Grantee Research Project Results
2005 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
EPA Project Officer: Aja, Hayley
Project Period: October 17, 2001 through October 16, 2002
Project Period Covered by this Report: October 17, 2004 through October 16, 2005
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 is to provide analytical support to both the epidemiological (R829390C002) and laboratory research projects (R829390C001 and R829390C004) of the FRIENDS Children’s Environmental Health Center. This core facility support involves the development of new and improved analytical techniques and analysis of human and animal study samples. All procedures are controlled by a strict QA/QC program. The core facility has extensive experience in analyzing specific congeners of polychlorinated biphenyls (PCBs), pesticides, and heavy metals at levels normally encountered in biological and experimental samples. The core facility also provides analysis of dioxin-like activity in biological samples using a reporter gene bioassay for quantitation of total dioxin (i.e., TCDD) toxic equivalents.
Progress Summary:
Advancements in Analytical Methods
Methods for analysis of specific PCB congeners and pesticides are being improved constantly to enhance sensitivity and efficiency of sample analysis. The following method improvements were accomplished during Year 4 of the project:
Accelerated Solvent Extraction (ASE). Methods and conditions were optimized for serum, rat brain tissues, and fish tissues. A new ASE method has been developed using acidified silica, which removes up to 0.5 g of oil in a sample. This procedure has been effective in extracting fish tissue and corn oil (dosing solution) samples. Disposable solid phase extraction cartridges also have been integrated into all ASE methods. The Dionex ASE has been upgraded to allow for programmed solvent switching, and we also have computerized the system, which gives us more programming capability and makes the unit more user-friendly to operate.
Some contamination problems were encountered following the analysis of fish tissue samples and have been determined to be attributed to the extraction cell frits. An ultrasonic clean-up procedure has been found to be successful in cleaning and removing contamination of the frits. Separate cells also are now being used for fish and serum samples. Additional work has been done to determine the elution characteristics of coplanar PCBs through the ASE system and Florisil® adsorption columns.
Development of Method for Polybrominated Diphenyl Ethers (PBDE) Analysis. Preliminary work has been done looking at gas chromatography-mass spectrometry (GCMS) conditions and determining sensitivity using electron impact GCMS. It appears as if the sensitivity of the GCMS operating in this mode may be sufficient for determining higher level PBDEs in fish samples but would be unable to determine the lower levels found in serum. To determine the lower levels in serum, the instrument would have to be operated in the negative chemical ionization mode, which would require an upgrade on the GCMS instrument. The software and electronics on this instrument have been upgraded to allow for faster scan rates and increased sensitivity in the SIM mode and is more inert for active compounds.
Evaluation of Apex Large Volume Injector (LVI). We have been evaluating the Apex Large Volume Injector (LVI), which will allow for larger sample volumes of extracts to be injected into the gas chromatograph. The objective is to increase the size of the sample extract being analyzed (10-20 μL vs. 1 μL), resulting in lower detection limits for samples. This is particularly important for rat brain tissue samples, where sample weights are only in the mg range. This also may reduce sample preparation time by decreasing the amount of solvent evaporation necessary for final sample extracts. Preliminary work evaluating LVI parameters with PCB mixed standards has shown that the injector is capable of increasing the sensitivity by allowing larger volume samples to be injected and that the response is linear. Problems with non-linearity of matrix samples (i.e., serum extracts) and broadening of chromatographic peaks have been encountered, however, and further experiments are being conducted to determine the cause.
Participation in the Artic Monitoring and Assessment Programme Ring Test for Persistent Organic Pollutants in Human Serum. The laboratory is participating in a proficiency test program for PCB congeners and selected pesticides (DDE and hexachlorobenzene) in human serum. The program is administered by the Centre de Toxicologie du Quebec. There are three rounds of proficiency samples/year. The laboratory has completed the first round and obtained the highest possible rating of excellent. Testing will be expanded to include lipids in serum.
Analysis of Center Samples
A total of approximately 1,200 analyses were performed in Year 4 of the project for specific PCB congeners and pesticides. This includes actual samples (human serum, rat brain tissues, and fish tissues) received from Center projects, as well as samples for the development of new methods, evaluation of new sample matrices and new equipment, and quality control/calibration samples. The number of study samples analyzed for PCB congeners and pesticides was 24 human serum samples, 71 rat tissue (brain, liver), 24 rat serum samples, 34 fish tissue/commodity samples, and 9 PCB dosing solutions in corn oil. Dosing solutions also were analyzed for TCDD-like activity using the luciferase reporter gene assay. Additionally, the human serum samples were analyzed for methylmercury and lipids.
Future Activities:
Sample Analysis
Analyze samples in support of Center projects, including human serum and rat brain, liver, and serum samples for specific PCB congeners, DDE, Mirex, and methylmercury.
Method Improvements
- Assess GCMS methods for determining PBDEs in human serum and fish samples. Determine extraction efficiency, chromatographic cleanup elution profiles, and gas chromatographic conditions for PDBEs. Methods then will be validated for specific sample matrices. The ultimate goal is to develop a method that will allow for simultaneous extraction/determination of both PCBs and PBDEs.
- Continue evaluation of LVI System operating parameters for the GC analysis of PCB congeners. The objective is to increase the size of the sample extract being analyzed, which will result in lower detection limits for samples.
- Establish more efficient means to transfer and process raw data/chromatograms from gas chromatographic software to Excel spread sheets.
- Publish analytical methods that have been developed at the State University of New York at Buffalo Translational Research Center analytical laboratory for the determination of PCBs in serum and fish tissues for the FRIENDS project.
- Develop methods for the extraction and determination of hydroxylated PCBs and metabolites in serum.
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 metalsProgress 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.