Effect of phospholipid insertion on arrayed polydiacetylene biosensors

Micro-arrayed polydiacetylene (PDA) vesicles mixed with phospholipids on glass slides were prepared for label-free detection of Escherichia coli. When E. coli bound to its antibodies chemically attached to polydiacetylene, the fluorescence of the vesicles was dramatically increased. The insertion of dimyristoyl phosphatidylcholine (DMPC) in the vesicles drastically reduced the response time for the fluorescence changes. Vesicles with 20-30% DMPC provided optimal results for bacterial detection. Fourier transform infrared (FTIR) spectra analysis suggested that DMPC insertion decreased the strength of hydrogen bonding among the amide and carboxylic acid groups of the polydiacetylene vesicles. Reduced bonding strength resulted in less rigid structure of the polydiacetylene polymer, allowing more rapid detection upon molecular recognition.     

Effect of amphiphilic molecules upon chromatic transitions of polydiacetylene vesicles in aqueous solutions

Effect of amphiphilic molecules upon the chromatic transitions of polymerized 10,12-pentacosadiynoic acid (PCDA) vesicles in aqueous solutions was reported. The colorimetric response of polymerized PCDA vesicles for 1-pentanol is higher than that for ethanol due to more hydrophobic property of 1-pentanol. The colorimetric response of polymerized PCDA vesicles for sodium dodecyl sulfate (SDS) and Triton X-100 is lower than that for cetyltrimethylammonium bromide (CTAB). The strong ability of CTAB to induce chromatic transition of the vesicles is related to the positively charged headgroups of CTAB, which favors approach of CTAB to the negatively charged carboxylate groups at the vesicle surface. The insertion of alkyl chain of CTAB into the hydrophobic domain perturbs the conformation of the conjugated polymer backbone and induces color change of polydiacetylene vesicles. For a series of alkylamine hydrochloric salts, the longer the alkyl chain, the stronger the ability of alkylamine to induce chromatic transition of polydiacetylene vesicles.     

Assembly of polydiacetylene vesicles on solid substrates

A method to create assembly of polydiacetylene vesicles on solid substrates in a novel fashion is described. The formation of the assembly is based on electrostatic layer-by-layer deposition using negatively charged 10,12-pentacosadiynoic acid (PCDA) vesicles and polyelectrolyte polyethylenimine, or positively charged PCDA-2(')-aminoethylamide (PCDANH2) vesicles. This is an efficient method for preparing the chromatic sensor films of polydiacetylene vesicles after ultraviolet light irradiation. The strategy would be useful in the development of polydiacetylene-based chemosensors and biosensors.     

Color fingerprinting of proteins by calixarenes embedded in lipid/polydiacetylene vesicles

"Naked eye" color detection of proteins was achieved by embedding calixarene receptors within vesicles comprising phospholipids and the chromatic polymer polydiacetylene. Dramatic visible absorbance changes were induced through electrostatic interactions between the protein surface and the vesicle-incorporated hosts. The colorimetric responses could be induced by micromolar protein concentrations, and furthermore, specific protein fingerprints could be obtained by incorporating different receptors within the vesicles. Fluorescence and circular dichroism experiments confirmed the relationship between the colorimetric phenomena and protein docking on the surface of the chromatic vesicles. The colorimetric assay constitutes a generic platform for high-sensitivity detection of soluble proteins and for evaluation of protein surface charge distribution.     

聚联乙炔囊泡负载的Bola型两亲分子识别三聚氰胺过程中Chaotrope促进型显色机制及热力学

用聚联乙炔囊泡为载体,将bola型两亲分子1,12-二乳清酸十二胺盐(DDO)对三聚氰胺的分子识别作用用肉眼可见的颜色变化显示出来.通过比较不同碳链长度的聚联乙炔囊泡对分子识别过程的反映,发现二十三烷基-2.4-二炔酸(TCDA)囊泡的显色灵敏度较高.研究表明,TCDA肉眼可见的颜色变化来自于DDO与三聚氰胺多重氢键的形成以及溶液环境中水结构的变化.为了更好地理解显色机理,用差示扫描量热(DSC)仪详细研究了分子识别过程中聚联乙炔囊泡的相变行为及热力学参数.结果表明:TCDA囊泡和带有识别分子的DDO/TCDA囊泡在三聚氰胺存在下,相变温度Tm均向高温方向移动,并且,随三聚氰胺浓度的增加,Tm值逐渐增大直至囊泡瓦解;但是Tm值的变化没有与囊泡变色必然关联,仅仅DDO/TCDA囊泡具有变色现象,而且,只有当三聚氰胺的浓度超过分子识别氢键形成所需理论量时,肉眼才能可见明显的由蓝到红的颜色变化.为了理解溶液中过量的三聚氰胺对囊泡变色的作用,选用蔗糖和尿素作为典型的水结构促进剂和水结构破坏剂(chaotrope),详细研究了它们对聚联乙炔囊泡反映分子识别过程中相变温度的影响及显色规律.结果表明,过量的三聚氰胺在溶液中起到类似尿素水结构破坏剂的作用.这种作用和分子识别过程中多重氢键的形成对聚联乙炔囊泡的变色缺一不可.本研究首次揭示了由水结构破坏剂参与的聚联乙炔囊泡变色机理,有助于理解共轭聚合物热相变过程中的Hofmeister效应.     

Colorimetric detection and fingerprinting of bacteria by glass-supported lipid/polydiacetylene films

Glass-supported films of lipids and polydiacetylene were applied for visual detection and colorimetric fingerprinting of bacteria. The sensor films comprise polydiacetylene domains serving as the chromatic reporter interspersed within lipid monolayers that function as a biomimetic membrane platform. The detection schemes are based on either visible blue-red transitions or fluorescence transformations of polydiacetylene, induced by amphiphilic molecules secreted by proliferating bacteria. An important feature of the new film platform is the feasibility of either naked-eye detection of bacteria or color analysis using conventional scanners. Furthermore, we find that the degrees of bacterially induced color transformations depend both on the bacterial strains examined and the lipid compositions of the films. Accordingly, bacterial fingerprinting can be achieved through pattern recognition obtained by recording the chromatic transformations in an array of lipid/PDA films having different lipid components.     

Induced Color Change of Conjugated Polymeric Vesicles by Interfacial Catalysis of Phospholipase A2

A blue to red color change is induced on addition of phospholipase A₂ to modified PDA vesicles 1 (PDA=polydiacetylene). This bathochromic transition results from chemical modification of the vesicles by hydrolysis of the enzyme substrate embedded in the PDA matrix. Addition of a known phospholipase inhibitor or removal of Ca²⁺ ions suppresses the color change, which suggests the potential for applications in high‐throughput screening assays.     

Direct colorimetric study on the interaction of Escherichia coli with mannose in polydiacetylene Langmuir–Blodgett films

The membranes of polydiacetylene backbone decorated with mannose assembled by Langmuir–Blodgett technology can interact with Escherichia coli. The interactions lead to the color transition of the membranes which was readily visible to the naked eyes and could be quantified by visible absorption spectroscopy. To understand the mechanism of the chromatic transition, the affinochromism properties of polydiacetylene were examined by resonance Raman spectroscopy. The results demonstrated that the side chains of polymer backbone performed rearrangement, and the electronic structure in the polymer backbone changed from acetylene to butatriene form when the chromatic transformation from blue to red. The direct colorimetric detection by polydiacetylene membranes not only opens a new path for the use of these membranes in the area of biosensor development but also offers new possibilities for diagnostic applications and screening for binding ligand.     

Selective detection of catecholamines by synthetic receptors embedded in chromatic polydiacetylene vesicles

A new detection scheme for catecholamines was constructed through embedding synthetic receptors within vesicles comprising phospholipids and polydiacetylene. Fluorescence emission of the polydiacetylene was induced through specific interactions between the soluble ligands and the vesicle-incorporated hosts. The system demonstrated remarkable selectivity among structurally similar ligands and achieved much lower detection thresholds compared to that of other reported catecholamine sensors. The chromatic assembly provides a generic route for high sensitivity detection of ligand-receptor interactions.     

Size effect of polydiacetylene vesicles functionalized with glycolipids on their colorimetric detection ability

In this paper, the size effect of the polydiacetylene vesicles functionalized with glycolipids on their colorimetric detection ability has been studied. Polydiacetylene vesicles in which were incorporated glycolipids acted as a model system for the affinochromatic property. Visible color changes from blue to red could be observed to the naked eye owing to Con A binding to the sugar moiety and be detected quantitatively by the visible absorption spectrum. In the experiment, small and uniform vesicles were obtained after extrusion through membranes with different pore sizes. The morphology and mean size distribution of the extruded vesicles were studied by transmission electron microscopy (TEM) and dynamic light scattering (DLS), respectively. Our work shows that the smaller the vesicles are, the stronger is the effect, making the detection of Con A easier. The results may apply to the sensitivity enhancement of polydiacetylene biosensors for the recognition of other biomolecules.     

Effect of cholesterol or phospholipid incorporation on the chemical stability of the muramyldpeptide derivative B30-MDP in mixed vesicles

Application of the muramyldipeptide derivative B30-MDP to liposomal vaccines will aid in the development of improved high immunogenicity vaccines. To give full play to the effectiveness of B30-MDP as a liposomal vaccine, it is important to evaluate the effect of cholesterol, dimyristoylphosphatidylcholine (DMPC) or distearoylphosphatidylcholine (DSPC) incorporation on the chemical stability of B30-MDP and physicochemical properties of B30-MDP/lipid mixed vesicles from the view point of pharmaceutics.The observed degradation rate constants of B30-MDP by hydrolysis in B30-MDP/cholesterol mixed vesicles were increased with increasing concentration of cholesterol, however, those in B30-MDP/DMPC and B30-MDP/DSPC mixed vesicles were unchanged with increasing concentration of DMPC and DSPC. The degradation behavior of B30-MDP was then compared with physicochemical properties of B30-MDP/lipid mixed vesicles, such as membrane fluidity and particle size. It was apparent that the degradation of B30-MDP in B30-MDP/cholesterol mixed vesicles was influenced by the particle size, but not by the fluidity of the membranes. In the case of B30-MDP/phospholipid mixed vesicles, MDP/phospholipid mixed vesicles, the degradation of B30-MDP was not influenced by either the membranes' fluidity or the particle size of the mixed vesicles.It is considered that the degradation of B30-MDP in the mixed vesicles is dependent on the membrane state, and the addition of cholesterol to B30-MDP vesicle inhibits the mutual interaction of MDP regions, whereas the addition of phospholipids hardly influences the mutual interaction of MDP regions, possibly owing to phase separation between B30-MDP and phospholipids.     

以物理力将糖脂引入聚联乙炴基质脂的变色囊泡

囊泡具有与生物膜结构类似的封闭双分子层结构,具有模拟生物膜结构的突出优点,因此受到人们的极大重视．因为它具有胶体粒子的大小尺寸,其表面膜的性质易于控制,具有较大的输送客最以及生物相容性,因此,囊泡在药物的传输、基因治疗、癌症的化学治疗、分子识别、分子光学器件的组装、超细颗粒的制备、太阳能转化及反应性能的控制等领域都具有十分重要的意义．然而,在分子识别及药物输送过程中,由丁蟹池的稳定性差而严重影响其应用．聚合表面活性剂囊泡的出现较好地解决了其稳定性差的问题．在众多的可聚合的表面活性剂单体中,具有联…     

Colorimetric detection of oligonucleotides using a polydiacetylene vesicle sensor

A new system for the colorimetric detection of oligonucleotides was developed using polydiacetylene vesicles, which play the dual role of an indicator of color transition and an amplification tag. The results are of significance in understanding the mechanism of color transition of biological recognition in polydiacetylene systems and in designing new biosensors.     

STM studies of fusion of cholesterol suspensions and mixed 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC)/cholesterol vesicles onto a Au(111) electrode surface

Electrochemical scanning tunneling microscopy (EC-STM) has been applied to study the structure of the film formed by fusion of cholesterol suspensions and mixed dimyristoylphosphatidylcholine (DMPC)/cholesterol vesicles on a Au(111) electrode surface. It has been demonstrated that cholesterol molecules assemble at the gold support into several structures templated by the crystallography of the metal surface and involving flat or edge-on adsorbed molecules. Studies of the film formed by fusion of mixed DMPC/cholesterol vesicles revealed that ordered domains of either pure DMPC or pure cholesterol were formed. These results indicate that, at the metal surface, the molecules released by the rupture of a vesicle initially self-assemble into a well-ordered monolayer. The self-assembly is controlled by the hydrocarbon skeleton-metal surface interaction. In the case of mixed DMPC/cholesterol vesicles, the molecule-metal interactions induce segregation of the two components into single component domains. However, the molecule-metal interaction induced monolayer is a transient phenomenon. When more molecules accumulate at the surface, the molecule-molecule interactions dominate the assembly, and the monolayer is transformed into a bilayer.     

Vesicular polydiacetylene sensor for colorimetric signaling of bacterial pore-forming toxin

A vesicle-based polydiacetylene biosensor for colorimetric detection of bacterial pore-forming toxin streptolysin O (SLO) is reported. The sensor was constructed with three lipid constituents: glycine-terminated diacetylene lipid Gly-PCDA, cell membrane-mimicking component PC-DIYNE, and cholesterol (CHO), which serves as the bait molecule. UV irradiation led to photopolymerization of the diacetylene lipids that gave the material a blue appearance. Incubation of the vesicles with SLO from Streptococcus pyrogenes turned the vesicle solution red, and the color change was found to be correlated to SLO concentration. The optimal sensing performance was found with vesicles consisting of 71% Gly-PCDA, 25% CHO, and 4% PC-DIYNE, and a correlation relationship was obtained for 20 HU to 500 HU/mL, or 100 pM to 6.3 nM of SLO toxin. Transmission electron microscopy and dynamic light scattering was used for further characterization of the vesicular assemblies. Transmembrane pores (holes) with diameter around 30 nm were observed on the vesicle membranes, in particular on the peripheral of the membrane structures, suggesting pore formation by SLO toxin provides the driving force for the color change of the functional vesicles.     

联乙炔囊泡——一种基于分子组装的生物分子识别器件

有序排列的联乙炔分子,在紫外光照射下发生聚合。聚联乙炔（PDA）线性骨架的离域π电子,在可见光区产生π-π* 跃迁,显示特有的蓝色。聚联乙炔囊泡表面的分子探针（受体）在遇到可识别的生物大分子（如DNA,抗体-抗原和细菌等）的配体时,可使其颜色从蓝色转变为红色,有时能产生荧光。这种囊泡将检测与显示集为一体,是一种典型的生物分子器件,近年来已被用于很多物质的测定,其中包括病毒、细菌、亲脂性酶、抗菌肽类、哺乳动物肽类、离子、抗体、蛋白质和寡核苷酸等。此外,聚合囊泡还可以与金结合形成空心金球,可以作为一种三维纳米金的载体,对DNA的固定、识别和分离具有极为诱人的应用前景。通过对各种影响因素的研究和检测条件的改进,聚联乙炔生物传感器的灵敏度和选择性还能进一步提高,具有广阔的应用前景。     

Electrochemical and PM-IRRAS studies of the effect of cholesterol on the properties of the headgroup region of a DMPC bilayer supported at a Au(111) electrode

Polarization modulation infrared reflection absorption spectroscopy (PM-IRRAS) was employed to investigate the interaction of cholesterol with the headgroups of dimyristoylphosphatidycholine (DMPC) molecules under a static electric field. DMPC/cholesterol (7:3 molar ratio) mixtures form a bilayer on a Au(111) electrode surface by fusion and spreading of small unilamellar vesicles. PM-IRRAS experiments provided detailed information concerning the conformation and hydration of headgroups of DMPC bilayers in the presence and absence of 30% cholesterol. The presence of 30% cholesterol increases the space between the headgroups of DMPC molecules and hence increases the hydration of the DMPC/cholesterol mixed bilayer. The conformational state of the headgroups of DMPC molecules in the mixed bilayer is also significantly changed. The phosphate group is closer to the surface compared with the pure DMPC bilayer. The conformation of the -O-C-C-N moiety changes from gauche to trans in the presence of cholesterol.     

A colorimetric and fluorescent chemosensor for detection of Hg2+ using counterion exchange of cationic polydiacetylene

In this study, a colorimetric and fluorescent chemosensor for mercury ions (Hg²⁺) was developed. Cationic polydiacetylene (PDA) vesicles with a quaternary ammonium cation and iodide as a counterion show a blue-to-red color transition; the color change is accompanied by a fluorescence enhancement in selective response to Hg²⁺ ions because of a perturbation of the ene–yne conjugated backbone induced by counterion exchange. It allows for selective detection of Hg²⁺ with the naked eye and the sensor is used to determine Hg²⁺ concentrations in tap water samples.     

大分子拥挤环境中脂肪酸影响磷脂囊泡相变的差示扫描量热研究

脂肪酸诱导的磷脂膜的热力学行为对于认识细胞内复杂的机制有着重要意义,而前人在研究脂肪酸与磷脂膜相互作用时大都在稀溶液中进行;拥挤环境下脂肪酸诱导磷脂膜的相变行为还未见报道。本文以二肉豆蔻酰磷脂酰胆碱(DMPC)构建囊泡模型,采用差示扫描量热法系统地研究了在不同浓度、不同分子量的聚乙二醇(PEG)拥挤环境中不同结构的脂肪酸对DMPC磷脂囊泡相变的影响。研究结果表明,在拥挤环境中,PEG对纯的磷脂囊泡相变的影响与大分子的分子量和浓度相关。对于脂肪酸/磷脂囊泡(FA/DMPC),PEG的存在对囊泡相变产生显著影响。在所考察的分子量和浓度范围内,PEG使FA/DMPC囊泡相变增加。短链饱和脂肪酸、不饱和脂肪酸原本使DPMC囊泡相变降低,但PEG缩小了降低幅度,甚至导致相变增加。进一步的研究表明,在大多数情况下,PEG对FA/DMPC的相变具有协作增强效应,且其影响均与大分子的分子量和浓度相关。另外,随着PEG浓度的升高,磷脂囊泡的协同单位数逐渐降低,表明拥挤环境会影响磷脂双分子层的均一性,使协同发生相变的分子数降低。本文的研究表明,大分子拥挤环境能够对扰动的磷脂双分子层起到一定的修复作用,这一现象在生物膜相关领域不可忽视。     

Thin wetting films from aqueous solutions of surfactants and phospholipid dispersions

The microscopic thin wetting film method was used to study the stability of wetting films from aqueous solution of surfactants and phospholipid dispersions on a solid surface. In the case of tetradecyltrimethylammonium bromide (C(14)TAB) films the experimental data for the receding contact angle, film lifetime, surface potential at the vapor/solution and solution/silica interface were used to analyze the stability of the studied films. It is shown that with increasing C(14)TAB concentration charge reversal occurs at both (vapor/solution and solution/silica) interfaces, which affects the thin-film stability. The spontaneous rupture of the thin aqueous film was interpreted in terms of the earlier proposed heterocoagulation mechanism. The presence of the mixed cationic/anionic surfactants was found to lower contact angles and suppresses the thin aqueous film rupture, thus inducing longer film lifetime, as compared to the pure amine system. In the case of mixed surfactants hetero-coagulation could arise through the formation of ionic surfactant complexes. The influence of the melting phase-transition temperature T(c) of the dimyristoylphosphatiddylcholine (DMPC) on the stability of thin films from dispersions of DMPC small unilamellar vesicles on a silica surface was studied by measuring the film lifetime and the TPC expansion rate. The stability of thin wetting films formed from dispersions of DMPC small unilamellar vesicles was investigated by the microinterferometric method. The formation of wetting films from diluted dispersions of DMPC multilamellar vesicles was studied in the temperature range 25-32 degrees C. The stability of thin film of lipid vesicles was explained on the basis of hydrophobic interactions. The results obtained show that the stability of wetting films from aqueous solutions of single cationic and mixed cationic-anionic surfactants has electrostatic origin, whereas the stability of the phospholipid film is due to hydrophobic interaction.