Asymmetric-flow field-flow fractionation for measuring particle size, drug loading and (in)stability of nanopharmaceuticals. The joint view of European Union Nanomedicine Characterization Laboratory and National Cancer Institute - Nanotechnology Characterization Laboratory.

Asymmetric-flow field-flow fractionation (AF4) has been recognized as an invaluable tool for the characterisation of particle size, polydispersity, drug loading and stability of nanopharmaceuticals. However, the application of robust and high quality standard operating procedures (SOPs) is critical for accurate measurements, especially as these complex drug nanoformulations are most often inherently polydisperse. In this review we describe a unique international collaboration that lead to the development of a robust SOP for the measurement of physical-chemical properties of nanopharmaceuticals by multi-detector AF4 (MD-AF4) involving two state of the art infrastructures in the field of nanomedicine, the European Union Nanomedicine Characterization Laboratory (EUNCL) and the National Cancer Institute-Nanotechnology Characterisation Laboratory (NCI-NCL). We present examples of how MD-AF4 has been used for the analysis of key quality attributes, such as particle size, shape, drug loading and stability of complex nanomedicine formulations. The results highlight that MD-AF4 is a very versatile analytical technique to obtain critical information on a material particle size distribution, polydispersity and qualitative information on drug loading. The ability to conduct analysis in complex physiological matrices is an additional very important advantage of MD-AF4 over many other analytical techniques used in the field for stability studies. Overall, the joint NCI-NCL/EUNCL experience demonstrates the ability to implement a powerful and highly complex analytical technique such as MD-AF4 to the demanding quality standards set by the regulatory authorities for the pre-clinical safety characterization of nanomedicines.

Squaraine as a bright, stable and environment-sensitive far-red label for receptor-specific cellular imaging.

Herein, we show that a far-red arylidene-squaraine dye is stable against nucleophiles, in contrast to arene-squaraines. Owing to the fluorescence enhancement in apolar media together with high brightness and photostability, this dye was successfully applied to detect the oxytocin G protein-coupled receptor and monitor its internalization in living cells.

Pseudo-prolines: induction of cis/trans-conformational interconversion by decreased transition state barriers.

Molecular events such as cis/trans isomerization of Xaa-Pro tertiary amide bonds in peptides and proteins are slow on the overall time scale of the formation of a final biostructure and are, therefore, rate limiting. In order to pursue a better understanding of the molecular events underlying such slow interconversions, we applied the recently introduced pseudo-proline (PsiPro) concept as a tool to study the dynamics of Xaa-Pro bonds by determining the kinetics and thermodynamics of cis/trans isomerism. We show that enhanced isomerization rates of tertiary amide bonds prior to a PsiPro unit in short model peptides is due to lowered transition state barriers. In addition, pronounced effects upon the dynamics of the reversible transition between helix I and II of oligoprolines containing one or several PsiPro units were observed.