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Imidazolium-based dicationic ILs (DILs) presenting antimicrobial activity and relatively low toxicity are highly desirable and are envisioned for use in live tissue to prevent bacterial or fungal infections. In this context, we present here DILs with dicarboxylate anions [Cn(MIM)2[Cn(MIM)2][CO2-(CH2)mCO2], in which n = 4, 6, 8, and 10, and m = 0, 1, 2, 3, 4, and 5. The results showed that DILs with an alkyl chain spacer of ten carbons were active against yeasts and the bacterial strains tested. However, most of the DILs were cytotoxic and toxic at 1 mM. By contrast, DILs with alkyl chains possessing less than ten carbons were active against some specific Candidas and bacteria (mainly S. aureus), and they showed moderate cytotoxicity. The best activity against Gram-positive bacteria was observed for [C4(MIM)2][Pim] toward MRSA. For the DILs described herein, their level of toxicity against C. elegans was lower than that of most of the mono- and dicationic IL analogs with other anions. Our results showed that the presence of carboxylate anions reduces the toxicity of DILs compared to DILs containing halide anions, which is particularly significant to the means of designing biologically active compounds in antimicrobial formulations.

Automation in solid state NMR (ssNMR) requires appropriate hardware, from rotor loading mechanisms over highly stable rf-transmitters and probe circuitry to automatic tuning and matching capabilities including automatic magic angle adjustment for ssNMR probes. While these hardware capabilities are highly desirable and are, to various degrees, provided by manufacturers, we focus herein on automating experiment setup using radio frequency (rf) fields, which are key parameters in solid state NMR experiments. Specifically, these include spinlock fields during cross polarization (CP), or rf-fields for homo- or heteronuclear spin recoupling or decoupling. Often, these fields have specific relationships to the magic angle spinning (MAS) frequency. Relying on a well-maintained spectrometer, the experiment setup shifts from traditionally required optimization of rf-power values for each element of an experiment sequence to automatically setting all parameters correctly without any need for optimization. The proposed approach allows executing an experiment by reading its rf-amplitude requirements based on the actual MAS rotation frequency just before starting data acquisition, while all other hardware-related parameters are automatically provided through global tables and scripts. Under modest MAS frequencies, no further rf-power optimization is required while providing optimal sensitivity of better than 90% of the optimal signal to noise. Any optional parameter optimization relates only to adjusting rf-nutation frequencies to the requirements of the sample and the sample rotation frequency rather than the spectrometer hardware. Fast MAS CP experiments with MAS frequencies above 40 kHz require a semi-automated setup by optimizing Hartmann-Hahn (HH) matched rf-fields that are synchronously varied relative to the MAS-frequency. This allows for a significant reduction of setup steps by up to one order of magnitude for such experiments, avoiding the traditional grid search for optimal CPMAS conditions. The approach presented here can also be applied to decoupling or recoupling sequences, requiring rotor synchronized rf-fields, reducing the setup to a few steps addressing the spin system's properties rather than the spectrometer hardware. Our approach permits automating all basic solid state NMR experiments for high throughput analytical tasks., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

A new comprehensive analytical approach based on single-particle inductively coupled plasma-sector field mass spectrometry (spICP-SFMS) and electrical asymmetric-flow field-flow-fractionation combined with multi-angle light scattering detection (EAF4-MALS) has been examined for the characterization of galactosamine-terminated poly(N-hydroxyethyl acrylamide)-coated gold nanorods (GNRs) in two different degrees of polymerization (DP) by tuning the feed ratio (short: DP 35; long: DP 60). spICP-SFMS provided information on the particle number concentration, size and size distribution of the GNRs, and was found to be useful as an orthogonal method for fast characterization of GNRs. Glycoconjugated GNRs were separated and characterized via EAF4-MALS in terms of their size and charge and compared to the bare GNRs. In contrast to spICP-SFMS, EAF4-MALS was also able of providing an estimate of the thickness of the glycopolymer coating on the GNRs surface.

Lipid-based nanoparticles for RNA delivery (LNP-RNA) are revolutionizing the nanomedicine field, with one approved gene therapy formulation and two approved vaccines against COVID-19, as well as multiple ongoing clinical trials. As for other innovative nanopharmaceuticals (NPhs), the advancement of robust methods to assess their quality and safety profiles-in line with regulatory needs-is critical for facilitating their development and clinical translation. Asymmetric-flow field-flow fractionation coupled to multiple online optical detectors (MD-AF4) is considered a very versatile and robust approach for the physical characterisation of nanocarriers, and has been used successfully for measuring particle size, polydispersity and physical stability of lipid-based systems, including liposomes and solid lipid nanoparticles. However, the unique core structure of LNP-RNA, composed of ionizable lipids electrostatically complexed with RNA, and the relatively labile lipid-monolayer coating, is more prone to destabilization during focusing in MD-AF4 than previously characterised nanoparticles, resulting in particle aggregation and sample loss. Hence characterisation of LNP-RNA by MD-AF4 needs significant adaptation of the methods developed for liposomes. To improve the performance of MD-AF4 applied to LNP-RNA in a systematic and comprehensive manner, we have explored the use of the frit-inlet channel where, differently from the standard AF4 channel, the particles are relaxed hydrodynamically as they are injected. The absence of a focusing step minimizes contact between the particle and the membrane, reducing artefacts (e.g. sample loss, particle aggregation). Separation in a frit-inlet channel enables satisfactory reproducibility and acceptable sample recovery in the commercially available MD-AF4 instruments. In addition to slice-by-slice measurements of particle size, MD-AF4 also allows to determine particle concentration and the particle size distribution, demonstrating enhanced versatility beyond standard sizing measurements., (Copyright © 2021 The Author(s). Published by Elsevier B.V. All rights reserved.)

Background: Space use by animals is determined by the interplay between movement and the environment, and is thus mediated by habitat selection, biotic interactions and intrinsic factors of moving individuals. These processes ultimately determine home range size, but their relative contributions and dynamic nature remain less explored. We investigated the role of habitat selection, movement unrelated to habitat selection and intrinsic factors related to sex in driving space use and home range size in Iberian ibex, Capra pyrenaica . We used GPS collars to track ibex across the year in two different geographical areas of Sierra Nevada, Spain, and measured habitat variables related to forage and roost availability., Results: By using integrated step selection analysis (iSSA), we show that habitat selection was important to explain space use by ibex. As a consequence, movement was constrained by habitat selection, as observed displacement rate was shorter than expected under null selection. Selection-independent movement, selection strength and resource availability were important drivers of seasonal home range size. Both displacement rate and directional persistence had a positive relationship with home range size while accounting for habitat selection, suggesting that individual characteristics and state may also affect home range size. Ibex living at higher altitudes, where resource availability shows stronger altitudinal gradients across the year, had larger home ranges. Home range size was larger in spring and autumn, when ibex ascend and descend back, and smaller in summer and winter, when resources are more stable. Therefore, home range size decreased with resource availability. Finally, males had larger home ranges than females, which might be explained by differences in body size and reproductive behaviour., Conclusions: Movement, selection strength, resource availability and intrinsic factors related to sex determined home range size of Iberian ibex. Our results highlight the need to integrate and account for process dependencies, here the interdependence of movement and habitat selection, to understand how animals use space. This study contributes to understand how movement links environmental and geographical space use and determines home range behaviour in large herbivores., Competing Interests: All the procedures involving animal capturing and tagging were performed under the current ethical guidelines and approved by the pertinent authorities.Not applicable.The authors declare that they have no competing interests.Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Flow field-flow fractionation is a powerful method for the analysis of nanoparticle size distributions, but its widespread use has been hampered by large analyte losses, especially of metal nanoparticles. Here, we report on the colloidal mechanisms underlying the losses. We systematically studied gold nanoparticles (AuNPs) during asymmetrical flow field-flow fractionation (AF4) by systematic variation of the particle properties and the eluent composition. Recoveries of AuNPs (core diameter 12 nm) stabilized by citrate or polyethylene glycol (PEG) at different ionic strengths were determined. We used online UV-vis detection and off-line elementary analysis to follow particle losses during full analysis runs, runs without cross-flow, and runs with parts of the instrument bypassed. The combination allowed us to calculate relative and absolute analyte losses at different stages of the analytic protocol. We found different loss mechanisms depending on the ligand. Citrate-stabilized particles degraded during analysis and suffered large losses (up to 74%). PEG-stabilized particles had smaller relative losses at moderate ionic strengths (1-20%) that depended on PEG length. Long PEGs at higher ionic strengths (≥5 mM) caused particle loss due to bridging adsorption at the membrane. Bulk agglomeration was not a relevant loss mechanism at low ionic strengths ≤5 mM for any of the studied particles. An unexpectedly large fraction of particles was lost at tubing and other internal surfaces. We propose that the colloidal mechanisms observed here are relevant loss mechanisms in many particle analysis protocols and discuss strategies to avoid them.

Introduction: Low Carbohydrate High Protein diet represents a popular strategy to achieve weight loss., Objective: The aim of this study was to characterize effects of low carbohydrate, high protein diet (LCHP) on atherosclerotic plaque development in brachiocephalic artery (BCA) in apoE/LDLR-/- mice and to elucidate mechanisms of proatherogenic effects of LCHP diet., Materials and Methods: Atherosclerosis plaques in brachiocephalic artery (BCA) as well as in aortic roots, lipoprotein profile, inflammation biomarkers, expression of SREBP-1 in the liver as well as mortality were analyzed in Control diet (AIN-93G) or LCHP (Low Carbohydrate High Protein) diet fed mice., Results: Area of atherosclerotic plaques in aortic roots or BCA from LCHP diet fed mice was substantially increased as compared to mice fed control diet and was characterized by increased lipids and cholesterol contents (ORO staining, FT-IR analysis), increased macrophage infiltration (MOMA-2) and activity of MMPs (zymography). Pro-atherogenic phenotype of LCHP fed apoE/LDLR-/- mice was associated with increased plasma total cholesterol concentration, and in LDL and VLDL fractions, increased TG contents in VLDL, and a modest increase in plasma urea. LCHP diet increased SCD-1 index, activated SREBP-1 transcription factor in the liver and triggered acute phase response as evidence by an increased plasma concentration of haptoglobin, CRP or AGP. Finally, in long-term experiment survival of apoE/LDLR-/- mice fed LCHP diet was substantially reduced as compared to their counterparts fed control diet suggesting overall detrimental effects of LCHP diet on health., Conclusions: The pro-atherogenic effect of LCHP diet in apoE/LDLR-/- mice is associated with profound increase in LDL and VLDL cholesterol, VLDL triglicerides, liver SREBP-1 upregulation, and systemic inflammation.

A new FFF method is presented which combines asymmetrical flow-FFF (AF4) and electrical FFF (ElFFF) in one channel to electrical asymmetrical flow-FFF (EAF4) to overcome the restrictions of pure ElFFF. It allows for measuring electrophoretic mobility (μ) as a function of size. The method provides an absolute value and does not require calibration. Results of μ for two particle standards are in good agreement with values determined by phase analysis light scattering (PALS). There is no requirement for low ionic strength carriers with EAF4. This overcomes one of the main limitations of ElFFF, making it feasible to measure proteins under physiological conditions. EAF4 has the capability to determine μ for individual populations which are resolved into separate peaks. This is demonstrated for a mixture of three polystyrene latex particles with different sizes as well as for the monomer and dimer of BSA and an antibody. The experimental setup consists of an AF4 channel with added electrodes; one is placed beneath the frit at the bottom wall and the other covers the inside of the upper channel plate. This design minimizes contamination from the electrolysis reactions by keeping the particles distant from the electrodes. In addition the applied voltage range is low (1.5-5 V), which reduces the quantity of gaseous electrolysis products below a threshold that interferes with the laminar flow profile or detector signals. Besides measuring μ, the method can be useful to improve the separation between sample components compared to pure flow-FFF. For two proteins (BSA and a monoclonal antibody), enhanced resolution of the monomer and dimer is achieved by applying an electric field.

Post-infectious sequelea such as Guillain Barré syndrome (GBS), reactive arthritis (RA), and inflammatory bowel disease (IBD) may arise as a consequence of acute Campylobacter-enteritis (AE). However, reliable seroprevalence data of Campylobacter-associated sequelae has not been established. The objectives of this study were, first, to identify the most specific and sensitive test antigen in an optimized ELISA assay for diagnosing a previous Campylobacter-infection and, second, to compare the prevalence of anti-Campylobacter antibodies in cohorts of healthy blood donors (BD), AE, GBS, RA, and IBD patients with antibodies against known GBS, RA and IBD triggering pathogens. Optimized ELISAs of single and combined Campylobacter-proteins OMP18 and P39 as antigens were prepared and sera from AE, GBS, RA and IBD patients and BD were tested for Campylobcter-specific IgA and IgG antibodies. The results were compared with MIKROGEN™-recomLine Campylobacter IgA/IgG and whole cell lysate-immunoblot. Antibodies specific for Helicobacter pylori, Mycoplasma pneumoniae, Yersinia enterocolitica, and Borrelia afzelii were tested with commercial immunoblots. ROC plot analysis revealed AUC maxima in the combination of OMP18 and P39 for IgA and in the P39-antigen for IgG. As a result, 34-49 % GBS cases, 44-62 % RA cases and 23-40 % IBD cases were associated with Campylobacter-infection. These data show that Campylobcater-seropositivity in these patient groups is significantly higher than other triggering pathogens suggesting that it plays an important role in development of GBS and RA, and supports the hypothesis that recurrent acute campylobacteriosis triggers IBD.

Background: Toxigenic Clostridium difficile strains are known as the most common infectious cause of antibiotic-associated intestinal disease and nosocomial diarrhoea. The increased incidence of hypervirulent strains gives rise to worldwide concern. In particular, courses with multiple recurrences are observed in the presence of immunosuppression., Methods: In this retrospective controlled observational study we aimed to determine immunosuppression as an independent risk factor for symptomatic CDI and to identify characteristics and differences of immunocompromised patients with respect to disease severity, disease progression, intestinal manifestations, recurrence rates and other factors. We compared symptoms and clinical features of 55 immunosuppressed patients with confirmed CDI with those of 50 patients without immunosuppressive medication (mean age 59.3 years ± 16.2 vs. 69.2 years ± 15.0) who were treated at the Departments of Internal Medicine I and IV of the University Hospital Halle (Saale), Germany, between 2006 and 2009. Multivariate analysis using binary logistic regression was performed for a control group of 105 patients without CDI. In this group, there were 62 patients without evidence of immunosuppression and 43 immunosuppressed patients (mean age 66.9 years ± 12.4 vs. 56.0 years ± 13.7)., Results: The clinical courses of the two groups differed considerably. Immunosuppressed patients were significantly more frequently colonised with Clostridium difficile without clinically detectable manifestation or only mild clinical symptoms not requiring therapy (22 vs. 2 %, p = 0.003), while there were similar numbers of moderate (46 vs. 52 %, p = 0.503) but less severe CDI cases (27 vs. 40 %, p = 0.167). Relapses were observed more frequently in the group of immunosuppressed patients (15 vs. 6 %, p = 0.153). Multivariate analysis using logistic regression identified immunosuppression as an independent risk factor for CDI (OR = 2.75), in addition to prior antibiotic therapy (OR = 10.15) and PPI intake (OR = 2.93)., Conclusion: We conclude that immunosuppression has to be regarded as an independent risk factor for CDI. Immunosuppressive treatment increases the risk of colonisation and infection with Clostridium difficile and leads to a higher relapse rate in patients with CDI., (© Georg Thieme Verlag KG Stuttgart · New York.)

A new system design and setup are proposed for the combined use of asymmetrical flow field-flow fractionation (AF4) and hollow-fiber flow field-flow fractionation (HF5) within the same instrumentation. To this purpose, three innovations are presented: (a) a new flow control scheme where focusing flow rates are measured in real time allowing to adjust the flow rate ratio as desired; (b) a new HF5 channel design consisting of two sets of ferrule, gasket and cap nut used to mount the fiber inside a tube. This design provides a mechanism for effective and straightforward sealing of the fiber; (c) a new AF4 channel design with only two fluid connections on the upper plate. Only one pump is needed to deliver the necessary flow rates. In the focusing/relaxation step the two parts of the focusing flow and a bypass flow flushing the detectors are created with two splits of the flow from the pump. In the elution mode the cross-flow is measured and controlled with a flow controller device. This leads to reduced pressure pulsations in the channel and improves signal to noise ratio in the detectors. Experimental results of the separation of bovine serum albumin (BSA) and of a mix of four proteins demonstrate a significant improvement in the HF5 separation performance, in terms of efficiency, resolution, and run-to-run reproducibility compared to what has been reported in the literature. Separation performance in HF5 mode is shown to be comparable to the performance in AF4 mode using a channel with two connections in the upper plate., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Differential scanning calorimetry (DSC) was used to study the miscibility of phosphatidic acids (PAs) with phosphatidylethanolamines (PEs) as a function of chain length (n = 14, 16) and degree of ionization of PAs at pH 4, pH 7, and pH 12. Phase diagrams were constructed using temperature data for onset and end of the phase transition obtained from the direct simulation of the heat-capacity curves. The phase diagrams were analyzed by simulations of the coexistence curves utilizing a four-parameter regular solution model. For PA-PE mixtures, the non-ideality parameters are a function of composition indicating non-symmetric non-ideal mixing behavior. At pH 7, where the PA component is negatively charged, the systems DMPA:DMPE and DPPA:DPPE have positive non-ideality parameters ρ (1) in both phases, indicating a preferred aggregation of like molecules. In contrast, DMPA:DPPE and DPPA:DMPE mixtures had negative ρ (1) values. Measurements at pH 4 showed that mixed pair formation is favored when PA is protonated. At pH 12 where PA is doubly charged, highly positive ρ (l1) parameters are obtained for the liquid-crystalline phase except for the system DPPA:DPPE (ρ (1) < 0). This indicates clustering of like molecules and possibly domain formation in the liquid-crystalline phase. DPPA:DMPE at pH 12 even shows a miscibility gap in the liquid-crystalline phase. Obviously, despite the presence of doubly charged PA a fluid-fluid immiscibility is induced.

Synthesis and applications of new functional nanoparticles are topics of increasing interest in many fields of nanotechnology. Chemical modifications of inorganic nanoparticles are often necessary to improve their features as spectroscopic tracers or chemical sensors, and to increase water solubility and biocompatibility for applications in nano-biotechnology. Analysis and characterization of structured nanoparticles are then key steps for their synthesis optimization and final quality control. Many properties of structured nanoparticles are size-dependent. Particle size distribution analysis then provides fundamental analytical information. Asymmetrical flow field-flow fractionation (AF4) with multi-angle light scattering (MALS) detection is able to size-separate and to characterize nanosized analytes in dispersion. In this work we focus on the central role of AF4-MALS to analyze and characterize different types of structured nanoparticles that are finding increasing applications in nano-biotechnology and nanomedicine: polymer-coated gold nanoparticles, fluorescent silica nanoparticles, and quantum dots. AF4 not only size-fractionated these nanoparticles and measured their hydrodynamic radius (r(h)) distribution but it also separated them from the unbound, relatively low-M(r) components of the nanoparticle structures which were still present in the sample solution. On-line MALS detection on real-time gave the gyration radius (r(g)) distribution of the fractionated nanoparticles. Additional information on nanoparticle morphology was then obtained from the r(h)/r(g) index. Stability of the nanoparticle dispersions was finally investigated. Aggregation of the fluorescent silica nanoparticles was found to depend on the concentration at which they were dispersed. Partial release of the polymeric coating from water-soluble QDs was found when shear stress was induced by increasing flowrates during fractionation.

Alterations of lipoproteins (LPs) and related lipid levels in blood serum are correlated to the risk of coronary artery disease (CAD). Fast, possibly automated methods to obtain complete, multi-parametric LP profiles are therefore welcome to be developed for routine, clinical analysis practice. In this work, asymmetrical flow field-flow fractionation (AF4) with on-line, dual post-fractionation reaction detection (PFRD) is applied to develop a method for single-run, simultaneous quantification of cholesterol (CHOL) and triglycerides (TGs) in each fractionated LP class. The enzymatic reagents used for the post-fractionation reaction are available as commercial kits for certified, standard clinical protocols for the analysis of CHOL and TGs in serum. Using CHOL and glycerol as reference standards, a new procedure is applied to optimize the experimental conditions for PFRD-based, quantitative analysis. Upon optimized PFRD and AF4 conditions, results obtained for the determination of total CHOL (TC), TGs, HDL-cholesterol (HDL-C), and LDL-cholesterol (LDL-C) in a set of serum samples from healthy donors are found in agreement with the values provided by a clinical laboratory. The intra-day and inter-day precisions of the method were found always lower than 10% (CV). When the method was applied to serum samples from patients affected by sepsis, differences in CHOL and TG profiles between patients and healthy donors were observed.

Numerous studies have shown increasing item reliabilities as an effect of the item position in personality scales. Traditionally, these context effects are analyzed based on item-total correlations. This approach neglects that trends in item reliabilities can be caused either by an increase in true score variance or by a decrease in error variance. This article presents the Confirmatory Analysis of Item Reliability Trends (CAIRT) that allows estimating both trends separately within a structural equation modeling framework. Results of a simulation study prove the CAIRT method to provide reliable and independent parameter estimates; the power exceeds the analysis of item-total correlations. We present an empirical application to self- and peer ratings collected in an Internet-based experiment. Results show that reliability trends are caused by increasing true score variance in self-ratings and by decreasing error variance in peer ratings.

Background: Demonstration of recanalized coronary artery is mostly done by angiographic techniques. Early bedside demonstration of reperfusion after thrombolysis by transthoracic echocardiography (TTE) has important implications in the subsequent risk stratification and timing of coronary interventions., Methods: In this study, 12 patients with acute anterior myocardial infarction who received thrombolytic therapy were studied. Echocardiographic Doppler evaluation of left main coronary artery, proximal left anterior descending coronary artery (LAD), and proximal left circumflex coronary artery were studied before, during, and after thrombolytic therapy. Coronary flow in these arterial segments was assessed both by color flow and velocity measurements. These results were compared with coronary angiographic studies performed within 30 minutes to 48 hours of thrombolysis., Results: Blood flow in left main coronary artery, LAD, and left circumflex coronary artery could be assessed in 9 patients. There was no demonstrable flow in LAD in 6 patients before thrombolysis. In 7 patients flow could be demonstrated in LAD after thrombolysis within 15 minutes to 6 hours. The peak flow velocity in LAD at a localized area of turbulence postthrombolysis varied from 1.8 to 4.5 m/s. One patient showed mosaic color flow in left main coronary artery with a peak velocity of 1.9 m/s before thrombolysis that improved to a laminar flow with a peak velocity of 1.0 m/s after thrombolysis. Two patients showed normal flow in proximal LAD, but no flow in mid-LAD. Two patients did not show any flow in LAD even after 12 hours of thrombolysis. There was good correlation of site of critical narrowing in LAD by TTE with coronary angiography in 6 patients. In 3 patients absent flow in mid-LAD by TTE correlated with total occlusion of either proximal (one patient) or mid-LAD (two patients)., Conclusions: Demonstration of recanalized infarct-related left coronary artery soon after thrombolytic therapy is feasible. Locating the actual site of critical narrowing at bedside by TTE has important implications in the subsequent treatment of patients with acute anterior wall myocardial infarction.

Demonstration of coronary collaterals has been mostly done by coronary angiographic techniques. Coronary collaterals have been demonstrated by transthoracic echocardiography for patients with anomalous origin of coronary arteries from pulmonary artery. Indirect assessment of collaterals to an infarct-related artery has been done using myocardial contrast echocardiography. We describe in this article 3 patients with coronary artery disease in whom coronary collaterals were demonstrated by transthoracic echocardiography before angiography. To our knowledge, coronary collateral demonstration by transthoracic echocardiography before angiography in patients with coronary artery disease has not been previously reported.

Recently, colloidal dispersions made from mixtures of solid and liquid lipids were described to combine controlled release characteristics with higher drug loading capacities than solid lipid nanoparticles (SLN). It has been proposed that these nanostructured lipid carriers (NLC) are composed of oily droplets which solubilize the drug and which are embedded in a solid lipid matrix. The structures of SLN and NLC based on glyceryl behenate and medium chain triglycerides were characterized by photon correlation spectroscopy (PCS) and laser diffraction (LD), field-flow fractionation (FFF) with multi-angle light scattering detection (MALS), and cryo transmission electron microscopy (cryo TEM). PCS indicates that SLN and NLC differ from a nanoemulsion with respect to Brownian motion due to asymmetric particle shapes. Non-spherical particles, in case of SLN and NLC, lead to higher polydispersity indices compared to the nanoemulsion. In FFF, the nanodroplets elute much earlier than SLN- and NLC-platelets although their PCS and LD data show similar particle sizes. In TEM platelet (for SLN), oil loaded platelet ("nanospoons"; for NLC) and droplet (for nanoemulsion) structures were observed. In contrast to literature reports, the investigated SLN appear as thin platelets. NLC are found to be lipid platelets with oil spots sticking on the surface. Very short diffusion pathways in platelets, increased water-lipid interfaces and low drug incorporation in crystalline lipids are the drawback of SLN and NLC compared to conventional nanoemulsions.

The miscibilities of phosphatidic acids (PAs) and phosphatidylcholines (PCs) with different chain lengths (n = 14, 16) at pH 4, pH 7, and pH 12 were examined by differential scanning calorimetry. Simulation of heat capacity curves was performed using a new approach that incorporates changes of cooperativity of the transition in addition to nonideal mixing in the gel and the liquid-crystalline phase as a function of composition. From the simulations of the heat capacity curves, first estimates for the nonideality parameters for nonideal mixing as a function of composition were obtained, and phase diagrams were constructed using temperatures for onset and end of melting, which were corrected for the broadening effect caused by a decrease in cooperativity. In all cases the composition dependence of the nonideality parameters indicated nonsymmetrical mixing behavior. The phase diagrams were therefore further refined by simulations of the coexistence curves using a four-parameter approximation to account for nonideal and nonsymmetrical mixing in the gel and the liquid-crystalline phase. The mixing behavior was studied at three different pH values to investigate how changes in headgroup charge of the PA influences the miscibility. The experiments showed that at pH 7, where the PA component is negatively charged, the nonideality parameters are in most cases negative, indicating that electrostatic effects favor a mixing of the two components. Partial protonation of the PA component at pH 4 leads to strong changes in miscibility; the nonideality parameters for the liquid-crystalline phase are now in most cases positive, indicating clustering of like molecules. The phase diagram for 1,2-dimyristoyl-sn-glycero-3-phosphatidic acid:1,2-dipalmitoyl-sn-glycero-3-phosphorylcholine mixtures at pH 4 indicates that a fluid-fluid immiscibility is likely. The results show that a decrease in ionization of PAs can induce large changes in mixing behavior. This occurs because of a reduction in electrostatic repulsion between PA headgroups and a concomitant increase in attractive hydrogen bonding interactions.

A simulation program using least-squares minimization was developed to calculate and fit heat capacity (cp) curves to experimental thermograms of dilute aqueous dispersions of phospholipid mixtures determined by high-sensitivity differential scanning calorimetry. We analyzed cp curves and phase diagrams of the pseudobinary aqueous lipid systems 1,2-dimyristoyl-sn-glycero-3-phosphatidylglycerol/ 1,2-dipalmitoyl-sn-glycero-3phosphatidylcholine (DMPG/DPPC) and 1,2-dimyristoyl-sn-glycero-3-phosphatidic acid/1, 2-dipalmitoyl-sn-glycero-3-phosphatidylcholine (DMPA/DPPC) at pH 7. The simulation of the cp curves is based on regular solution theory using two nonideality parameters rho g and rho l for symmetric nonideal mixing in the gel and the liquid-crystalline phases. The broadening of the cp curves owing to limited cooperativity is incorporated into the simulation by convolution of the cp curves calculated for infinite cooperativity with a broadening function derived from a simple two-state transition model with the cooperative unit size n = delta HVH/delta Hcal as an adjustable parameter. The nonideality parameters and the cooperative unit size turn out to be functions of composition. In a second step, phase diagrams were calculated and fitted to the experimental data by use of regular solution theory with four different model assumptions. The best fits were obtained with a four-parameter model based on nonsymmetric, nonideal mixing in both phases. The simulations of the phase diagrams show that the absolute values of the nonideality parameters can be changed in a certain range without large effects on the shape of the phase diagram as long as the difference of the nonideality parameters for rho g for the gel and rho l for the liquid-crystalline phase remains constant. The miscibility in DMPG/DPPC and DMPA/DPPC mixtures differs remarkably because, for DMPG/DPPC, delta rho = rho l -rho g is negative, whereas for DMPA/DPPC this difference is positive. For DMPA/DPPC, this difference is interpreted as being caused by a negative rho g value, indicating complex formation of unlike molecules in the gel phase.