Viscosity of protein solution is one of the most troublesome issues for the high-concentration formulation of protein drugs. In this review, we summarize the practical methods that suppress the viscosi-ty of protein solution using small molecular additives. The small amount of salts decreases the viscosity that results from electrostatic repulsion and attraction. The chaotrope suppresses the hydrophobic attraction and cluster formation, which can lower the solution viscosity. Arginine hydrochloride (ArgHCl) also sup-presses the solution viscosity due to the hydrophobic and aromatic interactions between protein molecules. The small molecular additives are the simplest resolution of the high viscosity of protein solution as well as understanding of the primary cause in complex phenomena of protein interactions.

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Proteins have nonspecific adsorption capacities for solid surfaces. Although the nonspecific adsorption capacities are generally understood to be related to the hydrophobicity or charge density of the surfaces, little is known at the amino acid level about the interaction between proteins and polyaromatic surfaces such as carbon nanotubes, which have recently been used for biotechnology applications. In this study, we investigated the interaction between proteinogenic amino acids and carbon nanotubes using high-performance liquid chromatography on silica matrices coated by single-wall carbon nanotubes (SWCNTs). Among the amino acids used in this study, tryptophan, tyrosine, and phenylalanine showed exceptional affinity for the matrices. The characteristic affinities of these amino acids were ascribed to their unique interactions with the large polyaromatic surfaces of the SWCNTs. These results are useful for understanding and controlling protein adsorption onto aromatic surfaces.

Hen egg white contains more than 40 kinds of proteins with concentrations reaching 100 mg/mL. Highly concentrated protein mixtures are common in the food industry, but the effects of a crowded environment containing salts on protein stability and aggregation have only been investigated using pure protein solutions. Here, we investigated the thermal aggregation of hen egg white protein (EWP) at various concentrations in the presence of inorganic salts by solubility measurements and SDS-PAGE. EWP at 1 mg/mL formed aggregates with increasing temperature above 55 °C; the aggregation temperatures increased in the presence of inorganic salt with the Hofmeister series. Namely, the chaotrope 0.5 M NaSCN completely suppressed the thermal aggregation of 1 mg/mL EWP. As the protein concentration increased, NaSCN unexpectedly enhanced the protein aggregation; the aggregation temperature of 10 and 100 mg/mL EWP solutions were dramatically decreased at 62 and 47 °C, respectively. This decrease in aggregation temperatures due to the chaotrope was described by the excluded volume effect, based on a comparative experiment using Ficoll 70 as a neutral crowder. By contrast, the kosmotrope Na2SO4 did not affect the aggregation temperature at concentrations from 1 to 100 mg/mL EWPs. The unexpected fact that a chaotrope rather enhanced the protein aggregation at high concentration provides new insight into the aggregation phenomena with the Hofmeister effect as well as the crude state of highly concentrated proteins.