Reliable characterization of protein coronas (PCs) that form when nanomaterials are introduced into biological fluids is a critical step in the development of safe and efficient nanomedicine. We observed that bovine serum albumin (BSA)-coated multi-walled carbon nanotubes (MWCNTs) do not induce cytotoxicity, but have different cellular uptake rates depending on the BSA pretreatment concentration. To determine how these slight differences affect A549 cell responses and intracellular changes, we conducted spectroscopic (circular dichroism and Fourier-transform infrared) and spectrometric (nanoflow liquid chromatography-electrospray ionization-tandem mass spectrometry) analyses. The various characterization techniques conducted in this study reveal the following. (i) The composition ratio of PCs on MWCNTs differs depending on the BSA concentration. (ii) Analysis of the secondary structure of the proteins revealed that the α-helix structure increased with increasing BSA concentration. (iii) Proteomic analysis showed that different biological pathways were activated at levels higher and lower than 5 mg mL-1. Such combined spectroscopic and spectrometric approaches provide an integrated understanding of PC composition as well as how nano/bio-interface states are linked to cellular-level responses. Our results can support reliable and practical applications of nanomedicine development.
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