A Simple Millifluidic Benchtop Reactor System for the High-Throughput Synthesis and Functionalization of Gold Nanoparticles with Different Sizes and Shapes

Despite the continuing interest in the applications of functionalized nanomaterials, the controlled and reproducible synthesis of many important functionalized nanoparticles (NPs) above the milligram scale continues to be a significant challenge. The synthesis of functionalized NPs in automated reactors provides a viable approach to circumvent some of the shortcomings of traditional nanomaterial batch syntheses, providing superior control over reagent addition, improved reproducibility, the opportunity to interface real-time product monitoring, and a viable high-throughput synthetic approach. Here, we demonstrate the construction and operation of a simple millifluidic reactor assembled entirely from commercially available components found in almost any chemical laboratory. This reactor facilitates the aqueous gram-scale synthesis of a variety of functionalized gold nanoparticles, including the synthesis of gold nanospheres with tightly controlled core diameters and gold nanorods with controlled aspect ratios between 1.5 and 4.0. The absolute dimensions (<i>i.e.</i>, the transverse diameter) of gold nanorods synthesized within the reactor can also be tailored to produce different gold nanorod shapes, including “small” gold nanorods and gold nanocubes. In addition, we show that the reactor can interface with existing purification and monitoring techniques in order to enable the high-throughput functionalization/purification of gold nanorods and real-time monitoring of gold nanoparticle products for quality control. We anticipate that this millifluidic reactor will provide the blueprint for a versatile and portable approach to the gram-scale synthesis of monodisperse, hydrophilically functionalized metal NPs that can be realized in almost any chemistry research laboratory

Quantitative Determination of Ligand Densities on Nanomaterials by X‑ray Photoelectron Spectroscopy

X-ray photoelectron spectroscopy (XPS) is a nearly universal method for quantitative characterization of both organic and inorganic layers on surfaces. When applied to nanoparticles, the analysis is complicated by the strong curvature of the surface and by the fact that the electron attenuation length can be comparable to the diameter of the nanoparticles, making it necessary to explicitly include the shape of the nanoparticle to achieve quantitative analysis. We describe a combined experimental and computational analysis of XPS data for molecular ligands on gold nanoparticles. The analysis includes scattering in both Au core and organic shells and is valid even for nanoparticles having diameters comparable to the electron attenuation length (EAL). To test this model, we show experimentally how varying particle diameter from 1.3 to 6.3 nm leads to a change in the measured <i>A</i>_C/<i>A</i>_Au peak area ratio, changing by a factor of 15. By analyzing the data in a simple computational model, we demonstrate that ligand densities can be obtained, and, moreover, that the actual ligand densities for these nanoparticles are a constant value of 3.9 ± 0.2 molecules nm^–2. This model can be easily extended to a wide range of core–shell nanoparticles, providing a simple pathway to extend XPS quantitative analysis to a broader range of nanomaterials

Nanoparticle–Protein Interactions: A Thermodynamic and Kinetic Study of the Adsorption of Bovine Serum Albumin to Gold Nanoparticle Surfaces

Investigating the adsorption process of proteins on nanoparticle surfaces is essential to understand how to control the biological interactions of functionalized nanoparticles. In this work, a library of spherical and rod-shaped gold nanoparticles (GNPs) was used to evaluate the process of protein adsorption to their surfaces. The binding of a model protein (bovine serum albumin, BSA) to GNPs as a function of particle shape, size, and surface charge was investigated. Two independent comparative analytical methods were used to evaluate the adsorption process: steady-state fluorescence quenching titration and affinity capillary electrophoresis (ACE). Although under favorable electrostatic conditions kinetic analysis showed a faster adsorption of BSA to the surface of cationic GNPs, equilibrium binding constant determinations indicated that BSA has a comparable binding affinity to all of the GNPs tested, regardless of surface charge. BSA was even found to adsorb strongly to GNPs with a pegylated/neutral surface. However, these fluorescence titrations suffer from significant interference from the strong light absorption of the GNPs. The BSA–GNP equilibrium binding constants, as determined by the ACE method, were 10⁵ times lower than values determined using spectroscopic titrations. While both analytical methods could be suitable to determine the binding constants for protein adsorption to NP surfaces, both methods have limitations that complicate the determination of protein–GNP binding constants. The optical properties of GNPs interfere with <i>K</i>_a determinations by static fluorescence quenching analysis. ACE, in contrast, suffers from material compatibility issues, as positively charged GNPs adhere to the walls of the capillary during analysis. Researchers seeking to determine equilibrium binding constants for protein–GNP interactions should therefore utilize as many orthogonal techniques as possible to study a protein–GNP system

Lipopolysaccharide Density and Structure Govern the Extent and Distance of Nanoparticle Interaction with Actual and Model Bacterial Outer Membranes

Design of nanomedicines and nanoparticle-based antimicrobial and antifouling formulations and assessment of the potential implications of nanoparticle release into the environment requires understanding nanoparticle interaction with bacterial surfaces. Here we demonstrate the electrostatically driven association of functionalized nanoparticles with lipopolysaccharides of Gram-negative bacterial outer membranes and find that lipopolysaccharide structure influences the extent and location of binding relative to the outer leaflet-solution interface. By manipulating the lipopolysaccharide content in <i>Shewanella oneidensis</i> outer membranes, we observed the electrostatically driven interaction of cationic gold nanoparticles with the lipopolysaccharide-containing leaflet. We probed this interaction by quartz crystal microbalance with dissipation monitoring (QCM-D) and second harmonic generation (SHG) using solid-supported lipopolysaccharide-containing bilayers. The association of cationic nanoparticles increased with lipopolysaccharide content, while no association of anionic nanoparticles was observed. The harmonic-dependence of QCM-D measurements suggested that a population of the cationic nanoparticles was held at a distance from the outer leaflet-solution interface of bilayers containing smooth lipopolysaccharides (those bearing a long <i>O</i>-polysaccharide). Additionally, smooth lipopolysaccharides held the bulk of the associated cationic particles outside of the interfacial zone probed by SHG. Our results demonstrate that positively charged nanoparticles are more likely to interact with Gram-negative bacteria than are negatively charged particles, and this interaction occurs primarily through lipopolysaccharides

Direct Probes of 4 nm Diameter Gold Nanoparticles Interacting with Supported Lipid Bilayers

This work presents molecular-level investigations of how well-characterized silica-supported phospholipid bilayers formed from either pure DOPC or a 9:1 mixture of DOPC:DOTAP interact with positively and negatively charged 4 nm gold metal nanoparticles at pH 7.4 and NaCl concentrations ranging from 0.001 to 0.1 M. Second harmonic generation (SHG) charge screening measurements indicate the supported bilayers carry a negative interfacial potential. Resonantly enhanced SHG measurements probing electronic transitions within the gold core of the nanoparticles show the particles interact irreversibly with the supported bilayers at a range of concentrations. At 0.1 M NaCl, surface coverages for the particles functionalized with the negatively charged ligand mercaptopropionic acid (MPA) or wrapped in the cationic polyelectrolyte poly­(allylamine) hydrochloride (PAH) are estimated from a joint analysis of QCM-D, XPS, AFM, and ToF-SIMS to be roughly 1 × 10⁷ and 1 × 10¹¹ particles cm^–2, respectively. Results from complementary SHG charge screening experiments point to the possibility that the surface coverage of the MPA-coated particles is more limited by interparticle Coulomb repulsion due to the charges within their hydrodynamic volumes than with the PAH-wrapped particles. Yet, SHG adsorption isotherms indicate that the interaction strength per particle is independent of ionic strength and particle coating, highlighting the importance of multivalent interactions. ¹H NMR spectra of the lipids within vesicles suspended in solution show little change upon interaction with either particle type but indicate loosening of the gold-bound PAH polymer wrapping upon attachment to the vesicles. The thermodynamic, spectroscopic, and electrostatic data presented here may serve to benchmark experimental and computational studies of nanoparticle attachment processes at the nano–bio interface