Wastewater-based epidemiology? Unorthodox measurements of AGH UST scientists

Both drugs and polycyclic aromatic hydrocarbons (PAHs) are counted among the so-called xenobiotics (from Greek xenos – foreigner, stranger and bios – life), that is, substances which are not naturally produced within an organism, yet they do have an effect on it. The group comprises also, for example, medications that, contrary to the aforementioned chemical compounds, usually have a possitive effect on human organism. However, some PAHs, which are the main focus of the researchers from the Faculty of Energy and Fuels, are toxic substances that show carcinogenic or mutagenic activity – for example, they are responsible for DNA mutations, leukaemia, or lung cancer. Additionally, they pose threats not only to humans, but also the environment, to which – given their slow degradation – they are a long-lasting nuisance.

PAHs constitute an ingredient of particulate matter air pollution, and their source is primarily the incomplete combustion of organic materials. Given that they are part of the so-called small fraction PM1 and PM2.5, they can easily penetrate our respiratory and cardiovascular systems. The highest concentrations of these substances in the atmopshere are recorded in ubran areas, where, on the one hand, the traffic is considerably more intense, and on the other, the pollution is densely distributed. Nevertheless, PAHs can penetrate the organism not only from air. Their presence in the human body can result from diet or job that one does. Thus, measuring the environment is not enough if we want to know the full extent to which the population is exposed to these compounds.

‘We’re exposed to PAHs not only through air pollution, but also water contamination. High exposure to those toxic substances can also result from smoking cigarettes or the job one has. Another way that humans can come into contact with those harmful polycyclic aromatic hydrocarbons is through food contamination. The more fried or grilled foods we eat, the more we’re susceptible to consuming higher doses of PAHs’, explains Professor Katarzyna Styszko, leader of the project.

Sewage analysis

To determine the degree of exposure to those compounds, it might be helpful to combine measurements in the city with biomonitoring with respect to PAHs and their metabolites that are a result of chemical transformations occurring in the organism. To do this, an innovative method could be used that is based on a chemical analysis of specified products of human metabolism in sewage works, that is, wastewater-based epidemiology (WBE), which is able to provide an integrated image of citizen exposure to polycyclic aromatic hydrocarbons. In current biomonitoring studies, a very precise correlation has been shown between the population exposure to PAHs and finding their biomarkers in urine.

‘Classic epidemiology is based primarily on surveys or blood and urine tests of patients or volunteers who participate in such activities. This usually represents a narrow spectrum of the population; moreover, such studies are very expensive, require a lot of work, and you have to wait a long time before you get the results. In the case of wastewater-based epidemiology, we can practically assess the exposure of a specific population to selected substances in real time. We usually count it in mg/24h per one thousand citizens. Now, because of the pandemic, intense studied are being conducted towards detecting the virus in wastewater’, says Professor Styszko.

Therefore, WBE provides objective data on the aggreggate exposure to PAH xenobiotics of citizens from a specified area, which, after penetrating the human organism, are metabolised quickly and after a few days leave the body in the form of hydroxy derivatives. Wastewater-based epidemiology can therefore be called a collective urine test, based on which the analysts can assess the degree of population exposure to dangerous substances almost in real time. This method was used by the engineers from the Faculty of Energy and Fuels, who, during their comprehensive tests, have not only thoroughly analysed particulate matter, but also made precise measurements of wastewater, establishing a fruitful cooperation with the largest sewage works in Krakow, second largest in Poland.

Measurement campaign

As part of a grant from the IERU project, AGH UST researchers organised two measurement campaigns of wastewater – summer edition (2020) and winter edition (2021), to determine seasonal changes in the concentration of the compounds under investigation. The samples collected in the sewage works were transported to a laboratory, where, after appropriate treatment, were analysed with the use of a gas chromatography–mass spectrometry method.  The measurements did not exhaust the subject, because to assess the degree of exposure to PAHs, it was necessary to know the processes of PAH metabolism and adequate back-calculations that should allow scientists to find answers to the fundamental questions.

 

The procedure of wastewater sample preparation for analysis]

‘The combination of gas chromatography and mass spectrometry is an example of the so-called hybrid analytical techniques. Chromatography is an analytical technique which is aimed to separate particular fractions within a sample, while the application of mass spectrometry allows us to determine the substances contained therein and indicate their concentrations. The separation of compounds occurs in a chromatography column as a result of using an appropriate temperature gradient. The compounds then travel with the carrier gas to the mass spectrometer that acts as a detector’, describes Professor Styszko briefly.

Presentation of results

The initial results of laboratory tests that were published in four articles in high-impact journals and presented during the scientific conference held in December 2021 at the AGH UST pointed to a high seasonal correlation of Krakow citizens exposure to polycyclic aromatic hydrocarbons. Positive effects of the research carried out will allow scientists to expand wastewater analyses to other cities – not only in Poland but also abroad. The study also facilitates risk assessment related to penetration of PAH hydroxylic derivatives into water environment, which – as it turns out – are not completely filtered out in sewage works and can be, according to the researchers, even more toxic than the initial compound.

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The project was funded by a university grant within the framework of the “Initiative for Excellence – Research University” project (the AGH UST 2020–2022, PRA-2).