE) and DH-N,N-dimethyl-PE (DHMe2PE)) were observed by differential scanning calorimetry (DSC) and fluorometry under atmospheric pressure and light-transmittance measurements under high pressure. The DSC thermograms showed that the N-methylated DHPE bilayers underwent the phase transitions from the gel phase to the liquid crystalline (Lα) phase under atmospheric pressure. The gel phase was identified by fluorometry as the lamellar gel (Lβ) phase, and not interdigitated gel (LβI) phase. The gel/Lα transition temperature increased with pressure while decreased stepwise with increasing polar head-group size. This stepwise depression in the transition temperature may be caused by the inverse-proportional hydrogen-bonding capabilities of the head-group to the head-group size. The thermodynamic quantities of the gel/Lα transition were comparable for the N-methylated DHPE bilayers. The pressure-induced LβI phase was not found in these bilayers although the bilayer of di-O-hexadecylphosphatidylcholine (DHPC), which is a kind of N-methylated DHPEs, forms the LβI phase only by hydration under atmospheric pressure. Taking into account that the bilayers of diacyl-homologs of N-methylated DHPEs, N-methylated dipalmitoyl-PEs except for dipalmitoylphosphatidylcholine (DPPC), do not form the LβI phase in the whole pressure range investigated but the DPPC bilayer forms the LβI phase under high pressure, we can say that the interdigitation requires a weaker interaction between large-sized head groups like the bulky choline group.

極性基サイズの異なる3種類のエーテル型リン脂質N-メチル化ジヘキサデシルホスファチジルエタノールアミンが形成する二分子膜の高圧相挙動を調査した．極性基サイズの増加と共に相転移温度は段階的に降下したが，相転移熱力学量は極性基サイズにはほとんど依存しなかった．指組み構造ゲル相は，これら脂質二分子膜では常圧，高圧下共に観測されなかった．エステル型リン脂質N-メチル化ジパルミトイルホスファチジルエタノールアミン二分子膜系との結果の比較から，指組み構造形成にはコリン基のような嵩高い極性基が必須であることが明らかとなった．


Thermotropic and barotropic phase transitions on diacylphosphatidylethanolamine bilayer membranes

The bilayer phase transitions of four diacylphosphatidylethanolamines (PEs) with identical two saturated acyl chains (Cn = 12, 14, 16 and 18) and two PEs with identical two unsaturated acyl chains containing a different kind of double bonds were observed by differential scanning calorimetry under atmospheric pressure and light-transmittance measurements under high pressure. The temperature-pressure phase diagrams for these PE bilayer membranes were constructed from the obtained phase-transition data. The saturated PE bilayer membranes underwent two different phase transitions related to the liquid crystalline (Lα) phase, the transition from the hydrated crystalline (Lc) phase and the chain melting (main) transition, depending on the thermal history. Pressure induced the change in the gel-phase stability of the bilayer membranes of PEs with longer chains at a low pressure. Comparing the thermodynamic quantities of the saturated PE bilayer membranes with those of diacylphosphatidylcholines (PC) bilayer membranes, the PE bilayer membranes showed higher phase-transition temperatures and formed more stable Lc phase, which originates from the strong interaction between polar head groups of PE molecules. On the other hand, the unsaturated PE bilayer membranes underwent the transition from the Lα phase to the inverted hexagonal (HII) phase at a high temperature and this transition showed small thermodynamic quantities but high-pressure responsivity. It turned out that the kind of double bonds markedly affects both bilayer-bilayer and bilayer-nonbilayer transitions and the Lα/HII transition is a volume driven transition for the reconstruction of molecular packing. Further, the phase-transition behavior was explained by chemical potential profiles of bilayer phases.

4種類の飽和ホスファチジルエタノールアミン(PE)および2種類の不飽和PEが形成する二分子膜の温度および圧力誘起相転移を調査した．飽和PE二分子膜は熱履歴に依存して水和結晶相/液晶相転移およびゲル相/液晶相(主)転移を示し，長鎖飽和PE二分子膜においては高圧力下でゲル相の安定性が変化した．熱力学量から，飽和PE二分子膜は同鎖長のホスファチジルコリン(PC)二分子膜よりも顕著に安定な水和結晶相を形成することがわかった．他方，不飽和PE二分子膜は，液晶相から逆ヘキサゴナル相への二分子膜/非二分子膜転移を誘起した．熱力学的な考察から，この転移は体積駆動の転移であることを示した．

Biochimica et Biophysica Acta (BBA) - Biomembranes


Inactivation of foodborne pathogenic and spoilage microorganisms using ultraviolet-A light in combination with ferulic acid

The low energy of UV-A (315 400 nm) is insufficient for disinfection. To improve UV-A disinfection technology, the effect of ferulic acid (FA) addition on inactivation by UV-A light-emitting diode (LED) light (350 385 nm) was evaluated in the eliminating of suspensions of various food spoilers and pathogens (seven bacteria and four fungi). Photoantimicrobial assays were performed at FA concentrations below the MIC. The MIC of the isomerized FA, consisting of 93% cis-form and 7% trans-form, was very similar to that of the commercially available FA (trans-form). Irradiation with UV-A (1.0 J cm-2) in the presence of 100 mg l-1 FA resulted in enhanced reducing of all of the tested bacterial strains. A combination of UV-A (10 J cm-2) and 1000 mg l-1 FA resulted in enhanced reducing of Saccharomyces cerevisiae and one of the tested filamentous fungi. These results demonstrated that the combination of a short-term application of UV-A and FA at a low concentration yielded synergistic enhancement of antimicrobial activity, especially against bacteria.


Comprehensive characterization of temperature- and pressure-induced bilayer phase transitions for saturated phosphatidylcholines containing longer chain homologs

Complete elucidation of the phase behavior of phospholipid bilayers requires information on the subtransition from the lamellar crystal (Lc) phase to the gel phase. However, for bilayers of saturated diacylphosphatidylcholines (CnPCs), especially longer chain homologs, equilibration in the Lc phase is known to be very slow. In this study, bilayer phase transitions of three CnPCs with longer acyl chains, C19PC, C20PC and C21PC, were observed by differential scanning calorimetry under atmospheric pressure and by light-transmittance measurements under high pressure. Using lipid samples treated by thermal annealing enabled the observation of the sub-, pre- and main transitions of the C19PC and C20PC bilayers under atmospheric pressure. Only the pre- and main transitions could be observed for the C21PC bilayer due to very slow kinetics of the Lc phase formation for lipids with long acyl chains. The temperature and pressure phase diagrams constructed and phase-transitions quantities (enthalpy, entropy and volume changes) evaluated for these bilayers were compared with one another and with those of bilayers of the CnPC homologs examined in previous studies. These results allowed us (1) to clarify the temperature- and pressure-dependent phase sequence and phase stability of the CnPC (n = 12 22) bilayers as a function of the hydrophobicity of the molecules, (2) to prove the presence of a shorter and a longer limit (n = 13 and 21) in the acyl-chain length for the pressure-induced bilayer interdigitation and (3) to reveal the chain-length dependence of the thermodynamic quantities of the subtransitions including the volume change.

疎水鎖長が異なる長鎖飽和ジアシルホスファチジルコリン(CnPC: n = 19 - 21)の温度および圧力誘起二分子膜相転移を，特に水和結晶(Lc)相に着目して調査した．C20PCまではLc相を観測することができたが，C21PCでは観測できなかった．得られた結果を過去になされたCnPC同族体のものと比較して，CnPC二分子膜の相挙動を包括的に考察した．結果，CnPC(n = 12 - 22)二分子膜の温度-圧力相図，各種の相転移(副転移，前転移，主転移)に伴う熱力学量のアシル鎖長依存性および圧力誘起指組み構造形成の短鎖および長鎖限界など，CnPC二分子膜相挙動に関する有用な情報を得ることができた．


Pressure-induced interdigitation of lipid bilayer membranes: Dependence on Acyl-chain length and limitation of the formation

脂質膜の圧力誘起指組構造形成:疎水鎖長依存性と形成限界

We investigated the bilayer interdigitation of diacylphosphatidylcholines (CnPCs) with long acyl chains (n = 19 - 22) under high pressure. The temperature-pressure phase diagrams of the CnPC bilayers were constructed by the methods of differential scanning calorimetry and high-pressure light transmittance. We confirmed the bilayer interdigitation for CnPCs (n = 19 - 21) under high pressure. On the other hand, the C22PC bilayer showed peculiar behavior: two transitions were observed under atmospheric pressure while one transition under high pressure. To identify the low-temperature phase of the C22PC bilayer, we performed small angle neutron scattering measurements under atmospheric pressure and fluorescence measurements under high pressure. From these results, the phase was identified as the interdigitated gel (LβI) phase. Considering our previous results, it turned out that the acyl-chain length of CnPC that exhibits the pressure-induced interdigitation is limited within n = 14 - 21. The present results definitely indicate that the bilayer interdigitation is governed by the hydrophobic interaction as well as the hydrophilic interaction of the PC molecule in the bilayer.

Journal of the Society of Materials Science, Japan


Thermotropic phase behavior of hydrogenated soybean phosphatidylcholine-cholesterol binary liposome membrane.

By combination of differential scanning calorimetry (DSC) and fluorescence spectroscopy of 6-propionyl-2-(dimethylamino)naphthalene (Prodan), we elucidated the thermotropic phase behavior of hydrogenated soybean phosphatidylcholine (HSPC)-cholesterol binary liposome membrane which has similar lipid composition to Doxil®, the widely used liposome product in treatment of various tumors. We found that the characteristic points at cholesterol mole fraction (Xch)=0.023 and 0.077 correspond to the hexagonal lattice, in which cholesterol molecules are considered to be regularly distributed in all regions of HSPC lipid bilayer with 1 : 42 and 1 : 12 units, respectively, as static averaged structures. Apparent endothermic peak disappeared at Xch=0.40 in the DSC thermograms, indicating the existence of single liquid ordered phase at Xch>0.40. In addition, fluorescence measurements of Prodan and its lauroyl derivative in poly(ethylene glycol) (PEG)-modified liposomes indicated that PEG modification has a negligible effect on the phase behavior of HSPC-cholesterol binary liposome membrane. These results may provide useful information in developing novel liposome products whose stability and encapsulated drug release are controlled.


Possible role of inter-domain salt bridges in oligopeptidase B from Trypanosoma brucei: critical role of Glu172 of non-catalytic -propeller domain in catalytic activity and Glu490 of catalytic domain in stability of OPB.

Oligopeptidase B (OPB) is a member of the prolyl oligopeptidase (POP) family of serine proteases. OPB in trypanosomes is an important virulence factor and potential pharmaceutical target. Characteristic structural features of POP family members include lack of a propeptide and presence of a -propeller domain (PD), although the role of the -PD has yet to be fully understood. In this work, residues Glu(172), Glu(490), Glu(524) and Arg(689) in Trypanosoma brucei OPB (Tb OPB), which are predicted to form inter-domain salt bridges, were substituted for Gln and Ala, respectively. These mutants were evaluated in terms of catalytic properties and stability. A negative effect on kcat/Km was obtained following mutation of Glu(172) or Arg(689). In contrast, the E490Q mutant exhibited markedly decreased thermal stability, although this mutation had less effect on catalytic properties compared to the E172Q and R689A mutants. Trypsin digestion showed that the boundary regions between the -PD and catalytic domains (CDs) of the E490Q mutant are unfolded with heat treatment. These results indicated that Glu(490) in the CD plays a role in stabilization of Tb OPB, whereas Glu(172) in the -PD is critical for the catalytic activity of Tb OPB.


How Does Acyl-chain Length Affect Thermotropic Phase Behavior of Saturated Diacylphosphatidylcholine-chloresterol Binary Bilayers?

Thermotropic phase behavior of diacylphosphatidylcholine (CnPC) cholesterol binary bilayers (n = 14 16) was examined by fluorescence spectroscopy using 6-propionyl-2-(dimethylamino)naphthalene (Prodan) and differential scanning calorimetry. The former technique can detect structural changes of the bilayer in response to the changes in polarity around Prodan molecules partitioned in a relatively hydrophilic region of the bilayer, while the latter is sensitive to the conformational changes of the acyl chains. On the basis of the data from both techniques, we propose possible temperature T cholesterol composition Xch phase diagrams for these binary bilayers. A notable feature of our phase diagrams, including our previous results for C17PC and C18PC, is that there is a peritectic-like point around Xch = 0.15, which can be interpreted as indicating the formation of a 1:6-complex of cholesterol and CnPCs within the binary bilayer irrespective of the acyl chain length. We could give a reasonable explanation for such complex formation using the modified superlattice view. Our results also showed that the Xch value of the abolition of the main transition is almost constant for n = 14 17 (ca. 0.33), while it increases to ca.0.50 for n = 18. By contrast, a biphasic n-dependence of Xch was observed for the abolition of the pretransition, suggesting that there are at least two antagonistic n-dependent factors. We speculate that this could be explained by the enhancement of the van der Waals interaction with increases in n and the weakening of the repulsion between the neighboring headgroups with decreases in n.


An Attempt to Reveal Viscoelastic Behavior of Lipid Bilayer Membrane by Pressure Perturbation Calorimetry


Thermotropic Phase Behavior of Binary Bilayer Membrane of Dipalmitoylphosphatidylcholine and Ergosterol


Study on the Subgel-Phase Formation Using an Asymmetric Phospholipid Bilayer Membrane by High-Pressure Fluorometry

The myristoylpalmitoylphosphatidylcholine (MPPC) bilayer membrane shows a complicated temperature-pressure phase diagram. The large portion of the lamellar gel (L(β)'), ripple gel (P(β)'), and pressure-induced gel (L(β)I) phases exist as metastable phases due to the extremely stable subgel (L(c)) phase. The stable L(c) phase enables us to examine the properties of the L(c) phase. The phases of the MPPC bilayers under atmospheric and high pressures were studied by small-angle neutron scattering (SANS) and fluorescence spectroscopy using a polarity-sensitive fluorescent probe Prodan. The SANS measurements clearly demonstrated the existence of the metastable L(β)I phase with the smallest lamellar repeat distance. From a second-derivative analysis of the fluorescence data, the line shape for the L(c) phase under high pressure was characterized by a broad peak with a minimum of ca. 460 nm. The line shapes and the minimum intensity wavelength (λ''(min)) values changed with pressure, indicating that the L(c) phase has highly pressure-sensible structure. The λ''(min) values of the L(c) phase spectra were split into ca. 430 and 500 nm in the L(β)I phase region, which corresponds to the formation of a interdigitated subgel L(c) (L(c)I) phase. Moreover, the phase transitions related to the L(c) phase were reversible transitions under high pressure. Taking into account the fluorescence behavior of Prodan for the L(c) phase, we concluded that the structure of the L(c) phase is highly probably a staggered structure, which can transform into the L(c)I phase easily.


Phase Behavior of Dipalmitoylphosphatidylglycerol Bilayer Membrane in Saline Water under High Pressure

International Journal of Modern Physics: Conference Series


Volumetric Study on Dioleoylphosphatidylcholine Bilayer by Pressure Perturbation Calorimetry


Volumetric characterization of ester- and ether-linked lipid bilayers by pressure perturbation calorimetry and densitometry

We investigated the thermotropic volume behavior of dimyristoylphosphatidylcholine (DMPC), dipalmitoylphosphatidylcholine (DPPC) and dihexadecylphosphatidylcholine (DHPC) membranes using pressure perturbation calorimetry (PPC) and densitometry. The ln φ(2) vs temperature curves (φ(2): apparent molar volume of phospholipid) obtained from the PPC data using an analysis method that we developed agreed with the results from the density measurements for these lipids within the relative difference of about 0.62%. From those curves, the volume changes with the main transition were estimated at 18.0±0.49, 23.5±2.33 and 23.0±0.33 cm(3) mol(-1) for DMPC, DPPC and DHPC, respectively. For DPPC and DMPC, the average volume per methylene group of the hydrocarbon chains v(CH2) calculated by referring to the procedure by Nagle and Wilkinson was consistent with the previous result, which indicates that the DPPC bilayer in the gel state has denser hydrophobic bilayer core than the DMPC bilayer. For DHPC, the volume of the headgroup region v(H) was calculated to be 244 Å(3) by assuming that v(CH2) of DHPC equals that of DPPC above 45°C. This value was comparable to that of DPPC when the volume of the carbonyl groups was considered, which may signify that there is no significant conformational difference in the polar headgroups of both phospholipids. However, it was suggested from the consideration on v(H) of DHPC at 20°C that expansion of the headgroup region should occur as the interdigitated structure is formed, which means some conformational change of the headgroup region is induced by the interdigitation.


Morphological Change of Vesicle Particle can Produce a Peculir Stepwise Transition in Dipalmitoylphosphatidylglycerol Bilayer at High NaCl Concentration


Imaging of Phosphatidylcholine Bilayers by a High-Pressure Fluorescence Technique: Detection of the Packing Difference

The packing states in phosphatidylcholine (PC) bilayers under high pressure were studied from fluorescence measurements using a polarity-sensitive probe Prodan. By using the Prodan fluorescence data, three-dimensional image plots, which can visually evaluate the bilayer packing, were constructed for the C16PC bilayer. It turned out from the shape of contours in the image plot constructed by the second-derivative spectra that the Prodan molecules can be distributed into multiple sites in the bilayer and move around the head-group region depending on the phase state. Furthermore, comparing the plot to those for the C17PC and C18PC bilayers, the slight difference in hydrophobicity among the PC molecules produced a marked difference in quantity of the Prodan molecules distributed among the bilayer gel, nonbilayer gel, and liquid crystalline phases of the PC membranes. The present study shows that the imaging using Prodan is useful for the evaluation of lipid bilayer packing.

Bulletin of the Chemical Society of Japan


Intrinsic interaction mode of an inhalation anesthetic with globular proteins: a comparative study on ligand recognition


Thermotropic and Barotropic Phase Transitions of Dialkyldimethylammonium Bromide Bilayer Membranes: Effect of Chain Length

The bilayer phase transitions of dialkyldimethylammonium bromides (2C(n)Br; n = 12, 14, 16) were observed by differential scanning calorimetry and high-pressure light-transmittance measurements. Under atmospheric pressure, the 2C(12)Br bilayer membrane underwent the stable transition from the lamellar crystal (L(c)) phase to the liquid crystalline (L(α)) phase. The 2C(14)Br bilayer underwent the main transition from the metastable lamellar gel (L(β)) phase to the metastable L(α) phase in addition to the stable L(c)/L(α) transition. For the 2C(16)Br bilayer, moreover, three kinds of phase transitions were observed: the metastable main transition, the metastable transition from the metastable lamellar crystal (L(c(2))) phase to the metastable L(α) phase, and the stable lamellar crystal (L(c(1)))/L(α) transition. The temperatures of all the phase transitions elevated almost linearly with increasing pressure. The temperature (T)-pressure (p) phase diagrams of the 2C(12)Br and 2C(14)Br bilayers were simple, but that of the 2C(16)Br bilayer was complex; that is, the T-p curves for the metastable main transition and the L(c(2))/L(α) transition intersect at ca. 25 MPa, which means the inversion of the relative phase stability between the metastable phases of L(β) and L(c(2)) above and below the pressure. Moreover, the T-p curve of the L(c(2))/L(α) transition was separated into two curves under high pressure, and as a result, the pressure-induced L(c(2P)) phase appeared in between. Thermodynamic quantities for phase transitions of the 2C(n)Br bilayers increased with an increase in alkyl-chain length. The chain-length dependence of the phase-transition temperature for all kinds of transitions observed suggests that the stable L(c(1))/L(α) transition incorporates the metastable L(c(2))/L(α) transition in the bilayers of 2C(n)Br with shorter alkyl chains, and the main-transition of the 2C(12)Br bilayer would occur at a temperature below 0 °C.


Prodan fluorescence detects the bilayer packing of asymmetric phospholipids

The bilayer phase behavior of asymmetric phospholipids, palmitoylstearoylphosphatidylcholine (PSPC) and stearoylpalmitoylphosphatidylcholine (SPPC), with different vesicle sizes (large multilamellar vesicle (LMV) and giant multilamellar vesicle (GMV)) was investigated by fluorescence spectroscopy using a polarity-sensitive fluorescent probe Prodan under high pressure. The results were compared with those of a symmetric phospholipid, diheptadecanoyl PC (C17PC). The difference in phase transitions of the PSPC and SPPC bilayers and in thermodynamic quantities of the transitions was hardly observed between LMV and GMV as the case of the C17PC bilayer. On the other hand, the Prodan fluorescence showed clear differences between LMV and GMV of the asymmetric PC bilayers. From the second derivative of Prodan fluorescence spectra, the three dimensional image plots in which we can clearly see the location of Prodan in the bilayer membrane as blue valleys were constructed for LMV and GMV under high pressure. We revealed from the plots that the bilayer packing is significantly dependent on not only the vesicle size but also the acyl-chain asymmetry of PC molecule in addition to the phase states. It was found that the packing of the gel phases of the asymmetric PC bilayers is weaker than that of the symmetric PC bilayer, and the size of vesicle affects the packing of the interdigitated gel phase the most markedly among three gel phases. This study suggests that the Prodan molecules can detect the effect of vesicle size on the phase states for the asymmetric PC bilayers, and they become a useful indicator for various membrane properties, especially bilayer interdigitation.


Chain elongation of diacylphosphatidylcholine induces fully bilayer interdigitation under atmospheric pressure

The phase transitions of dibehenoylphosphatidylcholine (C22PC) bilayer membrane were observed by differential scanning calorimetry under atmospheric pressure and light-transmittance measurements under high pressure. The constructed temperature-pressure phase diagram suggests that the gel phase at low temperatures is the interdigitated gel phase. To confirm the phase state, we performed small-angle neutron scattering and fluorescence measurements using a polarity-sensitive probe Prodan for the C22PC bilayer membrane under atmospheric pressure. The peaks obtained in both measurements clearly showed the characteristic patterns of the fully interdigitated gel phase. Taking into account of previous studies on the gel phase for long-chain PC bilayers under atmospheric pressure and our studies on the pressure-induced bilayer interdigitaion of diacyl-PCs, it turned out that the interdigitation of diacyl-PC bilayer membranes occurs when the carbon number of acyl chain reaches at least 22. The present study revealed that the interdigitation of PC bilayer membranes occurs not only by weakening the attractive force of polar head groups but also by strengthening the cohesive force of acyl chains. When dominating the force of acyl chains, the interdigitation can be induced even in a diacyl-PC bilayer membrane by only hydration under atmospheric pressure.


Hydrostatic Pressure Reveals Bilayer Phase Behavior of Dioctadecyldimethylammonium Bromide and Chloride

Bilayer phase transitions of dioctadecyldimethylammonium bromide (2C(18)Br) and chloride (2C(18)Cl) were observed by differential scanning calorimetry and high-pressure light-transmittance measurements. The 2C(18)Br bilayer membrane showed different kinds of transitions depending on preparation methods of samples under atmospheric pressure. Under certain conditions, the 2C(18)Br bilayer underwent three kinds of transitions, the metastable transition from the metastable lamellar crystal (L(c(2))) phase to the metastable lamellar gel (L(β)) phase at 35.4 °C, the metastable main transition from the metastable L(β) phase to the metastable liquid crystalline (L(α)) phase at 44.5 °C, and the stable transition from the stable lamellar crystal (L(c(1))) phase to the stable L(α) phase at 52.8 °C. On the contrary, the 2C(18)Cl bilayer underwent two kinds of transitions, the stable transition from the stable L(c) phase to the stable L(β) phase at 19.7 °C and the stable main transition from the stable L(β) phase to the stable L(α) phase at 39.9 °C. The temperatures of the phase transitions of the 2C(18)Br and 2C(18)Cl bilayers were almost linearly elevated by applying pressure. It was found from the temperature (T)-pressure (p) phase diagram of the 2C(18)Br bilayer that the T-p curves for the main transition and the L(c(1))/L(α) transition intersect at ca. 130 MPa because of the larger slope of the former transition curve. On the other hand, the T-p phase diagram of the 2C(18)Cl bilayer took a simple shape. The thermodynamic properties for the main transition of the 2C(18)Br and 2C(18)Cl bilayers were comparable to each other, whereas those for the L(c(1))/L(α) transition of the 2C(18)Br bilayer showed considerably high values, signifying that the L(c(1)) phase of the 2C(18)Br bilayer is extremely stable. These differences observed in both bilayers are attributable to the difference in interaction between a surfactant and its counterion.


Application of pressure perturbation calorimetry to the aqueous system of phospholipid vesicle dispersion


Packing state in bilayer membranes of diacylphosphatidylcholines with varying acyl chain lengths under high pressure


Effect of Vesicle Size on the Prodan Fluorescence in Diheptadecanoylphosphatidylcholine Bilayer Membrane under Atmospheric and High Pressures

The bilayer phase behavior of diheptadecanoylphosphatidylcholine (C17PC) with different vesicle sizes (large multilamellar vesicle (LMV) and giant multilamellar vesicle (GMV)) was investigated by fluorescence spectroscopy using a polarity-sensitive fluorescent probe Prodan under atmospheric and high pressures. The difference in phase transitions and thermodynamic quantities of the transition was hardly observed between LMV and GMV used here. On the contrary, the Prodan fluorescence in the bilayer membranes changed depending on the size of vesicles as well as on the phase states. From the second derivative of fluorescence spectra, the three-dimensional image plots in which we can see the location of Prodan in the bilayer membrane as blue valleys were constructed for LMV and GMV under atmospheric pressure. The following characteristic behavior was found: (1) the Prodan molecules in GMV can be distributed to not only adjacent glycerol backbone region, but also near bulk-water region in the lamellar gel or ripple gel phase; (2) the blue valleys of GMV became deeper than those of LMV because of the greater surface density of the Prodan molecules per unit area of GMV than LMV; (3) the liquid crystalline phase of the bilayer excludes the Prodan molecules to a more hydrophilic region at the membrane surface with an increase in vesicle size; (4) the accurate information as to the phase transitions is gradually lost with increasing vesicle size. Under the high-pressure condition, the difference in Prodan fluorescence between LMV and GMV was essentially the same as the difference under atmospheric pressure except for the existence of the pressure-induced interdigitated gel phase. Further, we found that Prodan fluorescence spectra in the interdigitated gel phase were especially affected by the size of vesicles. This study revealed that the Prodan molecules can move around the headgroup region by responding not only to the phase state but also to the vesicle size, and they become a useful membrane probe, detecting important membrane properties such as the packing stress.


Structural and morphological transition of long-chain phospholipid vesicles induced by mixing with short-chain phospholipid

Effects of a short-chain phospholipid, dihexanoylphosphatidylcholine (DHPC), on the structure and morphology of membrane assemblies of a long-chain phospholipid, dimyristoylphosphatidylcholine (DMPC), were examined by fluorescence spectroscopy, differential scanning calorimetry (DSC), and cryogenic transmission electron microscopy (cryo-TEM). It was found by fluorescence measurements that DHPC affects on the gel and liquid crystalline state of DMPC vesicle membranes in different ways. Further, the result of DSC suggested that, along the transition process from DMPC vesicle to DMPC-DHPC mixed micelle, there are at least three different concentration regions which are characterized by the individual variation pattern of the transition temperature and enthalpy change. The cryo-TEM micrographs demonstrated the formation of thread-like assemblies in the second region and the coexistence of the assemblies and spherical micelles in the third region. Thus, it was concluded that the structural transition from DMPC vesicle to DMPC-DHPC mixed micelle could occur in a stepwise manner through the formation of the thread-like assembly, which cannot be described by the three-stage model of vesicle to micelle transition.


Recovery of Tobacco BY-2 Cells After Treatment of High Hydrostatic Pressure

The recovery of Nicotiana tabacum L. cv. Bright Yellow 2 (BY-2) cells in Linsmaire and Skoog medium after treatment at high hydrostatic pressure was investigated using an Evans Blue staining method to discriminate live from dead cells. The survival of BY-2 cells just after the high-pressure treatment at 5 degrees C and 25 degrees C decreased abruptly at pressures higher than 50 MPa and 100 MPa, respectively. Furthermore, almost all of the BY-2 cells treated at 5 degrees C and 25 degrees C recovered pressures below 25 MPa and 75 MPa, respectively. However, no BY-2 cells recovered at pressures above 100 MPa at either temperature.

Annals of the New York Academy of Sciences


Pressure Study on Symmetric and Asymmetric Phospholipid Bilayers: Effect of Vesicle Size on Prodan Fluorescence

The bilayer phase behavior of symmetric and asymmetric phosphatidylcholines (PCs), 1,2-diheptadecanoyl-PC (C17PC), 1-palmitoyl-2-stearoyl-PC (PSPC), and 1-stearoyl-2-palmitoyl-PC (SPPC), with different vesicle sizes were investigated by a high-pressure fluorescence method using the polarity-sensitive fluorescent probe Prodan. The second derivative of fluorescence spectra for all the PCs of small-sized vesicle showed four minima characteristic of four membrane states on the spectra irrespective of the acyl-chain symmetry, whereas those of large-sized vesicle had one more minimum originating from the most hydrophilic site at the membrane surface. These findings indicate that Prodan molecules can distribute into multiple sites in the bilayer and move around the head-group region depending on the vesicle size. The behavior of the spectra in the SPPC bilayer suggested that the interdigitated gel phase had a less polar "pocket" formed by a space between uneven terminal methyl ends of the sn-1 and sn-2 chains. It turned out that the curvature of vesicles affects the distribution of the Prodan molecules in all phases, more particularly in the interdigitated gel phase.


Pressure Effect on the Bilayer Phase Transition of Asymmetric Lipids with an Unsaturated Acyl Chain

The bilayer phase transitions of mixed-chain lipids with monounsaturated acyl chain in the sn-2 position, 1-myristoyl-2-oleoyl-sn-glycero-3-phosphocholine (MOPC), 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), and 1-stearoyl-2-oleoyl-sn-glycero-3-phosphocholine (SOPC), and with a polyunsaturated acyl chain in the sn-2 position, 1-stearoyl-2-linoleoyl-sn-glycero-3-phosphocholine (SLPC), 1-stearoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (SAPC), and 1-stearoyl-2-docosahexaenoyl-sn-glycero-3-phosphocholine (SDPC), were observed by differential scanning calorimetry (DSC) under ambient pressure and by light-transmittance measurements under high pressure. The DSC thermogram for each lipid bilayer showed only one transition between the lamellar gel and liquid crystalline phases. The introduction of one or two cis double bonds into the sn-2 acyl chain caused the significant depression of the main-transition temperature and an obvious decrease of enthalpy and volume changes associated with the transition. These features are attributable to loose packing of saturated and unsaturated acyl chains in the bilayer gel phase. The existence of four or six double bonds in the sn-2 chain produced no further decrease in the transition temperature, and in fact six double bonds caused a slight increase in the transition temperature. Thermodynamic properties associated with the bilayer phase transition were discussed.


A critical role for highly conserved Glu610 residue of oligopeptidase B Tryoanosoma brucei in thermal stability

Oligopeptidase B from Trypanosoma brucei (Tb OPB) is a virulence factor and therapeutic target in African sleeping sickness. Three glutamic acid residues at positions 607, 609 and 610 of the catalytic domain are highly conserved in the OPB subfamily. In this study, the roles of Glu(607), Glu(609) and Glu(610) in Tb OPB were investigated by site-directed mutagenesis. A striking effect on k(cat)/K(m) was obtained following mutation of Glu(607) to glutamine. In contrast, the heat stability of Tb OPB decreased markedly following the single mutation of Glu(610) to glutamine, although this mutation had significantly less effect on catalytic properties compared with the Glu(607) mutation. Although no differences were found in the tertiary and secondary structures between wild-type (WT) OPB and the E610Q mutant prior to heat treatment, the E610Q mutant is inactivated more rapidly than WT OPB following heat treatment in a manner correlating with its attendant structural changes. Trypsin digestion showed that the boundary regions between the beta-propeller and catalytic domain of the E610Q mutant are unfolded with heat treatment. It is concluded that Glu(607) is essential for the catalytic activity of Tb OPB and that Glu(610) plays a critical role in stabilization rather than catalytic activity despite their close proximity.


Inhibition of anti-fluorescent probe monoclonal antibody by long-chain amphiphiles

Inhibitions of amphiphiles with a long chain (alcohols with carbon number of 10, 12, 14, 16 and the corresponding fatty acids) on the antigen-antibody reaction between fluorescent probe (fluorescein (FL)) and anti-fluorescent probe monoclonal antibody (immunoglobulin G (IgG)-49) were investigated by fluorescence spectroscopy. The resulting inhibition modes were compared with those of specific inhibitors, 5- and 6-aminofluoresceins (5- and 6-FLNH(2)). In the presence of all inhibitors, the fluorescence quenching of FL as the result of inhibitor binding to IgG-49 was hindered in an inhibitor concentration-dependent manner. The values of IC(50) and Hill number for the inhibitors were determined from dose-response curves of the quenching inhibition. The IC(50) values of the alcohols were much more dependent on the chain length than those of the fatty acids although all the values were in orders of microM. The Hill numbers of both kinds of inhibitors became larger values than 1, but they had different values for a series of alcohols and fatty acids, respectively. By contrast, the IC(50) values of 5- and 6-FLNH(2) were in orders of nM and the Hill numbers became smaller values than 1. Since the Lineweaver-Burk plots produced an intersection point on the intercept of the X-axis for the long-chain inhibitors with carbon number of 14 while did that on the intercept of the Y-axis for 6-FLNH(2), respectively, we judged the inhibition modes of the former as non-competitive ones and those of the latter as a competitive one. Further, the effect of pressure on the inhibition by C(14)OH, C(13)COOH and 5-FLNH(2) was examined, and the standard volume changes of IgG-49 for the inhibitor binding were calculated from the Johnson-Eyring plots. The volume change for 5-FLNH(2) was smaller than those for C(14)OH and C(13)COOH although they became all negative. This suggests that the volume of IgG-49 shrinks by the addition of these inhibitors, and IgG-49 is not relevant to a model protein for the molecular mechanism of anesthesia.


A Peculiar Phase Transition of Plasmalogen Bilayer Membrane under High Pressure

The bilayer phase transition of plasmalogen, monounsaturated plasmenylcholine 1-O-1'-(Z)-octadecenyl-2-oleoyl-sn-glycero-3-phosphocholine (Plg-SOPC), was examined by differential scanning calorimetry, high-pressure transmittance, and fluorescence techniques. The bilayer properties of Plg-SOPC such as the temperature-pressure phase diagram, the thermodynamic quantities of the transition, and the location of a fluorescent membrane probe in the bilayer, were compared with those of a similar phospholipid 1-stearoyl-2-oleoyl-phosphatidylcholine (SOPC). It turned out that a vinyl-ether bond in the sn-1 position of the glycerol backbone in the Plg-SOPC molecule produces a peculiar phase transition under high pressure and significantly affects the membrane properties.


Chain asymmetry alters thermotropic and barotropic properties of phospholipid bilayer membranes

The alignment of the sn-1 and sn-2 acyl chains at the terminal methyl ends generally produces significant influence on the thermodynamic properties of the bilayer phase transitions. We investigated the bilayer phase behavior of asymmetric phospholipids, myristoylpalmitoylphosphatidylcholine and palmitoylmyristoylphosphatidylcholine, by high-pressure light-transmittance and Prodan-fluorescence techniques and differential scanning calorimetry. Constructed temperature-pressure phase diagrams revealed that no stable Lbeta' phase can exist in the whole pressure range because of the formation of the most stable Lc phase. Nevertheless, the pretransition, the detection of which is severely hampered by the exceptionally prompt formation of the Lc phase, was successfully observed. Moreover, the effect of the total and difference of the sn-1 and sn-2 acyl chain lengths on minimal interdigitation pressure (MIP) was summarized in a MIP vs. chain-length inequivalence parameter plot, where the effect was proved to be classified in three zones depending on the alignment of both terminal methyl ends.


A comparative study on specific and nonspecific interactions in bovine serum albumin: thermal and volume effects of halothane and palmitic acid


Interdigitation and vesicle-to-micelle transformation induced by a local anesthetic tetracaine in phospholipids bilayers

The phase transitions of distearoyl- (DSPC), dipalmitoyl- (DPPC) and dimyristoyl-phosphatidylcholine (DMPC) bilayer membranes were observed by means of differential scanning calorimetry as a function of the concentration of a local anesthetic tetracaine hydrochloride (TC.HCl). The depression of both temperatures of the main- and pre-transition, which is accompanied by a decrease in enthalpy changes for both transitions, was observed initially by the addition of TC.HCl. Bilayer interdigitation, which is accompanied by an increase in enthalpy change for the main transition from the interdigitated gel phase to the liquid crystalline phase, was followed by disappearance of the pretransition. The TC.HCl concentration necessary for the bilayer interdigitation was found to be 10, 21 and 6 mmol kg(-1) for DSPC, DPPC and DMPC bilayers, respectively, which was not consistent with the order of acyl-chain length of lipids. Biphasic interactions for the interdigitation, that is, repulsive interaction between polar head groups and van der Waals attractive interaction between hydrophobic chains of lipids, were discussed. On the other hand, vesicle-to-micelle transformation, which is accompanied by a cooperative decrease in enthalpy change for the main transition, was observed at higher concentration of TC.HCl and was confirmed by the vesicle size determined by the dynamic light scattering. The longer the acyl-chain length of lipids, the higher the TC.HCl concentration necessary for the vesicle-to-micelle transformation.


Effect of a Local Anesthetic Lidocaine Hydrochloride on the Bilayer Structure of Phospholipids

The effect of a local anesthetic, lidocaine hydrochloride (LC x HCl), on the bilayer systems of purified egg phosphatidylcholine (EPC), dipalmitoylphosphatidylcholine (DPPC) and dioleoylphosphatidylcholine (DOPC) was studied by means of small-angle X-ray scattering (SAXS), Prodan fluorescence and electrophoretic light scattering. In the liquid crystalline phase of EPC and DOPC bilayers, the contraction of lamellar distance by ca. 0.8-1.0 nm and the decrease of average vesicle size were observed in the presence of LC x HCl. The adsorption of LC x HCl on the vesicle interface brought about the lateral expansion of bilayers and the decrease in the radius of curvature of vesicles. The contraction in the lamellar distance of EPC bilayer caused by high concentration of LC x HCl is attributable to the chain folding in the liquid crystalline state. In the gel phase of DPPC bilayer, the contraction of the lamellar distance in the presence of 0.37 M LC x HCl amounts to 1.6 nm, and the emission maximum of Prodan fluorescence was red-shifted from 440 nm to 518 nm. These phenomena are attributed to the formation of LC x HCl-induced interdigitated gel phase.


Barotropic and thermotropic bilayer phase behavior of positional isomers of unsaturated mixed-chain phosphatidylcholines

The bilayer phase transitions of six kinds of mixed-chain phosphatidylcholines (PCs) with an unsaturated acyl chain in the sn-1 or sn-2 position, 1-oleoyl-2-stearoyl- (OSPC), 1-stearoyl-2-oleoyl- (SOPC), 1-oleoyl-2-palmitoyl- (OPPC), 1-palmitoyl-2-oleoyl- (POPC), 1-oleoyl-2-myristoyl- (OMPC) and 1-myristoyl-2-oleoyl-sn-glycero-3-phosphocholine (MOPC), were observed by means of differential scanning calorimetry (DSC) and high-pressure light transmittance measurements. Bilayer membranes of SOPC, POPC and MOPC with an unsaturated acyl chain in the sn-2 position exhibited only one phase transition, which was identified as the main transition between the lamellar gel (L(beta)) and liquid crystalline (L(alpha)) phases. On the other hand, the bilayer membranes of OSPC, OPPC and OMPC with an unsaturated acyl chain in the sn-1 position exhibited not only the main transition but also a transition from the lamellar crystal (L(c)) to the L(beta) (or L(alpha)) phase. The stability of their gel phases was markedly affected by pressure and chain length of the saturated acyl chain in the sn-2 position. Considering the effective chain lengths of unsaturated mixed-chain PCs, the difference in the effective chain length between the sn-1 and sn-2 acyl chains was proven to be closely related to the temperature difference of the main transition. That is, a mismatch of the effective chain length promotes a temperature difference of the main transition between the positional isomers. Anomalously small volume changes of the L(c)/L(alpha) transition for the OPPC and OMPC bilayers were found despite their large enthalpy changes. This behavior is attributable to the existence of a cis double bond and to significant inequivalence between the sn-1 and sn-2 acyl chains, which brings about a small volume change for chain melting due to loose chain packing, corresponding to a large partial molar volume, even in the L(c) phase. Further, the bilayer behavior of unsaturated mixed-chain PCs containing an unsaturated acyl chain in the sn-1 or sn-2 position was well explained by the chemical-potential diagram of a lipid in each phase.

Biochimica et Biophysica Acta (BBA) - Biomembranes


Interaction modes of long-chain fatty acids in dipalmitoylphosphatidylcholine bilayer membrane: contrast to mode of inhalation anesthetics

The effects of long-chain fatty acids (four saturated and two unsaturated fatty acids, one derivative) on phase transitions of dipalmitoylphosphatidylcholine (DPPC) bilayer membranes were examined in the low concentration region, and the results were compared with those for an inhalation anesthetic. The effects of all fatty acids on the pre- and main-transition temperatures of the DPPC bilayer membrane appeared in the concentration range of microM order while that of the anesthetic appeared in the mM order. The appearance modes of these ligand actions were significantly different from one another. The three differential partition coefficients of the ligands between two phases of the DPPC bilayer membrane were evaluated by applying the thermodynamic equation to the variation of the phase-transition temperatures. The DPPC bilayer membranes showed the different receptivity for the ligands; the saturated fatty acids had an affinity for gel phase whereas unsaturated fatty acids and an anesthetic had an affinity for liquid-crystalline phase to the contrary. In particular, the receptivity for the ligands in the gel phase markedly changed depending on kinds of ligands. The interaction modes between the DPPC and fatty acid molecules in the gel phase were considered from the hexagonal lattice model. The disappearance compositions of the pretransition by the fatty acids coincided with the compositions at which the membrane is all covered by the units in each of which two fatty acids molecules are regularly distributed in the hexagonal lattice in a different way, and the distribution depended on the chain length and existence of a double bond for the fatty acids. The interpretation did not hold for the case of the anesthetic at all, which proved that a number of anesthetic molecules act the surface region of the bilayer membrane nonspecifically. The present study clearly implies that DPPC bilayer membranes have high ability to recognize kinds of ligand molecules and can discriminate among them with specific interaction by the membrane states.


High-pressure study on bilayer phase behavior of oleoylmyristoyl- and myristoyloleoyl-phosphatidylcholines

We investigated the thermotropic and barotropic bilayer phase behavior of 1-myristoyl-2-oleoyl-sn-glycero-3-phosphocholine (MOPC) and 1-oleoyl-2-myristoyl-sn-glycero-3-phosphocholine (OMPC) by means of the differential scanning calorimetry (DSC) and high-pressure light-transmittance technique. Water could be used as a solvent for measurements at high pressures because of the elevation of the transition temperatures above 0 degrees C by pressurization, whereas aqueous 50 wt.% ethylene glycol solution was used mainly for those at low pressures. Only one phase transition was observed in the DSC thermogram of the MOPC bilayer membrane as an endothermic peak, and also observed at high pressures as an abrupt change of the light-transmittance. The transition was assigned as a main transition between the lamellar gel (L(beta)) and liquid-crystalline (L(alpha)) phases on the basis of the values of enthalpy change (DeltaH) and slope of the transition temperature with respect to pressure (dT/dP). The DSC thermogram of the OMPC bilayer membrane similarly showed a single endothermic peak but two kinds of phase transitions were observed at different temperatures in the light-transmittance profile at high pressures. The extrapolation of the lower-temperature transition in the high-pressure range to an ambient pressure coincided with the transition observed in the DSC thermogram. This transition was identified as a transition between the lamellar crystal (L(c)) and L(alpha) (or L(beta)) phases from the DeltaH and dT/dP values. The higher-temperature transition, appearing only at high pressures, was identified as the L(beta)/L(alpha) transition considering the topological resemblance of its temperature-pressure phase diagram as that of the dioleoylphosphatidylcholine bilayer membrane. The phase diagram of the OMPC bilayer membrane demonstrated that the L(beta) phase cannot exist at pressures below ca. 190 MPa while it can exist stably in a finite temperature range at pressures above the pressure.


Lateral phase separation in cholesterol/diheptadecanoylphosphatidylcholine binary bilayer membrane

We investigated the phase behavior of cholesterol/diheptadecanoylphosphatidylcholine (C17:0-PC) binary bilayer membrane as a function of the cholesterol composition (X(ch)) by fluorescence spectroscopy using 6-propionyl-2-(dimethylamino)naphthalene (Prodan) and differential scanning calorimetry (DSC). The fluorescence spectra showed that the wavelength at the maximum intensity (lambda(max)) changed depending on the bilayer state: ca. 440 nm for the lamellar gel (L'(beta) or L(beta)) and the liquid ordered (L(o)) phases and ca. 490 nm for the liquid-crystalline (L(alpha)) phase. The transition temperatures were determined from the temperature dependence of lambda(max) and endothermic peaks of the DSC thermograms. Both measurements showed that the pre- and main transition disappear around X(ch)=0.05 and 0.30, respectively. The constructed temperature-X(ch) phase diagram resembled a typical phase diagram for a eutectic binary mixture containing a peritectic point. The presence of a peritectic point at X(ch)=0.15 suggested that a complex of cholesterol and C17:0-PC is stoichiometrically formed in the gel phase. Consideration based on the hexagonal lattice model revealed that the compositions of 0.05 and 0.15 correspond to the bilayer states where cholesterol molecules are regularly distributed in different ways. The former is nearly equal to the composition for the membrane occupied entirely with Units (1:18), composed of a cholesterol and 18 surrounding C17:0-PC molecules within the next-next nearest neighbor sites. The latter is represented by a Unit (1:6), including a cholesterol and 6 surrounding C17:0-PC molecules. Further, the disappearance of the main transition at X(ch)=0.30 indicates that the pure L(o) phase can exist in X(ch)>0.30. The eutectic behavior observed in the phase diagram was explainable in terms of phase separation between two different types of regions with different types of regular distributions of cholesterol.


エーテル結合型リン脂質における指組み構造形成

エーテル結合型リン脂質における指組み構造形成

High Pressure Bioscience and Biotechnology

高圧バイオサイエンスとバイオテクノロジー


高圧力下におけるジノナデカノイルホスファチジルコリン二分子膜の相挙動

高圧力下におけるジノナデカノイルホスファチジルコリン二分子膜の相挙動

高圧バイオサイエンスとバイオテクノロジー


Thermodynamic Consideration for Nonbilayer Formation of Phospholipid Membranes

リン脂質膜の非二分子膜形成に対する熱力学的考察

高圧バイオサイエンスとバイオテクノロジー


A new interpretation of eutectic behavior for distearoylphosphatidylcholine-cholesterol binary bilayer membrane

We investigated the thermotropic phase behavior of the distearoylphosphatidylcholine (DSPC)-cholesterol binary bilayer membrane as a function of the cholesterol composition (X(ch)) by fluorescence spectroscopy using 6-propionyl-2-(dimethylamino)naphthalene (Prodan) and differential scanning calorimetry (DSC). The fluorescence spectra, each of which has a single maximum, showed that the wavelength at the maximum intensity (lambda(max)) changed depending on the bilayer state: ca. 440 nm for the lamellar gel (L(beta)' or L(beta)) and the liquid ordered (L(o)) phases, ca. 470 nm for the ripple gel (P(beta)') phase and ca. 490 nm for the liquid crystalline (L(alpha)) phase, respectively. The transition temperatures were determined from the temperature dependences of the lambda(max) and endothermic peaks of the DSC thermograms. Both measurements showed that the pretransition disappears around X(ch)=0.035. The constructed temperature-X(ch) phase diagram indicated that the phase behavior of the binary bilayer membrane at X(ch)< or =0.15 is similar to that of general liquid-solid equilibrium for a binary system where both components are completely miscible in the liquid phase and completely immiscible in the solid phase. It was also revealed that the diagram has two characteristic points: a congruent melting point at X(ch)=0.08 and a peritectic-like point at X(ch)=0.15. The hexagonal lattice model was used for the interpretation of the phase behavior of the binary bilayer membrane. These characteristic compositions well correspond to the bilayer states in each of which cholesterol molecules are regularly distributed in the hexagonal lattice in a different way. That is, each composition of 0.035, 0.08 and 0.15 is nearly equal to that for the binary bilayer membrane which is entirely occupied with units, each composed of a cholesterol and 30 surrounding DSPC molecules within the next-next-next nearest neighbor sites (Unit (1:30): L(beta)(1:30)), with units, each of a cholesterol and 12 surrounding DSPC molecules within the next nearest sites (Unit (1:12): L(beta)(1:12)) or with units, each of a cholesterol and 6 surrounding DSPC molecules at the nearest neighbor sites (Unit (1:6): L(beta)(1:6)), respectively. Therefore, the eutectic behavior observed in the phase diagram was fully explainable in terms of a kind of phase separation between two different types of regions with different types of regular distributions of cholesterol. Further, the L(o) phase was found in the higher X(ch)-region (X(ch)>0.15). No endothermic peak over the temperature range from 10 to 80 degrees C at X(ch)=0.50 suggested that the single L(o) phase can exist at X(ch)>0.50.


Thermotropic and barotropic phase transitions of dilauroylphosphatidylcholine bilayer

The bilayer phase transitions of dilauroylphosphatidylcholine (DLPC), containing two linear acyl chains with 12 carbon atoms, were observed by means of differential scanning calorimetry (DSC) under ambient pressure and light transmittance under high pressure. When the heating scan for the DLPC bilayer in 50 wt.% aqueous ethylene glycol (EG) solution began at -30 degrees C after cold storage, the DSC thermogram showed two endothermic peaks at 1.7 and 4.5 degrees C, which correspond to the transition from the lamellar crystalline (Lc) phase to the intermediate liquid crystalline (Lx) phase and the transition from the Lx phase to the liquid crystalline (L) phase, respectively. Extremely large enthalpy change (32.9 kJ mol(-1)) is characteristic of the Lc/Lx phase transition. The DSC thermogram for the heating scan beginning from -10 degrees C showed a single endothermic peak with 9.2 kJ mol(-1) at -0.4 degrees C, which was assigned as the so-called main transition between the metastable ripple gel (P'(beta)) and metastable Lalpha phases. The DLPC bilayer under high pressure underwent three kinds of transitions in EG solution, whereas only one transition was observed in water under high pressure. The middle-temperature transition in EG solution could be assigned to the main transition because of its consistency with the main transition in water. The lower-temperature transition is probably assigned as transition from the Lc phase to the P'(beta) phase. Since the slope (dT/dp) of the Lc/P'(beta) phase boundary is smaller than that for the main transition, the Lc/P'(beta) phase boundary and the main transition curves crossed each other at 40 MPa on the temperature-pressure phase diagram. The higher-temperature transition in EG solution refers to the transition from the Lx phase to the Lalpha phase. The Lx phase disappeared at about 180 MPa, and the direct transition from the P'(beta) phase to the Lalpha phase was observed at high pressures above 180 MPa.


Cholesterol Suppresses Pressure-induced Interdigitation of Dipalmitoylphosphatidylcholine Bilayer Membrane


Effect of hydrostatic pressure on the bilayer phase behavior of symmetric and asymmetric phospholipids with the same total chain length

The bilayer phase transitions of palmitoylstearoyl-phosphatidylcholine (PSPC), diheptadecanoyl-PC (C17PC) and stearoylpalmitoyl-PC (SPPC) which have the same total carbon numbers in the two acyl chains were observed by differential scanning calorimetry and high-pressure optical method. As the temperature increased, these bilayers exhibited four phases of the subgel (Lc), lamellar gel (L beta'), ripple gel (P beta') and liquid crystal (L alpha), in turn. The Lc phase was observed only in the first heating scan after cold storage. The temperatures of the phase transitions were almost linearly elevated by applying pressure. The temperature-pressure phase diagrams and the thermodynamic quantities associated with the phase transitions were compared among the lipid bilayers. For all the bilayers studied, the pressure-induced interdigitated gel (L beta I) phase appeared above the critical interdigitation pressure (CIP) between the L beta' and P beta' phases. The CIPs for the PSPC, C17PC and SPPC bilayers were found to be 50.6, 79.1 and 93.0 MPa, respectively. Contribution of two acyl chains to thermodynamic properties for the phase transitions of asymmetric PSPC and SPPC bilayers was not even. The sn-2 acyl chain lengths of asymmetric PCs governed primarily the bilayer properties. The fluorescence spectra of Prodan in lipid bilayers showed the emission maxima characteristic of bilayer phases, which were dependent on the location of Prodan in the bilayers. Second derivative of fluorescent spectrum exhibited the original emission spectrum of Prodan to be composed of the distribution of Prodan into multiple locations in the lipid bilayer. The F''497/F''430 value, a ratio of second derivative of fluorescence intensity at 497 nm to that at 430 nm, is decisive evidence whether bilayer interdigitation will occur. With respect to the L beta'/L beta I phase transition in the SPPC bilayer, the emission maximum of Prodan exhibited the narrow-range red-shift from 441 to 449 nm, indicating that the L beta I phase in the SPPC bilayer has a less polar "pocket" formed by a space between uneven terminal methyl ends of the sn-1 and sn-2 chains, in which the Prodan molecule remains stably.


Miscibility and interaction between 1-alkanol and short-chain phosphocholine in the adsorbed film and micelles

The miscibility and interaction of 1-hexanol (C6OH) and 1-heptanol (C7OH) with 1,2-dihexanoyl-sn-glycero-3-phosphocholine (DHPC) in the adsorbed films and micelles were investigated by measuring the surface tension of aqueous C6OH-DHPC and aqueous C7OH-DHPC solutions. The surface density, the mean molecular area, the composition of the adsorbed film, and the excess Gibbs energy of adsorption g(H,E), were estimated. Further, the critical micelle concentration of the mixtures was determined from the surface tension versus molality curves; the micellar composition was calculated. The miscibility of the 1-alkanols and DHPC molecules in the adsorbed film and micelles was examined using the phase diagram of adsorption (PDA) and that of micellization (PDM). The PDA and the composition dependence of g(H,E) indicated the non-ideal mixing of the 1-alkanols and DHPC molecules due to the attractive interaction between the molecules in the adsorbed film, while the PDM indicated that the 1-alkanol molecules were not incorporated in the micelles within DHPC rich region. The dependence of the mean molecular area of the mixtures on the surface composition suggested that the packing property of the adsorbed film depends on the chain length of 1-alkanol: C6OH expands the DHPC adsorbed film more than C7OH.


Volume study on the exclusion of lithium naphthylsulfonate from lithium decylsulfonate micelles


Thermotropic and barotropic phase transitions in bilayer membranes of ether-linked phospholipids with varying alkyl chain lengths

疎水鎖長の異なる一連のエーテル型リン脂質ジアルキルホスファチジルコリン二分子膜の相転移に関する熱力学諸量と温度-圧力相図を対応するエステル型リン脂質の結果と比較した．主転移に関する熱力学量の詳細な比較から，ゲル相中におけるエーテル型リン脂質の相互作用はエステル型リン脂質よりも弱いことを示した．さらにエーテル型リン脂質の相図はエステル型リン脂質の相図の高温，高圧領域に対応することを明らかにした．

Biochimica et Biophysica Acta (BBA) - Biomembranes


Adsorption-Equilibrium Surface Tension of Surfactant Solutions -Examination by the Drop Volume Method-

Bulletin of the Chemical Society of Japan


Bilayer Phase Transitions of N-methylated Dioleoylphosphatidylethanolamines under High Pressure

The bilayer phase transitions of four kinds of unsaturated phospholipids with different-sized polar head groups, dioleoylphosphatidylethanolamine (DOPE), dioleoylphosphatidyl-N-methylethanolamine (DOMePE), dioleoylphosphatidyl-N,N-dimethylethanolamine (DOMe2PE) and dioleoylphosphatidylcholine (DOPC), were observed by means of differential scanning calorimetry (DSC) and high-pressure light-transmittance. DSC thermogram and light-transmittance curve for each phospholipid vesicle solution exhibited only one phase transition under ambient pressure, respectively. The light-transmittance of DOPC solution at pressure higher than 234 MPa abruptly increased stepwise at two temperatures, which corresponds to the appearance of stable subgel and lamellar gel phases under high pressure in addition to the liquid crystal phase. The constructed temperature (T)-pressure (p) phase diagrams were compared among these phospholipids. The phase-transition temperatures of the phospholipids decreased stepwise by N-methylation of the head group. The slops of the T-p phase boundary (dT/dp) of DOPE, DOMePE and DOMe2PE bilayers (0.127-0.145 K MPa-1) were found to be close to that of the transition from the lamellar crystal (or subgel; Lc) phase to the liquid crystal (Lalpha) phase for DOPC bilayer (0.131 K MPa-1). On the other hand, the dT/dp value of the main transition from the lamellar gel (Lbeta) phase to the Lalpha phase for DOPC bilayer (0.233 K MPa-1) was significantly different from that of the Lc/Lalpha transition, hence both curves intersected with each other at 234 MPa. The thermodynamic quantities associated with the phase transition of DOPE, DOMePE and DOMe2PE bilayers had also similar values to those for the Lc/Lalpha transition of DOPC bilayer. Taking into account of the values of transition temperature, dT/dp and thermodynamic quantities compared with the corresponding results of saturated phospholipids, we identified the phase transitions observed in the DOPE, DOMePE and DOMe2PE bilayers as the transition from the Lc phase to the Lalpha phase although they have been regarded as the main transition in the previous studies. The Lbeta phase is probably unstable for DOPE, DOMePE and DOMe2PE bilayers at all pressures, it exists as a metastable phase at pressures below 234 MPa while as a stable phase at pressures above 234 MPa in DOPC bilayer. The difference in phase stability among the phospholipid bilayers is originated from that in hydration structure of the polar head groups.


Barotropic phase transition between the lamellar liquid crystal phase and the inverted hexagonal phase of dioleoylphosphatidylethanolamine

The phase transition between the lamellar liquid crystal (Lalpha) phase and the inverted hexagonal (H(II)) phase of dioleoylphosphatidylethanolamine (DOPE) in aqueous NaCl solutions was observed by means of differential scanning calorimetry (DSC) under ambient pressure and light-transmittance technique under high pressure. The pressure dependence of the transition temperature (dT/dp) and the thermodynamic quantities for the Lalpha/H(II) transition were compared with those of another phase transition found in the DOPE bilayer membrane, which is the transition from the lamellar crystal (Lc) phase to the Lalpha phase. The dT/dp value of the Lalpha/H(II) transition was about 3.5 times as large as that of the Lc/Lalpha transition while the thermodynamic quantities were significantly smaller than those of the latter to the contrary. Comparing the enthalpy and volume behavior of the Lalpha/H(II) transition with that of the Lc/Lalpha transition, we concluded that the Lalpha/H(II) transition can be regarded as the volume-controlled transition for the reconstruction of molecular packing.


Pressure-Induced Phase Transitions of Lipid Bilayers Observed by Fluorescent Probes Prodan and Laurdan

ジステアロイルホスファチジルコリン二重膜中における2種類の蛍光プローブ色素プロダンとローダンの蛍光スペクトルを温度一定下，圧力の関数として観測した．二重膜の相状態に対応して両プローブの蛍光スペクトル極大波長の変化を見出し，各相状態における両プローブ分子の配向位置を推定した．ゲル相から液晶相への主転移においては，両プローブ共に極大波長に鋭敏な変化が観測されたが，指組み構造相への転移に関してはプロダンの極大波長に変化が見られたが，ローダンの極大波長には変化が見られなかった．ローダン分子はプロダン分子とは異なり，その長いアシル鎖のために二重膜の疎水性中心にアンカーとして固定され，指組み構造形成の際に脂質分子のグリセロール骨格から排除されないことが示唆された．


Observation of the lipid bilayer phase transition using the ion sensor

イオン選択電極法による脂質二分子膜の相転移観測

Some aromatic amine local anesthetics are a kind of amphiphilic ion. Binding of local anesthetics, dibucaine (DC), bupivacaine (BC), and lidocaine (LC) to a dipalmitoylphosphatidylcholine (DPPC) bilayer membrane was studied by potentiometry with ion-selective electrodes sensitive to DC, BC and LC cations, respectively. The DPPC membrane-buffer partition coefficient was directly determined as a function of the anesthetic concentration and the temperature. The DPPC bilayer underwent a thermotropic pretransition from the lamellar gel phase to the ripple gel phase as well as a main transition from the ripple gel phase to the liquid crystal phase. The partition coefficients of anesthetics at pH 5.5 varied with the temperature, namely the bilayer phases. Potentiometry using an ion-selective electrode sensitive to amphiphilic ions is effective for the detection of a bilayer phase transition through partition coefficients.


Effect of deuterium oxide on the thermodynamic quantities associated with phase transitions of phosphatidylcholine bilayer membranes

重水および軽水中における3種類のホスファチジルコリン(ジパルミトイル，ジステアロイル，ジヘキサデシル)二分子膜の相転移を示差走査熱量測定および高圧光透過率測定で追跡した．重水置換の効果は主転移よりも前転移を含むゲル相間転移に大きく現れ，指組み構造形成を不安定化した．両溶媒中における二分子膜の界面自由エネルギーの相違を考察し，重水置換は二分子膜界面において分子占有面積の縮小を引き起こし，より小さな分子占有面積を有する膜状態がその影響を顕著に受けることを明らかにした．


Effect of Pressure on the Prodan Fluorescence in Bilayer Membranes of Phospholipids with Varying Acyl Chain Lengths

4種類のアシル鎖長が異なるホスファチジルコリン(ジラウロイル，ジミリストイル，ジパルミトイル，ジステアロイル)二分子膜中における蛍光プローブ色素プロダンの蛍光スペクトルを圧力の関数として観測した．二分子膜の相状態に対応して蛍光スペクトル極大波長の変化を見出し，各相状態におけるプロダン分子の配向位置を推定した．リップルゲル相におけるプロダンの蛍光強度にはアシル鎖長および圧力依存性を観測した．ゲル相および液晶相の極大波長における蛍光強度比は相転移に伴い鋭敏に変化し，特に指組み構造ゲル相形成の検出に役立つことを示した．


Effect of cholesterol on partition coefficients of local anesthetic into bilayer membranes of dimyristoylphosphatidylcholine

脂質二分子膜への局所麻酔薬の分配に及ぼすコレステロールの影響

リン脂質二分子膜への局所麻酔薬の分配に対するコレステロールの影響を調べるためにコレステロール濃度の異なるジミリストイルホスファチジルコリン-コレステロール混合二分子膜へのジブカイン塩酸塩の取り込みを局所麻酔薬選択性電極法により調べた．コレステロール低組成領域では，ゲル相における分配係数は増加し，液晶相の分配係数は減少する二相性を観測した．他方，コレステロール高組成領域では分配係数は相状態によらず一定値となり，コレステロール濃度の増加に伴って減少した．上記の結果はコレステロールは麻酔薬の分配に対して制御能を有することを示唆する．


Partition coefficients of local anesthetic into lipid bilayer membranes containing unsaturated Acyl chain

不飽和アシル鎖を持つリン脂質二分子膜への局所麻酔薬の分配

リン脂質アシル鎖の飽和および不飽和形式が麻酔薬の生体膜への分配に与える影響を調べる目的で，4種類の不飽和ホスファチジルコリン(1-オレオイル-2-ステアロイル，1-ステアロイル-2-オレオイル，ジオレオイル，ジエライドイル)二分子膜へのジブカイン塩酸塩の取り込みを局所麻酔薬選択性電極法により調べた．算出された分配係数の値はジエライドイル<1-オレオイル-2-ステアロイル≒1-ステアロイル-2-オレオイル<ジオレオイルの順序で大きくなり，脂質膜への結合は脂質分子のアシル鎖構造にも依存することを示した．


Effect of halothane on the viscosity of aqueous bovine serum albumin solutions

牛血清アルブミン水溶液の粘度におよぼすハロセンの影響

溶液粘度の迅速，高精度且つ自動測定を目的として，オストワルド粘度計を使用した自動粘度測定装置を開発し，牛血清アルブミン水溶液の粘度測定を行った．中性pH領域において吸入麻酔薬ハロセン存在下における牛血清アルブミン水溶液の固有粘度は非存在下のものよりも小さな値となり，ハロセン添加により溶液内で牛血清アルブミンの体積が収縮することが示唆された．上記の結果より，非常に親水性の強いタンパク質は麻酔作用のモデルタンパク質としては適当ではないと推察した．


Inhibition modes of amphiphilic ligands with a long chain on the anti-fluorescent probe monoclonal antibody

蛍光プローブ特異的モノクローナル抗体への長鎖両親媒性リガンドの阻害様式

蛍光プローブフルオレセインとその特異的モノクローナル抗体との抗原抗体反応におよぼす長鎖両親媒性リガンド(炭素数10，12，14，16のアルコールおよび対応する脂肪酸)の作用様式を蛍光分光法で調べた．消光阻害の用量応答曲線はアルコール系列および脂肪酸系列でそれぞれ異なり，両リガンドの阻害様式の違いが示唆された．また，上記リガンドの阻害反応に対する圧力効果を調べ，両リガンド存在下ではフルオレセイン特異的モノクローナル抗体の体積は収縮することがわかった．


Thermotropic and barotropic phase transitions of N-methylated dipalmitoylphosphatidylethanolamine bilayers

極性基の大きさの異なる4種類のN-メチル化ジパルミトイルホスファチジルエタノールアミン二分子膜の相転移を高圧光透過法で観測した．ゲル相から液晶相への主転移および結晶相からゲル相への副転移，両転移温度は共に加圧により上昇し，極性基サイズの増大につれて降下した．ジパルミトイルホスファチジルエタノールアミン二分子膜の場合には21.8 MPaにおいて主転移温度曲線と副転移温度曲線が交差し，ゲル相は21.8 MPa以下の圧力では準安定相として存在し，21.8 MPa以上の圧力においてのみ安定相として存在することがわかった．指組み構造ゲル相に代表されるゲル相の多形現象はジパルミトイルホスファチジルコリン二分子膜のみに見出された．

Biochimica et Biophysica Acta (BBA) - Biomembranes


Dissociation Equilibrium between Uncharged and Charged Local Anesthetic Lidocaine in a Surface-Adsorbed Film

表面吸着膜中における非解離型—解離型局所麻酔薬の分配平衡を記述する熱力学関係式を導出し，非解離型局所麻酔薬リドカイン-塩酸混合水溶液の表面張力およびpHデータに適用した．計算結果の両者の表面密度から，両者の共存組成においては非解離型麻酔薬が解離型麻酔薬よりも多く存在し，吸着膜中における両者の分配挙動は3つの組成領域で特徴づけられることを示した．非解離型麻酔薬は解離型麻酔薬よりも細胞膜が提供する疎水性環境へ移行し易いことを結論づけた．


Barotropic Phase Transitions of 1-Palmitoyl-2-stearoylphosphatidylcholine Bilayer Membrane

非対称型リン脂質1-パルミトイル-2-ステアロイルホスファチジルコリン二分子膜の圧力誘起相転移を高圧光透過率測定および示差走査熱量測定により観測した．決定した相転移点に基づいて温度—圧力相図を構築し，3種類の対称型ホスファチジルコリン二分子膜の相図と比較した．1-パルミトイル-2-ステアロイルホスファチジルコリン二分子膜の相挙動は対称型ホスファチジルコリン二分子膜とは大きく異なり，指組み構造形成の臨界圧力は全ての脂質のうちで最も小さくなった．


Biological and physicochemical properties of gemini quaternary ammonium compounds in which the positions of a cross-linking sulfur in the spacer differ

Gemini型第四アンモニウム化合物(gemini QAC)の架橋構造のスルフィド結合の位置に着目した．メチレンスルフィド，エチレンジスルフィド，スルフィドがピリジン環に連結したgemini QACの3種のを合成し，抗菌活性，ヒト赤血球溶血毒性，界面活性能に関するスルフィド結合位置の影響について検討した．ピリジン環に連結したスルフィド結合を有する化合物は，抗菌力，溶血毒性，界面活性能いずれにおいても，他の2種薬剤よりも高いことが分かった．その強い抗菌力と溶血毒性は，スルフィド結合の共鳴効果によるアンモニウムカチオン電子密度の上昇，スルフィドがテトラメチレンスペーサ鎖両末端に存在することによるスペーサ鎖の高い伸縮性に起因した小さい分子占有面積，そして高い薬剤分子疎水性に寄与すると考え，スペーサ鎖の立体構造に影響し，スルフィドの位置がアンモニウムカチオン電子密度に影響を及ぼすと結論した．


Preferential partitioning of uncharged local anesthetics into the surface-adsorbed film

非解離型および解離型局所麻酔薬の競合表面吸着を調べるために3つの塩酸—非解離型局所麻酔薬(メピバカイン，ブピバカイン，ジブカイン)混合水溶液の表面張力とpHを全溶質濃度と麻酔薬組成の関数として測定した．非解離型および解離型麻酔薬の表面吸着膜中への分配量を熱力学的に計算した個々の表面密度で評価した．両者の共存領域では非解離型麻酔薬の優先的吸着が観測され，吸着膜への移行のし易さは麻酔薬分子疎水性に比例し，麻酔ポテンシーと相関した．本研究は生理的pHにおける表面活性分子は非解離型麻酔薬であることを明示した．


Anesthetic receptivity of lipid membranes with different bilayer phases

リン脂質二分子膜 の相状態と麻酔薬感受性

リン脂質二分子膜の相状態が局所麻酔薬の二分子膜中への分配に与える影響を調べるためにエステル結合型リン脂質ジパルミトイルホスファチジルコリンおよびエーテル結合型リン脂質ジヘキサデシルホスファチジルコリン二分子膜中へのジブカイン塩酸塩の取り込みを局所麻酔薬選択性電極法により調査した．決定された両二分子膜各相へのジブカインの分配係数値から，両二分子膜ともリップルゲル相および液晶相での分配係数は，ほぼ同等の値を示したが，ゲル相間ではラメラゲル相 < リップルゲル相 < 指組み構造ゲル相の順序となり差異が見られた．この差異は脂質分子の極性基当たりの占有面積とよい相関を示した．


Recognition for long-chian fatty acids by dipalmitoylphosphatidylcholine bilayer membranes

ジパルミトイルホスファチジルコリン二分子膜の長鎖脂肪酸認識

リン脂質ジパルミトイルホスファチジルコリン二分子膜相転移におよぼす4種類の長鎖脂肪酸(パルミチン酸，パルミトレイン酸，ステアリン酸，オレイン酸)と脂肪酸化合物(オレアミド)の影響を示差走査熱量法で調べた．熱力学的解析から得られた分配係数差と主転移エンタルピーから，飽和脂肪酸は脂質二分子膜ゲル相に強い親和性を有し，不飽和脂肪酸およびオレアミドは液晶相に強い親和性を有することがわかった．脂質二分子膜は高いリガンド感受性を有し，長鎖脂肪酸の鎖長や不飽和結合の有無を認識可能である．


Partition coefficients of the local anesthetic dibucaine into a lipid bilayer membrane determined with a dibucaine cation sensor

局所麻酔薬ジブカインカチオンセンサーを用いる脂質二分子膜へのジブカインの分配測定

レシチンベシクル膜中への局所麻酔薬ジブカインの分配をジブカインカチオン選択性電極を用いて調べ，その分配係数を決定した．分配係数はpH増加に伴い増大する傾向を示した．分配係数のpH依存性の解析から麻酔薬の解離型および非解離型それぞれの分配係数への分割を行い，レシチンベシクル膜中への非解離型麻酔薬の大きな分配を明確にした．生体膜中における麻酔薬分配計測への局所麻酔薬電極の有用性を示した．


Effect of myristic acid on the phase transition temperatures of dipalmitoylphosphatidylcholine bilayer membranes

ジパルミトイルホスファチジルコリン二分子膜の相転移温度におよぼすミリスチン酸の影響

リン脂質ジパルミトイルホスファチジルコリン二分子膜相転移におよぼす長鎖脂肪酸ミリスチン酸の影響を光透過法および示差走査熱量法を用いて調べ，結果を吸入麻酔薬イソフルランのものと比較した．相転移温度変化に熱力学関係式を適用し，脂質二分子膜における両リガンドの3種類の2相間分配係数差を計算した．ミリスチン酸は脂質二分子膜の流動性を下げ，ゲル相を安定化させるが，イソフルランは逆に二分子膜の流動性を上げ，ゲル相を不安定化させることを結論づけた．


局所麻酔薬によるリン脂質二分子膜 の相転移温度降下 -リン脂質極性基の効果-

局所麻酔薬によるリン脂質二分子膜 の相転移温度降下 -リン脂質極性基の効果-

ジパルミトイルホスファチジルコリンとジミリストイルホスファチジルエタノールアミン二分子膜の相転移温度におよぼす3種類の局所麻酔薬塩酸塩(テトラカイン，リドカインおよびプロカイン)の影響を示差走査熱量測定法と光透過法を用いて調べた．麻酔薬添加による相転移温度降下度から計算したホスファチジルエタノールアミン二分子膜に対する分配係数はホスファチジルコリン二分子膜のものより小さくなった．この麻酔薬分配の相違はホスファチジルエタノールアミン二分子膜中における極性基間の強い引力的相互作用に起因している．


非解離型および解離型局所麻酔薬の競合表面吸着

非解離型および解離型局所麻酔薬の競合表面吸着

3種類の非解離型局所麻酔薬(メピバカイン，ブピバカイン，ジブカイン)を塩酸でプロトン付加させ，それら麻酔薬の非解離型および解離型両者の競合表面吸着を非解離型麻酔薬-塩酸混合水溶液の表面張力とpH測定から調べた．混合組成が0.5以上の場合においてのみ両者が水溶液中で共存し，この組成領域では，非解離型麻酔薬の方が解離型麻酔薬より優先的に表面吸着膜へ移行した．また，表面吸着膜への移行のし易さは麻酔薬分子の有する疎水性に比例し，麻酔ポテンシーと相関していることがわかった．


Partition coefficients of charged and uncharged local anesthetics into dipalmitoylphosphatidylcholine bilayer membrane: estimation from pH dependence on the depression of phase transition temperatures

Effects of the local anesthetics, dibucaine, bupivacaine and lidocaine on the phase transition temperatures of dipalmitoylphosphatidylcholine (DPPC) bilayer membrane were studied by the optical method. We focus our attention on pH dependence of the depression of main transition and pretransition temperatures. The temperatures of both transitions of DPPC bilayer membrane were depressed by the addition of anesthetics; the higher the value of pH, the larger the depression of main transition temperature and/or pretransition temperature by anesthetics. By extending the colligative thermodynamic framework to the depression of main transition temperature by an anesthetic, we can estimate the differential partition coefficient, which is defined by the difference in partition coefficients of an anesthetic into the ripple gel and liquid crystal phases. The difference in partition coefficient between the lamellar and ripple gel phases can also be estimated from the depression of pretransition temperature. Since the differential partition coefficients include both contributions of the charged and uncharged anesthetics, we could estimate the partition coefficients of the charged and uncharged anesthetic into the membranes from the pH dependence of differential partition coefficients. The liquid crystalline membrane of DPPC bilayer was more receptive to the uncharged local anesthetics than the charged species. The partition coefficients of the charged and uncharged anesthetics into the liquid crystalline phase of DPPC bilayer membrane were 3540 and 249000 (for dibucaine), 1120 and 83900 (for bupivacaine), 256 and 11700 (for lidocaine), respectively. The transfer free energy of uncharged anesthetics from the aqueous phase to the liquid crystalline membrane was well correlated to the local anesthetic potency.

リン脂質ジパルミトイルホスファチジルコリン二分子膜の相転移温度におよぼす3種類の局所麻酔薬(ジブカイン，ブピバカイン，リドカイン)の影響を光学的手法で追跡した．脂質二分子膜の主転移および前転移温度は麻酔薬添加により降下し，pH増加につれて降下度は大きくなった．束一性の熱力学関係式より各局所麻酔薬の二相間分配係数を計算し，さらに分配係数のpH依存性から解離型および非解離型麻酔薬それぞれの分配係数を見積もった．脂質二分子膜は非解離型麻酔薬に強い感受性を示し，麻酔薬の取り込まれ易さは麻酔ポテンシーによく相関した．


Partitioning of uncharged local anesthetic benzocaine into model biomembranes

The partitioning of uncharged local anesthetic benzocaine (BzC) into molecular aggregates formed by cationic surfactant decylammonium chloride (DeAC) and phospholipid dipalmitoylphosphatidylcholine (DPPC) was studied from the surface tension and light transmittance measurements. The quantities concerning the partitioning of BzC, the compositions of BzC in the surface-adsorbed film and micelle and three kinds of differential partition coefficients corresponding to phase transitions of the DPPC bilayer membrane were evaluated from thermodynamic analysis of the experimental data. The surface-adsorbed film and micelle were more abundant in BzC than the aqueous solution and significantly large differential partition coefficients for the DPPC membranes were observed. The results clearly showed that the BzC molecules greatly partitioned into hydrophobic environments produced by surfactant-monolayer and phospholipid-bilayer membranes. The partitioning behavior of BzC was also compared with that of charged local anesthetic procaine hydrochloride (PC.HCl). It was shown that the PC.HCl molecule did not or hardly partition into such hydrophobic environments. The contrasting results of the partitioning between BzC and PC.HCl are attributable to the drastic decrease of hydrophilicity of BzC due to the lacking of ionic polar head group in comparison with PC.HCl.

陽イオン性界面活性剤塩化デシルアンモニウムとリン脂質ジパルミトイルホスファチジルコリンが形成する分子集合体中への非解離型局所麻酔薬ベンゾカインの分配を表面張力および光透過率測定から調べた．表面吸着膜とミセルは水溶液中に比べてベンゾカインに富み，脂質二分子膜に対する非常に大きな分配係数が得られた．上記ベンゾカインの分配挙動を構造が類似した解離型局所麻酔薬プロカイン塩酸塩のものと比較し，両者の分配差はイオン性極性基の有無による親水性の変化に起因することを示した．


Solubilization Study of Local Anesthetics into Sodium Dodecyl Sulfate Micelle Using Anesthetic Cation Selective Electrodes

The free concentrations of local anesthetic cations in equilibrium with sodium dodecyl sulfate (SDS) micelle which solubilized the anesthetic were determined by using ion-selective electrodes sensitive to local anesthetics, procaine (PC), lidocaine (LC), and mepivacaine (MC). Solubilizate distribution between water and SDS micelle was analyzed by means of the stepwise mass-action model. Association constant, K(1), was found to depend upon the anesthetic concentration, which decreased exponentially as the concentration of free anesthetic increased. Therefore, K(1) should include the interaction function φ(A) as K(1)=K(int)exp{-φ(A)} where K(int) is an intrinsic association constant. φ(A) is an increasing function of the anesthetic concentration, which means that occupation of a solubilization site by a local anesthetic cation makes sequential solubilization more difficult. The binding affinity of an anesthetic with SDS micelle increased in the following order PC<LC<MC.The critical micelle concentration (CMC) of mixed micelle was determined as a function of the concentration of free anesthetic. The CMC decreased with an increasing amount of anesthetics solubilized. All the anesthetic compositions in the micelle calculated thermodynamically from the CMC data were larger than the corresponding ones in the aqueous phase. Although the local anesthetics used here do not form micelles by themselves, the CMC vs composition curve can be regarded as a part of a micellar phase diagram showing the negative azeotropic behavior, which reflects the attractive interaction between the anionic surfactant micelle and the local anesthetic cation.

ドデシル硫酸ナトリウムミセルに可溶化した3種類の局所麻酔薬(プロカイン，リドカイン，メピバカイン)と水溶液中に平衡に存在するそれぞれの遊離局所麻酔薬カチオン濃度を局所麻酔薬選択性電極を用いて決定した．水中とミセル中の可溶化物質の分布を段階的質量作用モデルを用いて解析した．ミセルとの結合親和性はプロカイン<リドカイン<メピバカインの順序で増大した．さらに臨界ミセル濃度の組成依存性からミセル中の麻酔薬組成を求めた．ミセル形成の相図は負の共沸型となり，陰イオン性界面活性剤ミセルと陽イオン性局所麻酔薬カチオン間の強い引力的相互作用を示した．


Surface Tension Measurements with High Accuracy: Investigation of Accuracy for Automatic Measurements by the Drop Volume Method

表面張力の高精度測定のために必要と考えられる条件を純水の表面張力測定を通して探索した．純水の表面張力を測定誤差の原因となる様々な要因を設定した条件下で行い，それらの要因が表面張力値におよぼす影響を調べた．誤差要因の検討結果をもとにして，考えられる誤差要因を最小限に押さえた装置を製作した．この装置を用いて純水，界面活性剤水溶液および揮発性有機液体の表面張力を測定し，偏差が少なく且つ再現性ある表面張力の吸着平衡値を得ることができた．


Miscibility of Hydrocarbon and Fluorocarbon Surfactants in Adsorbed Film and Micelle

We investigated the miscibility of nonionic hydrocarbon and fluorocarbon surfactants in the adsorbed film and the micelle by surface tension measurements of the aqueous solution. The combination of tetraethyleneglycol monodecyl ether (C10E4) and tetraethyleneglycol mono-1,1,7-trihydrododecafluoroheptyl ether (FC7E4) was chosen because they have the same hydrophilic groups and about the same surface activity. The extent of nonideal mixing was estimated quantitatively in terms of the excess Gibbs energy in the adsorbed film g(H,E) and that in the micelle g(M,E). The excess area per adsorbed molecule, A(H,E), was also evaluated and discussed. The ionic hydrocarbon and fluorocarbon mixed surfactant systems, lithium dodecyl sulfate (LiDS)/lithium perfluorooctane sulfonate (LiFOS) and lithium tetradecyl sulfate (LiTS)/LiFOS systems are also investigated from the standpoint of excess Gibbs energy and excess area. It is also clearly shown that the regular solution approach does not fit in the systems that contain ionic species. Copyright 2001 Academic Press.

非イオン性の炭化水素系界面活性剤(テトラエチレングリコールモノデシルエーテル)とフッ素系界面活性剤(テトラエチレングリコールモノ-1,1,7-トリハイドロドデカフルオロヘプチルエーテル)の吸着膜とミセル中における混和性を表面張力測定から調べた．熱力学的解析から会合体形成の過剰熱力学量を求め，イオン性の炭化水素系界面活性剤-フッ素系界面活性剤混合系のものと比較し，両系の相違を議論した．さらに正則溶液理論はイオン性界面活性剤を含む系には適用できないことを示した．

Journal of Colloid and Interface Science


Interdigitation of Phospholipid Bilayer Membrane and Vesicle-Micelle Transition Induced by Local Anesthetic

局所麻酔薬が誘起するリン脂質二分子膜の指組構造化とベシクル-ミセル転移

局所麻酔薬テトラカイン塩酸塩存在下におけるジパルミトイルフォスファチジルコリン二分子膜の相挙動を示差走査熱量法で調べた．テトラカイン塩酸塩の添加により，二分子膜の指組構造化が観測された．二分子膜相挙動はテトラカイン塩酸塩濃度に依存し，テトラカイン塩酸塩を可溶化したベシクル領域，可溶化ベシクルと両者の混合ミセルの共存領域，混合ミセルのみの領域の3つの会合体領域に分けられた．さらに混合ミセル形成に関連した2種類の新しい転移を見出した．