In the present work, the immobilization of enzymes within poly-N-isopropylacrylamide (p-NIPAM) microgels using the method of solvent exchange is applied to the enzyme horseradish peroxidase (HRP). When the solvent is changed from water to isopropanol, HRP is embedded within the polymer structure. After the determination of the immobilized amount of enzyme, an enhanced specific activity of the biocatalyst in isopropanol can be observed. Karl Fischer titration is used to determine the amount of water within the microgel particles before and after solvent exchange, leading to the conclusion that an "aqueous cage" remains within the polymer structure. This represents the driving force for the immobilization due to the high affinity of HRP for water. Beside, confocal laser scanning microscopy (CLSM) images show that HRP is located within the microgel network after immobilization. This gives the best conditions for HRP to be protected against chemical and mechanical stress. We were able to transfer a water-soluble enzyme to an organic phase by reaching a high catalytic activity. Hence, the method of solvent exchange displays a general method for immobilizing enzymes within p-NIPAM microgels for use in organic solvents. With this strategy, enzymes that are not soluble in organic solvents such as HRP can be used in such polar organic solvents.

We introduce a novel class of membrane active peptidomimetics, the amphiphilic cationic β(3R3)-peptides, and evaluate their potential as antimicrobial agents. The design criteria, the building block and oligomer synthesis as well as a detailed structure-activity relationship (SAR) study are reported. Specifically, infrared reflection absorption spectroscopy (IRRAS) was employed to investigate structural features of amphiphilic cationic β(3R3)-peptide sequences at the hydrophobic/hydrophilic air/liquid interface. Furthermore, Langmuir monolayers of anionic and zwitterionic phospholipids have been used to model the interactions of amphiphilic cationic β(3R3)-peptides with prokaryotic and eukaryotic cellular membranes in order to predict their membrane selectivity and elucidate their mechanism of action. Lastly, antimicrobial activity was tested against Gram-positive M. luteus and S. aureus as well as against Gram-negative E. coli and P. aeruginosa bacteria along with testing hemolytic activity and cytotoxicity. We found that amphiphilic cationic β(3R3)-peptide sequences combine high and selective antimicrobial activity with exceptionally low cytotoxicity in comparison to values reported in the literature. Overall, this study provides further insights into the SAR of antimicrobial peptides and peptidomimetics and indicates that amphiphilic cationic β(3R3)-peptides are strong candidates for further development as antimicrobial agents with high therapeutic index.

Polyelectrolyte multilayers (PEMs) deposited on flexible supports, such as silicone rubber, show interesting properties upon elongation and release, like controlled wrinkling, elongation-based wetting, or dewetting and stimuli responsive nanovalves. To understand the underlying physical effects of PEM experiencing linear elongation, the orientation change of molecular groups within PEM experiencing linear elongation was investigated. The model PEM consists of polystyrenesulphonate and polydimethyldiallyl chloride. The investigation method was infrared attenuated total reflectance. In the study, the orientation change of the benzene and the sulfate groups of polystyrenesulphonate upon elongation of the PEM, prepared at high and low ionic strengths, was tracked. The gained results show that the benzene group shows no sign of orientation change upon elongation, whereas the sulfate group does, whereby the reorientation depends on the ionic strength of the preparation solution. Upon release of elongation, the PEM prepared at low ionic strength shows no orientation change, whereas PEM prepared at high ionic strength does show further orientation change, indicating that the formed wrinkles elongate the PEM on the top of the wrinkles.

Polyelectrolyte multilayers (PEM) loaded with bioactive molecules such as proteins serve as excellent mimics of an extracellular matrix and may find applications in fields such as biomedicine and cell biology. A question which is crucial to the successful employment of PEMs is whether conformation and bioactivity of the loaded proteins is preserved. In this work, the polarized attenuated total reflection Fourier transform infrared (ATR-FTIR) technique is applied to investigate the conformation of the protein lysozyme (Lys) loaded into the poly(L-lysine)/hyaluronic acid (PLL/HA) multilayers. Spectra are taken from the protein in the PEMs coated onto an ATR crystal during protein adsorption and desorption. For comparison, a similar investigation is performed for the case of Lys in contact with the uncoated crystal. The study highlights the presence of both “tightly” and “poorly bound” Lys fractions in the PEM. These fractions differ in their conformation and release behavior from the PEM upon washing. Comparison of spectra recorded with different polarizations suggests preferential orientation of alpha helical structures, beta sheets and turns in the “tightly bound” Lys. In contrast, the “poorly bound” fraction shows isotropic orientation and its conformation is well preserved.