Nanoparticles are ubiquitously used for signal enhancement in (bio)sensors, but their true possible performance is typically hampered by non-specific binding. A better understanding of the nature and the prevention of non-specific binding through surface engineering of the particles and sensor surfaces is needed to intelligently design (bio)sensors and potentially avoid bulk blocking methods. Hence, two types of liposomes were used as model for signal-enhancing nanoparticles. Their surface was engineered to bear negative surface charge. One type was synthesized with additional 6 mol% -COOH groups. Their interaction with four typical chemically modified sensor surfaces was then mechanistically characterized by surface plasmon resonance (SPR) spectroscopy. It was shown that the non-specific binding can be described with Langmuir isotherms providing quantitative information of dissociation constants and surface loading with especially high correlation coefficients (>0.97) for all the studied sensor surfaces modified with hydrophilic alkane thiols. By tailoring the sensor surface chemistry, non-specific binding was significantly minimized. Here, carboxyl- or methyl-terminated surfaces performed best. In fact, the pairing of -COOH groups on the sensor surface with -COOH groups on the liposomes almost completely eliminated non-specific binding, resulting in a SPR signal change of only 1 mRIU (refractive index unit) at 100 μM phospholipid concentration. Surprisingly though, -OH groups on the surface, which are also commonly used in sensing applications, did not lead to decreased adsorption, but caused significant signal changes (4 mRIU at 100 μM phospholipid) due to non-specific binding. Overall, the mechanistic studies presented here demonstrate that by careful design of the nanoparticle surface and by choosing sensor surfaces with terminal -CH3 or -COOH groups, improved sensing (micro)systems with very low non-specific adsorption can be obtained.

Harmane and norharmane are representative members of the large group of natural β-carboline alkaloids featured with diverse pharmacological activities. In blood, these agents are transported by human serum albumin (HSA) which has a profound impact on the pharmacokinetic and pharmacodynamic properties of many therapeutic drugs and xenobiotics. By combination of various spectroscopic methods, the present contribution is aimed to elucidate how nonesterified fatty acids (FAs), the primary endogenous ligands of HSA, affect the binding properties of harmane and norharmane. Analysis of induced circular dichroism (CD) and fluorescence spectroscopic data indicates the inclusion of the neutral form of both molecules into the binding pocket of subdomain IIIA, which hosts two FA binding sites, too. The induced CD and UV absorption spectra of harmane and norharmane exhibit peculiar changes upon addition of FAs, suggesting the formation of ternary complexes in which the lipid ligands significantly alter the binding mode of the alkaloids via cooperative allosteric mechanism. To our knowledge, it is the first instance of the demonstration of drug-FA cobinding at site IIIA. In line with these results, molecular docking calculations showed two distinct binding positions of norharmane within subdomain IIIA. The profound increase in the affinity constants of β-carbolines estimated in the presence of FAs predicts that the unbound, pharmacologically active serum fraction of these compounds strongly depends on the actual lipid binding profile of HSA.

Spectroscopic studies combined with computational analysis indicate the inherent conformational flexibility of the β-carbolin derivative evodiamine (EVD) featured with diverse pharmacological activities. Qualitative evaluation of the circular dichroism (CD) spectra of EVD enantiomers complemented with quantum chemical calculations reveals a chiral exciton signature that can be assigned to the folded, L-shaped conformation of the molecule. Changes of the exciton couplet measured in different solvents and the near-UV CD profile upon binding to human serum albumin (HSA) refer to the structural adaptability of EVD. The enantioselectivity of EVD-HSA interaction is demonstrated showing the binding preference of the (R)-enantiomer. Comparison of experimental and calculated CD spectra of various conformers of EVD as well as the results of molecular docking data suggest that the (R)-antipode is accomodated within the IIA subdomain of HSA in ridge-tile conformation. Rutaecarpine (RTC), the close congener of EVD, forms much tighter association complexes both with HSA and α1-acid glycoprotein. In contrast to EVD, the nearly planar geometry of the indoloquinazoline ring system of RTC allows its stacked dimeric binding to the HSA.