BODIPY-based fluorescent liposomes with sesquiterpene lactone trilobolide.

Like thapsigargin, which is undergoing clinical trials, trilobolide is a natural product with promising anticancer and anti-inflammatory properties. Similar to thapsigargin, it has limited aqueous solubility that strongly reduces its potential medicinal applications. The targeted delivery of hydrophobic drugs can be achieved using liposome-based carriers. Therefore, we designed a traceable liposomal drug delivery system for trilobolide. The fluorescent green-emitting dye BODIPY, cholesterol and trilobolide were used to create construct 6. The liposomes were composed of dipalmitoyl-3-trimethylammoniumpropane and phosphatidylethanolamine. The whole system was characterized by atomic force microscopy, the average size of the liposomes was 150 nm in width and 30 nm in height. We evaluated the biological activity of construct 6 and its liposomal formulation, both of which showed immunomodulatory properties in primary rat macrophages. The uptake and intracellular distribution of construct 6 and its liposomal formulation was monitored by means of live-cell fluorescence microscopy in two cancer cell lines. The encapsulation of construct 6 into the liposomes improved the drug distribution in cancer cells and was followed by cell death. This new liposomal trilobolide derivative not only retains the biological properties of pure trilobolide, but also enhances the bioavailability, and thus has potential for the use in theranostic applications.

Effect of ionic liquids on the interaction between liposomes and common wastewater pollutants investigated by capillary electrophoresis.

The effect of three phosphonium and imidazolium ionic liquids (ILs) on the interaction between liposomes and common pharmaceuticals found in wastewaters was studied. The liposomes comprised zwitterionic phosphatidyl choline and negatively charged phosphatidyl glycerol. A set of common cationic, anionic, and neutral compounds with varying chemical composition and unique structures were included in the study. The electrophoretic mobilities of the analytes were determined using conventional capillary electrophoresis (CE), using CE under reversed electroosmotic flow mobility conditions, and in the presence of ILs in the background electrolyte (BGE) solution by electrokinetic chromatography (EKC). In order to evaluate the impact of ILs on the interaction between the compounds and the liposomes, EKC was performed with liposome dispersions, with and without ILs. The retention factors of the compounds were calculated using BGEs including liposome dispersions with and without ILs. Two phosphonium based ILs, namely tributyl(tetradecyl)phosphonium chloride ([P14444]Cl) and octyltributylphosphonium chloride ([P8444]Cl), were chosen due to their long alkyl chains and their low aggregation concentrations. Another IL, i.e. 1-ethyl-3-methylimidazolium acetate ([emim][OAc]), was chosen based on our previous study, which suggests that it has a minimal or even nonexistent effect on liposomes at the used concentrations. The results indicate that the studied ILs have an effect on the interactions between wastewater compounds and liposomes, but the effect is highly dependent on the concentration of the IL and on the IL alkyl chain lengths. Most of the ILs hindered the interactions between the liposomes and the compounds, indicating strong interaction between ILs and liposomes. In addition, the nature of the studied compounds themselves affected the interactions.

Impact of amphiphilic biomass-dissolving ionic liquids on biological cells and liposomes.

The toxicity of some promising biomass-dissolving amidinium-, imidazolium-, and phosphonium-based ionic liquids (ILs), toward two different cell lines, human corneal epithelial cells and Escherichia coli bacterial cells, was investigated. In addition, dynamic light scattering (DLS) and ζ potential measurements were used to study the effect of the ILs on the size and surface charge of some model liposomes. Capillary electrophoresis (CE) was used for determination of the electrophoretic mobilities of the liposomes and for determination of the critical micelle concentration (cmc) of the ILs. The toxicity of the phosphonium ILs was highly dependent on the longest linear chain of the IL, due to increasing hydrophobicity, with the long-chain phosphonium ILs being toxic while the shorter-chain versions were significantly less toxic or not toxic at all. Amidinium and imidazolium ILs showed no significant effect on the cells, within the concentration range used. Moreover, the more hydrophobic ILs were found to have a major effect on the surface charges and size distributions of the model liposomes, which can lead to disruption of the lipid bilayer. This indicates that the cytotoxicity is at least to some extent dependent on direct interactions between ILs and the biomembrane.

Plasmonic nanodiamonds: targeted core-shell type nanoparticles for cancer cell thermoablation.

Targeted biocompatible nanostructures with controlled plasmonic and morphological parameters are promising materials for cancer treatment based on selective thermal ablation of cells. Here, core-shell plasmonic nanodiamonds consisting of a silica-encapsulated diamond nanocrystal coated in a gold shell are designed and synthesized. The architecture of particles is analyzed and confirmed in detail using electron tomography. The particles are biocompatibilized using a PEG polymer terminated with bioorthogonally reactive alkyne groups. Azide-modified transferrin is attached to these particles, and their high colloidal stability and successful targeting to cancer cells overexpressing the transferrin receptor are demonstrated. The particles are nontoxic to the cells and they are readily internalized upon binding to the transferrin receptor. The high plasmonic cross section of the particles in the near-infrared region is utilized to quantitatively ablate the cancer cells with a short, one-minute irradiation by a pulse 750-nm laser.

Determination of the distribution constants of aromatic compounds and steroids in biphasic micellar phosphonium ionic liquid/aqueous buffer systems by capillary electrokinetic chromatography.

The distribution constants of some analytes, closely connected to the petrochemical industry, between an aqueous phase and a phosphonium ionic liquid phase, were determined by ionic liquid micellar electrokinetic chromatography (MEKC). The phosphonium ionic liquids studied were the water-soluble tributyl(tetradecyl)phosphonium with chloride or acetate as the counter ion. The retention factors were calculated and used for determination of the distribution constants. For calculating the retention factors the electrophoretic mobilities of the ionic liquids were required, thus, we adopted the iterative process, based on a homologous series of alkyl benzoates. Calculation of the distribution constants required information on the phase-ratio of the systems. For this the critical micelle concentrations (CMC) of the ionic liquids were needed. The CMCs were calculated using a method based on PeakMaster simulations, using the electrophoretic mobilities of system peaks. The resulting distribution constants for the neutral analytes between the ionic liquid and the aqueous (buffer) phase were compared with octanol-water partitioning coefficients. The results indicate that there are other factors affecting the distribution of analytes between phases, than just simple hydrophobic interactions.

Capillary electromigration techniques for studying interactions between analytes and lipid dispersions.

CE has matured into a well-known and much used separation technique, with applications covering more or less all types of samples. EKC, which originally was developed for the separation of uncharged compounds, is still today under much development, with main focus on finding the perfect or ideal carriers (pseudo-stationary phase) for a broad range of analytes. In this review, the use of lipid dispersions as pseudostationary phases in EKC performed in capillaries and microchips, in addition to CE frontal analysis and partial filling EKC using lipid dispersions is discussed. Various types of lipid dispersions including liposomes, PEG-stabilized aggregates, proteoliposomes, lipid-based nanoparticles, and commercial lipid emulsions are described. The purpose of the review is to give the reader an overview of how EKC, CE frontal analysis, and partial filling EKC, have been applied to the study of interactions between analytes and lipid membranes.

Comparison of lipid sinks in sequestering common intoxicating drugs.

Intravenous lipid emulsion is a recommended treatment for local anesthetic intoxication. The lipid sink theory hypothesizes that the mechanism behind the lipid treatment is the entrapment of toxic drugs in plasma, preventing them from reaching target receptors. Lipid sink treatment has also been used as a last refuge treatment for severe tricyclic antidepressant intoxication with seemingly beneficial results. We selected three drugs, i.e. amiodarone, ketamine, and amitriptyline, that can cause severe intoxication and compared their interactions with two commercial fat emulsions (Intralipid® and ClinOleic®) and one synthetic liposome (80:20 mol% phosphatidylcholine/phosphatidylglycerol) dispersion. The interaction studies were carried out by capillary electrokinetic chromatography and the retention factors and distributions constants of the drugs were calculated. The results demonstrate that there is stronger interaction between the drugs and the synthetic liposome dispersion than with the commercial emulsions.

Phospholipids covalently attached to silica particles as stationary phase in nano-liquid chromatography.

Silica particles were covalently modified with phospholipids and used as packing material for nano-liquid chromatography (nano-LC). This modification involved aminopropylsilylation of the raw silica particles using 3-(aminopropyl)-triethoxysilane, covalent binding of glutaraldehyde molecules to the aminopropylsilylated particles, and finally covalent binding of different phospholipid vesicles containing primary amino groups to the iminoaldehyde silica particles. Capillaries with an inner diameter of 100µm were packed with phospholipid-coated silica particles using a slurry packing method. The packed capillaries were tested in nano-LC with UV-detection for the separation of acidic, neutral, and basic model analytes. The effect of the buffer ion on the retention factor of the analytes was evaluated using buffer solutions with constant ionic strength and pH. In addition, the effect of the volume of methanol in the mobile phase was studied. The calculated distribution coefficients (logK(D)) of the model compounds were in agreement with those reported in the literature. A good correlation between logK(D) values and octanol/water partitioning coefficients (P(o/w)) for neutral hydrophobic analytes was obtained proving the applicability of the method for predicting partitioning of the compounds with the biomembranes.

In vitro and in vivo entrapment of bupivacaine by lipid dispersions.

Intravenous lipid emulsion is recommended as treatment for local anesthetic intoxication based on the hypothesis that the lipophilic drug is entrapped by the lipid phase created in plasma. We compared a 15.6 mM 80/20 mol% phosphatidyl choline (PC)/phosphatidyl glycerol (PG)-based liposome dispersion with the commercially available Intralipid® emulsion in a pig model of local anesthetic intoxication. Bupivacaine-lipid interactions were studied by electrokinetic capillary chromatography. Multilamellar vesicles were used in the first in vivo experiment series. This series was interrupted when the liposome dispersion was discovered to cause cardiovascular collapse. The toxicity was decreased by an optimized sonication of the 50% diluted liposome dispersion (7.8 mM). Twenty anesthetized pigs were then infused with either sonicated PC/PG liposome dispersion or Intralipid®, following infusion of a toxic dose of bupivacaine which decreased the mean arterial pressure by 50% from baseline. Bupivacaine concentrations were quantified in blood samples using liquid chromatography/mass spectrometry. No significant difference in the context-sensitive plasma half-life of bupivacaine was detected (p=0.932). After 30 min of lipid infusion, the bupivacaine concentration was 8.2±1.5 mg/L in the PC/PG group and 7.8±1.8 mg/L in the Intralipid® group, with no difference between groups (p=0.591). No difference in hemodynamic recovery was detected between groups (p > 0.05).

In vitro capturing of various lipophilic illicit drugs by lipid dispersions. An electrokinetic capillary chromatography and fluorescence polarization study.

Fatal drug overdoses are a cause for concern all over the world. We present here a lipid-based formulation which has a strong affinity for some common illicit street drugs and can be used in vivo as a lipid 'sink'. In this study, the in vitro interactions of nine lipophilic drugs and three lipid dispersions were determined by electrokinetic capillary chromatography and fluorescence polarization. Two lipid dispersions, zwitterionic 1-palmitoyl-2-oleyl-sn-glycero-3-phosphocholine (POPC) and an anionic mixture of POPC and 1-palmitoyl-2-oleyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (POPG) were tested and compared with a commercial lipid dispersion Intralipid(®), which has been successfully used for resuscitation of patients in cases of anesthetic overdoses. The interactions between dispersions and the drugs were quantified by means of retention factors and distribution constants, which makes the results highly comparable to those obtained from any other formulation of lipids. The results demonstrate a stronger interaction between the drugs and an artificial liposome dispersion than with the commercial Intralipid dispersion. The liposome dispersion composed of POPC and POPG functions as a lipid 'sink' for efficient entrapment of various lipophilic drugs.

Liposomes for entrapping local anesthetics: a liposome electrokinetic chromatographic study.

Bupivacaine is a lipophilic, long-acting, amide class local anesthetic commonly used in clinical practice to provide local anesthesia during surgical procedures. Several cases of accidental overdose with cardiac arrest and death have been reported since bupivacaine was introduced to human use. Recent case reports have suggested that Intralipid (Fresenius Kabi) is an effective therapy for cardiac toxicity from high systemic concentrations of, e.g. bupivacaine, even though the mechanism behind the interaction is not fully clear yet. Our long-term aim is to develop a sensitive, efficient, and non-harmful lipid-based formulation to specifically trap harmful substances in vivo. In this study, the in vitro interaction of local anesthetics (bupivacaine, prilocaine, and lidocaine) with Intralipid or lipid vesicles containing phosphatidylglycerol, phosphatidylcholine, cardiolipin, cholesterol, and N-palmitoyl-D-erythro-sphingosine (ceramide) was determined by liposome electrokinetic chromatography. The interactions were evaluated by calculating the retention factors and distribution constants. Atomic force microscopy measurements were carried out to confirm that the interaction mechanism was solely due to interactions between the analytes and the moving pseudostationary phase and not by interactions with a stationary lipid phase adsorbed to the fused-silica wall. The heterogeneity of the liposomes was also studied by atomic force microscopy. The liposome electrokinetic chromatography results demonstrate that there is higher interaction between the drugs and negatively charged liposome dispersion than with the commercial Intralipid dispersion.

Interaction of a commercial lipid dispersion and local anesthetics in human plasma: implications for drug trapping by "lipid-sinks".

Interactions between Intralipid dispersion and local anesthetics (bupivacaine, prilocaine, and lidocaine) were investigated. The amount of bupivacaine (the most cardiotoxic analyte of the local anesthetics studied) entrapped in Intralipid in the presence of plasma was studied using an off-line filtration and solid phase extraction method combined with capillary zone electrophoresis for quantification of free unbound bupivacaine. To confirm interactions between the analytes and Intralipid at lower concentrations, direct injection mass spectrometry was used. The use of immobilized Intralipid chromatography-atmospheric pressure ionization-ion trap mass spectrometry in the study of interactions between drugs and Intralipid dispersion is demonstrated. Finally, interactions between Intralipid dispersion and local anesthetics were investigated by electrokinetic capillary chromatography. The electrophoretic mobility of the Intralipid dispersed phase was calculated using the iterative procedure and a homologous series of alkyl phenyl benzoates (C(1)-C(6)), and the retention factors for the analytes were determined.

Marker compounds for the determination of retention factors in EKC.

EKC and its sub-techniques, such as MEKC and microemulsion EKC, have attracted wide interest in recent years. Investigations on this topic have covered several analytical applications, but attention has also been paid more and more to basic studies. This review provides an overview of the different approaches to calculating retention factors, which express the ratio of the amount of sample component in the pseudostationary and mobile phases. Special attention is given to the selection of markers for the determination of the electrophoretic mobility or migration time of a marker describing the behavior of the pseudostationary phase in EKC. Introduction of a hydrophobic marker is by far the most common approach, but the use of a homologous series of compounds is also quite popular. In addition, other possible approaches found in the literature will be described.

System peaks in micellar electrophoresis: I. Utilization of system peaks for determination of critical micelle concentration.

A new way to determine the critical micelle concentration (CMC) based on the mobilities of system peaks is presented. A general approach for the CMC determination is based on the change of the slope or on finding the inflection point in the plot of a physical property of solution as a function of surfactant concentration. The determination of CMC by system peaks in CE utilizes a "jump" instead of a continuous change in the measured quantity. This phenomenon was predicted by the program PeakMaster, which was modified for simulation of micellar systems. The simulation of the steep change in mobilities of the anionic system peaks showing the CMC value was verified experimentally in a set of measurements, where the concentration of the surfactant was varied while the ionic strength was kept constant. The experimental work fully proved our model. A comparative electric current measurement was carried out. The proposed method seems to offer easier CMC determination as compared to the standard methods.

Interpolymer complexes based on the core/shell micelles. interaction of polystyrene-block-poly(methacrylic acid) micelles with linear poly(2-vinylpyridine) in 1,4-dioxane water mixtures and in aqueous media.

The size and structural changes of nanoparticles formed after the addition of poly(2-vinylpyridine), PVP, to block copolymer micelles of polystyrene-block-poly(methacrylic acid), PS-PMA, were studied by light scattering and atomic force microscopy. Due to the strong hydrogen bonding between PVP and PMA segments, complex structures based on the core/shell micelles form in mixed selective solvents. As proven by a combination of light scattering and atomic force microscopy, individual PS-PMA micelles are "glued" together by PVP chains. The dialysis against solvents with a high content of water results in transient increase in polydispersity and turbidity of originally clear solutions. However, the precipitated polymer material dissolves in basic buffers and stable soluble nanoparticles reform in aqueous media. The behavior of their solutions was studied in a broad pH range by light scattering, atomic force microscopy and capillary zone electrophoresis.

Eigenmobilities in background electrolytes for CZE. V. Intensity (amplitudes) of system peaks.

We present a mathematical model of CZE based on the concept of eigenmobilities - the eigenvalues of matrix M tied to the linearized governing equations of electromigration, and the spectral decomposition of matrix M into matrices of amplitudes P(j). Any peak in an electropherogram, regardless of whether it is an analyte peak or a system peak (system zone), is matched with its matrix P(j). This enables calculation of the peak parameters, such as the transfer ratio and the molar conductivity detection response (which give the indirect detection signal and the conductivity detection signal, respectively), when the initial disturbance caused by the injection of the sample is known. We also introduce new quantities, such as the generalized transfer ratio and the conductivity response of system zones, and show how the amplitude (intensity, area) of the analyte peaks and the system peaks can be calculated. We offer a free software, PeakMaster (http://www.natur.cuni.cz/gas), which yields this information in a user-friendly way.

Linear free energy relationship as a tool for characterization of three teicoplanin-based chiral stationary phases under various mobile phase compositions.

Teicoplanin, teicoplanin aglycon, and methylated teicoplanin aglycon chiral stationary phases (CSPs) have been compared on the basis of the regression coefficients calculated from the linear free energy relationship (LFER) equation. The parameters have been obtained from the measurements of a set of 34 structurally diverse solutes. Influence of mobile phase composition - variation of methanol (MeOH) content - on the participation of different interactions types in the retention mechanism has been evaluated. Retention of the various interaction forces in analytes differs with both the CSP and the mobile phase composition. Hydrophobic interactions play a major role in mobile phases for high buffer contents. The more hydrophobic the CSP, the more important are they in the retention mechanism. With increase of MeOH contents in the mobile phase the major role in the interaction mechanism is shifted to more polar forces in which basicity and dipolarity/polarizability dominate. Although the LFER model does not address chiral aspects, we have attempted to explore the importance of the individual interactions in chiral discrimination of amino acids and their N-tert-butyloxycarbonyl derivatives.

Comparative study of three teicoplanin-based chiral stationary phases using the linear free energy relationship model.

Teicoplanin (T) is a macrocyclic glycopeptide that is highly effective as a chiral selector for enantiomeric separations. In this study, we used three teicoplanin-based chiral stationary phases (CSPs) - native teicoplanin, teicoplanin aglycon (TAG) and recently synthesized methylated teicoplanin aglycon (MTAG). In order to examine the importance of various interaction types in the chiral recognition mechanism the three related CSPs were evaluated and compared using a linear free energy relationship (LFER). The capacity factors of 19 widely different solutes, with known solvation parameters, were determined on each of the columns under the same mobile phase conditions used for the chiral separations. The regression coefficients obtained revealed the magnitude of the contribution of individual interaction types to the retention on the compared columns under those specific experimental conditions. Statistically derived standardized regression coefficients were used to evaluate the contribution of individual molecular interactions within one stationary phase. It has been concluded that intermolecular interactions of the hydrophobic type significantly contribute to retention on all the CSPs studied here. Other retention increasing factors are n- and pi-electron interactions and dipole-dipole or dipole-induced dipole ones, while hydrogen donating or accepting interactions are more predominant with the mobile phase than with the stationary phases. However, these types of interactions are not equally significant for all the CSPs studied.