Vegetation-facilitated removal kinetics and transformation of organic biocides

Claire Penrose Muerdter

doi:10.17077/etd.005807

The purpose of this study was to determine the vegetative uptake rates and relevant plant transformation processes of water-soluble organic biocides, to inform public health and maintain environmental quality. My central hypothesis is that plant uptake and metabolism are critical to determining the environmental fate of small, nitrogenous, water-soluble organic biocides. This work was addressed through a critical review paper and three main research objectives.The critical review publication addresses plant uptake and metabolism of a wide variety of pollutants in the setting of bioretention treatment of stormwater, an important real-world example of plant processing of humanmade chemicals. Vegetation can be a critical part of the removal of pollutants in stormwater, e.g., nitrogen and trace organic contaminants. This chapter describes the latest knowledge on this topic, as well as other roles of vegetation in bioretention such as hydrologic impacts and ancillary benefits, to provide application context for the remainder of the dissertation. As of submission of this thesis, the critical review publication has been cited over 60 times (per Google Scholar). The first research objective was to quantify the impact of duckweed and its associated microbes on neonicotinoid pesticide removal from water. I collected duckweed from a local agricultural pond and quantified its impact on dissolved neonicotinoid concentration and transformation. Neonicotinoids are the most widely used insecticides in the world and are commonly measured in aquatic environments, including freshwater wetlands. We report for the first time the synergistic transformation of neonicotinoids by a Lemna duckweed and associated microbes. Imidacloprid and thiacloprid were removed at statistically indistinguishable rates (0.63±0.07 and 0.62±0.05 day−1, respectively) from hydroponic medium only when in the presence of both duckweed and its associated microbial community. As evidence for this duckweed−microbial synergy, experiments with surface-sterilized duckweed, duckweed-associated microbes, pond water microbes alone, and two other plant species (Typha sp. and Ceratophyllum demersum) did not yield significant neonicotinoid removal beyond initial biomass sorption. Degradation of imidacloprid and thiacloprid by the duckweed−microbial system generated multiple, known neonicotinoid metabolites (desnitro-imidacloprid, imidacloprid urea, thiacloprid amide, and 6-chloronicotinic acid). Measured metabolites with increased insect or vertebrate toxicity were either absent (imidacloprid olefin) or present only in small amounts (desnitro-imidacloprid; <1% of the parent). The neonicotinoid parent and metabolite mass balance did not fully account for total neonicotinoid removal, suggesting mineralization and/or other unidentified transformation products with unknown toxicity. This novel duckweed- and microbe-facilitated neonicotinoid degradation may represent an important contribution to the environmental fate of neonicotinoids. This chapter has been published. The second research objective was to quantify the rapid uptake of several isothiazolinone biocides by hydroponically grown Arabidopsis plants, and the plant metabolism of one of the compounds, BIT. Isothiazolinones biocides are water-soluble, low molecular weight, nitrogenous compounds widely-used to prevent microbial growth in a variety of applications including personal care products and building façade materials. Because isothiazolinones from buildings wash off and enter stormwater, interactions with terrestrial plants may represent an important part of the environmental fate of these compounds. Using the model plant Arabidopsis thaliana under hydroponic conditions, we observed rapid (within 24 hours), plant-driven removal of four commonly used isothiazolinones: benzisothiazolinone (BIT), chloromethylisothiazolinone (CMI), methylisothiazolinone (MIT), and octylisothiazolinone (OIT). No significant differences in uptake rate occurred between the four compounds; therefore, BIT was used for further investigation. BIT uptake by Arabidopsis was concentration-dependent and decreased over the measured range (8–2,127 µg/L), indicating transporter-mediated substrate inhibition, instead of rising to a steady reaction rate as in classical Michaelis-Menten kinetics. BIT uptake was also minimally impacted by multiple BIT spikes (25% increase with the second spike), suggesting that uptake is constituently active. BIT plant uptake was robust, with uptake rate unaffected by multiple inhibitors (competitive, plant uptake, or plant metabolism). Following Arabidopsis uptake, the major metabolites in the plant tissue followed known detoxification pathways, generating multiple novel metabolites: hydroxylated BIT, an amino acid conjugate, and a glutathione conjugate. Nicotinic acid production also increased in BIT-exposed plant tissue, indicating that BIT impacted normal plant metabolic functions beyond direct formation of xenobiotic conjugates. The hydroxylated BIT and glutathione-BIT conjugates also were present in the hydroponic medium, likely through plant excretion. The rapid plant-driven isothiazolinone removal in this work indicates that plant-isothiazolinone processes may be relevant to the environmental fate of these compounds in stormwater, and may represent overall detoxification. The final research objective was to describe the plant uptake and metabolism of a variety of benzimidazole fungicides and two benzotriazole anti-corrosive compounds. Experimentally determining plant uptake for every relevant human chemical in the environment is impractical. There is a critical need to illuminate the role of specific functional groups on contaminants of emerging concern to enhance predictive power for compounds that may be taken up by plant transporters, an emerging area. In this work, we used benzimidazole as a representative molecule with a suite of derivatives that differ by a single functional group, to probe the impact of functional group electrostatic nature and position on plant uptake and metabolism using the hydroponic model plant Arabidopsis thaliana. Two benzotriazole molecules were also used for comparison, as the base chemical structure of benzotriazoles only differs by one nitrogen from the base chemical structure of benzimidazoles. The greatest plant uptake rates occurred with an electron-withdrawing functional group at the 2-position and no functional group at the 1-position. Electron-donating groups at the 2-position still generated some plant uptake, indicating possible steric effects between the chemical and transporter protein. An electron-donating group at the 1-position significantly slowed uptake for both base molecules (benzimidazole and benzotriazole). For the unsubstituted base molecules, the additional heterocyclic nitrogen in benzotriazole increased plant uptake compared to benzimidazole. Compounds with significantly different uptake rates followed similar metabolic pathways, including hydroxylation, glutathione conjugation, and amino-acid conjugation to generate auxin-like metabolites. Endogenous plant molecules involved in glutathione production also increased. This study provides novel insights into the impact of specific functional groups on plant uptake, with implications for environmental fate and consumer exposure. Taken together, this work demonstrates the importance of vegetation in removing small, organic, nitrogenous biocide molecules from water. The new information about plant uptake rates for a variety of compounds, including often overlooked protein-mediated transport and the importance of different functional groups in plant uptake, can inform anthropogenic chemical plant uptake models and understanding of the environmental and agricultural fate of these compounds. The plant metabolites of these compounds generally represent detoxification vs. the parent compound. Multiple novel metabolites are presented, and molecules with similar chemical structure but different plant uptake rates are shown to follow similar metabolic pathways in the plant. Taken together, these results suggest that plants can take up and detoxify these biocides, whether in an environmental context or in agricultural settings with wastewater reused for irrigation.

Vegetation-facilitated removal kinetics and transformation of organic biocides

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