Oxidative potential of particulate matter and organic air pollution markers – OxAirTracers

Project duration: 36 months
Project start: 12 January 2026

General

Air quality is now a major public health issue, responsible for 4.2 million premature deaths worldwide each year (WHO), and this number is expected to double by 2050. The strongest evidence of public health concern relates to particulate matter (PM), which reduces life expectancy, causes and exacerbates numerous diseases, and negatively impacts ecosystems, climate stability, and the environment. Short-term exposure to PM can lead to various forms of cardiovascular and respiratory diseases, while long-term exposure can result in congenital heart defects and adverse effects on the cardiovascular system. Exposure to PM is also associated with neurological disorders, obesity, metabolic disorders such as diabetes, cognitive decline, and adverse pregnancy outcomes.

In air quality science, PM is classified by size, with the most common categories being PM10 (aerodynamic diameter < 10 μm), PM2.5 (< 2.5 μm), PM1 (< 1 μm) and PM0.1 (< 0.1 μm). The size of the particles affects how they enter the human body – coarse particles (e.g. PM2.5–10) can be filtered in the upper respiratory tract, while finer particles (PM2.5 or PM1) penetrate deeper, even to the cardiovascular and nervous systems, as is the case with PM0.1. PM is a mixture of particles of different chemical composition, size and shape, which depends on the emission sources and transformation processes in the atmosphere. Consequently, the toxicity of PM varies considerably. The health effects of PM depend on the sources and chemical composition of the aerosol, and some components can seriously damage lung cells by causing inflammation. Although the mass concentration of PM is often used to assess risk, it does not reflect their actual toxicity, which depends on their physicochemical properties. Exposure to PM induces inflammatory reactions in the lungs, and oxidative stress, caused by an imbalance between reactive oxygen species (ROS) and the antioxidant defenses of cells, is a key mechanism of their toxicity.

Oxidative potential (OP), which indicates the ability of PM to transport ROS or to induce their synthesis in vitro, is increasingly used as an indicator of toxic effects. PM can increase ROS levels by direct transfer or by interaction with biological molecules, disrupting the redox balance of the organism, which can lead to respiratory, cardiovascular and neurological disorders, including asthma and chronic bronchitis. The OP of airborne particles depends on their chemical composition, but the complete determination of all PM components remains a challenge. OP is considered a more precise indicator of toxicity than the mass of PM alone because it estimates the ability to generate ROS, taking into account particle size, specific surface area and chemical composition. The PM2.5 fraction is particularly important due to the higher oxidative stress it induces. Although methods exist for determining OP in the laboratory, there are still numerous challenges in comparing different testing methods.

Various acellular assays have been developed to measure OP, which are divided into those that measure oxidant production and those that monitor antioxidant depletion. Oxidant production assays include hydroxyl radical (OH) assays and electron paramagnetic resonance (EPR) assays, while depletion assays include ascorbic acid (AA), glutathione (GSH) and dithiothreitol (DTT) assays. While AA and GSH are crucial for cellular and extracellular defenses, DTT is often used as a surrogate for biological reductants. It has been observed that there is a significant difference between the results obtained by different methods, which also depends on the characteristics of the measurement location itself, i.e. the dominant sources of air pollution present. Comparing the spatial distribution of OP with PM sources has proven useful for their identification. Many unknowns remain regarding OP assays, including which assays are most reliable in predicting health outcomes and which PM components generate the signals recorded in these assays. The chemical composition of PM and the determination of their OP are closely related to the biological response of the organism.

The project plans to introduce two commonly used methods, including DTT analysis and AA analysis. AA is the predominant natural antioxidant in the lung, while DTT acts as a chemical surrogate for cellular reductants, such as nicotinamide adenine dinucleotide (NADH) and nicotinamide adenine dinucleotide phosphate (NADPH). These antioxidants are consumed when components present in PM catalytically transfer one electron from an AA or DTT molecule to molecular oxygen, generating superoxide anion and mimicking the key initial step in the in vivo generation of ROS. DTT is oxidized to its disulfide form upon interaction with redox active compounds present in PM, and the linear rate of DTT decay is used as an index of the oxidative capacity of PM. AA analysis is performed similarly to DTT analysis. After controlled incubation of AA in an aqueous extract, the measurement of the decrease in AA concentration over time is directly monitored by the decrease in UV-Vis absorbance of ascorbic ion. Although different methods for OP analysis are sensitive to the same redox active species, they differ in their relationship to the chemical components of PM that contribute to the total OP. DTT analysis is mainly sensitive to organic compounds that accumulate mostly in the fine fraction of PM, while AA mainly reacts to metals that accumulate mostly in coarse particles. When determining OP, a synergistic approach is proposed that uses multiple methods to collect the most complete information on the reactivity of PM. The methods developed in this project would be used to conduct the first, indicative measurements of OP of suspended particles in the Republic of Croatia (RH) at multiple locations.