Development and Applications of Novel Exposure Biomarkers for Polycyclic Aromatic Hydrocarbons (PAHs) Reflecting Long-term Exposures or Wood Smoke Exposure

Abstract

Polycyclic aromatic hydrocarbons (PAHs), commonly present in air pollution mixtures, are linked to adverse human health effects, including oxidative stress, inflammation, and chronic diseases such as respiratory and cardiovascular conditions, as well as cancer. PAH metabolites, serving as biomarkers for PAH exposure, provide a quantitative method to assess individual exposure levels and evaluate dose-response relationships between air pollution and associated health effects. While urinary hydroxylated PAHs (OH-PAHs) are established biomarkers for assessing short-term PAH exposures, their effectiveness under contemporary conditions of lower air pollution levels and their ability to capture source-specific exposures (e.g., from wildfires) remain uncertain. Additionally, because many chronic diseases result from long-term exposures, biomarkers reflecting longer-term exposure are needed. However, the short half-life of OH-PAHs limits their utility in evaluating health risks associated with prolonged exposures.This dissertation research addresses these issues by examining urinary 1-hydroxypyrene (1-OHP) in relation to air pollution and oxidative stress in a small United States city with air pollution levels typically below National Ambient Air Quality Standards and developing and evaluating novel PAH biomarkers for long-term and wood smoke exposures, respectively. Specifically, this research focuses on PAH-hemoglobin adducts as biomarkers of long-term exposure and urinary PAH carboxylic acids (PAH-CAs) as indicators of source-specific exposure, such as wood smoke. These will be addressed through the following specific aims: Aim 1 is to investigate the association between urinary 1-OHP and oxidative stress in a low-level air pollution setting. A cohort of 305 pregnant individuals in western New York was studied to evaluate the relationships between 1-OHP and oxidative stress biomarkers, including malondialdehyde (MDA) and 8-hydroxy-2'-deoxyguanosine (8-OHdG). In a previous study, 1-OHP was found to be associated with ambient PM2.5. Thus, a significant association between 1-OHP and MDA or 8-OHdG would suggest that 1-OHP serves as a biomarker of PAH inhalation exposure capable of reflecting increased oxidative stress under these conditions. Biomarkers in urine samples collected across trimesters were analyzed using linear mixed-effects models, with adjustments for gestational stage, maternal age, lifestyle, and socioeconomic factors. Aim 2 focuses on developing and validating biomarkers for assessing longer-term air pollution exposure. An analytical method for detecting hemoglobin adducts of benzo[a]pyrene (BaP) and phenanthrene (PHE) was developed and validated. This was accomplished by analyzing blood samples from 235 pregnant individuals from the same cohort studied in Aim 1. The hydrolysis products of these adducts, benzo[a]pyrene-tetrols (BaPT) and phenanthrene-tetrols (PHET), were concentrated, purified, and quantified using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Cumulative ambient exposure to PM2.5 and NO2 during the gestational weeks preceding blood collection was estimated, and associations with PAH biomarkers were assessed using linear mixed-effects models, controlling for relevant covariates. Aim 3 focuses on developing and validating an LC-MS/MS method for the simultaneous quantification of OH-PAHs and PAH-CAs. Validation was conducted in a field study involving eight healthy participants exposed to a 4-hour woodsmoke session. Urine samples were collected at baseline and 2, 4, 6, 8, 12, 24 hours post-exposure. PAH metabolites were quantified, and their temporal changes were analyzed. Correlations between urinary metabolites and external exposure measures, including black carbon in the air and PAHs measured in wristbands, were also evaluated. The findings of Aim 1 indicate that at an average PM2.5 exposure level of 6.60 µg/m³ (well below the current 24-hour NAAQS standard of 35 µg/m³ and annual standard of 9 µg/m³), 1-OHP was significantly associated with PM2.5 exposure and increased MDA levels, with each interquartile range (IQR) increase corresponding to a 7.73% rise (95% CI: 3.18%, 12.3%), particularly during the first and second trimesters. In this low-level exposure cohort, 1-OHP can serve as a biomarker for identifying critical windows of air pollution exposure linked to oxidative stress effects. In the same cohort, we observed positive associations between PHET concentrations and cumulative PM2.5 exposure during gestational weeks 12–17, as well as between BaPT concentrations and cumulative PM2.5 exposure during gestational weeks 3–16 prior to sample collection. Each IQR increase in 14-week PM2.5 exposure (1.26 μg/m³) was associated with a 9.02% increase in PHET levels (95% CI: 0.30%, 17.7%) and a 12.8% increase in BaPT levels (95% CI: 1.09%, 23.5%). These findings highlight the potential of PAH-hemoglobin adducts as biomarkers for longer-term exposure (spanning weeks to four months) to ambient PM2.5, establishing their value in assessing long-term exposure to air pollution containing PAHs. In the Aim 3 field study, urine samples were collected from eight participants exposed to a 4-hour woodsmoke session; urinary 1-pyrene-carboxylic acid (1-PYRCA) levels significantly increased at 4 hours (96.9%, 95% CI: 2.60–101%), 6 hours (96.8%, 95% CI: 5.85–107%), and 8 hours (92.5%, 95% CI: 3.59–99.2%) post-exposure, returning to baseline by 24 hours. Strong correlations were identified between 1-PYRCA and ambient black carbon (r = 0.69) and between 1-PYRCA and wristband PAH levels (r = 0.54). In contrast, other urinary PAH metabolites showed no significant changes following woodsmoke exposure. These findings highlight the specificity and sensitivity of 1-PYRCA as a biomarker for assessing short-term woodsmoke exposure. In conclusion, the findings of this dissertation (Aim 1) support the use of urinary 1-hydroxypyrene as an exposure biomarker that is associated with oxidative stress and with low levels of ambient PM2.5. This research introduces two novel types of PAH exposure biomarkers, demonstrating their effectiveness in assessing long-term exposure to low-level ambient air pollution (Aim 2) and wood smoke exposure (Aim 3). These results not only highlight the potential of PAH metabolites as valuable quantitative tools for exposure assessment but also expand their application in investigating health effects under contemporary conditions.