The Impact of Virgin and Aged Microstructured Plastics on Proteins: The Case of Hemoglobin Adsorption and Oxygenation

Abstract

Plastic particles, particularly micro- and nanoparticles, are emerging pollutants due to the ever-growing amount of plastics produced across a wide variety of sectors. When plastic particles enter a biological medium, they become surrounded by a corona, giving them their biological identity and determining their interactions in the living environment and their biological effects. Here, we studied the interactions of microstructured plastics with hemoglobin (Hb). Virgin polyethylene microparticles (PEMPs) and polypropylene microparticles (PPMPs) as well as heat- or irradiation-aged microparticles (ag-PEMPs and ag-PPMPs) were used to quantify Hb adsorption. Polypropylene filters (PP-filters) were used to measure the oxygenation of adsorbed Hb. Microstructured plastics were characterized using optical microscopy, SAXS, ATR-FTIR, XPS, and Raman spectroscopy. Adsorption isotherms showed that the Hb corona thickness is larger on PPMPs than on PEMPs and Hb has a higher affinity for PPMPs than for PEMPs. Hb had a lower affinity for ag-PEMPs and ag-PPMPs, but they can be adsorbed in larger amounts. The presence of partial charges on the plastic surface and the oxidation rate of microplastics may explain these differences. Tonometry experiments using an original method, the diffuse reflection of light, showed that adsorbed Hb on PP-filters retains its cooperativity, but its affinity for O₂ decreases significantly.

Keywords: hemoglobin; hemoglobin activity; hemoglobin adsorption; microstructured plastics; oxygenation.

Figure 1

Microscopic visible images of aged PE microparticles (left) and aged PP microparticles (right).

Figure 2

The adsorption isotherms of oxyhemoglobin on virgin polyethylene microparticles (PEMPs) (red circles) and on polypropylene microparticles (PPMPs) (green circles) in 0.1 mol.L⁻¹ phosphate buffer pH 7.0. Experimental data (circles) and error bars correspond to the average and standard deviation of three biological replicates. Data were fitted using the Langmuir adsorption model (solid lines).

Figure 3

The adsorption isotherms of oxyhemoglobin on aged polyethylene (PE) microparticles (ag-PEMPs) (orange circles) and polyethylene (PP) microparticles (ag-PPMPs) (teal circles) in 0.1 mol.L⁻¹ phosphate buffer pH 7.0. Experimental data (circles) and error bars correspond to the average and standard deviation of three biological replicates. Data were fitted using the Langmuir adsorption model (solid lines).

Figure 4

The Kubelka–Munk transformation of the UV–visible spectra of hemoglobin adsorbed on the polypropylene filter obtained using diffuse reflection. Oxygen additions and spectrum measurements were repeated until the fully oxygenated form was reached. Arrows are oriented from the deoxyhemoglobin spectrum to the oxyhemoglobin spectrum.

Figure 5

Oxygen binding curves: native hemoglobin (gray circles), hemoglobin adsorbed on the polypropylene filter (PP-filter) (light blue squares), and hemoglobin after contact with the PP-filter (dark blue triangles) in 0.1 mol.L⁻¹ phosphate buffer pH 7.4. The fit using the Hill equation is shown (solid lines).

Figure 6

Oxygen binding curves: native hemoglobin (Hb) (gray circles), Hb after contact with polyethylene microparticles (PEMPs) (red squares), and Hb after contact with polypropylene microparticles (PPMPs) (green triangles) in 0.1 mol.L⁻¹ phosphate buffer pH 7.4. The data were fit to the Hill equation (solid lines).

Figure 7

A polypropylene filter in a tonometer with 100 µL of 0.1 mol.L⁻¹ phosphate buffer. The red color of adsorbed hemoglobin can barely be seen with the naked eye, but it can be detected and analyzed through diffuse reflection symbolized by the red arrows. The white arrow represents incident light.