The physical state of lipid nanoparticles influences their effect on in vitro cell viability.

Although lipid nanoparticles represent potent drug carriers, for many formulations toxicity data are rare. Thus, in this study, the effect of different lipid nanoparticles on the cell viability of L929 mouse fibroblasts was systematically investigated using the MTT assay. The formulations were composed of trimyristin, tristearin or cholesteryl myristate stabilized with poloxamer 188, polysorbate 80, polyvinyl alcohol or a blend of soybean phospholipid and sodium glycocholate. Depending on lipid and storage conditions, the nanoparticles were prepared in different physical states or crystal modifications leading to different particle shapes. The cell viability was influenced considerably by the physical state of the particle matrix with crystalline nanoparticles causing a stronger decrease in viability than the corresponding liquid or liquid crystalline particles. Effects on the cell viability were also related to the type of matrix lipid, stabilizer and the particle shape. However, the effects of differently shaped particles of different polymorphic modifications of crystalline tristearin were comparable. The low viability caused by poloxamer 188-stabilized particles could be correlated with a strong cell uptake which was investigated by confocal laser scanning microscopy.

Assessment of fluidity of different invasomes by electron spin resonance and differential scanning calorimetry.

The aim of this study was to investigate the influence of membrane-softening components (terpenes/terpene mixtures, ethanol) on fluidity of phospholipid membranes in invasomes, which contain besides phosphatidylcholine and water, also ethanol and terpenes. Also mTHPC was incorporated into invasomes in order to study its molecular interaction with phospholipids in vesicular membranes. Fluidity of bilayers was investigated by electron spin resonance (ESR) using spin labels 5- and 16-doxyl stearic acid and by differential scanning calorimetry (DSC). Addition of 1% of a single terpene/terpene mixture led to significant fluidity increase around the C16 atom of phospholipid acyl chains comprising the vesicles. However, it was not possible to differentiate between the influences of single terpenes or terpene mixtures. Incorporation of mTHPC into the bilayer of vesicles decreased fluidity near the C16 atom of acyl chains, indicating its localization in the inner hydrophobic zone of bilayers. These results are in agreement with DSC measurements, which showed that terpenes increased fluidity of bilayers, while mTHPC decreased fluidity. Thus, invasomes represent vesicles with very high membrane fluidity. However, no direct correlation between fluidity of invasomes and their penetration enhancing ability was found, indicating that besides fluidity also other phenomena might be responsible for improved skin delivery of mTHPC.

Flow cytometry as a new approach to investigate drug transfer between lipid particles.

Lipid nanoparticles and liposomal carrier systems are of growing interest for intravenous drug delivery due to their biocompatibility and targetability. It is, however, difficult to investigate their release behavior for lipophilic drugs under physiological conditions. This study describes a novel flow cytometric method studying drug transfer from such carrier systems to particles simulating physiological receptor sites. For this purpose, liquid and solid trimyristin nanoparticles or soybean phospholipid liposomes were loaded with the lipophilic fluorescent substances Nile red, temoporfin, and DiI. The transfer of these model drugs to large emulsion droplets was examined by flow cytometry. Transfer of DiI to differently sized acceptor emulsions was also monitored by separating donor and acceptor particles using ultracentrifugation. Flow cytometry revealed a completion of transfer within a few minutes for Nile red and temoporfin at considerable amounts of transferred dye. In contrast, the highly lipophilic DiI transferred over a period of weeks only for a small percentage of the dye. Ultracentrifugation results confirmed this for DiI and indicated a dependence of transfer characteristics on the acceptor surface area. Nile red transfer into a bulk oil phase as alternative acceptor system was also very slow. Flow cytometry seems to be well suited to study the intrinsic transfer of fluorescent lipophilic substances, as no kinetic hindrances like dialysis bags nor separation steps are required. Additional detailed experiments will, however, be necessary to elucidate the prevalent transfer mechanisms completely.

Micro-structured smart hydrogels with enhanced protein loading and release efficiency.

A series of temperature-responsive poly(N-isopropylacrylamide) (PNIPAAm) hydrogels with highly porous microstructures were successfully prepared by using hydrophobic polydimethylsiloxane (PDMS) and sodium dodecyl sulfate as liquid template and stabilizer, respectively. These newly prepared hydrogels possess highly porous structures. In contrast to the conventional PNIPAAm hydrogel, the swelling ratios of the porous gels at room temperature were higher, and their response rates were significantly faster as the temperature was raised above the lower critical solution temperature. For example, the novel hydrogel prepared with 40% PDMS template lost over 95% water within 5 min, while the conventional PNIPAAm gel only lost approximately 14% water in the same time. The improved properties are achieved due to the presence of liquid PDMS templates in the reaction solutions, which lead to the formation of porous structures during the polymerization/crosslinking. Lysozyme and bovine serum albumin (BSA) as protein models were for the first time loaded into these micro-structured smart hydrogels through a physical absorption method. The experimental results show that the loading efficiency of BSA with a higher molecular weight is lower than that of lysozyme due to the size exclusion effect, and the loading efficiencies of both proteins in the porous hydrogel are much higher than those in the conventional PNIPAAm hydrogel. For example, the loading efficiency of BSA in porous hydrogel is 0.114, approximately 200% higher than that in conventional hydrogel (0.035). Both lysozyme and BSA were completely released from the porous hydrogel at 22 degrees C. Furthermore, the release kinetics of the proteins from the porous hydrogel could be modulated by tuning the environmental temperature. These newly prepared porous materials provide an avenue to increase the loading efficiency and to control the release patterns of macromolecular drugs from hydrogels, and show great promise for application in protein or gene delivery.