Dawn Reinhold

Assessment of plant-driven removal of emerging organic pollutants by duckweed

Constructed treatment wetlands have the potential to reclaim wastewaters through removal of trace concentrations of emerging organic pollutants, including pharmaceuticals, personal care products, and pesticides. Flask-scale assessments incorporating active and inactivated duckweed were used to screen for plant-associated removal of emerging organic pollutants in aquatic plant systems. Removals of four of eight pollutants, specifically atrazine, meta-N,N-diethyl toluamide (DEET), picloram, and clofibric acid, were negligible in all experimental systems, while duckweed actively increased aqueous depletion of fluoxetine, ibuprofen, 2,4-dichlorophenoxyacetic acid, and triclosan. Active plant processes affecting depletion of experimental pollutants included enhancement of microbial degradation of ibuprofen, uptake of fluoxetine, and uptake of degradation products of triclosan and 2,4-dichlorophenoxyacetic acid. Passive plant processes, particularly sorption, also contributed to aqueous depletion of fluoxetine and triclosan. Overall, studies demonstrated that aquatic plants contribute directly and indirectly to the aqueous depletion of emerging organic pollutants in wetland systems through both active and passive processes.

Phytoaccumulation of antimicrobials from biosolids: impacts on environmental fate and relevance to human exposure.

Triclocarban and triclosan, two antimicrobials widely used in consumer products, can adversely affect ecosystems and potentially impact human health. The application of biosolids to agricultural fields introduces triclocarban and triclosan to soil and water resources. This research examined the phytoaccumulation of antimicrobials, effects of plant growth on migration of antimicrobials to water resources, and relevance of phytoaccumulation in human exposure to antimicrobials. Pumpkin, zucchini, and switch grass were grown in soil columns to which biosolids were applied. Leachate from soil columns was assessed every other week for triclocarban and triclosan. At the end of the trial, concentrations of triclocarban and triclosan were determined for soil, roots, stems, and leaves. Results indicated that plants can reduce leaching of antimicrobials to water resources. Pumpkin and zucchini growth significantly reduced soil concentrations of triclosan to less than 0.001 mg/kg, while zucchini significantly reduced soil concentrations of triclocarban to 0.04 mg/kg. Pumpkin, zucchini, and switch grass accumulated triclocarban and triclosan in mg per kg (dry) concentrations. Potential human exposure to triclocarban from consumption of pumpkin or zucchini was substantially less than exposure from product use, but was greater than exposure from drinking water consumption. Consequently, research indicated that pumpkin and zucchini may beneficially impact the fate of antimicrobials in agricultural fields, while presenting minimal acute risk to human health.

Biosolid-borne tetracyclines and sulfonamides in plants

Tetracyclines and sulfonamides used in human and animal medicine are released to terrestrial ecosystems from wastewater treatment plants or by direct manure application. The interactions between plants and these antibiotics are numerous and complex, including uptake and accumulation, phytometabolism, toxicity responses, and degradation in the rhizosphere. Uptake and accumulation of antibiotics have been studied in plants such as wheat, maize, potato, vegetables, and ornamentals. Once accumulated in plant tissue, organic contaminants can be metabolized through a sequential process of transformation, conjugation through glycosylation and glutathione pathways, and ultimately sequestration into plant tissue. While studies have yet to fully elucidate the phytometabolism of tetracyclines and sulfonamides, an in-depth review of plant and mammalian studies suggest multiple potential transformation and conjugation pathways for tetracyclines and sulfonamides. The presence of contaminants in the vicinity or within the plants can elicit stress responses and defense mechanisms that can help tolerate the negative effects of contaminants. Antibiotics can change microbial communities and enzyme activity in the rhizosphere, potentially inducing microbial antibiotic resistance. On the other hand, the interaction of microbes and root exudates on pharmaceuticals in the rhizosphere can result in degradation of the parent molecule to less toxic compounds. To fully characterize the environmental impacts of increased antibiotic use in human medicine and animal production, further research is essential to understand the effects of different antibiotics on plant physiology and productivity, uptake, translocation, and phytometabolism of antibiotics, and the role of antibiotics in the rhizosphere.

Phytoremediation of Fluorinated Agrochemicals by Duckweed

In natural and engineered systems, aquatic plants actively remediate agricultural wastes and chemicals. Characterization of agrochemical removal by aquatic plants is essential for improved design of phytoremediation systems for polluted surface waters. To measure removal of fluorinated agrochemicals by duckweed, duckweed was exposed to fluorinated phenols in batch reactors. Aqueous fluorinated phenol concentration was regularly sampled and analyzed (HPLC/DAD/MS). After a 50 h exposure, duckweed activity was quantified using oxygen production assessment to determine whether duckweed was inhibited. Removal of fluorinated phenols by duckweed was rapid, with pseudo-first-order uptake rates of 0.21 to 0.85 L d-1 for fluoro- and trifluoromethylphenols. Uptake rates of fluorinated phenols were compound-specific and appeared to depend on factors affecting rates of enzymatic processing. However, attempts to correlate removal rates with chemical parameters were unsuccessful. The positioning and type of fluoro-substituents on the phenol ring were the most indicative parameters of uptake rates. TFM, a lampricide used in the Great Lakes, was the only fluorinated phenol that was not uptaken by duckweed. Negligible uptake of TFM by duckweed was not attributed to presence of the nitro- or trifluoromethyl- groups, as 2,4,6-trinitrotoluene and 3-trifluoromethylphenol were uptaken. Although uptake rates were not easily predicted by known chemical parameters, uptake rates indicated that duckweed plays an important role in phytoremediation of fluorinated agrochemicals in surface waters.

Modeling Escherichia coli removal in constructed wetlands under pulse loading

Manure-borne pathogens are a threat to water quality and have resulted in disease outbreaks globally. Land application of livestock manure to croplands may result in pathogen transport through surface runoff and tile drains, eventually entering water bodies such as rivers and wetlands. The goal of this study was to develop a robust model for estimating the pathogen removal in surface flow wetlands under pulse loading conditions. A new modeling approach was used to describe Escherichia coli removal in pulse-loaded constructed wetlands using adaptive neuro-fuzzy inference systems (ANFIS). Several ANFIS models were developed and validated using experimental data under pulse loading over two seasons (winter and summer). In addition to ANFIS, a mechanistic fecal coliform removal model was validated using the same sets of experimental data. The results showed that the ANFIS model significantly improved the ability to describe the dynamics of E. coli removal under pulse loading. The mechanistic model performed poorly as demonstrated by lower coefficient of determination and higher root mean squared error compared to the ANFIS models. The E. coli concentrations corresponding to the inflection points on the tracer study were keys to improving the predictability of the E. coli removal model.

Phytoaccumulation of antimicrobials by hydroponic Cucurbita pepo.

Consumer use of antimicrobial-containing products continuously introduces triclocarban and triclosan into the environment. Triclocarban and triclosan adversely affect plants and animals and have the potential to affect human health. Research examined the phytoaccumulation of triclocarban and triclosan by pumpkin (Cucurbita pepo cultivar Howden) and zucchini (Cucurbita pepo cultivar Gold Rush) grown hydroponically. Pumpkin and zucchini were grown in nutrient solution spiked with 0.315 μg/mL triclocarban and 0.289 μg/mL triclosan for two months. Concentrations of triclocarban and triclosan in nutrient solutions were monitored weekly. At the end of the trial, roots and shoots were analyzed for triclocarban and triclosan. Research demonstrated that pumpkin and zucchini accumulated triclocarban and triclosan. Root accumulation factors were 1.78 and 0.64 and translocation factors were 0.001 and 0.082 for triclocarban and triclosan, respectively. The results of this experiment were compared with a previous soil column study that represented environmentally relevant exposure of antimicrobials from biosolids and had similar root mass. Plants were not as efficient in removing triclocarban and triclosan in hydroponic systems as in soil systems. Shoot concentrations of antimicrobials were the same or lower in hydroponic systems than in soil columns, indicating that hydroponic system does not overpredict the concentrations of antimicrobials.

Development and application of oxygen production rate assessment to uptake of fluorinated organics by Lemna minor.

The effects of contaminant concentration on contaminant removal by Lemna minor were examined under elevated plant densities typical of full surface coverage. Uptake of 3-fluorophenol and 3-trifluoromethylphenol by L. minor was determined at concentrations from 10 to 1750 microM. Because standardized toxicity tests typically do not measure plant activity of L. minor at high plant densities, an oxygen production rate (OP) assessment was developed with consideration of experimental parameters that may inhibit OP by L. minor, including concentrations of carbonate and phosphate buffer in the media, time of assessment, and mass of L. minor. The OP by L. minor was inhibited by increasing media pH and mass of L. minor. The developed assessment used 0.5 g of L. minor per 60 ml of modified Standard Methods medium. A total of 30 h was needed to determine toxicity of fluorinated phenols, with an exposure period of 24 h, an incubation time of 6 h, and negligible analysis time. At concentrations from 10 to 1750 microM, 3-trifluoromethylphenol exhibited a sigmoidal toxicity response. However, 3-fluorophenol was neither toxic nor inhibitory at the experimental concentrations tested. Substantial decreases in uptake rate constants were observed for increasing concentrations of both contaminants, indicating that concentration may be a more important indicator of uptake by L. minor compared with decreasing plant activity because of toxicity.

Phytoremediation of fluorinated pollutants by duckweed

In natural and engineered systems, aquatic plants actively remediate agricultural wastes and pollutants. Characterization of pollutant removal by aquatic plants is essential for improved design of phytoremediation systems for polluted surface waters. To measure removal of fluorinated pollutants by duckweed, duckweed was exposed to fluorinated pollutants in batch reactors. Aqueous pollutant concentration was regularly sampled and analyzed (HPLC/DAD/MS). After a 50-hr exposure, duckweed activity was quantified using oxygen production assessment to determine whether duckweed was inhibited. Removal of fluorinated pollutants by duckweed was rapid, with pseudo-first-order uptake rates of 0.21 – 0.85 d-1 for fluoro- and trifluoromethyl- phenols. Uptake rates of fluorinated pollutants were pollutant-specific and appeared to depend on factors affecting rates of enzymatic processing. However, attempts to correlate removal rates with chemical parameters were unsuccessful. The positioning and type of fluoro- substituents on the phenol ring was the most indicative parameter of uptake rates. TFM, a lampricide used in the Great Lakes, was the only fluorinated pollutant that was not uptaken by duckweed. Negligible uptake of TFM by duckweed was not attributed to presence of the nitro- or trifluoromethyl- groups, as 2,4,6- trinitrotoluene and 3-trifluoromethylphenol were actively uptaken. Although uptake rates were not easily predicted by known chemical parameters, uptake rates indicated that duckweed plays an important role in phytoremediation of fluorinated pollutants in surface waters.

Callus Cultures for Phytometabolism Studies: Phytometabolites of 3-Trifluoromethylphenol in Lemnaceae Plants and Callus Cultures

Plant callus cultures have the potential to advance phytoremediation science by allowing study of cellular phytometabolism in absence of sorption, translocation, microbial degradation, and other phytoremediation processes; however, studies demonstrating the applicability of results from callus cultures to whole plants are limited. The aim of this study was to evaluate the feasability and applicability of using callus cultures to study phytometabolism. This aim was accomplished through evaluation of induction and growth of Lemnaceae callus cultures and comparison of phytometabolism in callus cultures and whole plants. Four out of eight published methods for callus culture of Lemnaceae successfully induced callus cultures that exhibited doubling times of 1.7 to 23 wks. Callus cultures and whole plants of Landoltia punctata and Lemna minor metabolized 3-trifluoromethylphenol (3-TFMP) through conjugation with glucopyranoside, malonyl-glucopyranoside, and glucopyranosyl-apiofuranoside. However, concentrations of metabolites were approximately 10 times less in callus cultures than in plants. While results demonstrated applicability of callus cultures results to whole plants, the low success rate of callus induction procedures, length of time required to produce substantial callus mass, and the low accumulation of metabolites in callus cultures may limit the feasibility of callus cultures for assessing phytometabolism.

Anaerobic Digestion of Dairy Manure Combined with Duckweed (Lemnaceae)

Anaerobic digestion (AD) has become an increasingly popular method of treating wastewater or sludges from animal feeding operations. Enhancement of biogas production in anaerobic digesters, through addition of commonly-available and under-utilized biomass, could benefit sustainability of farm-scale anaerobic digesters. Duckweed is a common aquatic plant that aggressively grows in farm ponds, lagoons, and other water bodies that receive agricultural runoff. As such, duckweed is a readily-available biomass that could be easily added to farm-scale anaerobic digesters. Therefore, research aimed to determine if biogas (methane) production could be improved by supplementing digesters with duckweed (e.g., Landoltia punctata). Increases in biogas production and rate of attaining peak biogas production were assessed in batch continuously-stirred reactors at 35oC. Varying concentrations of duckweed were added to dairy manure slurries and gas production was observed for 20-40 days. Additionally, subsequent research will assess changes in chemical oxygen demand (COD), pH, and fatty acids within manure/duckweed slurries in parallel with analyses of biogas production, with time. Preliminary results indicate that addition of duckweed, in the range of 0.5 to 2% (dry mass), enhanced methane and total gas production in dairy manure slurries; however, subsequent increases in methane and total gas production at >2% duckweed were not observed. In conclusion, addition of duckweed biomass, produced during treatment of agricultural wastewaters and runoff, to anaerobic digesters is a promising approach to enhancement of biogas production.