Capillary Electrophoresis Mass Spectrometry Approaches for Characterization of the Protein and Metabolite Corona Acquired by Nanomaterials

Abstract

The adsorption of biomolecules from surrounding biological matrices to the surface of nanomaterials (NMs) to form the corona has been of interest for the past decade. Interest in the bio-nano interface arises from the fact that the biomolecular corona confers a biological identity to NMs and thus causes the body to identify them as "self". For example, previous studies have demonstrated that the proteins in the corona are capable of interacting with membrane receptors to influence cellular uptake and established that the corona is responsible for cellular trafficking of NMs and their eventual toxicity. To date, most research has focused upon the protein corona and overlooked the possible impacts of the metabolites included in the corona or synergistic effects between components in the complete biomolecular corona. As such, this work demonstrates methodologies to characterize both the protein and metabolite components of the biomolecular corona using bottom-up proteomics and metabolomics approaches in parallel. This includes an on-particle digest of the protein corona with a surfactant used to increase protein recovery, and a passive characterization of the metabolite corona by analyzing metabolite matrices before and after NM exposures. This work introduces capillary electrophoresis-mass spectrometry (CESI-MS) as a new technique for NM corona characterization. The protocols outlined here demonstrate how CESI-MS can be used for the reliable characterization of both the protein and metabolite corona acquired by NMs. The move to CESI-MS greatly decreases the volume of sample required (compared to traditional liquid chromatography-mass spectrometry (LC-MS) approaches) with multiple injections possible from as little as 5 µL of sample, making it ideal for volume limited samples. Furthermore, the environmental consequences of analysis

Coronas of micro/nano plastics: a key determinant in their risk assessments

“…Fadare et al [163] exploited this technique to analyze the coronas on M/ NPs after interacting with humic substances and identified a total of 1004 proteins, with 281 and 723 unique proteins for eco-corona and bio-corona, respectively. In addition, CE-MS has recently been introduced as a new technique for the characterization of protein and metabolite coronas on nanomaterials [271]. Nevertheless, full interpretation of coronas on M/NPs is still a major challenge, which may require a rational combination of several analytical techniques.…”

Understanding the Significance of Sample Preparation in Studies of the Nanoparticle Metabolite Corona

“…Electrospray ionization (ESI) was performed in positive ion mode for cations and negative ion mode for anions, with the porous tip of the capillary positioned 3–4 mm from the entrance of the MS inlet. Detailed descriptions of the analytical methods employed were provided in previous studies. ,, Optimal IonSpray Voltage Floating values were determined with manual tuning to be in the range of 1350–1550 V for ESI-MS positive ion mode and 1400–1600 V for negative ion mode. The conditioning of new capillaries and rinsing between runs were performed as previously described. , The dried samples (except for samples prepared with plasma) were reconstituted in 35 μL of ISTD solutions, and 20 μL of supernatants after centrifugation at 21,000 g at 4 °C for 10 min were placed into nanoVials (SCIEX).…”

“…Detailed descriptions of the analytical methods employed were provided in previous studies. 10,29,33 Optimal IonSpray Voltage Floating values were determined with manual tuning to be in the range of 1350−1550 V for ESI-MS positive ion mode and 1400− 1600 V for negative ion mode. The conditioning of new capillaries and rinsing between runs were performed as previously described.…”

“…The conditioning of new capillaries and rinsing between runs were performed as previously described. 10,29 The dried samples (except for samples prepared with plasma) were reconstituted in 35 μL of ISTD solutions, and 20 μL of supernatants after centrifugation at 21,000 g at 4 °C for 10 min were placed into nanoVials (SCIEX). The sample tray was thermostatted at 10 °C.…”

“…These encompass monomers of proteins and nucleic acids, which are known to adsorb to NMs − in addition to other metabolites encompassing a wide chemical space, which have either previously been seen to adsorb to NMs or cover key metabolite classes. Due to the highly charged and polar nature of the metabolites, they were analyzed using sheathless capillary zone electrophoresis mass spectrometry (CE-MS). , This approach is well-suited for the selective and sensitive profiling of polar ionogenic compounds in a wide range of matrices , and has been successfully applied to NM research and protein , and metabolite corona , analysis, with recent work showing excellent reproducibility for metabolomics . This work offers a basis for future studies investigating the polar metabolites present in the biomolecular corona acquired by NMs in complex biofluids, enabling high reproducibility and high recoveries, to ensure that accurate and reliable metabolite corona analysis can be performed.…”