Document Outline

• Introduction
• Scope and Application
• Summary of Method
• Definitions
• Contamination and Interferences
• Safety
• Equipment and Supplies
• Reagents and Standards
• Sample Collection, Preservation, and Storage
• Quality Control
• Calibration and Standardization
• Procedure
• Data Analysis and Calculations
• Method Performance
• Pollution Prevention
• Waste Management
• References
• Tables and Diagrams
• Glossary
• Appendix A

Method 1630

Methyl Mercury in Water by Distillation, Aqueous Ethylation, Purge and Trap, and Cold Vapor Atomic Fluorescence Spectrometry

August, 1998

U.S. Environmental Protection Agency
Office of Water
Office of Science and Technology
Engineering and Analysis Division (4303)
401 M Street SW
Washington, D.C. 20460

Acknowledgments

This method was prepared under the direction of William A. Telliard of the Engineering and Analysis Division (EAD) within the U.S. Environmental Agency's (EPA's) Office of Science and Technology (OST). The method was prepared by Nicholas Bloom of Frontier Geosciences under EPA Contract 68-C3-0337 with the DynCorp Environmental Programs Division. Additional assistance in preparing the method was provided by DynCorp Environmental and Interface, Inc.

Disclaimer

This draft method has been reviewed and approved for publication by the Analytical Methods Staff within the Engineering and Analysis Division of the U.S. Environmental Protection Agency. Mention of trade names or commercial products does not constitute endorsement or recommendation for use. EPA plans further validation of this draft method. The method may be revised following validation to reflect results of the study.

EPA welcomes suggestions for improvement of this method. Suggestions and questions concerning this method or its application should be addressed to:

W.A. Telliard
Engineering and Analysis Division (4303)
U.S. Environmental Protection Agency
401 M Street SW
Washington, D.C. 20460
Phone: 202/260-7134
Fax: 202/260-7185

Introduction

This analytical method supports water quality monitoring programs authorized under the Clean Water Act (CWA, the "Act"). CWA Section 304(a) requires EPA to publish water quality criteria that reflect the latest scientific knowledge concerning the physical fate (e.g., concentration and dispersal) of pollutants, the effects of pollutants on ecological and human health, and the effect of pollutants on biological community diversity, productivity, and stability.

CWA Section 303 requires each state to set a water quality standard for each body of water within its boundaries. A state water quality standard consists of a designated use or uses of a water body or a segment of a water body, the water quality criteria that are necessary to protect the designated use or uses, and an antidegradation policy. These water quality standards serve two purposes: (1) they establish the water quality goals for a specific water body, and (2) they are the basis for establishing water quality-based treatment controls and strategies beyond the technology-based controls required by CWA Sections 301(b) and 306.

In defining water quality standards, the state may use narrative criteria, numeric criteria, or both. However, the 1987 amendments to CWA required states to adopt numeric criteria for toxic pollutants (designated in Section 307(a) of the Act) based on EPA Section 304(a) criteria or other scientific data, when the discharge or presence of those toxic pollutants could reasonably be expected to interfere with designated uses.

In some cases, these water quality criteria are as much as 280 times lower than those achievable using existing EPA methods and required to support technology-based permits. Therefore, EPA developed new sampling and analysis methods to specifically address state needs for measuring toxic metals at water quality criteria levels, when such measurements are necessary to protect designated uses in state water quality standards. The latest criteria published by EPA are those listed in the National Toxics Rule (58 FR 60848) and the Stay of Federal Water Quality Criteria for Metals (60 FR 22228). These rules include water quality criteria for 13 metals, and it is these criteria on which the new sampling and analysis methods are based. Method 1630 was specifically developed to provide reliable measurements of methyl mercury at EPA WQC levels.

In developing methods for determination of trace metals, EPA found that one of the greatest difficulties was precluding sample contamination during collection, transport, and analysis. The degree of difficulty, however, is highly dependent on the metal and site-specific conditions. This method is designed to preclude contamination in nearly all situations. It also contains procedures necessary to produce reliable results at the lowest ambient water quality criteria published by EPA. In recognition of the variety of situations to which this method may be applied, and in recognition of continuing technological advances, Method 1630 is performance based. Alternative procedures may be used so long as those procedures are demonstrated to yield reliable results.

Requests for additional copies of this method should be directed to:

U.S. EPA NCEPI
11209 Kenwood Road
Cincinnati, OH 45242
513/489-8190

Method 1630
Methyl Mercury in Water by Distillation, Aqueous Ethylation,
Purge and Trap, and Cold Vapor Atomic Fluorescence Spectrometry

Note: This method is performance based. The laboratory is permitted to omit any step or modify any procedure provided that all performance requirements in this method are met. The laboratory may not omit any quality control analyses. The terms “shall,” “must,” and “may not” define procedures required for producing reliable data at water quality criteria levels. The terms “should” and “may” indicate optional steps that may be modified or omitted if the laboratory can demonstrate that the modified method produces results equivalent or superior to results produced by this method.

1.0 Scope and Application

1.1 This method is for determination of methyl mercury (CH3Hg3) in filtered and unfiltered water by distillation, aqueous ethylation, purge and trap, desorption, and cold vapor atomic fluorescence spectrometry (CVAFS). This method is for use in EPA's data gathering and monitoring programs associated with the Clean Water Act, the Resource Conservation and Recovery Act, the Comprehensive Environmental Response, Compensation and Liability Act, and the Safe Drinking Water Act. The method is based on a contractor-developed method (Reference 1) and on peer-reviewed, published procedures for the determination of CH3Hg in aqueous samples, ranging from sea water to sewage effluent (References 2-7).

1.2 This method is accompanied by Method 1669: Sampling Ambient Water for Determination of Trace Metals at EPA Water Quality Criteria Levels (Sampling Method). The Sampling Method is necessary to preclude contamination during the sampling process.

1.3 This method is designed for determination of CH3Hg3 in the range of 0.02-5 ng/L and may be extended to higher levels by selection of a smaller sample size.

1.4 The ease of contaminating ambient water samples with the metal(s) of interest and interfering substances cannot be overemphasized. This method includes suggestions for improvements in facilities and analytical techniques that should maximize the ability of the laboratory to make reliable trace metal determinations and minimize contamination (Section 4.0).

1.5 The detection limit and minimum level of quantitation in this method are usually dependent on the level of background elements rather than instrumental limitations. The method detection limit (MDL; 40 CFR 136, Appendix B) for CH3Hg has been determined to be 0.02 ng/L when no background elements or interferences are present. The minimum level (ML) has been established as 0.06 ng/L. An MDL as low as 0.009 ng/L can be achieved for low CH3Hg samples by using extra caution in sample handling and reagent selection, particularly the use of “for ultra-low level only” distillation equipment.

1.6 Clean and ultraclean—The terms "clean" and "ultraclean" have been applied to the techniques needed to reduce or eliminate contamination in trace metal determinations. These terms are not used in this method because they lack an exact definition. However, the information provided in this method is consistent with the summary guidance on clean and ultraclean techniques.

1.7 This method follows the EPA Environmental Methods Management Council's "Format for Method Documentation."

1.8 This method is "performance based." The analyst is permitted to modify the method to overcome interferences or lower the cost of measurements if all performance criteria are met. Section 9.1.2 gives the requirements for establishing method equivalency.

1.9 Any modification of this method, beyond those expressly permitted, shall be considered a major modification subject to application and approval of alternate test procedures under 40 CFR 136.4 and 136.5.

1.10 This method should be used only by analysts who are experienced in the use of CVAFS techniques and who are thoroughly trained in the sample handling and instrumental techniques described in this method. Each analyst who uses this method must demonstrate the ability to generate acceptable results using the procedure in Section 9.2.

1.11 This method is accompanied by a data verification and validation guidance document, Guidance on the Documentation and Evaluation of Trace Metals Data Collected for CWA Compliance Monitoring. Data users should state data quality objectives (DQOs) required for a project before this method is used.

2.0 Summary of Method

2.1 A 100-2000 mL sample is collected directly into specially cleaned, pretested, fluoropolymer or borosilicate bottle(s) using sample handling techniques specially designed for collection of metals at trace levels (Reference 6).

2.2 For dissolved CH3Hg, samples are filtered through a 0.45-µm capsule filter.

2.3 Fresh water samples are preserved by adding 4 mL/L of pretested 11.6 M HCl, while saline samples ([Cl-] > 500 ppm) are preserved with 2 mL/L of 9 M H2SO4solution, to avoid distillation interferences caused by excess chloride.

2.4 Prior to analysis, a 45-mL sample aliquot is placed in a specially designed fluoropolymer distillation vessel, and 35 mL of the water is distilled into the receiving vessel at 125°C under N2 flow.

2.5 After distillation, the sample is adjusted to pH 4.9 with an acetate buffer and ethylated in a closed purge vessel by the addition of sodium tetraethyl borate (NaBEt ).

2.6 The ethyl analog of CH3Hg, methylethyl mercury (CH3CH3CH2Hg), is separated from solution by purging with N2 onto a graphitic carbon (Carbotrap®) trap.

2.7 The trapped methylethyl mercury is thermally desorbed from the Carbotrap® trap into an inert gas stream that carries the released methylethyl mercury first through a pyrolytic decomposition column, which converts organo mercury forms to elemental mercury (Hg0), and then into the cell of a cold-vapor atomic fluorescence spectrometer (CVAFS) for detection.

2.8 Quality is ensured through calibration and testing of the distillation, ethylation, purging, and detection systems.

3.0 Definitions

3.1 Apparatus: Throughout this method, the sample containers, sampling devices, instrumentation, and all other materials and devices used in sample collection, sample processing, and sample analysis that come in contact with the sample and therefore require careful cleaning will be referred to collectively as the Apparatus.

3.2 Dissolved methyl mercury: All distillable CH3Hg forms and species found in the filtrate of an aqueous solution that has been filtered through a 0.45 micron filter.

3.3 Methyl mercury: All acid-distillable Hg, which, upon reaction with NaBEt4 yields methylethyl mercury. This includes, but is not limited to, CH3Hg+, strongly organo-complexed CH3Hg compounds, adsorbed particulate CH3Hg, and CH3Hg bound in microorganisms. In freshly collected samples, dimethyl mercury ((CH3) Hg) will not be recovered as CH3Hg, but in samples which have been acidified for several days, most (CH3)2Hg has broken down to CH3Hg. In this method, CH3Hg and total recoverable CH3Hg are synonymous.

3.4 Definitions of other terms used in this method are given in the glossary at the end of the method.

4.0 Contamination and Interferences

4.1 Preventing ambient water samples from becoming contaminated during the sampling and analysis process constitutes one of the greatest difficulties encountered in trace metals determinations. Over the last two decades, marine chemists have come to recognize that much of the historical data on the concentrations of dissolved trace metals in seawater are erroneously high because the concentrations reflect contamination from sampling and analysis rather than ambient levels. Therefore, it is imperative that extreme care be taken to avoid contamination when collecting and analyzing ambient water samples for trace metals.

4.2 Samples may become contaminated by numerous routes. Potential sources of trace metal contamination during sampling include: metallic or metal-containing labware (e.g., talc gloves that contain high levels of zinc), containers, sampling equipment, reagents, and reagent water; improperly cleaned and stored equipment, labware, and reagents; and atmospheric inputs such as dirt and dust. Even human contact can be a source of trace metal contamination. For example, it has been demonstrated that dental work (e.g., mercury amalgam fillings) in the mouths of laboratory personnel can contaminate samples that are directly exposed to exhalation (Reference 5).

4.3 Contamination control

4.3.1 Philosophy—The philosophy behind contamination control is to ensure that any object or substance that contacts the sample is metal free and free from any material that may contain Hg or CH3Hg.

4.3.1.1 The integrity of the results produced cannot be compromised by
contamination of samples. This method and the Sampling Method give requirements and suggestions for control of sample contamination.

4.3.1.2 Substances in a sample cannot be allowed to contaminate the laboratory work area or instrumentation used for trace metals measurements. This method gives requirements and suggestions for protecting the laboratory.

4.3.1.3 Although contamination control is essential, personnel health and safety remain the highest priority. The Sampling Method and Section 5 of this method give requirements and suggestions for personnel safety.

4.3.2 Avoid contamination—The best way to control contamination is to completely avoid exposure of the sample to contamination in the first place. Avoiding exposure means performing operations in an area known to be free from contamination. Two of the most important factors in avoiding/reducing sample contamination are (1) an awareness of potential sources of contamination and (2) strict attention to the work being done. Therefore, it is imperative that the procedures described in this method be carried out by well-trained, experienced personnel.

4.3.3 Use a clean environment—The ideal environment for processing samples is a class 100 clean room. If a clean room is not available, all sample preparation should be performed in a class 100 clean bench or a nonmetal glove box fed by mercury-free and particle-free air or nitrogen. Digestions should be performed in a nonmetal fume hood situated, ideally in the clean room.

4.3.4 Minimize exposure—The Apparatus that will contact samples, blanks, or standard solutions should be opened or exposed only in a clean room, clean bench, or glove box so that exposure to an uncontrolled atmosphere is minimized. When not in use, the Apparatus should be covered with clean plastic wrap, stored in the clean bench or in a plastic box or glove box, or bagged in clean zip-type bags. Minimizing the time between cleaning and use will also minimize contamination.

4.3.5 Clean work surfaces—Before a given batch of samples is processed, all work surfaces in the hood, clean bench, or glove box in which the samples will be processed should be cleaned by wiping with a lint-free cloth or wipe soaked with reagent water.

4.3.6 Wear gloves—Sampling personnel must wear clean, non talc gloves during all operations involving handling of the Apparatus, samples, and blanks. Only clean gloves may touch the Apparatus. If another object or substance is touched, the glove(s) must be changed before again handling the Apparatus. If it is even suspected that gloves have become contaminated, work must be halted, the contaminated gloves removed, and a new pair of clean gloves put on. Wearing multiple layers of clean gloves will allow the old pair to be quickly stripped with minimal disruption to the work activity.

4.3.7 Use metal-free Apparatus—All Apparatus used for determination of CH3Hg at ambient water quality criteria levels must be nonmetallic and free of material that may contain metals.

4.3.7.1 Construction materials—Only fluoropolymer or borosilicate glass
containers should be used for samples that will be analyzed for Hg because Hg vapors can diffuse in or out of other materials, resulting in results that are biased low or high. All materials, regardless of
construction, that will directly or indirectly contact the sample must be cleaned using the procedures in this method and must be known to be clean and mercury free before proceeding.

4.3.7.2 Serialization—It is recommended that serial numbers be indelibly marked or etched on each piece of Apparatus so that contamination can be traced, and logbooks should be maintained to track the sample from the container through the labware to introduction into the instrument. It may be useful to dedicate separate sets of labware to different sample types; e.g., receiving waters vs. effluents. However, the Apparatus used for processing blanks and standards must be mixed with the Apparatus used to process samples so that contamination of all labware can be detected.

4.3.7.3 The laboratory or cleaning facility is responsible for cleaning the Apparatus used by the sampling team. If there are any indications that the Apparatus is not clean when received by the sampling team (e.g., ripped storage bags), an assessment of the likelihood of contamination must be
made. Sampling must not proceed if it is possible that the Apparatus is contaminated. If the Apparatus is contaminated, it must be returned to the
laboratory or cleaning facility for proper cleaning before any sampling activity resumes.

4.3.8 Avoid sources of contamination—Avoid contamination by being aware of potential sources and routes of contamination.

4.3.8.1 Contamination by carryover—Contamination may occur when a sample containing a low concentration of CH3Hg is processed immediately after a sample containing a relatively high concentration. When an unusually
concentrated sample is encountered, a ethylation blank should be analyzed
immediately following the sample to check for carryover. Samples known or suspected to contain the lowest concentration of CH3Hg should be analyzed first followed by samples containing higher levels.

4.3.8.2 Contamination by samples—Significant laboratory or instrument contamination may result when untreated effluents, in-process waters,
landfill leachates, and other samples containing high concentrations of Hg
or CH3Hg are processed and analyzed. This method is not intended for application to these samples, and samples containing high concentrations of trace metals should not be permitted into the clean room and laboratory dedicated for processing trace metals samples.

4.3.8.3 Contamination by indirect contact—Apparatus that may not directly come in contact with the samples may still be a source of contamination. For
example, clean tubing placed in a dirty plastic bag may pick up contamination from the bag and subsequently transfer the contamination to the sample. Therefore, it is imperative that every piece of the Apparatus that is directly or indirectly used in the collection, processing, and analysis of samples be thoroughly cleaned (see Section 6.1.2).

4.3.8.4 Contamination by airborne particulate matter—Airborne particles are less obvious substances capable of contaminating samples. Samples may be contaminated by airborne dust, dirt, particles, or vapors from unfiltered air supplies; nearby corroded or rusted pipes, wires, or other fixtures; or metal-containing paint. Whenever possible, sample processing and analysis should occur as far as possible from sources of airborne contamination.

4.4 Interferences

4.4.1 When the method is properly applied, no significant interferences have been observed in the analysis of ambient waters.

4.4.2 Distillation of CH3Hg from solution requires a carefully controlled level of HCl in solution. Distillation will not be quantitative if too little HCl is added, but too much HCl results in co-distillation of HCl fumes, which interfere with the ethylation procedure. Therefore fresh water samples must be preserved only with between 0.3% and 0.5% (v/v) 11.6 M HCl, and salt water samples with between 0.1% and 0.2% (v/v) 9 M H2SO4.

4.4.3 Samples preserved with nitric acid (HNO3) cannot be analyzed for CH3Hg as the analyte is partially decomposed in the distillation step by this reagent.

4.4.4 The fluorescent intensity is strongly dependent upon the presence of molecular species in the carrier gas that can cause "quenching" of the excited atoms. The Carbotrap® trap eliminates quenching due to trace gases, but it still remains the analyst's responsibility to ensure high purity inert carrier gas and a leak-free analytical train. In some rare cases (such as oil polluted water) low molecular weight organic compounds may purge with the methylethyl mercury and collect on the Carbotrap® trap, subsequently resulting in signal quenching during elution. Such cases are best treated by sample dilution prior to distillation.

4.4.5 Recent investigations have shown that a positive artifact is possible with the distillation procedure in cases where high inorganic Hg concentrations are present (Reference 7). In natural waters, approximately 0.01 to 0.05% of the ambient inorganic Hg in solution may be methylated by ambient organic matter during the distillation step. In most waters, where the percent CH3Hg is 1-30% of the total, this effect is trivial. However, the analyst should be aware that in inorganic Hg contaminated waters, the fraction CH3Hg can be < 1% of the total, and so flagging of the data (as representing a maximum estimate of CH3Hg concentration) may be warranted. In samples with high levels of divalent mercury (Hg(II)), solvent extraction may be preferable to distillation (Reference 7).

5.0 Safety

5.1 The toxicity or carcinogenicity of each chemical used in this method has not been precisely determined; however, each compound should be treated as a potential health hazard. Exposure to these compounds should be reduced to the lowest possible level. It is suggested that the laboratory perform personal hygiene monitoring of each analyst using this method and that the results of this monitoring be made available to the analyst.

5.1.1 Chronic Hg exposure may cause kidney damage, muscle tremors, spasms, personality changes, depression, irritability, and nervousness. Organo-mercurials may cause permanent brain damage. Because of the toxicological and physical properties of CH3Hg, pure standards should be handled only by highly trained personnel thoroughly familiar with handling and cautionary procedures and the associated risks.

5.1.2 It is recommended that the laboratory purchase a dilute standard solution of CH3Hg for this method. If primary solutions are prepared, they shall be prepared in a hood, and a NIOSH/MESA-approved toxic gas respirator shall be worn when high concentrations are handled.

5.2 This method does not address all safety issues associated with its use. The laboratory is responsible for maintaining a current awareness file of OSHA regulations for the safe handling of the chemicals specified in this method. A reference file of material safety data sheets (MSDSs) should also be made available to all personnel involved in these analyses. Additional information on laboratory safety can be found in References 7-10. The references and bibliography at the end of Reference 10 are particularly comprehensive in dealing with the general subject of laboratory safety.

5.3 Samples suspected of containing high concentrations of CH3Hg are handled using essentially the same techniques employed in handling radioactive or infectious materials. Well-ventilated, controlled access laboratories are required. Assistance in evaluating the health hazards of particular laboratory conditions may be obtained from certain consulting laboratories and from State Departments of Health or Labor, many of which have an industrial health service. Each laboratory must develop a strict safety program for handling CH3Hg.

5.3.1 Facility—When samples known or suspected to contain high concentrations of CH Hg are handled, all operations (including removal of samples from sample containers, weighing, transferring, and mixing) should be performed in a glove box demonstrated to be leakproof or in a fume hood demonstrated to have adequate airflow. Gross losses to the laboratory ventilation system must not be allowed. Handling of the dilute solutions normally used in analytical and animal work presents no inhalation hazard except in an accident.

5.3.2 Protective equipment—Disposable plastic gloves, apron or lab coat, safety glasses or mask, and a glove box or fume hood adequate for radioactive work should be used. During analytical operations that may give rise to aerosols or dusts, personnel should wear respirators equipped with activated carbon filters.

5.3.3 Training—Workers must be trained in the proper method of removing contaminated gloves and clothing without contacting the exterior surfaces.

5.3.4 Personal hygiene—Hands and forearms should be washed thoroughly after each manipulation and before breaks (coffee, lunch, and shift).

5.3.5 Confinement—Isolated work areas posted with signs, segregated glassware and tools, and plastic absorbent paper on bench tops will aid in confining contamination.

5.3.6 Effluent vapors—The effluent from the CVAFS should pass through either a column of activated charcoal or a trap containing gold or sulfur to amalgamate or react with Hg vapors.

5.3.7 Waste handling—Good technique includes minimizing contaminated waste. Plastic bag liners should be used in waste cans. Janitors and other personnel must be trained in the safe handling of waste.

5.3.8 Decontamination

5.3.8.1 Decontamination of personnel—Use any mild soap with plenty of scrubbing action.

5.3.8.2 Glassware, tools, and surfaces—Activated carbon powder will adsorb CH3Hg, eliminating the possible volatilization of CH3Hg. Satisfactory cleaning may be accomplished by dusting a surface lightly with activated carbon powder, then washing with any detergent and water.

5.3.9 Laundry—Clothing known to be contaminated should be collected in plastic bags. Persons who convey the bags and launder the clothing should be advised of the hazard and trained in proper handling. If the launderer knows of the potential problem, the clothing may be put into a washer without contact. The washer should be run through a cycle before being used again for other clothing.

5.3.10 Wipe tests—A useful method of determining cleanliness of work surfaces and tools is to wipe the surface with a piece of filter paper. Extraction and analysis by this method can achieve a limit of detection of less than 1 ng per wipe. Less than 0.1 µg per wipe indicates acceptable cleanliness; anything higher warrants further cleaning. More than 10 µg on a wipe constitutes an acute hazard, requires prompt cleaning before further use of the equipment or work space, and indicates that unacceptable work practices have been employed.

6.0 Equipment and Supplies

NOTE: The mention of trade names or commercial products in this method is for illustrative purposes only and does not constitute endorsement or recommendation for use by the Environmental Protection Agency. Equivalent performance may be achievable using apparatus, materials, or cleaning procedures other than those suggested here. The laboratory is responsible for demonstrating equivalent performance.

6.1 Sampling equipment

6.1.1 Sample collection bottles-fluoropolymer or borosilicate glass, 125- to 1000-mL, with fluoropolymer or fluoropolymer-lined cap.

6.1.2 Cleaning

6.1.2.1 New bottles are cleaned by heating to 65-75°C in 4 N HCl for at least 48 h. The bottles are cooled, rinsed three times with reagent water, and filled with reagent water containing 1% HCl. These bottles are capped and placed in a clean oven at 60-70°C overnight. After cooling, they are rinsed three more times with reagent water, filled with reagent water containing 0.4% (v/v) HCl, capped, and placed in a mercury-free class 100 clean bench until the outside of the bottle is dry. The caps are then tightened with a wrench and the bottles are double-bagged in new polyethylene zip-type bags. The capped bottles are stored in wooden or plastic boxes until use.

6.1.2.2 To avoid long-term accumulation of Hg or CH3Hg on the bottle walls due to trace organic coatings, used bottles are filled with reagent water containing 0.02 N BrCl solution and allowed to stand over night. The BrCl is neutralized with the addition of 0.2 mL of 20% NH2OH solution. The bottles are then cleaned exactly as in Section 6.1.2.1, except that they soak only 6-12 h in hot 4 N HCl.

6.1.2.3 Bottle blanks should be analyzed as described in Section 9.4.4.1 to verify the effectiveness of the cleaning procedures.

6.1.3 Filtration Apparatus

6.1.3.1 Filter—0.45-µm, 15-mm diameter capsule filter (Gelman Supor 12175, or equivalent)

6.1.3.2 Peristaltic pump—115-V a.c., 12-V d.c., internal battery, variable-speed, single-head (Cole-Parmer, portable, "Masterflex L/S," Catalog No. H- 07570-10 drive with Quick Load pump head, Catalog No. H-07021-24, or equivalent).

6.1.3.3 Tubing—styrene/ethylene/butylene/silicone (SEBS) resin for use with peristaltic pump, approx 3/8-in i.d. by approximately 3 ft (Cole-Parmer size 18, Catalog No. G-06464-18, or approximately 1/4-in i.d., Cole- Parmer size 17, Catalog No. G-06464-17, or equivalent). Tubing is cleaned by soaking in 5-10% HCl solution for 8-24 h. It is rinsed with reagent water on a clean bench in a clean room and dried on the clean bench by purging with metal-free air or nitrogen. After drying, the tubing is double-bagged in clear polyethylene bags, serialized with a unique number, and stored until use.

6.2 Equipment for bottle and glassware cleaning

6.2.1 Vat, 100-200 L, high-density polyethylene (HDPE), half filled with 4 N HCl in reagent water.

6.2.2 Panel immersion heater, 500-W, all-fluoropolymer coated, 120 vac (Cole-Parmer H- 03053-04, or equivalent)

NOTE: Read instructions carefully!! The heater will maintain steady state, without temperature feedback control, of 60-75°C in a vat of the size described. However, the equilibrium temperature will be higher (up to boiling) in a smaller vat. Also, the heater plate MUST be maintained in a vertical position, completely submerged and away from the vat walls to avoid melting the vat or burning out!

6.2.3 Laboratory sink in class 100 clean area, with high-flow reagent water (Section 7.1) for rinsing.

6.2.4 Clean bench, class 100, for drying rinsed bottles.

6.2.5 Oven, stainless steel, in class 100 clean area, capable of maintaining ± 5°C in the 60-70°C temperature range.

6.3 Cold vapor atomic fluorescence spectrometer (CVAFS): The CVAFS system used may either be purchased from a supplier, or built in the laboratory from commercially available components.

6.3.1 Commercially available: Tekran Model 2357 CVAFS, Brooks-Rand Model III CVAFS, or equivalent

6.3.2 Custom-built CVAFS (Reference 11). Figure 1 shows the schematic diagram. The system consists of the following:

6.3.2.1 Low-pressure 4-W mercury vapor lamp

6.3.2.2 Far UV quartz flow-through fluorescence cell—12 mm x 12 mm x 45
mm, with a 10-mm path length (NSG Cells, or equivalent).

6.3.2.3 UV-visible photomultiplier (PMT)—sensitive to < 230 nm. This PMT is
isolated from outside light with a 253.7-nm interference filter (Oriel Corp., or equivalent).

6.3.2.4 Photometer and PMT power supply (Oriel Corp., or equivalent), to convert PMT output (nanoamp) to millivolts

6.3.2.5 Black anodized aluminum optical block—holds fluorescence cell, PMT,
and light source at perpendicular angles, and provides collimation of incident and fluorescent beams (Frontier Geosciences Inc., or equivalent).

6.3.2.6 Flowmeter, with needle valve capable of keeping the carrier gas at a
reproducible flow rate of 30 mL/min

6.3.2.7 Ultra high-purity argon (grade 5.0)

6.4 Equipment for CH3Hg purging system—Figure 2a shows the schematic diagram for the purging system. The system consists of the following:

6.4.1 Flow meter/needle valve—capable of controlling and measuring gas flow rate to the purge vessel at 350 (± 50) mL/min.

6.4.2 Fluoropolymer fittings—connections between components and columns are made using 6.4-mm o.d. fluoropolymer tubing and fluoropolymer friction-fit or threaded tubing connectors. Connections between components requiring mobility are made with 3.2-mm o.d. fluoropolymer tubing because of its greater flexibility.

6.4.3 Cold vapor generator (bubbler)—200-mL borosilicate glass (15 cm high x 5.0 cm diameter) with standard taper 24/40 neck, fitted with a sparging stopper having a coarse glass frit that extends to within 0.2 cm of the bubbler bottom (Frontier Geosciences, Inc., or equivalent).

6.5 Equipment for the isothermal gas chromatography (GC) system.

6.5.1 Figure 1 shows the schematic for the interface of the GC with the CVAFS detector (Reference 6).

6.5.2 Figure 2b shows the orientation consideration for purging and desorbing CH3Hg from the Carbotrap® traps.

6.5.3 Carbotrap® traps—10-cm x 6.5-mm o.d. x 4-mm i.d. quartz tubing. The tube is filled with 3.4 cm of 30/45 mesh Carbotrap® graphitic carbon adsorbant (Supelco, Inc., or equivalent). The ends are plugged with silanized glass wool.

6.5.3.1 Traps are fitted with 6.5-mm i.d. fluoropolymer friction-fit sleeves for
making connection to the system. When traps are not in use, fluoropolymer end plugs are inserted in trap ends to eliminate contamination.

6.5.3.2 At least six traps are needed for efficient operation.

6.5.3.3 Because the direction of flow is important in this analysis, the crimped end of the Carbotrap® trap will be referred to as “side A,” while the uncrimped end will be referred to as “side B.”

6.5.4 Heating of Carbotrap® traps—To desorb CH3Hg collected on a trap, heat for 45 sec to 450-500°C (a barely visible red glow when the room is darkened) with a coil consisting of 75 cm of 24-gauge Nichrome wire at a potential of 16-20 vac. Potential is applied and finely adjusted with an autotransformer.

6.5.5 Timer—The heating interval is controlled by a timer-activated 120-V outlet, into which the heating coil autotransformer is plugged.

6.5.6 Isothermal GC—Consists of two parts, a custom fabricated packed GC column, and a custom fabricated constant temperature oven.

6.5.6.1 The column is 1 m long, made from 0.25 inch OD by 4 mm ID borosilicate glass GC column tubing. The column is formed into an 8 cm diameter coil, with 15 cm straight extensions from each end. The column is silanized, packed in the coiled portion with 60/80 mesh 15% OV-3 on acid-washed Chromasorb W, and then conditioned under inert gas flow at 200°C. A column meeting these specifications may be custom fabricated (Supelco Inc., or equivalent).

6.5.6.2 The GC oven consists of a 500-watt aluminum jacketed heating mantle, fitted with a custom machined fluoropolymer lid (14 cm OD by 1 cm thick). The lid is attached with stainless steel screws and contains three threaded holes (0.25 inch female NPT) in a triangular pattern in the top. The spacing of the holes conforms exactly to the spacing between the two 15 cm glass extensions of the GC column.

6.5.6.3 Fluoropolymer fittings, with 0.25-inch male NPT threads on the bottom and 0.25-inch compression fittings on top, are placed into the threaded holes. The GC column is secured into the oven by passing the glass extensions through two of the fluoropolymer fittings, so that 3 cm of the glass extensions protrude from the top, and tightening the compression fittings. The fluoropolymer lid holding the GC column is then screwed to the top of the oven.

6.5.6.4 Temperature feedback control (110 ± 2°C) is achieved through a
thermocouple temperature controller. The oven is plugged into the controller and the thermocouple probe is inserted through the third fluoropolymer fitting in the lid, such that the sensor is located near the center of the GC coil.

6.5.6.5 Several research groups have successfully interfaced the Carbotrap®/CVAFS system directly to a commercial gas chromatograph. The use of capillary column GC will result in better peak separation, although at higher cost.

6.5.7 Pyrolytic column—The output from the GC oven is connected directly to a high temperature column to decompose eluted organo-mercurial compounds to Hg0. The output of the pyrolytic column is connected to the inlet of the CVAFS system.

6.5.7.1 The column consists of a 20-cm length of quartz tubing, packed over the central 10 cm with quartz wool.

6.5.7.2 The column is heated to orange heat (~ 700°C) by a 1 m length of 22 gauge Nichrome wire, tightly wrapped around the quartz wool packed portion of the tube. The temperature of the coil is adjusted by visual inspection of the color, using a 0-120 volt autotransformer.

6.6 Recorder—Any multi-range millivolt chart recorder or integrator with a range compatible with the CVAFS is acceptable. By using a two pen recorder with pen sensitivity offset by a factor of 10, the dynamic range of the system is extended to 103.

6.7 Distillation unit—The distillation unit is a custom made temperature controlled aluminum block heater, as shown schematically in Figure 3 (Frontier Geosciences Inc., or equivalent).

6.7.1 Heating block insulation—Each heating block is encased first in refractory spun rock insulation (1 inch thickness) and then an exterior wood shell for rigidity.

6.7.2 Each heating block (10 cm wide x 20 cm long x 15 cm high) is bored with five 31 mm diameter holes (evenly spaced), 120 mm deep. A 3/8 inch diameter hole is bored to 90% of the block length, perpendicular to and behind the distillation tube holes, to accommodate a cylindrical heating element. A 2 mm diameter hole is bored parallel to the heating element hole, and 2 cm above it, to accommodate the temperature sensor.

6.7.3 Heating element—Each heating block is equipped with a 750 watt cylindrical heating element, 6 inches long by 3/8 inch diameter (Omega Inc.), immobilized in its respective hole by a dab of silicone glue.

6.7.4 Type J thermocouple probe—Each heating block is equipped with a type J thermocouple probe immobilized in its respective hole by a dab of silicone glue.

6.7.5 Digital temperature controller—The heating element and thermocouple are connected to a digital temperature controller.

6.7.6 Fluoropolymer vials with caps—The distillation unit is designed to accommodate 60-mL fluoropolymer vials (part number 0202, Savillex, or equivalent). The original caps are used to close the vials when distillate is to be stored until analysis.

6.7.6.1 For each distillation, two identical vials are needed: a distillation vessel and a receiving vessel. For convenience, each vial should be engraved with a line at 40.0 mL (obtained by weighing 40 g of water in the vial), and a unique identification number, both on the vial and the cap.

6.7.6.2 Fluoropolymer vials are acid cleaned initially as described for other
fluoropolymer ware and stored filled with 0.5% HCl. After use, receiving vials are rinsed with reagent water and filled with 0.5% HCL. The tubing is looped around the cap as described in Section 6.7.7.1, and the vials are placed in a 70°C (± 5°C) oven overnight. Cleaning is the same for the distillation vials, with the exception that first the vials, caps, and tubing are thoroughly scrubbed with an alkaline detergent and test tube brush to remove any residues from the samples.

6.7.7 Purge caps—The standard caps on the fluoropolymer vials are replaced with purge caps (part number 33-2-2, Savillex, or equivalent) for distillation purposes.

6.7.7.1 Fluoropolymer tubing—each purge cap is threaded with a piece of 1/8 inch fluoropolymer tubing, approximately 30-40 cm long. One end is pulled through one of the holes in the cap, down to a length that will allow it to reach the bottom of the distillation vial when the vial is screwed onto the cap. The bottom end of this tubing is cut at a 45° angle. The outside end of the tubing is cut perpendicularly and is looped around and inserted into the second cap hole when not in use (to keep the system closed and clean).

6.7.8 Aluminum distillation cover—The cover for the heating block consists of a 5 cm high