Effect of Ester Moiety on Structural Properties of Binary Mixed Monolayers of Alpha-Tocopherol Derivatives with DPPC

Phospholipid membranes are ubiquitous components of cells involved in physiological processes; thus, knowledge regarding their interactions with other molecules, including tocopherol ester derivatives, is of great importance. The surface pressure–area isotherms of pure α-tocopherol (Toc) and its derivatives (oxalate (OT), malonate (MT), succinate (ST), and carbo analog (CT)) were studied in Langmuir monolayers in order to evaluate phase formation, compressibility, packing, and ordering. The isotherms and compressibility results indicate that, under pressure, the ester derivatives and CT are able to form two-dimensional liquid-condensed (LC) ordered structures with collapse pressures ranging from 27 mN/m for CT to 44 mN/m for OT. Next, the effect of length of ester moiety on the surface behavior of DPPC/Toc derivatives’ binary monolayers at air–water interface was investigated. The average molecular area, elastic modulus, compressibility, and miscibility were calculated as a function of molar fraction of derivatives. Increasing the presence of Toc derivatives in DPPC monolayer induces expansion of isotherms, increased monolayer elasticity, interrupted packing, and lowered ordering in monolayer, leading to its fluidization. Decreasing collapse pressure with increasing molar ratio of derivatives indicates on the miscibility of Toc esters in DPPC monolayer. The interactions between components were analyzed using additivity rule and thermodynamic calculations of excess and total Gibbs energy of mixing. Calculated excess area and Gibbs energy indicated repulsion between components, confirming their partial mixing. In summary, the mechanism of the observed phenomena is mainly connected with interactions of ionized carboxyl groups of ester moieties with DPPC headgroup moieties where formed conformations perturb alignment of acyl chains, resulting in increasing mean area per molecule, leading to disordering and fluidization of mixed monolayer.     

不同固醇与DPPC二元体系的液态有序相

应用同步辐射X射线衍射和差示扫描量热法研究了由不同结构的固醇(胆固醇、脱氢胆固醇、豆固醇、谷固醇、麦角固醇以及固醇核)和二棕榈酰磷脂酰胆碱(DPPC)二元体系形成的液态有序相. 研究表明, 胆固醇比植物固醇(豆固醇和谷固醇)和真菌固醇(麦角固醇)能更有效地与DPPC形成液态有序相(Lo); 有胆固醇或者脱氢胆固醇参与的液态有序相能够在较宽的温度范围内保持稳定, 而由植物固醇和真菌固醇参与的液态有序相对温度有较强的依赖性, 在DPPC主相变温度附近有明显的热致相变过程, 因此这一液态有序相应该进一步区分为Loβ和Loα相. 研究结果有助于阐明固醇尾链在液态有序相以及脂筏中的作用, 也有助于理解在进化过程中动物细胞膜为何选择胆固醇作为主要固醇.     

二次谐波光谱和布鲁斯特角显微镜研究二棕榈酰磷脂酰胆碱和维生素B2的相互作用

二棕榈酰磷脂酰胆碱（DPPC）分子在气液界面上形成的Langmuir膜是一种重要的生物膜模拟体系,其手性结构及其与外来物质的相互作用一直是相关学科研究的前沿问题。维生素B₂（VB₂）是人体中一种重要的营养物质,它在代谢障碍引起的脂质沉积性类疾病中有大量的实例应用,经常在一些特殊的临床症状中有出乎意料的治疗奇效。目前,VB₂如何参与到膜上生物事件的过程和细胞乃至生命的作用过程中的研究报道较少,特别是VB₂分子与磷脂分子靶标的立体相互作用,其可能发生的手性分子识别现象会在许多生物事件中起着关键作用。综合二次谐波-线二色光谱（SHG-LD）、Langmuir膜天平和布鲁斯特角显微镜(BAM)技术初步研究了VB₂和DPPC分子在气液界面上的相互作用,分别从气液界面上介观水平和宏观水平上互补表征脂质分子在气液界面上的分子骨架自组装的结构。压缩等温线发现纯水界面L-DPPC和D-DPPC液态扩展相/液态凝聚相(LE/LC)共存阶段的膜压几乎不变,race-DPPC的共存相膜压区域稍微缩短,VB₂水溶液界面上race-DPPC的LE/LC共存相消失。此外,弹性模量研究表明VB₂分子可以提高L-DPPC单分子层膜的弹性模量,但降低D-DPPC和race-DPPC单层膜的弹性模量。结合SHG-LD研究发现,在膜压13 mN·m-1下,L-DPPC在纯水和VB₂水溶液界面上表面手性过量值（DCE）保持不变。与纯水界面相比较,D-DPPC在VB₂水溶液上DCE值出现反转,而race-DPPC的DCE值则不随亚相改变而变化。相同膜压下,BAM观察到单一手性相互作用使得L-DPPC和D-DPPC在纯水界面上各自组装成不同枝臂弯曲方向的手性三叶草微畴(microdomain)。VB₂诱导D-DPPC微畴,使其直径增大1～2倍。同时,VB₂也诱导了race-DPPC单层膜上近似圆形状的微畴伸展,并长出了三条有曲率的枝臂。对此可以解释为VB₂降低了非单一手性相互作用的能量,使得race-DPPC出现手性相分离。与此同时,VB₂也诱导了race-DPPC单层膜微畴的手性结构发生变化。该研究有助于理解VB₂调节磷脂膜横向组织结构的分子机理,在细胞膜界面发生的过程中,脂层单层的二维特性和生物分子之间的相互作用可能决定了生物分子的亲和力。     

Interaction of dipalmitoyl phosphatidylcholine (DPPC) liposomes and insulin

Insulin, a peptide that has been used for decades in the treatment of diabetes, has well-defined properties and delivery requirements. Liposomes, which are lipid bilayer vesicles, have gained increasing attention as drug carriers which reduce the toxicity and increase the pharmacological activity of various drugs. The molecular interaction between (uncharged lipid) dipalmitoyl phosphatidylcholine (DPPC) liposomes and insulin has been characterized by using Fourier transform infrared spectroscopy (FTIR) and X-ray diffraction. The characteristic protein absorption band peaks, Amide I (at about 1660?cm^?1) and Amide II band (at about 1546?cm^?1) are potentially reduced in the liposome insulin complex. Wide-angle x-ray scattering measurements showed that the association of insulin with DPPC lipid of liposomes still maintains the characteristic DPPC diffraction peaks with almost no change in relative intensities or change in peak positions. The absence of any shift in protein peak positions after insulin being associated with DPPC liposomes indicates that insulin is successfully forming complex with DPPC liposomes with possibly no pronounced alterations in the structure of insulin molecule.     

Phase behaviour of polymer-grafted DPPC membranes for drug delivery systems design

Summary DPPC dispersions containing DPPE with attached PEG of molecular masses 350, 2000 and 5000 were investigated by DSC in order to determine their phase behaviour and potential use as drug delivery systems. In comparison with previously obtained ESR data, DSC provided a definition of the lipid composition and temperature at which the vesicles are in a liquid crystalline phase. For DPPC DPPE-PEG 350 the composition range is at molar fractions 0<_&khgr;PEG350<0.5.For DPPC DPPE-PEG 2000 the range of applicability is 0<_{&khgr;PEG2000}<0.07 and for DPPC/DPPE-PEG 5000 system it is 0<_{&khgr;PEG5000}<0.05.     

Effect of tetracaine on model and erythrocyte membranes by DSC and EPR

Summary DSC and EPR experiments were performed on human erythrocyte membranes and DPPC vesicles in order to study the effect of the anaesthetic drug tetracaine on structure and dynamics of the lipid region. Experiments using spin label technique showed that tetracaine induced fluidity changes of the lipid region in the environment of the fatty acid probe molecules incorporated into the membranes in the vicinity of the lipid-water interface. Similarly to EPR observations, DSC measurements reported decrease of the main melting and the pretransition temperature in comparison to control DPPC vesicles, which is the sign of destabilisation of the structure in the head group region of the lipids. Similar effect was observed in the case of erythrocytes where the protein conformation was also controlled in the presence of drug. A separated membrane melting with well distinguished membrane protein phase transition was found that was affected significantly by tetracaine. These results suggest that tetracaine is able to modify not only the internal dynamics of erythrocyte membranes and produce destabilisation of the lipid structure, but the protein system as well. These might lead to further damage of the biological functions.     

Calorimetric and spin-label ESR studies of PEG:2000-DPPE containing DPPC/lyso-PPC mixtures

The thermotropic behavior of dipalmitoylphosphatidylcholine (DPPC) multibilayers containing up to 10 mol% of lyso-palmitoylphosphatidylcholine (lyso-PPC) with and without low content of poly(ethylene glycol:2000)-grafted dipalmitoylphosphatidylethanolamine (PEG:2000-DPPE) has been studied by high sensitivity differential scanning calorimetry (DSC) and electron spin resonance (ESR) using the spin probe di-tert-butyl-nitroxide (DTBN). The three lipids, dispersed in buffer at appropriate concentrations, form thermosensitive liposomes used as site-specific drug-delivery systems. Without polymer–lipids, the DPPC main transition temperature is downshifted of 1.2–1.3 °C at the highest lyso-PPC content. The molar enthalpy and the cooperative unit of the DPPC main transition first decrease rapidly, then more slowly and finally slightly increase with lyso-PPC content. Moreover, in the mixed dispersions, the membrane fluidity increases at any temperature. The addition up to 5 mol% of PEG:2000-DPPE to DPPC/10 mol% lyso-PPC mixtures does not affect neither the thermotropic phase behavior nor the transition cooperativity and the fluidity of the dispersions.     

Surface forces in foam films from DPPC and lung surfactant phospholipid fraction

Surface properties of foam films formed from aqueous dispersions of dipalmitoyl-phosphatidylcholine (DPPC) and from solutions of a phospholipid fraction of lung surfactant (TPL) are studied employing the foam film method. Experiments are carried out within a wide range of NaCl concentrations (C_el) and the ranges of C_el determining formation of common films (CF), common black films (CBF) and bilayer Newton black films (NBF) are found. The thickness (h) of the CF and CBF decreases with the increase of C_el until the critical electrolyte concentrations (C_{el, cr}) is reached. The determined C_{el, cr} that characterize the transition to NBF show that C_{el, cr} of the TPL films is an order of magnitude higher than that of the DPPC films. The measured h of the TPL films is higher than that of the DPPC films in the whole C_el range. Besides, only the h(C_el) curve of the DPPC films outlines a metastable C_el range where both CF and NBF are obtained. Both the h(C_el) curves and the direct measurements of the disjoining pressure isotherms of the DPPC films (Π(h) isotherms) demonstrate the role of electrostatic repulsive forces for the stability of the phospholipid films The obtained results are compared with the DLVO theory equations and the evaluated potentials of the diffuse electric layer φ₀  20 mV for the DPPC films and φ₀  100 mV for TPL films show the strong effect of the charged phospholipids in the TPL mixture on the electric properties at the film interfaces.     

Lung surfactant dysfunction in tuberculosis: effect of mycobacterial tubercular lipids on dipalmitoylphosphatidylcholine surface activity

In pulmonary tuberculosis, Mycobacterium tuberculosis bacteria reside in the alveoli and are in close proximity with the alveolar surfactant. Mycolic acid in its free form and as cord factor, constitute the major lipids of the mycobacterial cell wall. They can detach from the bacteria easily and are known to be moderately surface active. We hypothesize that these surface-active mycobacterial cell wall lipids could interact with the pulmonary surfactant and result in lung surfactant dysfunction. In this study, the major phospholipid of the lung surfactant, dipalmitoylphosphatidylcholine (DPPC) and binary mixtures of DPPC:phosphatidylglycerol (PG) in 9:1 and 7:3 ratios were modelled as lung surfactant monolayers and the inhibitory potential of mycolic acid and cord factor on the surface activity of DPPC and DPPC:PG mixtures was evaluated using Langmuir monolayers. The mycobacterial lipids caused common profile changes in all the isotherms: increase in minimum surface tension, compressibility and percentage area change required for change in surface tension from 30 to 10 mN/m. Higher minimum surface tension values were achieved in the presence of mycolic acid (18.2 ± 0.7 mN/m) and cord factor (13.28 ± 1.2 mN/m) as compared to 0 mN/m, achieved by pure DPPC film. Similarly higher values of compressibility (0.375 ± 0.005 m/mN for mycolic acid:DPPC and 0.197 ± 0.003 m/mN for cord factor:DPPC monolayers) were obtained in presence of mycolic acid and cord factor. Thus, mycolic acid and cord factor were said to be inhibitory towards lung surfactant phospholipids. Higher surface tension and compressibility values in presence of tubercular lipids are suggestive of an unstable and fluid surfactant film, which will fail to achieve low surface tensions and can contribute to alveolar collapse in patients suffering from pulmonary tuberculosis. In conclusion a biophysical inhibition of lung surfactant may play a role in the pathogenesis of tuberculosis and may serve as a target for the development of new drug loaded surfactants for this condition.     

The interaction of a peptide with a scrambled hydrophobic/hydrophilic sequence (Pro-Asp-Ala-Asp-Ala-His-Ala-His-Ala-His-Ala-Ala-Ala-His-Gly) (PADH) with DPPC model membranes: a DSC study

Depending on their hydrophobicity, peptides can interact differently with lipid membranes inducing dramatic modifications into their host systems. In the present paper, the interaction of a synthetic peptide with a scrambled hydrophobic/hydrophilic sequence (Pro-Asp-Ala-Asp-Ala-His-Ala-His-Ala-His-Ala-Ala-Ala-His-Gly) (PADH) with 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) model membranes has been investigated by differential scanning calorimetry (DSC), adopting three different experimental approaches. In the first, the peptide is forced to be included into the hydrocarbon region of the lipid bilayer, by codissolving it with the lipid giving rise to mixed multilamellar vesicles–peptide systems; in the second, this system is passed through an extruder, thus producing large unilamellar vesicles–peptide systems; in the third, it is allowed to interact with the external surface of the membrane.

The whole of the DSC results obtained have shown that the incorporation of the peptide into the lipid bilayer by means of the first method induces a decrease in the enthalpy of the gel–liquid crystal transition of the membrane and a shift of the transition to the lower temperatures, thus resembling, in spite of its prevalently hydrophilic nature, the behavior of transbilayer hydrophobic peptides. The extrusion of these systems creates unilamellar vesicles free of peptides but of smaller size as evidenced by the decreased cooperativity of the transition. The peptide, added externally to the DPPC model membrane, has no effect on the phase behavior of the bilayer.

These findings suggest that the effect of the interaction of scrambled hydrophobic/hydrophilic peptides into lipid bilayers strongly affects the thermotropic behavior of the host membrane depending on the preparation method of the lipid/peptide systems. The whole of the results obtained in the present paper can be useful in approaching studies of bioactive peptides/lipids systems.