Standard Operating Procedure for
Analysis of Sediment for Total Mercury Using
the Cold Vapor Technique with the Leeman
Labs, Inc. Automated Mercury System
Theresa Uscinowicz, A & O Chemical Company1
and
Ronald Rossmann, USEPA
Large Lakes Research Station
9311 Groh Road
Grosselle, MI48138
Mid-Continent Ecology Division - Duluth
National Health and Environmental Effects Research Laboratory
Octobers, 1996
Revision 1
1 Current affiliation is SoBran, Inc./Pathology Associates International.
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Standard Operating Procedure for Analysis of Sediment for Total
Mercury Using the Cold Vapor Technique with the Leeman Labs, Inc.
Automated Mercury System
1.0 Introduction
Elemental concentrations of mercury in sediment and water are determined by the PS200 system,
and its operation is based upon cold vapor AAAS. The prepared sample enters the system in the
divalent phase, and is mixed with stannous chloride to form elemental mercury vapor. This
mixture moves to the liquid gas separator, and argon carries the mercury vapor through a drying
tube for vapor removal. The vapor enters one path of the cell optimized. The mercury lamp emits
light at 254 nm, and absorbance is measured by the detector.
2.0 Materials Required
2.1 Chemicals
Reagents needed include the following ultra pure grade chemicals:
2.1.1 Leeman Labs 1 OOppm Mercury Standard
2.1.2 Leeman Labs Hydrochloric, Nitric Acids
2.1.3 Leeman Labs Ultra Pure Water
2.1.4 Liquinox
2.1.5 J.T. Baker Brand Hydrochloric, Nitric Acids
2.1.6 J.T. Baker Brand Stannous Chloride or Leeman Labs Stannous Chloride
2.1.7 Hydroxylamine Hydrochloride
2.1.8 Magnesium Perchlorate 10-20 size mesh
2.1.9 Potassium Permanganate
2.2 Equipment and Supplies
2.2.1 Supplies Needed for analyses include:
2.2.1.1 PS200 Automated Mercury Analyzer with autosampler,
with
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Using the Cold Vapor Technique with the
Leeman Labs, Inc. Automated Mercury System Volume 2, Chapter 2
associated data acquisition system
2.2.1.2 AP200 Automated Preparation System with associated data
acquisition system
2.2.1.3 Analytical Balance in Biology Lab, Mettler 2100T
2.2.1.4 EDP pipettors and associated disposable tips
2.2.1.5 Associated pump tubing for sample drainage, tin chloride
2.2.1.6 Polyethylene tubes 12 mL capacity, and caps
2.2.1.7 Polyethylene tubes 45mL capacity
2.2.1.8 Teflon or polyethylene beakers
2.2.1.9 Teflon wash bottles
2.2.1.10 Teflon bottles (60 mL)
2.2.1.11 Electronic balance
2.2.1.12 PVC gloves
2.2.1.13 Paper towels, clean wipes
2.2.1.14 Lubricating oil for autosampler
2.2.1.15 Quartz wool and quartz glass drying tubes
2.2.1.16 Teflon spatula
2.2.2 Supplies needed before sample analyses if not using automated preparation system:
2.2.2.1 CEM microwave digester with associated Teflon PFA vessels
2.2.2.2 Low density 30-mL polyethylene bottles for sample storage
2.3 Reference Documents
The user of this method must be familiar with the following established standard
operation procedures:
LLRS-MET-SOP-001 Standard Operating Procedures for the Preparation of Materials
used for Ultra-low Trace Element Analyses
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Using the Cold Vapor Technique with the
Volume 2, Chapter 2 Leeman Labs, Inc. Automated Mercury
System
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SOP for Analysis of Sediment for Total Mercury
Using the Cold Vapor Technique with the
Leeman Labs, Inc. Automated Mercury System Volume 2, Chapter 2
LLRS-MET-SOP-003 Standard Operating Procedures for the Maintenance of the LLRS
Trace Metal Laboratories
LLRS-MET-SOP-010 Standard Operating Procedures for Analysis of Total Mercury in
Tissue and Sediment using the Cold Vapor Technique with the
Perkin-Elmer Model MHS-20 Gold Amalgam System
LLRS-QA-001 Minimum Analytical Quality Assurance Objectives for U. S. EPA
Large Lakes Research Station
LLRS-QA-SOP-001 Standard Operating Procedures for the Release of Data
LLRS-QA-SOP-002 Standard Operating Procedures for the Routine Review of Data
Quality and Quantity
3.0 Reagent Preparation
3.1 5% Nitric Acid Solution
This acid solution is used for rinsing materials used for cleaning materials used in the
analysis of samples.
3.4.1 Rinse a pre-cleaned graduated cylinder (100 mL) with three rinses of MSQ.
3.4.2 To the graduated cylinder, add 95 mL of MSQ.
3.4.3 Carefully add 5 mL of concentrated reagent grade nitric acid.
3.4.4 Transfer the solution to a squirt bottle.
3.4.5 Repeat steps 3.4.1 through 3.4.3 but add five milliliters of reagent grade acid. (For
preparation of 5% nitric acid rinses).
3.4.6 Repeat step 3.4.4 but add to separate precleaned teflon bottle.
3.2 10% Hydrochloric Acid Rinse Solution
Use reagent grade J.T. Baker or Leeman Labs hydrochloric acid. The amount needed for
a one day run is 300 mL. Approximately 2 L will be needed to be prepared weekly.
WARNING
Hydrochloric acid is highly corrosive and incompatible with metals, hydroxides,
amines, and alkalis. Handle it while wearing personal protection gear. A full face
shield is recommended. Always use the concentrated acid under a fume hood. Store it
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Using the Cold Vapor Technique with the
Volume 2, Chapter 2 Leeman Labs, Inc. Automated Mercury
System
in an appropriate place. Store concentrated acid in a corrosives cabinet.
3.2.1 Use a precleaned 1000 mL polyethylene graduated cylinder. Carefully rinse the
inside of the cylinder with 5% reagent grade nitric acid using a Teflon (precleaned)
squirt bottle. Follow this rinse with three rinses of Millipore Super-Q water
(MSQ).
3.2.2 Add 900 mL of MSQ to the graduated cylinder.
3.2.3 Carefully add 100 mL of reagent grade hydrochloric acid to the cylinder.
3.2.4 Transfer the mixture to a high density polyethylene bottle from Leeman
Labs.
3.3 1:1 Nitric Acid Solution
As recommended by the manufacturer, polyethylene autosampler cups must soak in 1:1
nitric acid before running to ensure acceptable results. Soak the cups for at least two
hours. Use J.T. Baker trace metal grade nitric acid for preparation. This solution can be
recycled, provided the autosampler cups are adequately rinsed with MSQ a minimum of
ten times before and after use. Dispose of 1:1 nitric after one month to eliminate the
possibility of any residual contamination.
WARNING
Nitric acid is corrosive and incompatible with combustible materials, metallic powders,
hydrogen sulftde, carbides, and alcohols. Handle it with personal protection. A full
face shield is recommended for the concentrated acid. Always use the acid under a
fume hood. Store the concentrated acid in a corrosives cabinet.
3.3.1 Use an appropriate pre-cleaned container. An empty Suprapure hydrochloric acid
bottle has been used. Rinse the container at least three times with MSQ before
addition of nitric acid.
3.3.2 Using the graduations on the glass bottle, add 400 mL MSQ.
3.3.3 Carefully add 400mL of J.T. Baker nitric acid to the glass bottle.
3.3.4 Swirl the contents of the bottle and , if necessary, label.
3.4 10%(w/v) Stannous Chloride Solution
The volume of tin chloride solution required to run is dependent upon the daily run time.
To run for six hours approximately 250 mL are required.
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Using the Cold Vapor Technique with the
Leeman Labs, Inc. Automated Mercury System Volume 2, Chapter 2
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Using the Cold Vapor Technique with the
Volume 2, Chapter 2 Leeman Labs, Inc. Automated Mercury
System
WARNING
Stannous chloride should be handled with care. Avoid contact with eyes, skin, and
clothing. Avoid breathing its dust. Handle solid chemical under a fume hood.
Handle while wearing personal protection gear for eyes and skin.
3.4.1 Rinse a precleaned wide mouth teflon bottle three times with quartz distilled water.
3.4.2 Tare the bottle and add 25 g of tin chloride to the bottle.
3.4.3 Rinse a precleaned graduated cylinder with 5% reagent grade nitric acid solution
followed by three rinses of MSQ.
3.4.4 Carefully add 25 mL of reagent grade hydrochloric acid to the graduated cylinder.
Carefully pour the cylinder's contents into the bottle containing the stannous
chloride.
3.4.5 Swirl the contents of the bottle vigorously to dissolve the stannous
chloride.
3.4.6 After the stannous chloride has been dissolved, add 200 mL of MSQ using the
same graduated cylinder.
3.4.7 Replace the cover and vigorously shake the bottle to ensure all of the tin chloride
is dissolved.
3.5 10% Nitric Acid Solution
This acid solution is used for preparation of standards and for dilution of samples.
Historically, this solution is used for samples that have undergone microwave digestion.
3.5.1. Rinse a pre-cleaned graduated cylinder (100 mL) with 5% reagent grade nitric
acid solution followed by three rinses of MSQ.
3.5.2 To the graduated cylinder, add 90 mL of MSQ.
3.5.3 Carefully add 10 mL of concentrated Seastar or reagent grade nitric acid
(whichever matches the matrix of samples).
3.5.4 Transfer the solution to a teflon bottle.
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Using the Cold Vapor Technique with the
Leeman Labs, Inc. Automated Mercury System Volume 2, Chapter 2
3.6 Working Mercury Standard Solution
Commercial 100 ppm Leeman Labs mercury standard is used to prepare the working
standard. The working standard is prepared at a concentration of 100 ppb or 0.1 jig
Hg/mL and is made fresh weekly.
WARNING
Mercury is a poison. Handle it, its compounds, and its solutions with personal
protection. Mercury can form a vapor and be inhaled. It also is absorbed through the
skin. Use this material in a fume hood. Always wear personal protection gear.
3.6.1 Rinse a pre-cleaned teflon bottle (LLRS-MET-SOP-001) three times with MSQ
and air dry.
3.6.2 Using a precleaned graduated cylinder, rinse three times with 5% nitric
and three times with MSQ.
3.6.3 Using two EDP separate automatic pipettors, rinse 2-1000 mL tips three times
with a 5% reagent grade nitric solution followed by three rinses with MSQ.
3.6.4 Tare a dry bottle on the electronic scale.
3.6.5 Using an additional EDP pipettor, rinse a 100 jiL tip in the same manner as 3.6.2
3.6.6 Using the cleaned 100 jiL tip, carefully add 50 jiL of the commercial 100 ppm
Leeman Labs mercury standard solution to the bottle.
3.6.7 Transfer 45mLs of the 10% nitric acid solution using the cleaned graduated
cylinder.
3.6.8 Using an EDP pipettor, with a 1000 jiL precleaned tip, add 10% nitric acid
solution to the bottle until the weight is 50 g.
3.7 Preparation of Recommended Range of Calibration Standards
The calibration standards are prepared fresh semi weekly.
3.7.1 Microwave Digestion Standard Range
The following range of standards has been used for analyses of Green Bay
Sediment samples and bracket the samples well. 0.250 ppb, 0.500 ppb, 1.00 ppb
2.00 ppb, 5.00 ppb are used (0.00025 |ig/mL, 0.00050 |ig/mL, 0.001 |ig/mL,
0.002 |ig/mL, 0.005 jig/mL). It may be possible to go below 0.250 ppb depending
upon instrument performance. The lowest concentration above background noise
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SOP for Analysis of Sediment for Total Mercury
Using the Cold Vapor Technique with the
Volume 2, Chapter 2 Leeman Labs, Inc. Automated Mercury
System
is approximately 0.087 ppb (0.000087 jig/ mL). 60 mL of each calibration
standard will last for two daily runs. Prepare double of the desired standard that
will be run as a check standard. The autosampler cups must be filled to at least 40
milliliters, (60 mL total capacity).
3.7.1.1 Rinse six pre-cleaned teflon bottles three times with MSQ and
allow to air dry.
3.7.1.2 Prepare two separate EDP pipettors each with a precleaned 1000
|iL tip. (Rinse each pipette tip three times with 5% nitric acid
followed by three rinses with MSQ) .
3.7.1.3 Rinse a precleaned graduated cylinder with 5% nitric acid
followed by three rinses of MSQ.
3.7.1.4 For each standard, tare each bottle individually.
3.7.1.5 For the 0.250 ppb standard, pipette 150 pL of the 100 ppb
working standard to the bottle.
3.7.1.6 Using the graduated cylinder, bring the total volume up to 55 mL
by the careful addition of prepared Seastar 10% nitric acid. The
weight of the bottles contents should now be roughly 55 g.
3.7.1.7 Add prepared Seastar 10% nitric acid with the other EDP
pipettor and precleaned tip until the total weight is 60 g.
3.7.1.8 Repeat steps 3.7.1.6 - 3.7.1.9 for the remaining standards by the
addition of 300 \iL of the 100 ppb standard for a 0.500 ppb
calibration standard, 600 jiL of the 100 ppb standard for the 1.00
ppb calibration standard, 1200 jiL of the 100 ppb standard for the
2.00 ppb calibration standard, and 3000 jiL of the 100 ppb
standard for the 5.00 ppb calibration standard.
3.7.1.9 Do not recycle the standards remaining in the autosampler cups at
the end of the day. Properly dispose of these daily. Attempts to
recycle the standards from autosampler cups have given
diminished intensities.
3.7.2 Automated Digester Standard Range
Use the following range of standards: 0.000 ppb, 0.125 ppb, 0.250 ppb, 0.500 ppb,
1.00 ppb, 2.00 ppb. Prepare in a 2% hydrochloric acid matrix in precleaned Teflon
bottles. Use Leeman Labs or J.T. Baker hydrochloric acid. Prepare 50mL of each
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Using the Cold Vapor Technique with the
Leeman Labs, Inc. Automated Mercury System Volume 2, Chapter 2
standard weekly or as needed.
3.7.2.1 0 ppb Standard
3.7.2.1.1 Rinse a precleaned graduated cylinder with 5% nitric acid
followed by three rinses of MSQ. Dispose of waste in an
appropriate container.
3.7.2.1.2 Rinse a precleaned SOOmL teflon bottle three times with
MSQ.
3.7.2.1.3 Add 245 mL MSQ to the bottle.
3.7.2.1.4 Carefully add 5 mL of concentrated hydrochloric acid to the
bottle.
3.7.2.1.5 Transferthe solution to a teflon bottle.
3.7.2.2 0.125 ppb Standard
3.7.2.2.1 Prepare one EDP pipettor with a precleaned 100 jiL
tip.
3.7.2.2.2 Prepare two separate EDP pipettors each with a precleaned
1000 pLtip.
3.7.2.2.3 Rinse a teflon bottle three times with MSQ that will be used
for each standard.
3.7.2.2.4 Tare the bottle on the balance.
3.7.2.2.5 Pipette 62.5 jiL of the 100 ppb working standard into the
bottle
3.7.2.2.6 Use the rinsed graduated cylinder to transfer 45 mL of the 0
ppb standard to the bottle.
3.7.2.2.7 Slowly pipette 5 mL of 0 ppb standard to the bottle until the
bottle contents weight 50 g.
3.7.2.3 0.250 ppb Standard
For the 0.250 ppb std, follow steps 3.7.2.2.1 through 3.7.2.2.4. In step
3.7.2.2.5, substitute 125 jiL of the 100 ppb working standard using a new
1000 jiL precleaned tip. Follow steps 3.7.2.2.6 through 3.7.2.2.7.
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Using the Cold Vapor Technique with the
Volume 2, Chapter 2 Leeman Labs, Inc. Automated Mercury
System
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SOP for Analysis of Sediment for Total Mercury
Using the Cold Vapor Technique with the
Leeman Labs, Inc. Automated Mercury System Volume 2, Chapter 2
3.7.2.4 0.500 ppb Standard
For the 0.500 ppb std, follow steps 3.7.2.2.1 through 3.7.2.2.4. In step
3.7.2.2.5, substitute 250 jiL of the 100 ppb working standard using a new
1000 jiL precleaned tip. Follow steps 3.7.2.2.6 through 3.7.2.2.7.
3.7.2.5 1.00 ppb Standard
For the 1.00 ppb std, follow steps 3.7.2.2.1 through 3.7.2.2.4. In step
3.7.2.2.5, substitute 500 jiL of the 100 ppb working standard using a new
1000 jiL precleaned tip. Follow steps 3.7.2.2.6 through 3.7.2.2.7.
3.7.2.6 2.00 ppb Standard
For the 2.00 ppb std, follow steps 3.7.2.2.1 through 3.7.2.2.4. In step
3.7.2.2.5, substitute 1000 jiL of the 100 ppb working standard using a new
1000 jiL precleaned tip. Follow steps 3.7.2.2.6 through 3.7.2.2.7.
3.8 Reagents Needed for Preparation of Sediment Samples using Protocol PRP7471
3.8.1 50% Aqua Regia (3:1 Hydrochloric : Nitric)
3.8.1.1 Use the glass container that previously held the Suprapure
hydrochloric acid, this is supplied with graduations. Rinse three
times with MSQ to eliminate any residual contamination.
3.8.1.2 To this glass container add 400 mL of MSQ water.
3.8.1.3 To the glass container add 300 mL of Leeman Labs hydrochloric
acid to the bottle.
3.8.1.4 Finally add to the bottle 100 mL of Leeman Labs nitric acid.
3.8.1.5 Carefully transfer solution to instrument bottle.
3.8.2 Potassium Permanganate Solution
3.8.2.1 Rinse a 100 mL precleaned finely graduated cylinder three times
with 5% reagent grade nitric acid followed by three rinses of
MSQ.
3.8.2.2 Dispose of the instrument bottle's contents if applicable and rinse
three times with MSQ.
3.8.2.3 Add 40mLs of the Leeman Labs potassium permanganate
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Using the Cold Vapor Technique with the
Volume 2, Chapter 2 Leeman Labs, Inc. Automated Mercury
System
solution to the rinsed 100 mL graduated cylinder.
WARNING
Potassium permanganate is a strong oxidizer. Keep it from contact with clothing or
combustible materials. Avoid contact with eyes or skin. Avoid breathing the dust.
Handle solid chemical under a fume hood. Handle it while wearing personal
protection gear.
3.8.2.4 Transfer to the instrument bottle.
3.8.2.5 Add 760 mL of MSQ to the bottle.
3.8.2.6 Swirl the contents of the bottle to ensure a mixed solution.
3.8.2.7 Attach to the instrument.
3.8.3 Hydroxylamine Sulfate Solution
3.8.3.1 Rinse a 100 mL precleaned finely graduated cylinder three times
with 5% reagent grade nitric acid followed by three rinses of
MSQ.
3.8.3.2 Dispose of the instrument bottle's contents if applicable and rinse
three times with MSQ.
WARNING
Hydroxylamine sulfate is an eye, skin, inhalation, and ingestion hazard. It will
cause skin irritation and may be absorbed through the skin. Always wear eye
and skin protection.
3.8.3.3 Add 96mL of the Leeman Labs Hydroxylamine Sulfate solution
to the lOOmL graduated cylinder.
3.8.3.4 Transfer the Hydroxylamine Sulfate to the instrument bottle and
continue to add 704 mL of MSQ to the instrument bottle.
3.8.3.5 Swirl bottle to ensure a mixed solution.
3.8.3.6 Attach bottle to instrument.
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Using the Cold Vapor Technique with the
Leeman Labs, Inc. Automated Mercury System Volume 2, Chapter 2
NOTE
This is a modification of the protocol that calls for full strength concentrations
of Potassium Permanganate and Hydroxylamine Sulfate. Full strength
concentrations of these chemicals were attempted and the fumes generated
were very strong. There was no difference in instrument performance in using
some higher concentrations of reagents in the protocol. Both gave acceptable
recoveries on the SRM material and similar instrumental intensities.
4.0 Microwave Sample Preparation
4.1 Preparation of Teflon Digester Vessels
Teflon digester vessels are cleaned following the procedure in section 4 of LLRS-MET-
SOP-010. An alternate method proposed by the manufacturer to increase the life of the
digestion vessels is under consideration. It may not be used until it is verified that blanks
are equally low for the two methods. Upon verification, the following alternate steps may
be substituted for steps 4.4 through 4.7 in LLRS-MET-SOP-010. Steps are taken directly
from CEM manual with slight variations to fit constraints of laboratories (Oilman 1988).
4.1.1 Add 20 mL of concentrated nitric acid to the digestion flask. Place the safety disk
on the vessel and tighten finger tight only. Place the vessel in the turntable, and
attach a venting tube.
4.1.2 Repeat step until the turntable contains 12 vessels.
4.1.3 Turn the MDS-8ID exhaust onto the maximum fan speed. Ensure the turntable is
rotating.
4.1.4 Program the instrument for five minutes and 100% power. Depress the start key
and allow the acid to heat.
4.1.5 Allow the acid to cool to room temperature and manually vent each vessel. Open
vessels and pour the acid into an appropriate waste container.
4.1.6 Rinse the vessels three times with MSQ water and allow them to dry in a clean
area.
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SOP for Analysis of Sediment for Total Mercury
Using the Cold Vapor Technique with the
Leeman Labs, Inc. Automated Mercury
System
4.2 Extraction of Samples
The maximum weight of samples extracted in CEM vessels without venting and still
obtaining acceptable recoveries for all metals has been two grams. For mercury, an
average weight of O.SOOg has been used. The total volume is 25mL in 10% Seastar nitric
acid. Follow steps 5.1.1 through 5.1.11 in LLRS-MET-SOP-010 for preparation of
sediments
5.0 AUTOMATED DIGESTER (AP200) SAMPLE PREPARATION
5.1 Daily Instrument Setup
5.1.1 Clean autosampler rails with isopropyl alcohol.
5.1.2 Lubricate rail s with oil daily
NOTE
Only if reagents are low or instrument was in shutdown will following steps
need to be followed (5.1.3 through 5.)
5.1.3 Open cover and carefully disconnect each bottle separately from the interior of the
instrument.
5.1.4 Rinse each bottle three times with MSQ
5.1.5 Fill bottles numbered 1, 5, 6 with MSQ up to the 800ml mark.
5.1.6 Prepare and fill bottle #3 with the Potassium Permanganate Solution.
5.1.7 Prepare and fill bottle #4 with Hydroxylamine Sulfate Solution
5.1.8 Carefully prepare and fill bottle #2 with the 3:1 Aquaregia solution
5.1.9 Check conditions of all fittings, caps and bottles
5.1.10 Close cover and pressurize system (turn gas on).
5.1.11 Set gas pressure = 20 psi
CAUTION
Do not exceed 25 psi
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5.1.12 Turn instrument on
5.1.13 Turn computer and monitor on.
5.1.14 Turn printer on and press the online button.
5.1.15 After the computer has booted up, at the C: prompt, type apps.
5.1.16 Follow instructions on p4-4 of manual revision c.
5.2 Software and Instrument Setup
Follow the instruction in system startup section 4-3, What follows is a summary. The user
should review the following sections prior to analyses.
References to AP200 Manual
Section 3- System Testing
Section 4- System Operation
Section 5- Routine Maintenance
Section 6- Troubleshooting
5.2.1 Select Protocal and Get PRP7471, defines method, (sec 4-4 of manual)
5.2.2 Check reagent pressure of system, ensure it is > then 5.5psi and within 6.5.
(section 2-10 of manual)
5.2.3 Go to menu, Fl, select Utility, select Diagnostics
5.2.4 Select Reagent Pressure
5.2.5 Run the change rinse solution macro, @CF£RTNS
5.2.6 Rinse the autosampler rinse tray 3-4 times with MSQ
5.2.7 Replace the rinse tray
5.2.8 Run the wake up macro @WAKEUP (section 4-3 of manual)
5.2.9 Check centering of autosampler tip over cups, adjust if necessary (section 3-1)
5.2.10 Check precision of dispenser IX/week (section 3-3), or if there is a pressure
change of .2 psi in system at step 14.3.3
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Using the Cold Vapor Technique with the
Volume 2, Chapter 2 Leeman Labs, Inc. Automated Mercury
System
5.2.11 Set up autosampler sequence, and start finish sequence (section 4.6)
5.3 Preparation for Sample Digestion
5.3.1 Soak the 45mL standard cups and sample cups in 1:1 HNO3 for at least 2.0 hours.
5.3.2 Recycle the acid from the cups back into the Suprapure Acid bottle.
5.3.3 Rinse each cup three to six times with MSQ and allow to air dry.
5.4 Preparation of Standards for Automated Digestion
5.4.1 Prepare an EDP pipettor with a precleaned lOOOul tip.
5.4.2 Using the electronic scale, tare the beaker, and then the standard cup within the
beaker.
5.4.3 Record the weight of the empty cup on the extraction sheet.
5.4.4 Under the laboratory hood, pipette 5mls of the 0 ppb std to the cup.
5.4.6 Weigh the cup to confirm volume delivered is 5mls, and adjust accordingly with
the EDP pipettor.
5.4.7 Repeat steps 16.2 - 16.5 to extract multiple 0 standards.
5.4.8 Repeat steps 16.1 to 16.5 for the 0.125ppb std, 0.250ppb std, O.SOOppbstd.
l.OOppb std, and 2.00ppb std. Each time use a new precleaned lOOOul tip.
5.4.9 Rinse polyethylene vapor covers 3x with MSQ and place over sample cups.
5.4.10 Snap in place aluminum guard over polyethylene vapor barrier.
5.5 Preparation of Sediment Samples for Automated Digestion
Note
Please read the instruction manual for the Mettler Analytical Balance before
proceeding.
5.5.1 Rinse a precleaned teflon spatula with 5% nitric acid.
5.5.2 Rinse the spatula three times with MSQ.
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5.5.3 Use the analytical balance in the Biology Lab, Mettler 21 GOT
5.5.4 Tare the lOOmL polyethylene beaker provided.
5.5.5 Add the 45mL sample cup to the beaker and tare the scale again.
5.5.6 Record the weight of the sample cup on the digestion sheet.
5.5.7 Carefully open the Whirlpak bag of sediment.
5.5.8 Open the Whirlpak bag partially, so only a small circular opening exists (about the
size of a nickel). Caution: if the bag is open all the way, dust from the bag tends to
migrate upwards.
5.5.9 Using the precleaned teflon spatula, carefully scoop an aliquot of sample = 0. lOOg.
5.5.10 Transfer this aliquot to the sample cup. Note: sand like samples will require a
very small aliquot, and silty samples will require a larger aliquot.
5.5.11 Record the weight of the sample in the most significant digits available on the
extraction log sheet.
5.5.12 Place the cup in its designated slot in a sample rack.
5.5.13 Zero the scale with the polyethylene beaker on it.
5.5.14 Rinse the teflon spatula with MSQ water three times.
5.5.15 Wipe teflon spatula dry with a fresh clean wipe square between samples.
5.5.16 Repeat steps 16.6.5 till 16.6.15 for the desired amount of samples, usually
fourteen.
5.5.17 Load dummy cups into any space not occupied by actual sample.
5.5.18 Rinse polyethylene vapor covers 3x with MSQ and place over sample cups.
5.5.19 Snap in place aluminum guard over polyethylene vapor barrier.
5.6 Initiation of the Digestion Procedure
5.6.1 Load autosampler racks into PS200
5.6.2 Confirm autosampler start to finish sequence is correct.
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System
5.6.3 Go to Main Menu, Fl, press MACRO key, and type in @PRP7471
5.6.4 Method will begin to run at this point.
5.6.5 User will be prompted the following: Wait before reducing with KMNO4,
answer Y
CAUTION
User will be prompted near end of 5 hr procedure: Add Hydroxylamine Sulfate, answer Y.
Method will not proceed without this input
5.7 Shutdown of AP200
5.7.1 After protocol is completed, wipe down bath to remove any remaining water.
5.7.2 Run an @APNAP two or three times to clean the dispenser with water.
5.7.3 Remove chemicals and replace reagents with MSQ if it will not be run for an
extended period, and run an @CLEAN twice. This cleans not only the dispenser,
but all reagent lines.
6.0 Automated Analysis of Digested Extracts
6.1 Preparation of the Instrument
6.1.1 Preparation of Drying Tube
6.1.1.1 Rinse a quartz drying tube three times with MSQ, followed by a dilute
rinse of Liquinox, if it previously contained perchlorate.
6.1.1.2 Rinse several times with MSQ to eliminate Liquinox residuals.
6.1.1.3 Allow tube to air dry.
6.1.1.4 Rinse the teflon spatula three times with MSQ and dry with a fresh clean
wipe.
6.1.1.5 Gently place a small plug of quartz wool into one end of drying tube.
6.1.1.6 Carefully pour the Leeman Labs Magnesium Perchlorate into the
plugged drying tube. Try to fill with coarse grained perchlorate. Do not
overfill the drying tube. Overfilling will cause the drying tube to become
blocked more easily once it becomes moistened by the gas stream. When
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SOP for Analysis of Sediment for Total Mercury
Using the Cold Vapor Technique with the
Leeman Labs, Inc. Automated Mercury System Volume 2, Chapter 2
filled, the perchlorate should be able to move within the tube when gently
moving the drying tube from side to side.
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SOP for Analysis of Sediment for Total Mercury
Using the Cold Vapor Technique with the
Volume 2, Chapter 2 Leeman Labs, Inc. Automated Mercury
System
WARNING
Magnesium perchlomte is moderately toxic and a strong oxidizing material. It
is a dangerous fire and explosion risk in contact with organic materials. It is
an inhalation hazard, and contact with skin or eyes can cause irritation. Work
with it in a fume hood while wearing skin and eye protection.
6.1.2 Autosampler Tray Rinse with 10% Hydrochloric Acid
6.1.2.1 Rinse the autosampler rinse tray three times with MSQ.
6.1.2.2 Fill the tray with the 10% hydrochloric rinse prepared in step 3.1.
6.1.3 Tin Chloride Rinse
6.1.3.1 Rinse the tin chloride bottle out three times with MSQ.
6.1.3.2 Fill it with the 10% tin chloride prepared in step 3.3.
6.1.4 Check Tubing Condition and Adjust to Appropriate Tension
6.1.4.1 Check condition of tubing for flattening, abrasion or other signs of wear.
If flattened, replace it.
6.1.4.2 Adjust tension on clamps to a halfway point. The sample line should be
halfway minus one notch.
CAUTION
Do not over tighten clamps. Too much tension will cause tube flattening and a
decrease in overall sensitivity.
6.1.5 Clean and Oil Autosampler Rails
6.1.5.1 Using a clean wipe or clean a paper towel, wipe the autosampler rails with
isopropyl alcohol.
6.1.5.2 Place a small amount of oil on bottom of each rail.
6.1.5.3 Complete a high stress maintenance cleaning monthly.
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SOP for Analysis of Sediment for Total Mercury
Using the Cold Vapor Technique with the
Leeman Labs, Inc. Automated Mercury System Volume 2, Chapter 2
6.1.6 Warmup Period
6.1.6.1 Allow the instrument to warmup for at least one hour time before
analyses. If the instrument was in SHUTDOWN mode for an extended
period allow it to warm up for several hours.
6.1.6.2 Software commands for warmstrt or coldstrt are found on page 16 of the
manual.
6.1.6.3 Perform a COLDSTRT or WARMSTRT
6.1.7 Optimize Optics with an Aperture Test
6.1.7.1 Go to Main Menu, Fl
6.1.7.2 Select Diagnostics, Select Aperture Test
6.1.7.3 Unscrew screw which is furthest out until the minimum absorbance is
obtained. An acceptable value is 0-100.
6.1.7.4 Select Test Optics from Diagnostics menu, and confirm gain, or intensities
are in the range of 500000-1100000 (= to voltage on lamp).
6.1.7.5 Difference between the two beams must be less than 100,000.
6.2 Software Setup for Routine Analysis
Consult Leeman Labs Automated Mercury Analyzer Manual pp 17-29 for guidance on the
software setup. What follows are the software and instrumental parameters used to date.
Complete the following steps before analyses.
6.2.1 Establishing a Protocol = Method file that Contains All Instrumental Parameters
6.2.1.1 From the Main Menu, select Protocol and then select Get.
6.2.1.2 To create a new Protocol, enter its name.
6.2.1.3 Suggested protocol naming is as follows: YYMMDD (Year, Year,
Month, Month, Day, Day. Example 960916F = September 16, 1996.
Note
Limitation is 8 Characters
6.2.1.4 Computer will guide you through prompts (PS200 Manual pp. 18-19).
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SOP for Analysis of Sediment for Total Mercury
Using the Cold Vapor Technique with the
Volume 2, Chapter 2 Leeman Labs, Inc. Automated Mercury
System
6.2.2 Creating a Folder=Data File
6.2.2.1 Press Fl, to be at the Main Menu.
6.2.2.2 Select Data output and select Open Folder.
6.2.2.3 Type in a folder name, suggested name = same name as protocol to avoid
confusion.
6.2.2.4 Follow instructions on page 19 of the manual.
6.2.3 Entering Instrumental Parameters
6.2.3.1 Follow instructions in PS200 instrument manual on pp 20-21.
6.2.3.2 The following are parameters used to date:
Integrations: 1
Uptake Time: 10
Weight: Y
Dilution: Y
Percent Recovery: N
On/Off: 10 (higher integration time generates a nonlinear curve)
Flow Rate: 0.30L/min
6.2.4 Standard Concentration Calculations
For those standards that are prepared with the automated digestion system, the
appropriate concentration to be used for keying in standard concentration data
must be calculated. Do not do this for microwave digested samples.
6.2.4.1 Tare a 100ml polyethylene beaker on the electronic scale.
6.2.4.2 Weigh a sample cup.
6.2.4.3 Record the weight on the extraction log, and zero the scale.
6.2 A A Repeat 6.1.1 through 6.1.3 for all sample cups.
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SOP for Analysis of Sediment for Total Mercury
Using the Cold Vapor Technique with the
Leeman Labs, Inc. Automated Mercury System Volume 2, Chapter 2
6.2.4.5 Apply the following formula to calculate the final concentration of the
extracted standards:
c = r * v /v
^2 M v 1' V2 •>
where,
Cj is the prepared standard concentration,
C2 is the final concentration in the standard,
Vj is the volume of prepared standard used,
and V2 is the total volume of the extract
For example if 5 mL of 0.500 ppb standard was added to the digestion
tube and the final volume of the extract is 43 mL, the resulting standard
concentration is as follows:
C2 = 0.500 ppb * 5 mL/43 mL = 0.058 ppb.
6.2.4.6 Record this concentration on the extraction log.
6.2.5 Entering Standard Concentrations
6.2.5.1 Follow instructions on p. 21 of the manual. Enter units in ppb, not ppm
(i.e. 0.500 not 0.00050).
6.2.5.2 Calculations are only carried out to three decimal points, 0.00050 will be
truncated to 0.000.
6.2.5.3 Do not enter terms of units, i.e. ppb, ppm. The final calculation will be in
ug/g.
6.2.5.4 The following are ranges of standards used for analysis. These ranges
have been successful in bracketing low level samples.
Microwave Digestion Standards (ppb) Automated Prepared Standards (ppb)
(Dependent upon total volume of extract.
See section 6.2.4)
0.000 0.0000
0.250 -0.0140
0.500 -0.0280
1.000 -0.0570
2.000 -0.1140
4.000 -0.2280
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SOP for Analysis of Sediment for Total Mercury
Using the Cold Vapor Technique with the
Volume 2, Chapter 2 Leeman Labs, Inc. Automated Mercury
System
6.2.6 Reset Calibration Intensity Data
Follow instructions on page 22 of manual.
6.2.7 Autosampler Rinse Time
6.2.7.1 Follow instructions on page 23 of manual.
6.2.7.2 Use a rinse time of 60, not 50 seconds. This is the rinse time between
samples in the analyses mode.
6.2.8 Autosampler Rack Entry
6.2.8.1 For basic entry information, refer to page 25 of the manual. For extended
information on macros and advanced command see reference section A-
B-7.
6.2.8.2 What follows is an example of an autosampler rack file. It was used for
analyses of Green Bay sediment. Prepare the file before analyses.
NOTE
Actual sample weight must be multiplied by 1000 to obtain results in ug/g.
Total volume = extraction volume * dilution factor.
1 OPPB 1.000
2GII25F102SQ* 323.00 250.00 SRM diluted IQx
extracted in 25mls
3 GB88-71 280.00 125.00 Sample diluted 5x
extracted in 25mls
4GB89-73 241.00 125.00
5 GB89-74 5.700 125.00
A microwave digested SRM will need to be diluted 10 to 20 times
depending upon the weight of the sample to be within range of standards.
Certified value for SRM2704 = 1.47 ug/g. For a sample that has a
extraction weight of 0.250g in 25mL, (1.47*.250)/25=0.018375
ppm=18.37 ppb. This is diluted 20X = .918 ppb
6.3 Analysis of Extracts
6.3.1 Filling Autosampler Cups
6.3.1.1 Recycle the 50% acid rinse used in the autosampler cups. Place it in the
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SOP for Analysis of Sediment for Total Mercury
Using the Cold Vapor Technique with the
Leeman Labs, Inc. Automated Mercury System Volume 2, Chapter 2
glass SUPRAPURE acid bottle.
6.3.1.2 Rinse each cup 5-10 times with MSQ.
6.3.1.3 Allow cups to air dry
6.3.1.4 Fill each standard cup with its designated standard.
6.3.1.5 Using two separate EDP pipettors, prepare a clean lOmL tip and lOOOuL
tip. Rinse each tip three times with 5% nitric acid followed by three rinses
ofMSQ.
6.3.1.6 For samples and SRMS requiring dilution, dilute to at least half their
capacity of the autosampler tips (6mL). First add the required volume of
diluent with the lOmL tip and then the required amount of the sample. An
SRM will need to be diluted 10 to 20 times depending on weight of
sample. For dilutions of auto-digested samples use 0 ppb standard that
has undergone digestion. For microwave digested samples use 10%
Seastar nitric acid.
6.3.1.7 Mix the sample 3-5 times with the lOOOuL tip.
6.3.1.8 Use a new precleaned lOOOuL tip for each sample.
6.3.2 Calibrate the Instrument
Calibrate the instrument using the Macro CAL245 (p. 26 in PS200 instrument
manual). Use a 5-point calibration curve that includes a zero standard. If an
acceptable correlation coefficient is obtained (0.995) and a standard's intensity is
within a the range expected, continue with SRM analyses. See Appendix A for
historic performance of the instrument
6.3.3 Analyze the SRM
Analyze the SRM. Refer to page 25 in manual for autosampler start to finish
sequence and reference section A-B 7.
6.3.4 Check Standards
6.3.4.1 Run check standards every 10 samples to ensure the instrument has not
drifted from its calibration range.
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SOP for Analysis of Sediment for Total Mercury
Using the Cold Vapor Technique with the
Volume 2, Chapter 2 Leeman Labs, Inc. Automated Mercury
System
6.3.4.2 Acceptable Check Standard Ranges are:
Microwave Digestion Automated Digestion
0.250ppb = 15% ~0.014ppb = 15-20%
0.500ppb=10% -0.028 ppb = 15%
1.00ppb=10% ~0.058ppb = 10%
2.00ppb=10% -0.115 ppb = 10%
6.3.4.3 Refer to reference section c-11 for more information.
CAUTION
If check standards fail, recalibrate the instrument. Do not use update slope or
intercept.
6.4 Data File Preparation
Refer to the reference sections D-l, E-l-5, R-3, and R-5 for preparation of post-run data
and computer files in the AP200 Manual. These data references apply to digested and
samples prepared on the Automated Digester.
6.5 Instrument Shutdown
6.5.1 Dispose of 10% Hydrochloric acid rinse in an appropriate container.
6.5.2 Rinse the tray out three times with MSQ.
6.5.3 Transfer remaining tin chloride to teflon bottle in which it was earlier prepared.
6.5.4 Rinse out tin chloride bottle three times with MSQ.
6.5.5 Fill autosampler tray and tin chloride bottle with MSQ, flush for ten minutes.
6.5.6 Use OVERNITE, or SHUTDOWN modes to shutdown the instrument.
6.5.7 If using OVERNITE MODE, check condition of drying tube, to ensure it is not
saturated with moisture.
6.5.8 Repack a new drying tube if necessary.
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SOP for Analysis of Sediment for Total Mercury
Using the Cold Vapor Technique with the
Leeman Labs, Inc. Automated Mercury System Volume 2, Chapter 2
7.0 Suggestions for Successful Analyses
7.1 Allow the autosampler cups and standard cups to rest in 50% nitric acid for at least two hours
before analyses.
7.2 For best results prepare tin chloride and standards as described.
7.3 Prepare a loosely packed drying tube daily. After use, dispose of perchlorate in an appropriate
container. Rinse a drying tube with MSQ and flush lightly with Liquinox. Rinse several times to
eliminate any residual Liquinox.
7.4 Periodically check tin chloride line and liquid gas separator for any blockage.
7.5 Check at seals of teflon tubing of drying tube connection for gas leaks.
7.6 Change tin chloride line and sample line weekly or after four days of continuous use.
7.7 Change drain tubing every two weeks as needed.
7.8 Clean autosampler rails with isopropyl alcohol weekly and oil rails daily. If not sufficiently
lubricated, the autosampler arm will encounter snags or stops.
7.9 Biweekly calibrate the autosampler tip to ensure it is picking up more than three milliliters.
8.0 Literature Cited
Oilman, L. B., 1988. General Guidelines for Microwave Sample Preparation. Revision of July
1988. CEM Corporation, Matthews, NC.
Leeman Labs, Inc., 1991. PS200 Automated Mercury Analyzer Set-up and Operating Manual.
Version of November 1991.
Leeman Labs, Inc., 1993. AP200 AutomatedMercury Preparation System Manual. Revision C
(11/20/93).
Rossmann, R. And K. A. Rygwelski, 1996. Standard Operating Procedures for the Release of
Data. Revision 1 (1/18/96) LLRS-QA-SOP-001.
Rossmann. R., 1992. Standard Operating Procedures for the Routine Review of Data Quality
and Quantity. Revision 0 (12/2/92) LLRS-QA-SOP-002.
Rossmann, R., 1992. Minimum Analytical Quality Assurance Objectives for U. S. EPA Large
Lakes Research Station. Revision 1 (7/28/92) LLRS-QA-001.
Rossmann, R., 1993. Standard Operating Procedures for the Preparation of Materials used for
Ultra-low Trace Element Analyses. Revision 1 (8/9/93) LLRS-MET-SOP-001
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Using the Cold Vapor Technique with the
Volume 2, Chapter 2 Leeman Labs, Inc. Automated Mercury
System
Rossmann, R., 1994. Standard Operating Procedures for the Maintenance of the LLRS Trace
Metal Laboratories. Revision 1 (5/4/94) LLRS-MET-SOP-003.
Rossmann, R. and T. Uscinowicz, 1994. Standard Operating Procedures for Analysis of Total
Mercury in Tissue and Sediment using the Cold Vapor Technique with the Perkin-Elmer
Model MHS-20 Gold Amalgam System. Revision 1 (5/6/94) LLRS-MET-SOP-010.
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Volume 2, Chapter 2
SOP for Analysis of Sediment for Total Mercury
Using the Cold Vapor Technique with the
Leeman Labs, Inc. Automated Mercury
System
APPENDIX: HISTORIC PERFORMANCE FOR STANDARDS ON PS200.
Daily Standard Intensities and Percent Drift for Standards. Italicized dates are check standards.
Date of
Analysis
01/17/96
01/18/96
01/24/96
01/25/96
01/29/96
01/31/96
02/01/96
02/05/96
02/05/96
02/06/96
2/6/96
2/6/96
2/6/96
02/08/96
2/8/96
2/8/96
02/13/9
2/13/96
2/13/96b
2/13/96b
02/15/96
2/15/96
2/15/96b
Begin New Lamp
03/04/96
03/04/96
03/04/96
03/05/96
03/05/96
03/06/96
0.250
11567
15405
14521
12899
12153
13360
15429
13614
13675
13628
12642
32237
31507
29918
22484
22151
Standard Concentration (ppb)
0.500 1.000 2.000
30807
24577
27507
25177
30207
25133
26287
28271
28294
21563
25966
24807
23963
31433
32989
31222
27169
30254
26879
28977
24557
21928
44465
46352
41993
65754
64815
45823
62450
52846
62220
55494
54795
47097
52552
55116
57208
43415
47569
49621
48835
62122
62558
60391
59876
63307
58308
60081
57516
54759
53348
127713
130143
119879
124625
114069
90596
128462
112763
127897
117531
108733
110427
113649
107236
116260
95356
103997
102217
99624
124122
121403
120489
121738
125174
121548
122166
118139
111736
112076
233412
226759
217858
264984
259107
180965
5.000
319499
289297
309450
279308
283705
280385
286800
262538
287149
258884
263870
259967
251241
312746
316603
312282
299635
304689
297642
278297
286801
268327
270167
620712
626601
598765
628674
609113
422997
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SOP for Analysis of Sediment for Total Mercury
Using the Cold Vapor Technique with the
Leeman Labs, Inc. Automated Mercury System
Volume 2, Chapter 2
APPENDIX: Continued.
Daily Standard Intensities and Percent Drift for Standards. Italicized dates are check standards.
Date of
Analysis
03/07/96
03/07/96
03/12/96
03/14/96
03/14/96
03/14/96
03/14/96
03/15/96
03/15/96
03/19/96
03/22/96
03/26/96b
03/28/961
03/28/962
03/29/96
03/29/96
03/29/96
03/29/96
04/03/96
04/09/96
04/09/96
04/09/96
04/11/96
1 Old Tubing
2 New Tubing
Standard Concentration (ppb)
0.250
21132
19282
23058
24152
24206
24008
25333
23274
23430
22930
23663
27619
22912
21975
20843
20494
42012
40964
21043
0.500
44232
39037
37940
46864
47468
47189
47491
48936
48272
35727
43189
45156
44270
53606
47100
44362
44504
42173
41875
42323
1.000
95743
79923
79106
96048
95645
96993
94850
98592
98146
81247
90404
84083
79330
109573
96970
90593
85748
83552
81646
89779
2.000
194749
202894
166230
192849
188087
188049
187538
195526
197376
178190
194519
188764
184093
212636
194573
191256
196257
167910
169354
167949
160894
178299
5.000
478754
411516
457716
469761
454648
453447
485709
473520
469406
493374
465947
457756
523307
483750
474460
429912
419310
401227
444557
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Volume 2, Chapter 2
SOP for Analysis of Sediment for Total Mercury
Using the Cold Vapor Technique with the
Leeman Labs, Inc. Automated Mercury
System
APPENDIX: Continued.
Daily Standard Intensities and Percent Drift for Standards Prepared with Automated Digestion
System. Italicized dates are check standards.
Date of
Analysis
06/1 1/96
06/13/96
06/18/96
07/03/96
07/09/96
07/12/96a
07/12/96b
07/16/96
07/17/96
07/24/96
07/25/96
08/01/96
08/15/96
08/16/96
09/18/96
-0.014
927
1175
980
600
895
1069
816
Standard Concentration (ppb)
-0.028 -0.056 -0.114
1937
2162
1867
1973
2124
1923
1728
1678
1819
4278
4275
4138
5043
4098
4308
4457
4146
4492
3989
3910
4189
360
8331
8689
8979
8057
8369
8088
8118
8488
8454
8043
7546
7908
7977
8468
7382
-0.228
17993
17995
15743
18117
17318
17009
17285
16962
16190
17072
15964
15077
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