Analytical Methods
Select target contaminants and analytical methods when designing the contaminant monitoring program. Select analytical methods based on the following criteria:
- Technical merit - Methods should be specific for the target analytes of concern, be directly applicable to tissue samples, and based on current, validated analytical techniques that are widely accepted by the scientific community.
- Sensitivity - Method detection and quantitation limits should be sufficiently low to allow reliable quantitation of the target analytes of concern.
- Data quality - The accuracy and precision should be adequate to ensure that analytical data are of acceptable quality for program objectives.
- Cost-efficiency - Costs should not be unreasonably high.
Suggested Analytical Methods for Target Analytes
| Analyte Class | Analytical Method |
|---|---|
| Arsenic | EPA Method 1632A |
| Cadmium | EPA Method 200.8, Rev. 5.4, with sample preparation by SW-846 Method 3050B, or other suitable strong acid digestion procedure applicable to tissues |
| Lead | EPA Method 200.8, Rev. 5.4, with sample preparation by SW-846 Method 3050B, or other suitable strong acid digestion procedure applicable to tissues |
| Mercury* | EPA Method 1631E, with preparation by the appendix in EPA Method 1631B |
| Selenium | EPA Method 200.8, Rev. 5.4, with sample preparation by SW-846 Method 3050B, or other suitable strong acid digestion procedure applicable to tissues |
| Tributyltin | EPA Method 8323 |
*Method 1631E is used for total mercury, but the assumption is that it is all (or mostly) methylmercury. This is a conservative assumption and costs less than analyzing only for methylmercury. SW-846 Method 7473 could be used for mercury analysis, but QC limits would need to be provided by the jurisdiction since Method 7473 does not contain firm QC limits. Method 1631E (with the fish sample preparation procedure in 1631B) contains firm QC requirements and has been widely validated and used across many studies and laboratories.
| Analyte Class | Analytical Method |
|---|---|
| Amphetamine† (pharmaceutical) | EPA Method 1694 |
| Aroclors | EPA Method 1656A or SW-846 Method 8082A with extraction methods of 3540C, 3541, 3545A, or 3546 |
| BDE-47 | EPA Method 1614A |
| Dioxins/furans | EPA Method 1613B |
| Organochlorine pesticides | EPA Method 1656A or SW-846 Method 8081 with extraction methods of 3540C, 3541, 3545A, or 3546 |
| Organophosphorus pesticides | EPA Method 1657A or SW-846 Method 8141B with extraction methods of 3540C, 3541, 3545A, or 3546 |
| Oxyfluorfen‡ | |
| PAHs | SW-846 Method 8310, with sample preparation by SW-846 Method 3540C, 3541, 3545A, or 3546 |
| PCB congeners | EPA Method 1668C or EPA Method 1628 |
| PFAS | EPA Method 1633 |
†Amphetamines are controlled substances and a laboratory must have a license to possess controlled substances to perform the test.
‡There is not an EPA approved method for oxyfluorfen analysis. A research paper describes a QuEChERS multi-residue GC-MS/MS method developed for determining pesticides in tissue (Camara et al., 2020). Method SW-846 8151A does not list oxyfluorfen or tissue, but this could be used as a basis for a method for tissue analysis.
| Analyte Class | Analytical Method |
|---|---|
| Cyanotoxins (microcystins, BMAA, DABA) | For microcystins, a method using 2-methoxy-3-methyl-4-phenylbutyric acid (MMPB) procedure is under development by the EPA. |
The 1600-Series methods are recommended because many explicitly address the preparation and analysis of tissue samples, but other options may be appropriate as well. Alternative techniques for analyses or methods from other sources may be used if acceptable detection limits, accuracy, and precision can be demonstrated. Project planners should select analytical methods that are most appropriate for their programs based on available resources, experience, program objectives, and data quality requirements.
The National Environmental Method Index is a searchable database for identifying and comparing analytical and field methods for all phases of environmental monitoring. Some methods are available to download. Summary information for each method includes estimated sample costs and detection limits.
The selection of an analytical method is often influenced by its cost. Costs tend to increase with increased sensitivity (i.e., lower detection limits) and reliability (i.e., accuracy and precision). Analytical costs are also dependent on the number of samples, the requested turnaround time, the number and type of analytes requested, the level of QC effort, and the amount of support documentation requested.
Lipid Analysis
The decision to collect lipid data is a project-specific one. Some monitoring programs analyze the lipid content of fish because some important organic contaminants, such as PCBs, are lipophilic. Normalizing concentrations of lipophilic contaminants to the lipid content may be useful in studies of trophic transfer and biomagnification. However, it is not useful or advisable to lipid-normalize any contaminants monitored for programs geared towards human health and fish consumption advisories or for contaminants that are not lipophilic (e.g., metals, PFAS).
If lipid data are of interest to the project, the most commonly used procedures for lipid analysis are based on solvent extraction of tissue samples, evaporation of the solvent, and gravimetric determination (weighing) of the lipid residue. If lipid-normalized data will be compared to that from other studies, then it is important to know what procedures and solvents were used in those other studies to ensure consistency of reported results among fish contaminant monitoring programs. Dichloromethane (aka methylene chloride) is a commonly used solvent, but other solvents (e.g., hexane) have been used in published studies. If PCB congener analysis is performed using EPA Method 1668C, the method describes how lipid analysis can be performed using a small portion of the same dichloromethane sample extract that will be analyzed for the PCBs.
In addition, when developing and validating project-specific procedures for homogenizing fish tissue samples, lipid analysis can be used as a surrogate to confirm homogeneity of the samples by testing lipids in three or more aliquots of the bulk homogenized tissue. The relative standard deviation of the replicate lipid results can be used to assess the effectiveness of the homogenization process. This can be used as part of the “start-up tests” for a new sample preparation laboratory. When used only for the purpose of testing sample homogeneity, the choice of extraction solvent may not be a significant concern, provided that the same solvent is used through the course of the study.