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Rationale and Significance: Many options are being considered for the control of invasive aquatic species in ship's ballast water. Any adopted control measure must be effective, affordable, safe and environmentally benign. Chemical biocides show perhaps the best promise at satisfying the first three criteria. However residual chronic sub-lethal toxicity of the parent compounds and/or degradation products has yet to be fully investigated. In view of the fact that chemicals containing -OH groups are implicated as endocrine disruptors in some cases, this potential will be investigated herein. Until lower effects thresholds are determined a proper risk assessment of these compounds cannot be completed. Our project addresses the RFP priority regarding assessing the adverse environmental impacts of using biocides for Control and Mitigation. Research Goals and Objectives: We propose to investigate the chronic sub-lethal toxicities of three quinone compounds in order to characterize the environmental threat that their discharge might pose to non-target organisms if used as ballast treatment biocides. We will use the standard chronic test procedures to determine no observable effect concentration (NOEC) and lowest observable effect consideration (LOEC) levels for several endpoints in representative freshwater and estuarine vertebrate and invertebrate species. The data generated will be made available for future risk assessment models of biocide treated ballast water discharge. In addition we will chemically characterize the major residuals/degradates of the parent compounds (including half-life estimates) and expand upon our initial test procedures by using realistic exposure regimes (i.e. mimicking various ballast water holding times) and coupling chemical analyses with determination of chronic sub-lethal toxicities, NOEC and LOEC levels of the residual compounds / degradates. The possibility that residuals/degradates may retain sub-lethal toxicity is an area that has yet to be fully investigated. Summary of Work Proposed: Quinones selected for study include two currently proposed as ballast water treatment biocides, namely, menadione (2-methl-1,4-naphthoquinone) and juglone (5,hydroxy- 1,4-naphthaquinone) and a third, plumbagin (2-methyl-5-hydroxy-1,4-naphthoquinone) which is currently under investigation as an alternative biocide. We will conduct a series of laboratory assays using standard EPA Whole Effluent Toxicity (WET) test methods to examine the chronic, sub-lethal toxicities of the parent compounds and residuals/degradates and to determine LOCE and NOEC levels for several endpoints in four test species using a range of possible chemical discharge concentrations. Various biomarkers will be used from the molecular through organ systems, including investigations of endocrine disruption, oxidation stress and genetic damage. Detailed chemical analyses of residuals/degradates will be carried out by HPLC and GS-MS. Outcomes/Benefits: By employing freshwater and estuarine vertebrate and invertebrate species the suite of assays proposed will broadly characterize acceptable discharge levels of the various quinone biocides into coastal, estuarine and inland waters. We will ascertain if residuals/degradates of proposed ballast water treatment biocides retain chronic sub-lethal toxicity to any of the selected species. While we have extensive knowledge of acute toxicity for these products, their potential for chronic sub-lethal toxic effects has yet to be fully investigated. Ultimately NOEC and LOEC levels will be determined for the parent biocides and their degradates. Such lower effects thresholds will aid in future risk assessment modeling of biocide treated ballast water discharges. Education and Outreach / Technology Transfer: Information gained will be made available in peer-reviewed papers and presentations at appropriate scientific meetings and workshops as they accrue. The facilities of MDSG and the extension program (CBL) will be employed to reach non-peer users, using media such as the SG website and newsletter publications.