Grantee Research Project Results
2003 Progress Report: FRIENDS Analytical Toxicology Core Facility
EPA Grant Number: R829390C003Subproject: this is subproject number 003 , established and managed by the Center Director under grant R829390
(EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).
Center: Water Innovation Network for Sustainable Small Systems
Center Director: Reckhow, David A.
Title: FRIENDS Analytical Toxicology Core Facility
Investigators: Kostyniak, Paul J.
Institution: The State University of New York at Buffalo , University of Illinois Urbana-Champaign
Current Institution: The State University of New York at Buffalo
EPA Project Officer: Aja, Hayley
Project Period: October 17, 2001 through October 16, 2002
Project Period Covered by this Report: October 17, 2002 through October 16, 2003
RFA: Centers for Children's Environmental Health and Disease Prevention Research (2001) RFA Text | Recipients Lists
Research Category: Human Health , Children's Health
Objective:
The objective of the Fox River Environment and Diet Study (FRIENDS) Analytical Toxicology Core Facility (ATCF) is to provide analytical support to both the epidemiological (R829390C002) and animal research projects (R829390C001 and R829390C004) of the FRIENDS Children’s Environmental Health Center. The ATCF support involves the development of new analytical techniques, improvement of existing methods, and analyses of study samples. All procedures are controlled by a strict QA/QC program. This laboratory has extensive experience determining specific congeners of polychlorinated biphenyls (PCBs), pesticides, and heavy metals at ppb levels in biological samples. The ATCF also performs a reporter gene bioassay for the quantification of total dioxin (i.e., 2,3,7,8-tetrachlorodibenzo-p-dioxin [TCDD]) toxic equivalents (TEQs).
Progress Summary:
Formulation and Evaluation of Dosing Solutions
The PCB congener mix to be used for the animal studies throughout the 5-year project was formulated, analyzed, and compared to the PCB congener profiles found in Fox River fish. Fish tended to have a congener profile with a more concentrated Aroclor 1242-like pattern, in which there was a higher concentration of lower chlorinated congeners than normally is observed in fish in the eastern Great Lakes. We constructed various theoretical mixtures of PCBs from published congener contents of various Aroclors. The Aroclor mix, which predicted a profile most similar to that found in the Fox River fish, was a mixture containing 35 percent Aroclor 1242, 35 percent Aroclor 1248, 15 percent Aroclor 1254, and 15 percent Aroclor 1260, on a weight basis. PCB congener profiles of the final dosing solution were compared to the theoretical Aroclor construct and the Fox River fish profiles. As seen in Figure 1, there was good agreement in the congener profiles of the three groups.
Figure 1. Fish Versus Theoretical Versus Actual Dosing Solution Mix of Aroclors 1242/1248/1254/1260
Advances in Analytical Methods
Existing methods for analysis of PCB congeners in serum and milk have been modified and extended to other sample matrices. A major modification in the method of solvent extraction and column cleanup has been accomplished by incorporating an automated extraction process using the Dionex ASE 300 system, allowing for rapid extraction and cleanup using elevated pressure and temperature. The introduction of this technology resulted in an improvement in lower blanks, lower limits of detection, and higher recoveries of individual PCB congeners in the assay. The mean recovery for 23 individual PCB congeners and pesticides from 4 g samples of serum was 103.8 (+6.6 percent). The method also has been applied to the analysis of fish and fish products. The method avoids an acid treatment step for removal of lipids, which is common to many methods, and results in high recoveries. The mean recovery of 12 individual PCB congeners and pesticides from 5 g samples of fish was 98.4 (+7.7 percent). Chromatograms indicated no breakdown products for any of the congeners of interest.
Analysis of Foodstuffs Commonly in the Diet of the Study Population
In an attempt to assess other potential vectors of PCB exposure in the study population, imported foodstuffs, primarily from Southeast Asia, were obtained from Hmong-operated specialty stores in Green Bay. Methods of extraction have been modified for each of these sample matrices to enable PCB congener and pesticide determination. Seven of the most popular brands of fish sauce, which is eaten on a daily basis, had levels of individual PCB congeners well below 1 ng/g. Thus, the fish sauces are not a significant vector for PCB and pesticide exposure. Ten varieties of pickled and canned fish, four varieties of crab and shrimp paste, and shrimp crackers have been extracted and analyzed for PCB congeners by the method adapted for fish analysis. Chromatographic profiles indicate a number of unidentified peaks, which are being assessed by gas chromatography/mass spectrometry. Once peaks have been confirmed, the congener-specific concentrations will be calculated. In addition, methods are being finalized to assay extracts from these samples using the reporter gene assay for dioxin equivalents.
Method Modifications for PCB Metabolites
The development of methods to include the hydroxylated metabolites of various PCB congeners has been initiated. Authentic standards have been located and ordered. Modification of extraction techniques are being performed to include these analytes in the congener-specific PCB analysis using the Dionex ASE 300 system.
Measurement of Dioxin Equivalents
The dosing solutions and the fish samples analyzed for individual PCB congeners are being analyzed for total dioxin equivalents using the reporter gene bioassay, which measures activity of dioxin-like compounds, including polychlorinated dibenzo-p-dioxins and dibenzofurans, with chlorine on at least the 2,3,7,8 positions and the coplanar (nonortho) and mono-ortho substituted PCBs that have biological and toxicological activities similar to TCDD, the most potent halogenated aromatic hydrocarbon. The bioassay requires that unknown samples (i.e., chemicals in dosing solution, fish, or other foodstuffs) be dissolved quantitatively in the cell culture media. In the case of the dosing solution, the complete extraction of the dosing solution is needed to assure that the culture media contains the dioxin-like chemicals, without residual corn oil vehicle. Although further optimization is needed for sample preparation, preliminary results in the bioassay indicated that the dosing solutions do contain dioxin TEQs of at least 60 ppb in the corn oil from the dosing solution D that contained 31.4 mg PCBs/g oil.
Future Activities:
The investigators did not report any future activities.
Supplemental Keywords:
children’s health, disease and cumulative effects, ecological risk assessment, susceptibility, sensitive population, toxicology, Fox River, PCBs, exposure assessment, heavy metals, methylmercury, pesticides, fish consumption,, RFA, Scientific Discipline, Health, ENVIRONMENTAL MANAGEMENT, Geographic Area, POLLUTANTS/TOXICS, Midwest, Toxicology, Health Risk Assessment, Epidemiology, Chemistry, Chemicals, Risk Assessments, Susceptibility/Sensitive Population/Genetic Susceptibility, Disease & Cumulative Effects, Ecological Risk Assessment, Molecular Biology/Genetics, Children's Health, genetic susceptability, Biology, Risk Assessment, human data, neurotoxic, behavioral assessment, PCBs, pesticides, animal model, electrochemical detection, children, neurotoxicity, motor development, behavioral deficits, methylmercury, PCB, cognitive development, human exposure, Wisconsin (WI), animal studies, reproductive health, exposure assessment, biomedical research, heavy metalsRelevant Websites:
http://www.cvm.uiuc.edu/vb/friends_center/ Exit
Progress and Final Reports:
Original AbstractMain Center Abstract and Reports:
R829390 Water Innovation Network for Sustainable Small Systems Subprojects under this Center: (EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).
R829390C001 Neurobehavioral Effects of PCBs and Methylmercury in Rats
R829390C002 Perinatal PCB Exposure and Neuropsychological/Auditory Function
R829390C003 FRIENDS Analytical Toxicology Core Facility
R829390C004 Developmental Effects of PCBs and Methylmercury
The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Conclusions drawn by the principal investigators have not been reviewed by the Agency.