NCD003200383_19901101_Koppers Co. Inc._FRBCERCLA SAP QAPP_Draft Fish Sampling Plan Addendum to RI-FS Work Plan-OCR

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J>RA-F1 ...
FISH SAMPLING PLAN '
ADDENDUM TO /
REMEDIAL INVESTIGATION/ .
FEASIBILIIT STUDY WORK PLAN
KOPPERS SUPERFUND SITE ..
MORRISVILLE, NORTH CA!lOLINA
Pjepare~•rd~:··•··
. BEt\ZiR EAST, INC. .... 436 SEVENTH A VENUE
PITTSBURGH, PA 15219
.. :.. ._::.:
Prepared by:
KF:)'.STONE ENVIRONMENTAL RESOURCES, INC.
······ 3000 TECH CENTER DRIVE
MONROEVILLE, PA 15146
PROJECT NO. 179280-03
NOVEMBER, 1990
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TABEL OF CONTENTS
1.0 INTRODUCTION ...................................................................................................... 1
2.0 BIOLOGICAL BACKGROUND ............................................................................. 1
3.0 PROPOSED FISH COLLECTION ........................................................................ 3
4.0
5.0
6.0
7.0
1-1
1-2
3-1
3-2
3.1 Fish Collection for Human Receptors ····:··:/L ........................................ .4
3.2 Fish Collection for Animal Receptors ,:E. .......... L ..................................... 4
3.3 Fish Sampling Methodology ......... '.;.'..:: ...... , ............. .'.i, ................................ 5
BACKGROUND FISH SAMPLING ........ L ............................................................ 8 ...... '••·
ANALYfICAL PROCEDURES ............ L ................................................................ 8
SCHEDULE ....... , ... : ... : ...... :: .... L, ...... '. ......................................................................... 8 <i:: .. _.:;:/::· ':::
DELIVERABLES ... • .................................................................................................... 9
TABLE OF FIGURES
Topographic•,Map ..................................................................................................... la
General Site Map .................................................................................................... 1 b
Proposed Fish Sampling Ponds .............................................................................. 3a
Fish Sampling Scheme ............................................................................................ 3b
APPENDICES
APPENDIX A Example of Keystones Field Data Sheet
APPENDIX B Example of Permit Reporting Form
APPEND IX C EPA -Extraction and Analysis of Organics in Biological Tissue
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1.0 INTRODUCTION
This report was prepared as an addendum to a previously approved work plan for
the former Koppers laminating plant site. The wood laminating portion of the site
is presently owned and operated by Unit Structures, Inc. and was previously owned
and operated by Koppers Company, Inc., who still owns a portion of the property.
The site is located approximately 1.5 miles northwest of Morrisville, North Carolina
in Cedar Fork Township of Wake County, specifically on Koppers Road, south of
North Carolina Route 54 (See Figure 1-1 ). Residential areas, light industry and
small businesses have developed around the facility. Surfai:~ drainage appears to be
south to southeasterly toward Crabtree Creek which in tum flows into the Neuse
River.
::/'_:: .::/:·
·-....
The plant has produced laminated wood productssince 1962. A general site map is
shown in Figure 1-2. In 1968 .a CELLQN wood treating process, using
pentachlorophenol (PCP), was constructed. atthe site to produce treated wood for
use in the laminated prod11ctt This process was discontinued and the equipment
dismantled in 1975, aftef \Vhich: tieated wood was imported from other facilities.
Unit Structures Inc .. c6nti11mf to p~oduce laminated products from this facility
today.
At the request of tqe EPA, Keystone has prepared a fish sampling. plan to
. ' ...
supplement existjng ecological data that has been collected at this site. A synopsis
of existing data and proposed data collection follows.
2.0 BIOLOGICAL BACKGROUND
A four acre pond is located on the site. This pond is known as the Fire Pond since it
was used to store water for the plant's fire protection system. It was made in about
1954 when the initial construction of the facility was completed. Water level is
maintained by two, side-by-side culverts located in the southwest corner of the Fire
Pond. Excess water flows from this pond via drainage ditches during storms into
Medlin Pond. Medlin Pond is approximately one acre in size.
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• ••
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1000
l-l PIGURJ!l
"15 ~ · AD RANG LE .... , .• " . '. CARY au OLIN A .
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In January, 1971, after a heavy rain, about 200 fingerling fish were found dead in
Medlin Pond. After the incident, the Fire Pond overflow was re-routed around
Medlin Pond until about 1980 when it was routed back to Medlin Pond.
Surface water samples taken in the Fire Pond in 1976 indicated that it could support
fish. The pond was then stocked with 500 catfish and 500 sunfish.
In May, 1977, the area received a strong rainfall which caused mixing of the Fire
Pond. At this time a large number of tadpoles in the pond died. No evidence of a
fish kill was noted. However, in September, 1979 a fish kiUwas reported in the Fire ... ·····
Pond after a heavy rainfall. This occurrence was. attributed to rainwater run-off .... ..,.
contaminated with calcium chloride, which wru; u~ed .as a du~tsuppressant on the
roads.
In September 1980 the Environmenta1Pfotectiqn Agency (Region IV), Surveillance
and Analysis Division (SAD), sampled fish iiib()th the Fire Pond and Medlin Pond
and performed tissue analysis. They C()ri<:luded that no PCP or other toxic organic
compounds were contained in the fish ;amples.
\\ .:/.: .:
In 1986, an ecol◊gi<:aJinv~tigation was conducted by Koppers and the State of
North Carolin~ concurrently with the surface water and sediment sampling in the
Fire Pond ~n.d Medlin P()nd. Results of this investigation indicated that the benthic
macroinvertebr3:te H::ommunity in the Fire Pond was stressed (benthic
macroinvertebrates were not sampled in Medlin Pond) and the fish in both ponds
were contaminated with PCP and some metals. Medlin Pond sediments were found
to contain 100 ug/kg to 892 ug/kg of PCP while the Fire Pond sediments contained
100 ug/kg to 2654 ug/kg of PCP. Oil and grease was detected in the Fire Pond at 73
mg/kg to 260 mg/kg and undetected in Medlin Pond.
Samplings in 1986 revealed four species of fish in the Fire Pond. Fish tissue analysis
of pumpkinseeds (Lepomis gibbosus) indicated PCP levels of 95-157 ug/kg for
whole body and 31.7 ug/kg for filets. In Medlin Pond a total number of fish species
was not obtained, however, whole body fish tissue analysis of bluegills (Lepomis
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macrochirus) indicated PCP levels of 31-200 ug/kg and largemouth bass
(Micropterus salmoides) at 28-65 ug/kg. These values represent results of analyses
from fish tissue collected by Keystone's and the State of North Carolina
laboratories.
In 1987, the EPA conducted additional fish tissue analysis from fish collected in
Medlin Pond. Polychlorinated dibenzo dioxins (PCDDs) were analyzed from a
composite fillet of largemouth bass and a composite whole~body analysis of
bluegills. Results were 4.4 parts per trillion (ppt) and 16.1 ppt, respectively.
3.0 PROPOSED FISH COLLECTION
.... ,.
Fish will be collected by Keystone to provide d;ita fo assess any potential human and
ecological impact from the ingestion of fish. J\ senior field biologist will be
responsible for obtaining a representative fish collection based on field
observations. Several methods riiay Qt: implemented for fish collection.
Electroshocking and gill-netting are the two•rilb~t1ikely methods.
The field sampling progra.111 :;L6egin at the control pond to assess its fish species
composition and density beforb proceeding to Fire Pond and Medlin Pond. If the
initially selecte<i coritrol p&~<i is not productive or does not support species found in
Fire and I\-iedlin PondsJnprevious surveys, another pond will be selected and
assessed bbfqre sampling is conducted. Figure 3-1 depicts two ponds which will be
considered fofuse.as control ponds.
A pre-survey of Fire and Medlin Ponds is not necessary since the dominant species
present are known from previous surveys. The Fire Pond is known to contain
bluegills (Lepornis macrochirus), pumpkinseeds (Lepornis gibbosus), mosquitofish
(Gambusia affinis) and a hybrid sunfish (Lepomis spp.) Medlin Pond is known to
contain bluegills and spotted bass (Micropterus punctulatus). A general fish
sampling scheme is presented as Figure 3-2.
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REFERENCE: ADC RALEIGH, NC STREET MAP
SCALE (FEET)
0 2000
3a
t ~e __ _
CONTROL ] . PONCA .
····---·-·-·-
"
FIGURE 3-1
PROPOSED
FISH SAMPLING PONDS
RALEIGH/MORRISVILLE RIIFS
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FILLET COMPOSITE
SAMPLE
# 1
IF SUFFICIENT SPORT FISH
ARE FOUND
REPLICATE
FILLET COMPOSITE SAMPLE
#2
FISH
FIGURE 3-2
SAMPL::tNG SCHEME
KOPPERS SUPERFUND SITE MORRISVILLE, NC
(FOR ANY GIVEN POND; )
HUMAN RECEPTOR
IF INSUFFICIENT QUANTITIES OF SPORT FISH ARE FOUND
SAMPLE ONLY
FOR ANIMAL RECEPTORS
REPLICATE
FILLET
COMPOSITE SAMPLE
#3
WHOLE
BODY
COMPOSITE
SAMPLE
# 1
POND
IF SUFFICIENT
QUANTITIES OF
BOTTOM DWELLERS
ARE FOUND
REPLICATE
WHOLE BODY COMPOSITE
SAMPLE
#2
ANIMAL RECEPTOR
_______ __J
REPLICATE WHOLE
BODY
COMPOSITE SAMPLE
#3
WHOLE BODY
COMPOSITE
SAMPLE
# 1
IF NO BOTTOM
DWELLERS ARE
FOUND, BUT OTHER NON-SPORT FISH ARE FOUND
USE OTHER
ABUNDANT
NON-SPORT FISH FOR SAMPLE
REPLICATE WHOLE
BODY COMPOSITE
SAMPLE
#2
STONE
IF INSUFFICIENT
NUMBERS OF
BOTTOM DWELLING
OR OTHER NON-SPORT FISH ARE FOUND
REPLICATE
WHOLE BODY COMPOSITE SAMPLE
#3
PERFORM RECONSTRUCTED
WHOLE BODY
ANALYSIS ON SPORT FISH
C68534
3-b
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An attempt will be made to obtain the following information form Medlin Pond and
the control pond: fish stocking dates, species stocked, source of stocked fish and
date pond was constructed.
3.1 Fish Collection for Human Receptors
Fish will be collected and analyzed for pentachlorophenol and polychlorinated
dibenzo dioxins (PCDDs) and polychlorinated dibenzo furans (PCDFs) from both
the Fire Pond and Medlin Pond. Pentachloroanisole may be substituted for
pentachlorophenol provided Beazer East agrees that the E.PA reference documents
relating to this compound are sufficient to allow risk .l!Ssessment calculations to be
performed. Three composite samples with a minimum of five adult fish of the same
species will be collected for the dominant sport species found in the ponds. A
dominant sport fish will be defined as the one that is present in more numbers than
other sport fish found in the same pond. In the field, Keystone's ecologist will
determine which species is domin~rif .Composite fish samples will consist of
individuals that are of similar legal si:ze to represent potential ingestion by humans.
Only the edible portion (fµtet) of theJish will be analyzed to represent that which
may be eaten by humans. ····· '. ... "'
·:::. .::··
If the required nufuber 6f_t!Je same species of sport fishes cannot be collected from
all ponds, a cl~sely r~fatecl species of sport fish will be collected. Those collected
from any b11e pond wiµ all be of the same species.
:::: .····
3.2 }.,i~h ~ollection for Animal Receptors
Fish will be analyzed for pentachlorophenol or pentachloroanisole as mentioned in
Section 3.1 and PCDDs and PCDFs from both the Fire Pond and Medlin Pond. An
attempt to assess a qualitative impact on bird and mammal receptors from fish
ingestion will be made. However, a quantitative assessment of adverse health
effects on the terrestrial biota is impossible to evaluate. Exposure frequency, source
contribution, and other important input variables do not exist for animal receptors
other than humans.
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l>;\IMO-.MF.-.Hl Rl.-4)1 M( L-..1~(
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Since animals may eat any portion of the fish of any species, for this assessment a
whole body analysis from a composite of a wide range of sizes of the same species
will be performed.
If catfish are not caught, another species such as Gambusia affinis or some other
abundant species will be collected. The species collected will not necessarily be a
bottom feeder if catfish are not present, however it will be a non-sport species to
enable an assessment of potential effects on vertebrate animal populations ( non-
human receptors). If no suitable non-sport fish are found, then both fillets and a
reconstructed whole-body analysis will be performed on the sport fish species. This
will provide data to assess both human and animalred:ptors. A reconstructed
whole body analysis is defined as the concentrati611 of the contaminant being
analyzed in the fillet plus the concentration ).rt the .. remainder of the fish after
filleting.
3.3 Fish Sampling Methodoli>!lY
Fish will be sampled from all po11ds lisirtg either a boat mounted electroshocking
device or gill nets, whicheV~r is 111hre appropriate. If gill nets are used they will be
checked on a frequent basis to avoid sample deterioration. If no catfish or other
suitable bottom dii~lling $ecies are obtained from these methods, then baited
hook-lines will be set ;t night based on the assumption that many bottom feeders
are night scayt:!ngers and may be obtained from this method.
Collected fish will be rinsed with distilled water so that no detritus, vegetation or
sediment are frozen with the sample. In the field, each fish will be weighed to the
nearest gram and measured before preparing for analysis. For the human receptor
study, the fillets will be removed from the fish in the field with a clean fillet knife.
The knife will be cleaned with isopropanol and organic-free water before and after
each use. The samples will then be wrapped in aluminum foil that will first be
rinsed with isopropanol, then organic-free water. Fish samples taken for animal
receptors will not be filleted, but sent to the laboratory as whole body samples.
Care will be taken not to expose the fish samples to contaminated surfaces during
processing. Disposable gloves will be worn while filleting the fish. The composited
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samples will be placed in a zip-lock freezer bag and frozen immediately using dry
ice. Each bag will contain a label with the appropriate information to identify the
sample.
The following information will be included on the field data sheets:
Date
Collector's Name
Sampling Location
Pond Name
Fish Species /
Individual Lengths and Weights of Fishfo. Sarriple
Sample Type (Whole or Fillet)
Comments or Unusual Conditiqris, i,e., tumors, sores, fin rot, blind,
etc.
::f::::;:::::::::;::: :.: .. __ ·_ :;;:
An example of the field data sheet can..be foulicl in. Appendix A.
_//\;::. '!\:/·"
Each individual fish £met b~mpClsite satnple will be securely wrapped in aluminum
foil, appropriately labeled, aricl placed in a plastic bag. The label will include:
species, sample nJriiber, cfa'te, location, length and weight of the fish and weight of
the fillets.
Samples will be p~cked in a cooler containing an amount of dry ice totaling 50% of
the sample biomass and shipped to Triangle Laboratory where it will be
homogenized and sent to the appropriate analytical laboratory and analyzed
according to EPA protocol. Sample coolers will be vented to prevent build-up of
carbon dioxide and chain-of-custody forms will accompany each cooler.
To reduce variation in sample preparation and handling, samples will be prepared
using the following procedures:
A.
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To initiate processing, each fish will be measured, weighed and external
condition noted. A fillet will include the flesh tissue, beginning at the mid-
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B.
C.
D.
E.
F.
G.
H.
dorsal line from the left side of each fish. A comparable fillet will be
obtained from the right side. Care will be exercised not to puncture any of
the internal organs. If the body cavity is entered, the fillet will be rinsed with
distilled water. Fish sex and condition of internal organs will be noted during
filleting. Each fillet will be individually weighed prior to wrapping.
Filleting will be conducted on cutting boards covered with heavy duty
aluminum foil, which will be changed between composite samples. Knives,
measurement boards, scales, and cutting board will be cleaned with reagent
grade isopropanol, followed by a rinse with distil.led water between each
composite sample.
Samples will be carefully handled to prevent contamiri.ation by sampler's
hands or by field or laboratory equipmenL Upon collection, fish will be
placed on wet ice in clean coolers prior to p~ocessing.
Excess aluminum foil, carefuliyfolde~, ~Ube used to wrap fillets.
.:,:::::;, ·-:'-,:::·
Pencil, not pen, iii. filling 1utI.6: labels; waterproof marker will be used in
filling out labels.
_./::::;;\:: ··:;;;;
FoHwr1pped sJple ~II be placed in a secured plastic bag.
·:::: :;::
···-.
SamplJwilhri.6~ be kept on ice for more than twenty-four hours.
Samples will be frozen solid as soon as possible and kept frozen until delivery
to the chemical analysis laboratory.
Keystones ecologist will be present at Triangle Labs to supervise the homogenizing
and splitting of fish tissue samples for chemical analysis.
Fish that are observed, but not sampled, will be identified and counted whenever
possible. All fish that are sampled will be recorded on Keystone's field data sheets
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and the permit reporting forms which are required as part of the collection permit
regulations issued by the State of North Carolina (see Appendix B)
4.0 CONTROL POND SAMPLING
An attempt will be made to sample either Pond A or B as noted in Figure 3-1. The
background pond will have the same general physical characteristics of the Fire
Pond and Medlin Pond. It should also have the same species of fish. One
composited sample of fillets from fish of one of the same species as collected in
either the Fire Pond of Medlin Pond will be collected as i>ackground. Likewise, a
composite of species for whole body analysis will be col.lectc:!.d for background. Both
samples will be analyzed for pentachlorophenol o~ pehtachl8ranisole,, PCDDs, and
PCDFs.
5.0 ANALYTICAL PROCEDURES
<_;;::>::,:,:::=' ·.:\.
Fish tissue samples for pentachlorophenoJor pe11tachloroanisole will be sent to a
certified analytical laboratory fbr'analy~is. EPA Method 8040 will be used.
/:iy-__ /\::<. \
Fish tissue samples for PCI)Ds and PCDFs will be sent to Triangle Labs for analysis
by EPA Method Si90>. . ....
... . ..
The EPA, Region IVreport on "Extraction and Analysis of Organics in Biological
Tissue" (Appdnclpc .C) will be used as a reference to aid in the analysis of the fish
samples
6.0 SCHEDULE
On-site fish sampling will commence during the week of November 26, 1990
pending approval of the revised RI/FS schedule and approval of this work plan.
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7.0 DELIVERABLES
A detailed report of findings will be prepared and submitted to Beazer East after
analytical results are obtained. This report will include but not be limited to a
tabulation of field notes, analytical data and lists of species observed and sampled.
Also, Permit Reporting Forms will be returned to the North Carolina Wildlife
Commission as required in the rules and procedures for the collection permit which
they issued for this study.
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APPENDIX A
Example of Keygtoifosf'ield Data Sheet
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FIELD DATA SHEET -FISH
Project Name _________ _ Sample Collector -----------
Project Number ---------Sample Time -------------
Sampling Date ________ _ Weather Description ---------
Sample Total
Location Species Lenoth (in) Weight (a) Comments
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/;: .:f"
.:.:. ···-
APPENDIXB ·-...
Example of Pifrriit R,epCl~ting Form
KEYSTONE 1,>1M0~"1[~T•I. MIMll'RCf_,, 1,,
I rc!CMI I ICCl'VIC """ l'UCM
I Nama of Lau ar 51111■:
I County:
Sample Loc:attaa:
------------------C.11101 No. _____ _ tNCWIIIC 111& ONl.'tl
Saple Date: I I I 7
lltltDIS C .... INe, NU ..... &a. lTAft ... Ta&ltl&CT, &O'fa O&alMATION. IITC.I
I Sa■pll111 !!!art:
•
I Collectlna Welbad: --Blamflahill1 __ Stbdq --N1tUn1 ·
--TN•Uns ---Tnpplna ---Rotenone
__ Otber lapeclfyl -------------------
I Collector:-------,--,------,--------------,-.,.,,,,.,.----------,N ........... ,,. OA COMl'AN1'1 AQCIIIIU I Penni! Nwnblr: ______________ _
Lill of Fl1be1:
I COIIIION IWII SCIINTIPIC IWII TOTAL NUIIIU TOTAL WIIC:HT
•----, _______ -------
1 -------------
1----
1--------------
•---------------
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APPENDIXC
EPA-Extractioriand Analysis of
Organics iri Biological Tissue
I
I Method OB 8/90
O,S. BNVl:RONMENTAL PROTECTION AGENCY
DIVIROHKENTAL SERVICES DIVISION, REGION IV
ANALYTICAL SUPPORT BRANCH
ATHENS, GA
U'l'RACTIOK AND ANALYSIS OF ORGANICS IM BIOLOGICAL TISSUE
SCOPB AND APPLICATION
I 1.1. This method covers the determination of semivolatile and
volatile organics, and pesticides/PCBs in biological tissue. See the
analytical methods for lists of analytes.
I 1.2. The limit of quantitation for this method is usually
dependent upon the level of interferences rather than instrumental
limitations. Where interferences are not a problem, the limit of l quantitation for moat compounds is as follows1 aemivolatiles-2 mg/kg;
volatiles-0.05 mg/kg; pesticides-0.01 mg/kg; and PCBs-0.0S mg/kg.
I 1. 3. Thia method is recommended for use only by experienced
residue analysts or under the close supervision of such qualified person~.
SUMMARY OF METHOD
2. l. The •biological tissue is ground to the consistency of
hamburger in a meat grinder. A portion of this is mixed with sodium
•
sulfate and extracted with a specified solvent using an ultrasonic prob~
(Method 3550). The extract is filtered, concentrated, cleaned up using
gel permeation chromatography, and again concentrated (Method 3640). The
l extract is split into two portions. One portion is exchanged into hexane,
cleaned up on a alumina microcolumn and analyzed on a gas chromatograph
/electron capture detector for pesticides and PCBs (Method 8080). 'l'he
other portion is analyzed by gas chromatograph/mass spectrometer (GC/MS)
l !or semivolatile organics (Method 8270). Another portion of ground tissue
is analyzed for volatile organics using GC/MS and a heated purge and trap
system (Method 8260). The Methods referred to are found in US EPA, Test
I Methoda for Evaluating solid waste, sw-846.
3. INTERFERENCES
I 3.1.
. chl:'omatography.
Most fish lipids are eliminated by gel permeation
I 3 ,2. Additional cleanup techniques may be necessary to remove
. other natural interferences when lower quantitation limits are required.
I samples.
3.3. The ultrasonic probe must be scrupulously cleaned between
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3. 4. Method interferences may be caused by contaminants in
solvent■, reagente, qla■eware, and other sample processing hardware that
lead to discrete artifact& or elevated baseline• in gas chromatograms.
J\ll theee material• must be routinely demonstrated to be free from
interferences under the condition& of the analyaie by analyzing laboratory
blanks.
4. APP.I\RATUS AND MATERW.S
4.1. Meat grinder. hand crank---for small fish and fillets (300
gm or less).
4.2. Food processor. Hobart Model 8181D or equivalent---for large
fish.
4.3. Sample preparation apparatus listed in SW-846 Method 3550,
Sonication Extraction,
4.4. Cleanup apparatus listed in SW-846 Method 3640, Gel
Permeation Cleanup.
4.5. Organochlorine Pesticide and PCB analysis apparatus listed
in SW-846 Method 8080.
4,6. Semivolatile Organic analysis apparatus listed in SW-846
Method 8270,
4.7. Volatile Organic analysis apparatus listed in SW-B46 Method
8260.
4. 8. Chromatographic column for alumina: 3-mL SPE disposable
filtration columns and bottom frit, J.T. Baker or equivalent.
4,9. Culture tubes for sulfuric acid cleanups 150 x 15 mm with
screw caps and Teflon liner.
5. REAGENTS1
5.1. Extraction solvent for,
5.1.1, Organic scan for semivolatiles and pesticides/PCBs-
--methylene chloride.
5.1.2, Organochlorine pesticides/PCBs---hexane.
5,2. Alumina-Neutral. Super I woelm or equivalent. (Available
fr0111 ICN Biomedicals, Inc.). Prepare activity III by adding 7\ (v/w)
reagent water to the Super I neutral alumina. This is normally done by
adding 5.25 mL of water to 75 gm of alumina in a 100 mL bottle. The bottle
is capped and shaken by hand until no lumps are observed. Tumble or shake
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I (in a wrist action shaker) overnight, There should be no lumps present.
Store in a tightly sealed glass container,
I ·5, 2, 1. Alumina Equivalency Check. Test the alumina by adding
the semivolatile surrogates in l I l acetone/hexane to the alumina and
following paragraph 7,4,2,3, The tribrornophenol should not be detected by
GC/EC if the alumina and its activation are acceptable. Also check
I recovery of all single component pesticides following the same procedure,
'The percent recovery for all single component pesticides must be> or a
BO\, except for endosulfan sulfate which must be > or = 60\, Endrin I aldehyde ia not recovered.
5,3. Sulfuric acid---reagent grade,
I 5,4. Lipid Spike, Prepare a solution containing 1 mg/mL of Cod
Liver Oil in hexane,
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5,5. See the specific SW-846 Method for other reagents,
SAMPLE COLLECTION, PRESERVJI.TION, AND HANDLING
I 6.1. Whole fish samples are submitted to the lab for projects
· where environmental effects are being investigated. Fillets only should
be submitted when human health effects are being investigated.
I• 6.2. Wrap the fish or tissue in aluminum foil (shiny side out),
Wide mouth glass jars with Teflon or foil lined lids are suitable for very
I small fish, organs, small organisms, or ground tissue. Plastic bags should
not be used for organic samples because of the possibility of contamination
by phthalates or other plasticizers, unless they are first sealed in aluminum foil,
I• · 6.3. Biological samples must be preserved on wet ice until hand .
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delivered to the lab to prevent deterioration of the tissue. They also
may be frozen in the field and shipped to the lab on dry ice. Store in
the freezer until the samples are ·prepared for analysis. Most organics
found in fish are relatively stable, however, emulsion problems increase
1 during sample preparation when stored for long periods of time.
6.4. Grinding of biological tissue is easier when the
, .. partially frozen. This is especially true when attempting to I skin from whole fish samples.
7 , PROCEDtJRB
tissue is
grind the
I 7.1. Organics are not evenly distributed throughout biological
tissue, especially fish. This is often true of even fish fillets.
Therefore, to obtain a homogeneous sample, the whole fish or the whole I fillet must be ground to a homogeneous consistency, '
7,1,1, Chop the samples into one inch chunks unless the
1 sample ia small enough to fit in the
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grinder. Then pass the whole sample
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through a meat grinder, Grinding of biological tissue is easier when the
tissue is partially frozen, Thia is especially true when attempting to
grind the akin from whole fish samples, (Chilling the grinder with a few
chips of dry ice will reduce the tendency of the tissue to stick to the
grinder. Do not freeze the grinder, since hard frozen tissue is difficult
to force through the chopper plate). The ground sample is divided into
quarters, opposite quarters are mixed by hand and thon the two halves are
mixed back together .. Repeat the mechanical grinding, quartering and hand
mixing two more tunes. No chunks of tissue should be present at this point
as they will not be ef fie lently extracted. If the ground fish ls to be
frozen prior to extraction and analysis, follow step 7.1.3.
7.1.2, Compositing fillets or whole fisha Grind each
individual sample separately as per step 7.1.l, Take equal amounts from
each sample to be composited to provide a total, equal to that required in
the method extraction step.
7,1.J, If the ground fish is to be frozen prior to extraction
and analysis, weigh out the exact amount for extraction into a small
container. Using a top loading balance, tare a 2 oz. glass jar (or a small
sheet of aluminum foil that can be formed into a sealed packet) to 0.0 gm
and carefully dispense a 20,0 gm portion of homogenized tissue into the
container. Tightly seal the container or foll packet. Repeat with a
second container to be used for duplicate analysis or stored for future
reanalysis. Lipid material tends to migrate during freezing, therefore,
storing a weighed portion ensures extraction of a representative portion
of the tissue.
7.1.4, Whenever a ground sample is to be split between two
or more labs, the ground sample from 7,1,1 or 7.1.2 must also be mixed with
anhydrous sodium sulfate to ensure the homogeneity of the sample prior to
splitting. Transfer 100 gm of ground tissue to a 600 mL beaker. Add 250
gm of anhydrous sodium sulfate and mix thoroughly with a stainless steel
spoon or a spatula, There should not be any lumps and the mixture should
appear homogeneous. Dispense exactly 70.0 gm of mixture to each lab and
note on the package that it contains 20 gm of tissue.
7.2. When preparing the tissue for volatile analysis, grind it
in an area free of volatile organic compounds. The meat grinder or food
processor must be heated in an oven for 30 min. at 105°C after solvent
rinsing, Immediately after grinding· the tissue, weigh duplicate 1 gm
portions into culture tubes with screw caps. Analyze immediately or store
.l.n a freezer, See Section 7,6 for volatile Analysis.
7.3. Sample preparations
7,3.1. Organochlorine Pesticide/PCB Analysis, Weigh 20 gm
of ground sample or quantitatively transfer the 20 gm portion weighed out
in 7,1,J, into a 400 mL beaker, Add 80 gm of anhydrous sodiwn sulfate and
mix thoroughly with a spatula until the sample has a free flowing, sandy
appearance. Drying normally takes 5 -10 minutes. Add surrogate spike
just prior to addition of solvent. Add 100 mL of hexane (add additional
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ntJif the sonicator horn is not below the surface of the solvent but , tne surface of the sample/sodium sulfate mixture). Follow the ,dure in Method 3550, Sonication Extraction. Stir the mixture with a 11~ when the 2nd and 3rd portion of solvent is added to ensure all ions of the sample are exposed to the ultrasonic energy. If any chunks isff/.are noted, carefully move the beaker so the chunk is close to the cJ:or horn while it is sonicating, It should disintegrate the tissue, O JU.LOW THE HORN TO TOUCH THE BEAKER WALL AS THE BEAKER WILL BREAK1
1 7.3,2. Organic Scan for Semivolatiles, (If Organochlorine ~c:,des and PCBs are also requested, they may be included in this ra.c ion) • Follow the same procedure as 7, 3, l except use methylene or e. Add surrogate spikes for both pesticides and semivolatile ~cs if both groups of analytes are requested, ·
.Ji 7,3,3. Lipid Spike, Add 25 mL of the Lipid Spike to a 400 ~Jker containing 80 gm of anhydrous sodium sulfate. Add 100 mL of the prQpriate solvent and take the Lipid Spike through the extraction oc· ureas specified in 7,3,l, Then proceed to the %Lipids Determination o0 tlined in 7,5, ,;
7,4, Extract Cleanup:
:,-7. 4. 1. All tissue extracts must be cleaned up with Gel e eation Chromatography· (GPC) unless the sulfuric acid cleanup is opriate. Where the analytes are amenable to sulfuric acid cleanup, ally no other cleanup is required, Follow Method 3640 for GPC cleanup.
7,4.l.l. Prior to GPC cleanup, concentrate the extracts 7,3,1 (Pesticides/PCB&) as close to l mL as possible. Follow the tructions on concentration found in Methods 3550 and 3640. Dilute back 10.0 mL with methylene chloride. Extracts from 7,3,2 (Organic scan) are centrated so that the final volume is 10, 0 mL, keeping methylene oride as the final solvent. One mL of the 10 mL is used for determina-n of% Lipids as per 7,5. The remaining volume is transferred to the system as per Method 3640,
7.4.l.2. Following GPC cleanup, concentrate as follows:
7.4.l.2.l. Pesticides/PCBs Only---Concentrate the C extract to approximately 10 mL, then add 20-30 mL of hexane and a fresh oiling chip. Concentrate to 0,5 mL and add 0,5 mL of acetone. The 1.0 final volume is ready for Micro Alumina cleanup (the final volwne is 2.0 considering only 5 of the 10 mL went through the GPC system).
7.4.l.2.2. Organic Scan including Pesticides/PCBs---Concentrate the GPC extract to a final volume of 1.0 mL. Transfer 0.5 mL of extract to a 2 mL GC vial and dilute with O. 5 mL of methylene chloride. Label this for Semivolatile analysis (the final volume is 4 mL considering only 5 of the 10 mL went through GPC plus the final 2 time dilution factor). This extract is ready for Semivolatile analysis by
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Method 8270, (High concentrations of natural organics from the fish tissue
often interfere with Semivolatile analysis. Additional adsorption cleanup
methods may be used to eliminate these interferences, however, the
base/acid compound• will also be removed), Reconnect the KD 500 mL flask
to the concentrator tube containing the other 0.S mL of extract and add 20-
30 mL of hexane along with a fresh boiling chip, Concentrate to 0,S mL and
add 0,S mL of acetone. This is for Pesticide/PCB analysis (also has a
final volwne of 4,0 mL) and is ready for Micro Alwnina cleanup. The Micro
Alwnina column removes the_ acid surrogates as well as additional lipid.
7.4.2. Micro Alwnina Cleanup (For Organochlorine Pes-
ticides/PCB& only),
7.4.2.1. Add 3 gm of activity III neutral alwnina to the
3 mL diaposable chromatographic column, Tap the column to settle the
alwnina, Do not pre-wet the alwnina.
7.4.2,2. Transfer the l mL of hexane/acetone extract from
paragraph 7, 4. 1. 2, 1 or 7. 4, 1. 2, 2 to the top of the alumina using a
disposable Pasteur pipet. Collect the eluate in a clean 10 mL concentrator
tube,
7.4,2,3, Add l mL of hexane to the original extract
concentrator tube to rinse it. Transfer these rinsings to the alwnina
column. Elute the column with an additional 9 mL of hexane,
7.4.2,4. Concentrate the extract to a final volume of
1.0 mL.
7.4.2,5, The pesticide/PCB fraction is ready for
analysis. Proceed to Method 8080.
7.4.3. Sulfuric acid cleanup. (This cleanup has only been
validated for PCBs, chlordane and the DDT series. Certain other or-
ganochlorine compounds may also be amenable to sulfuric acid cleanup,
however validation is required),
7,4,3,1, Combine all three hexane extracts, following
sonication and filtration, into a KD evaporator along with all rinsings.
Concentrate so that the final volwne is 10 mL,
7,4,3,2, Withdraw 5 mL into a culture tube. Add 10 mL
of concentrated sulfuric acid, Secure the screw cap and shake the tube
for l min, The lipids are extracted into the sulfuric acid leaving the
analytes in the cleaned up hexane layer. Quantitatively transfer tho top
(hexane) layer to a clean concentrator tube. Accurately redilute the
extract to 5,0 rnL if the 5.0 mL mark is accurate or concentrate to 1,0 mL
and pipet in 4,0 11\L, Thia dilution is required because the 5 mL taken for
acid cleanup contained l-2 mL. of lipid. Some samples may require
additional cleanup with additional 10 rnL portions of sulfuric acid,
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7.4.3.3. The extract is now ready for analysis by Method
8080.
7.5. petermination of, Lipids,
7.5.1. Transfer 1,0 mL of the 10,0 mL extract in 7.4.l.l to
a tared aluminum weighing boat, Thia will not effect the final volu.111.i as s mL is lost when loading the GPC,
7.S.2. Place the tared boats in a hood and allow the solvent
to evaporate overnight. Handle the boats with tongs and protect from
particulate causing inaccurate weights.
7.5.3. Determine the residue weight by reweighing the boats.
calculate\ lipids as follows:
\Lipids• Residue weight x 100 l mL/10 mL X Sample Weight
7.6. Volatile Organic Analysis,
7.6.1. Add 5.0 mL of organic free water already spiked with
the surrogate spike to the samples prepared in 7.2. Replace the cap and
shake the contents until the solids are dispersed throughout the water.
7.6.2. The samples are ready for purge and trap analysis as
outlined in Method 8240, using the low soil method of heating the purge
-device to 40°C,
8. QOALI'l'Y CONTROL
8.1. Any reagent blanks or matrix spike samples should be subject
to exactly the same analytical procedures as those used on actual samples.
8.2. Refer to Chapter One of SW-846 for specific quality control
procedures and Method 3500 for extraction and sample preparation
procedures,
9. MBTHOD PBRl"ORMAMCB
To be added,
10. RBl"BRENCBS
10.1. us EPA, Test Methods for Evaluating Solid waste, SW-846,
3rd Edition, 1986,
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