HICE and aeroHEALTH

Characterization of Reactive Organic Compounds in Gases and Particulate Matter

In WP I, aerosols from a variety of anthropogenic sources and model aerosols were characterized with respect to reactive organic compounds and made available for exposure experiments.

The composition of the pollutants depended strongly on the geographical location and atmospheric conditions and was therefore difficult to interpret. Therefore, emissions from typical anthropogenic combustion sources were studied. The consortium had extensive experience in operating emission sources and sampling systems. A wide range of technical equipment was available at the University of Western Finland and the University of Rostock: boiler burners, test setups for automotive engines and marine diesel engines. Flame soot and ash particles were produced for tests and standard reference measurements. Modern fuel formulations (biofuel), which are known to have higher oxygen content, were taken into account.

Modern methods of mass spectrometry and measurement techniques for chemical and physical processes were used to comprehensively characterize the emission aerosol. Mass spectrometric methods were additionally adapted to characterize the biological response. 

Both resonance-enhanced multiphoton ionization and single photon ionization formed the basis for the mass spectrometric methods.

For the analysis of aerosols, gas and particle phases were combined, but also used separately to account for the fact that reactive organic compounds often occur in a gaseous fraction as vapor in the gas phase, but also as a liquid or solid fraction condensed on the aerosol particles.

The reactive organic compounds (ROC) in the gas phase of the aerosol were characterized by photoionization mass spectrometry, and the particles were characterized by aerosol time-of-flight (ATOF) mass spectrometry. In addition, physical parameters of the particles such as number and mass concentration were determined online. Both the particles and the gas phase were collected and made available for offline analysis in WP III.
 
During the ongoing work, the analytical focus was shifted to the potential recurring markers based on the results obtained in WP II and WP III. The goal was an emission assessment based on (bio-)statistical approaches combining the results from WP I with those of the analytical approaches in WP II and WP III.

Health effects and toxicology of reactive organic compounds in aerosols

In WP II, the biological effects of the freshly generated combustion aerosol, the particulate phase alone (stripped with gas phase dust collectors), and the gas phase alone (filtered) were comparatively studied in human cell systems. Cells were exposed to the aerosol both in vitro and in an exposure facility at the air-liquid interface. In co-cultures, up to five cell types were combined to form a differentiated human lung tissue model. In vivo studies translated the results to human samples. The biological effects in cell cultures and tissues were studied at all hierarchical levels (genome, transcriptome, proteome and metabolome).

Since the emission sources were heavy stationary structures (test benches for diesel and gasoline engines, marine engines and boilers), test aerosol generators and exposure systems were mobile. All working groups involved in the HICE virtual institute met for several large-scale tests at the different sites in Europe.

The biological effects were investigated jointly with WP III. The biological endpoints were therefore e.g. cytotoxicity, metabolic activity, determination of functions, oxidative stress and inflammatory response markers. In addition, non-targeted effects were investigated e.g. using genome-wide gene expression, protein profiling and induction analyses. In selected experiments, stable isotope labeling was used to perform flux analyses in the proteome and metabolome.

Clinical partners provided human cells and tissues from lavages, biopsies, or pathological examinations of healthy and diseased individuals to screen for biomarkers found in laboratory experiments.
In collaboration with WP IV, the data were analyzed and screened for biomarkers associated with exposure and harm.

Comprehensive chemical analysis of small molecules in aerosols and biological systems

This work package focused on comprehensive chemical analysis. On the one hand, this was the characterization of the chemical composition of the different aerosol compartments. On the other hand, it was the study of the effects of chemically and physically well-characterized aerosols on biological test systems and their possible health effects. Therefore, the changes in the pattern of small molecules in biological test systems caused by exposure to aerosols were investigated.

Within HICE, there was a strong body of knowledge related to the chemical analysis of organic constituents of aerosols. A wide range of mass spectrometric and chromatographic techniques were established (LC, GC, TOFMS, FTMS, MALDI imaging). Much research was done to facilitate quantitative analysis of large time series and analysis of selected marker compounds. In recent years, concepts for comprehensive chemical profiling (GCxGC) and isotopic labeling "non-targeted tracer fate detection (NTFD)" had been developed and applied to environmental and/or biological applications. The NTFD concept was applied to cell cultures to study the effects of aerosols on biological systems. In a second step, the NTFD methodology was combined with comprehensive two-dimensional gas chromatography (GCxGC).

Targeted and non-targeted methods were used to reveal the complex chemical composition of the aerosol matrices from WP I as well as the biological system from WP II. FTMS and IRMS systems were used for marker identification and chemical profiling of aerosols and biological systems. Specific tools for data analysis were developed in close collaboration with WP I and IV. To identify similarities and correlations between gas and particle phase species and their potential interactions, analytical results were investigated and interpreted in conjunction with on-line data from WP I.

Scientific data management, chemometrics and biostatistics

WP IV was the central interface within the HICE. It was responsible for the combined statistical data analysis, which included all analytical work packages (WP I-III).

First, it had to set up and monitor the data management of all scientific data. Besides this permanent setup, WP IV supported the different projects within the WPs with the "Design of Experiments (DOE)" to ensure the applicability of the statistical methods and the documentation of the experiment. A third task was the central analysis and evaluation of the scientific data and a basic consistency check. For a comprehensive analysis of the data, the results from the working groups and within and between the working groups were combined into a biostatistical analysis.

The data from all measurement campaigns and exposure experiments were statistically evaluated and comprehensively interpreted with regard to possible biomarkers for aerosol-related health effects.