EPA
United States Environmental Protection Agency
Office of Water Regulations and Standards
Industrial Technology Division
Office of Water                 July 1989
 Method 1613:  Tetra-through
 Octa- Chlorinated Dioxins and
 Furans by Isotope Dilution

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Introduction
Method  1613  was   developed   by  the  Industrial  Technology
Division   (ITD)   within   the  United   States   Environmental
Protection Agency's  (USEPA) Office  of Water Regulations  and
Standards (OURS) to provide improved precision and accuracy of
analysis of pollutants in aqueous and solid matrices.   The ITD
is  responsible  for  development  and promulgation of nationwide
standards  setting   limits  on  pollutant  levels   in industrial
discharges.

Method  1613  is  a  high  resolution  capillary  column  gas
chromatography (HRGO/high resolution mass spectrometry (HRMS)
method  for  analysis  of  tetra-  through  octa-  chlorinated
dibenzo-p-dioxins  (PCDDs)  and  dibenzofurans  (PCDFs)  using
isotope dilution.   Specificity  is provided  for determination
of  the  2,3,7,8- substituted  isomers of tetrachlorodibenzo-p-
dioxin  (2,3,7,8-TCDD)  and tetrachlorodibenzofuran  (2,3,7.8-
TCOF).
  I
Questions concerning  the  method or  its  application  should be
addressed to:

U. A. Tel Hard
USEPA
Office of Water Regulations and Standards
401 M Street SW
Washington. DC  20460
202/382-7131

OR

USEPA OWRS
Sample Control Center
P.O. Box 1407
Alexandria, Virginia  22313
703/557-5040
Publication date:   July 1989
                                                                                   Printed on Recycled Paper

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Method  1613    July 1989
Tetra-  through  Octa- Chlorinated Dioxins and  Furans
by  Isotope  Dilution  HRGC/HRMS
        1    SCOPE AND APPLICATION

      1.1    This method  is designed to meet the survey
            requirements  of  the  USEPA   ITD.      The
            method  is  used  to  determine the  tetra-
            through   octa-   chlorinated   dibenzo-p-
            dioxins  (PCDDs)  and dibenzofurans (PCOFs)
            associated  with  the  Clean  Water Act  (as
            amended  1987);   the  Resource Conservation
            and Recovery Act  (as amended 1986);  and
            the Comprehensive  Environmental  Response,
            Compensation and Liability Act (as amended
            1986);   and   other   dioxin  and   furan
            compounds   amenable   to   high  resolution
          '  capillary    column    gas    chromatography
            (HRGC)/high  resolution  mass spectrometry
            (HRMS).     Specificity  is   provided  for
            determination of  the 2,3,7,8- substituted
            isomers   of   tetrachlorodibenzo-p-dioxin
            (2,3,7,8-TCDD) and  tetrachlorodibenzofuran
            (2,3,7,8-TCDF).

      1.2    The method  is  based on  EPA,  industry,
            commercial    laboratory,    and   academic
            methods  (References 1-6).

      1.3    The compounds listed in Table  1 may  be
            determined  in  waters, soils, sludges,  and
            other matrices by this method.

      1.4    The detection  limits of  the method  are
            usually   dependent   on   the   level   of
            interferences  rather  than  instrumental
            limitations.  The levels in  Table 2 typify
            the  minimum  quantities   that   can   be
            detected with no interferences present.

      1.5    The GCMS portions  of the method are  for
            use only   by  analysts  experienced  with
            HRGC/HRMS  or under  the  close supervision
            of  such    qualified  persons.       Each
            laboratory   that  uses  this  method  must
            demonstrate   the   ability    to   generate
         ,   acceptable  results  using  the procedure  in
         1   Section 8.2.

        2    SUMMARY OF METHOD

      2.1    Stable  isotopically  labeled  analogs  of  16
            of the  PCDDs and PCDFs  are  added to each
            sample.   Samples containing  coarse  solids
            are prepared for extraction  by grinding  or
            homogenization.       Water   samples   are
            filtered and then extracted  with methylene
            chloride     using     separator/     funnel
      procedures;   the  participates  from   the
      water  samples,  soils,  and  other  finely
      divided  solids  are  extracted   using  a
      combined   Soxhlet    extraction/Dean-Stark
      azeotropic  distillation  (Reference   7).
      Prior   to   cleanup   and   analysis,   the
      extracts of  the  filtered water  and  the
      particulates are combined.

2.2   After  extraction,    Cl^-labeled  2,3,7,8-
      TCDD is  added  to  each extract  to measure
      the  efficiency of  the  cleanup  process.
      Samples   cleanup    may   include    back
      extraction with acid and/or base,  and  gel
      permeation,   alumina,   silica  gel,   and
      activated  carbon  chromatography.    High
      performance  liquid  chromatography  (HPLC)
      can be  used  for further isolation  of  the
      2,3,7,8- isomers or other  specific isomers
      or congeners.

2.3   After cleanup,  the  extract is  concentrated
      to  near dryness.    Immediately  prior  to
      injection,   two  internal  standards   are
      added to each extract, and a  1  uL aliquot
      of  the  extract is  injected  into  the  gas
      chromatograph.   The analytes  are separated
      by   the  GC  and   detected   by   a   high
      resolution  (>10,000)   mass   spectrometer.
      The labeled compounds  serve to correct  for
      the   variability   of    the    analytical
      technique.

2.4   Dioxins  and   furans  are  identified   by
      comparing GC retention time ranges and  the
      ion abundance ratios of  the m/z's  with  the
      corresponding  retention  time  ranges   of
      authentic  standards and  the  theoretical
      ion abundance  ratios  of the   exact  m/z's.
      Isomers and congeners are identified  when
      the   retention  time   ranges   and   m/z
      abundance ratios agree  within pre-defined
      limits.  By  using  a GC column  or columns
      capable   of    resolving   the    2,3,7,8-
      substituted isomers from all   other tetra-
      isomers,  the  2,3,7,8-substituted  isomers
      are identified  when the retention  time  and
      m/z  abundance   ratios  agree  within  pre-
      defined limits  of the  retention  times  and
      exact m/z ratios of  authentic  standards.

2.5   Quantitative analysis  is performed by  GCMS
      using selected  ion  current profile (SICP)
      areas,   in  one  of  two ways:    1)   For  the

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        16  2.3,7,8-substituted  isomers  for which
        labeled  analogs are  available  (see Table
        1), the  GCMS  system is calibrated and the
        compound concentration  is determined using
        an  isotope dilution technique; 2) For non-
        2,3, 7,8-substituted isomers  and the total
        concentrations  of  all  isomers within  a
        level of  chlorination (i.e., total TCDO),
        concentrations   are  determined  assuming
        response  factors from  the  calibration of
        labeled  analogs  at   the  same  level  of
        chlorination.   Although  a  labeled analog
        of  the octachlorinated dibenzofuran (OCDF)
        is  available,  using  high  resolution mass
        spectrometry,  it  produces  an m/z that may
        interfere  with   the    identification  and
        quantisation of  the native octachlorinated
        dibenzo-p-dioxin  (OCOD).   Therefore,  this
        labeled  analog has not been  included in
        the calibration  standards,  and the native
        OCDF  is quantitated  against  the   labeled
        OCDD.

  2.6   The  quality  of  the  analysis  is  assured
        through   reproducible   calibration   and
        testing  of   the extraction,  cleanup,  and
        GCMS systems.

    3   CONTAMINATION AND INTERFERENCES

  3.1   Solvents,  reagents,   glassware,  and other
        sample   processing    hardware   may  yield
        artifacts    and/or    elevated , baselines
        causing misinterpretation of chromatograms
        (References 8-9).   Specific selection of
        reagents  and purification  of  solvents by
        distillation  in all-glass  systems  may be
        required.   Where possible,  reagents  are
        cleaned by extraction  or solvent rinse.

  3.2   Proper cleaning  of  glassware is extremely
        important because glassware may  not only
        contaminate  the  samples,   but  may  also
        remove   the   analytes  of   interest   by
        adsorption on the glass surface.

3.2.1   Glassware  should be   rinsed  with  solvent
        and  washed  with  a  detergent  solution as
        soon   after    use    as   is   practical.
        Sonication   of   glassware   containing  a
        detergent solution  for approximately 30 s
        may aid  in cleaning.

3.2.2   After detergent  washing, glassware should
        be  immediately  rinsed first  with methanol,
        then  with  hot  tap  water.   The tap water
        rinse  is  followed   by  another  methanol
        rinse,  and  then  acetone,   and methylene
        chloride.
3.2.3   Do not bake  reusable  glassware  in an oven.
        Repeated  baking  of  glassware  may  cause
        active  sites  on  the glass  surface  that
        will  irreversibly  adsorb PCDDs/PCDFs.

3.2.4   Immediately    prior   to   use,    Soxhlet
        extraction   glassware   should   be   pre-
        extracted with  toluene  for  approximately 3
        hours.   See  Section  11.1.2.3.   Separatory
        funnels  should  be shaken  with  methylene
        chloride for 2  minutes.

  3.3   All  materials  used  in  the analysis  shall
        be    demonstrated    to   be    free    from
        interferences  by running reference  matrix
        blanks  initially and with each  sample set
        (samples  started  through  the  extraction
        process  on  a  given  12-hour  shift,  to  a
        maximum  of  20).    The  reference  matrix
        blank   must    simulate,   as   closely   as
        possible,  the  sample matrix  under  test.
        Reagent  water  (Section 6.6.1)  is used  to
        simulate  water  samples;   playground  sand
        (Section  6.6.2)   or  white  quartz   sand
        (Section  6.5.4) can  be used  to  simulate
        soils;  filter  paper  (Section  6.6.3)  is
        used   to  simulate   papers  and   similar
        materials; other materials  (Section 6.6.4)
        can be used  to  simulate other matrices.

  3.4   Interferences   coextracted  from   samples
        will  vary   considerably   from   source  to
        source, depending  on the diversity  of  the
        site  being sampled.   Interfering compounds
        may  be present  at concentrations  several
        orders of  magnitude higher than  the PCDDs
        and    PCDFs.       The   most    frequently
        encountered  interferences  are  chlorinated-
        biphenyls,        methoxy        biphenyls,
        hydroxydiphenyl     ethers,     benzylphenyl
        ethers,    polynuclear    aromatics,     and
        pesticides.   Because very low levels  of
        PCDDs  and   PCDFs   are  measured  by  this
        method,  the  elimination  of  interferences
        is essential.   The cleanup steps  given  in
        Section  12  can   be  used  to   reduce  or
        eliminate  these interferences  and  thereby
        permit reliable determination of the PCDDs
        and PCDFs the at levels shown in Table 2.

    4   SAFETY

  4.1   The  toxicity  or  carcinogenicity  of  each
        compound  or  reagent  used  in  this  method
        has   not    been   precisely    determined;
        however, each  chemical  compound  should  be
        treated  as  a   potential   health   hazard.
        Exposure  to  these  compounds   should  be
        reduced to the  lowest possible  level.

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4.1.1   The  2,3,7,8-TCDD  isomer has been found  to
        be     acnegenic,     carcinogenic,     and
        teratogenic  in laboratory animal studies.
        It  is soluble in  water  to  approximately
        200   parts-per-trillion  and  in   organic
        solvents to 0.14 percent.  On  the basis  of
        the  available  toxicological  and physical
        properties  of  2,3,7,8-TCDD,  all   of the
        PCDDs  and  PCDFs should be handled  only  by
        highly   trained   personnel    thoroughly
        familiar   with  handling  and   cautionary
        procedures,    and   who   understand  the
        associated risks.

4.1.2   It   is recommended  that   the  laboratory
        purchase dilute standard solutions  of the
        analytes  in   this  method.    However,   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.

  4.2   The    laboratory   is   responsible  for
        maintaining  a  current awareness  file  of
        OSHA  regulations   regarding   the  safe
        handling  of   the  chemicals  specified   in
        this method.   A  reference  file  of data
        handling   sheets   should   also  be made
        available  to   all  personnel  involved   in
        these  analyses.   Additional  information  on
        laboratory   safety  can   be.  found   in
        References  10  -  13.    The references and
        bibliography  at  the  end  of  Reference  13
        are  particularly comprehensive  in  dealing
        with  the  general  subject  of  laboratory
        safety.

  4.3   The  PCDDs  and PCDFs and samples suspected
        to   contain  these  compounds  are  handled
        using  essentially  the  same techniques  as
        those  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 of  Labor,  many  of
        which  have an industrial  health service.
        The  PCDDs and  PCDFs are extremely toxic  to
        laboratory  animals.    However,  they have
        been handled  for years without injury  in
        analytical  and  biological   laboratories.
        Each  laboratory  must  develop   a   strict
        safety program for  handling the PCDDs and
        PCDFs.  The  following laboratory practices
        are  recommended (References 2  and 14):

4.3.1   Facility  --   When  finely  divided  samples
        (dusts, soils, dry  chemicals)  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 leak  tight or fume  hood  demonstrated  to
          have  adequate  air  flow.   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 hazards
          except in  the case of  an accident.

  4.3.2   Protective equipment  -- Throwaway plastic
          gloves,  apron  or  lab coat, safety glasses
          or  mask,  and   a  glove  box or  fume hood
          adequate  for  radioactive  work.     During
          analytical operations  which may  give rise
          to  aerosols  or dusts,   personnel  should
          wear  respirators  equipped  with  activated
          carbon filters.   Eye  protection  equipment
          (preferably  full   face  shields)   must   be
          worn  while working with  exposed samples  or
          pure  analytical standards.   Latex  gloves
          are commonly used  to  reduce  exposure  of
          the   hands.      When   handling    samples
          suspected   or   known   to   contain  high
          concentrations  of  the PCDDs  or PCDFs,   an
          additional set  of  gloves can also be worn
          beneath  the latex gloves.

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

  4.3.4   Personal  hygiene  --   Thorough  washing   of
          hands and  forearms  after  each  manipulation
          and   before  breaks   (coffee,   lunch,  and
          shift).

  4.3.5   Confinement --  Isolated work area,  posted
          with   signs,    segregated   glassware  and
          tools,  plastic absorbent   paper  on  bench
          tops.

  4.3.6   Effluent vapors -- The effluents  of sample
          splitters  for   the  gas  chromatograph and
          roughing  pumps on  the  GC/MS  should pass
          through  either a   column  of   activated
          charcoal  or  be  bubbled  through  a trap
          containing oil  or high-boiling alcohols.

  4.3.7   Waste

4.3.7.1   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.

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 4.3.7.2    Disposal

4.3.7.2.1   The PCDDs  and PCDFs  decompose above  800
            C.   Lou-level  waste  such  as  absorbent
            paper,    tissues,   animal   remains,   and
            plastic   gloves  may   be  burned   in   an
            appropriate incinerator.  Gross quantities
            (milligrams) should  be  packaged  securely
            and   disposed   through   commercial   or
            governmental channels which are capable of
            handling  extremely  toxic wastes.

4.3.7.2.2   Liquid   or   soluble    waste   should   be
            dissolved  in  methanol  or   ethanol  and
            irradiated with  ultraviolet  light  with  a
            wavelength greater  than 290 nm for several
            days.   (Use F 40 BL  lamps  or equivalent.)
            Analyze   liquid wastes  and dispose  of  the
            solutions when the PCDDs and  PCDFs  can no
            longer be detected.

   4.3.8    Decontamination

 4.3.8.1    Personal  decontamination  --  Use  any mild
            soap with plenty  of  scrubbing  action.

 4.3.8.2    Glassware,   tools,     and    surfaces
            Chlorothene NU Solvent (Trademark  of  the
            Dow Chemical Company)  is the  least toxic
            solvent     shown     to    be    effective.
            Satisfactory cleaning  may  be  accomplished
            by rinsing with  Chlorothene,  then washing
            with  any   detergent   and   water.      If
            glassware is  first  rinsed with  solvent,
            then the  dish water may be disposed of in
            the sewer.  Given the cost of disposal, it
            is prudent to minimize solvent  wastes.

   4.3.9    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.  The clothing may be put  into a
            washer without  contact  if  the  launderer
            knows of  the potential  problem.     The
            washer should be   run  through  a  cycle
            before   being    used    again   for   other
            clothing.

  4.3.10    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 GC  can  achieve   a   limit  of
            detection of 0.1 ug  per wipe.   Less than
            0.1  ug   per   wipe   indicates   acceptable
            cleanliness;   anything  higher   warrants
            further   cleaning.   More than  10  ug on  a
            wipe  constitutes  an   acute   hazard  and
            requires  prompt  cleaning  before  further
            use of the equipment  or work  space,  and
            indicates that unacceptable work practices
            have been employed  in the past.

  4.3.11    Accidents --  Remove  contaminated clothing
            immediately,   taking  precautions  not  to
            contaminate skin or other  articles.   Wash
            exposed  skin  vigorously  and  repeatedly
            until  medical  attention is obtained.

       5    APPARATUS AND MATERIALS

     5.1    Sampling    equipment   for   discrete   or
            composite sampling.

   5.1.1    Sample bottles and  caps

 5.1.1.1    Liquid   samples   (waters,   sludges   and
            similar materials  that contain  less  than
            five  percent  solids)  --  Sample  bottle,
            amber   glass,   1.1   liters  minimum,   with
            screw  cap.

 5.1.1.2    Solid  samples (soils,  sediments,  sludges,
            paper   pulps,  filter  cake,  compost,  and
            similar materials  that contain  more  than
            five  percent  solids)  --  Sample  bottle,
            wide mouth,  amber glass,  500 mL  minimum.

 5.1.1.3    If   amber   bottles   are   not   available,
            samples shall  be  protected from  light.

 5.1.1.4    Bottle caps   --  Threaded  to  fit  sample
            bottles.   Caps shall  be lined with  Teflon.

 5.1.1.5    Cleaning

5.1.1.5.1    Bottles are  detergent  water  washed,  then
            solvent rinsed before use.

5.1.1.5.2   Liners are  detergent  water  washed,  then
            rinsed with reagent water  (Section 6.6.1)
            and   then    solvent,    and    baked    at
            approximately 200 C for  one hour  minimum
            prior  to  use.

   5.1.2    Compositing  equipment   --   Automatic   or
            manual compositing  system  incorporating
            glass   containers   cleaned   per   bottle
            cleaning  procedure  above.  Glass or Teflon
            tubing only shall be used.   If the sampler
            uses a peristaltic  pump,  a minimum length
            of  compressible silicone rubber  tubing may
            be  used in the pump only.  Before use,  the
            tubing shall  be thoroughly  rinsed  with
            methanol,   followed  by   repeated  rinsings
            with  reagent  water  to  minimize  sample
            contamination.  An  integrating  flow  meter
            is  used to collect  proportional composite
            samples.

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    5.2   Equipment for glassware cleaning

  5.2.1   Laboratory sink with overhead fume hood

    5.3   Equipment for sample preparation

  5.3.1   Laboratory fume hood of sufficient size to
          contain  the  sample  preparation equipment
          listed below

  5.3.2   Glove box (optional)

  5.3.3   Tissue  homogenizer  --  VirTis  Model  45
          Macro  homogenizer   (American  Scientific
          Products   H-3515,   or   equivalent)   with
          stainless  steel  Macro-shaft  and  Turbo-
          shear blade.

  5.3.4   Meat  grinder  --   Hobart,  or  equivalent,
          with 3 - 5 mm holes in inner plate.

  5.3.5   Equipment for determining percent moisture

5.3.5.1   Oven, capable of maintaining a temperature
          of 110 5 C.

5.3.5.2   Dessicator

  5.3.6   Balances

5.3.6.1   Analytical -- Capable of weighing 0.1 mg.

5.3.6.2  , Top  loading -- Capable of weighing 10 mg.

    5.4  ' Extraction apparatus

  5.4.1   Water samples

5.4.1.1   pH    meter,    with    combination   glass
          electrode.

5.4.1.2   pH paper,  wide range  (Hydrion Papers, or
          equivalent).

5.4.1.3   Graduated cylinder, 1 L capacity

5.4.1.4   1  L  filtration flasks with  side arm, for
          use  in vacuum filtration of water samples.

5.4.1.5   Separatory funnels  --  250,  500,  and 2000
          mL,  with Teflon stop cocks.

  5.4.2   Soxhlet/Dean-Stark     (SOS)     extractor
          (Figure 1)
        i
5.4.2.1 !  Soxhlet  --  50  mm  i.d.,  200  mL  capacity
          with  500  mL  flask  (Cal-Glass  LG-6900, or
          equivalent, except substitute 500 mL round
          bottom  flask   for   300   mL  flat  bottom
          flask).
       FIGURE 1  Soxhlet/Dean-Stark Extractor
5.4.2.2   Thimble  --  43 x 123  to  fit Soxhlet (Cal-
          Glass LG-6901-122,  or equivalent).

5.4.2.3   Moisture trap -- Dean Stark or Barret with
          Teflon stopcock, to fit Soxhlet.

5.4.2.4   Heating  mantle   --  Hemispherical,  to  fit
          500 mL  round bottom  flask  (Cal-Glass  LG-
          8801-112, or equivalent).

5.4.2.5   Variable  transformer   --   Powerstat  (or
          equivalent), 110 volt, 10 amp.

  5.4.3   Beakers, 400 - 500 mL

  5.4.4   Spatulas -- Stainless steel

    5.5   Filtration apparatus

  5.5.1   Pyrex glass  wool --  Solvent  extracted or
          baked at 450 C for four hours minimum.

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  5.5.2    Glass funnel  --  125 -  250 mL

  5.5.3    Glass fiber filter paper (Whatman GF/D,  or
           equivalent)

  5.5.A    Drying column --  15  to  20 mm  i.d.  Pyrex
           chromatographic    column   equipped   with
           coarse glass  frit  or glass wool  plug.

  5.5.5    Buchner funnel,  15 cm.

  5.5.6    Glass fiber filter paper for above.

  5.5.7    Pressure filtration apparatus -- Hillipore
           YT30 142 HW,  or  equivalent.

    5.6    Centrifuge apparatus

  5.6.1    Centrifuge -- Capable of  rotating  500  mL
           centrifuge bottles or   15  ml  centrifuge
           tubes at 5,000 rpm minimum

  5.6.2    Centrifuge bottles  -- 500 ml,  with  screw
           caps, to fit  centrifuge

  5.6.3    Centrifuge tubes  --  12-15 mL,  with  screw
           caps, to fit  centrifuge

    5.7    Cleanup apparatus

  5.7.1    Automated  gel  permeation  chromatograph
           (Analytical   Biochemical    Labs,    Inc,
           Columbia, MO, Model GPC  Autoprep 1002,  or
.           equivalent).

5.7.1.1    Column  --  600  -   700 mm  x  25 mm  i.d.,
           packed with  70  g  of  SX-3  Bio-beads  (Bio-
           Rad   Laboratories,   Richmond,    CA,    or
           equivalent).

5.7.1.2    Syringe, 10 mL,  with Luer fitting.

5.7.1.3    Syringe  filter  holder,  stainless  steel,
           and glass  fiber or Teflon filters (Gelman
           4310, or equivalent).

5.7.1.4    UV  detectors --   254-mu,  preparative  or
           semi-prep flow cell:  (Isco,  Inc.,  Type  6;
           Schmadzu, 5  mm  path  length; Beckman-Altex
           152W,  8 uL  micro-prep  flow cell,  2  mm
           path; Pharmacia UV-1, 3  mm flow cell;  LDC
           Milton-Roy   UV-3,   monitor   #1203;    or
           equivalent).

  5.7.2    Reverse  phase  high   performance  liquid
           chromatograph

5.7.2.1    Column  oven  and detector  --  Perkin-Elmer
           Model  LC-65T  (or   equivalent) operated  at
           0.02 AUFS at  235 nm.
5.7.2.2   Injector  --  Rheodyne 7120  (or  equivalent)
          with 50 uL sample  loop.

5.7.2.3   Column  --  Two 6.2  x  250  mm  Zorbax-ODS
          columns   in   series  (DuPont   Instruments
          Division, Wilmington,  DE, or  equivalent),
          operated at 50 C  with 2.0  mL/min methanol
          isocratic effluent.

5.7.2.4   Pump -- Altex 110A (or equivalent).

  5.7.3   Pipets

5.7.3.1   Disposable,  Pasteur,  150  mm x  5 mm  i.d.
          (Fisher     Sceintific     13-678-6A,     or
          equivalent).

5.7.3.2   Disposable,  serological,   10  mL   (6   mm
          i.d.).

  5.7.4   Chromatographic columns

5.7.4.1   150 mm  x 8 mm  i.d.,  (Kontes K-420155,  or
          equivalent)  with  coarse   glass   frit   or
          glass wool plug and 250 mL  reservoir.

5.7.4.2   200  mm x  15 mm  i.d.,  with  coarse  glass
          frit  or   glass   wool   plug  and  250   mL
          reservoir.

  5.7.5   Oven -- For storage of adsorbents,  capable
          of maintaining a temperature of 130  5 C.

    5.8   Concentration apparatus

  5.8.1   Rotary   evaporator   --    Buchi/Brinkman-
          American   Scientific   No.   E5045-10   or
          equivalent,   equipped  with   a   variable
          temperature water  bath.

5.8.1.1   A vacuum source is required for use  of  the
          rotary  evaporator.   It  must  be  equipped
          with  a shutoff  valve at  the  evaporator,
          and preferably, have a vacuum gauge.

5.8.1.2   A recirculating water pump and  chiller  are
          recommended,  as   use  of   tap   water  for
          cooling   the   evaporator   wastes   large
          volumes   of   water   and  can   lead   to
          inconsistent    performance    as    water
          temperatures and pressures  vary.

5.8.1.3   Round  bottom flask  --  500  mL  or  larger,
          with ground  glass fitting  compatible with
          the rotary evaporator.

  5.8.2   Nitrogen  blowdown  apparatus  --   Equipped
          with water  bath controlled  at  35 -  40  C
          (N-Evap,  Organomation  Associates,   Inc.,
          South    Berlin,    MA,    or    equivalent),
          installed  in a fume hood.

-------
 5.8.3
   5.9
Sample vials -- Amber glass, 2
Teflon-lined screw cap.
 5.9.1
 5.9.2
  5.10
5.10.1
5.10.2
  5.11
Gas chromatograph  --  Shall  have split less
or on-column  injection  port for capillary
column,    temperature     program    with
isothermal hold, and shall meet all of the
performance specifications in Section 14.

GC  Column for  PCDDs  and  PCDFs   and  for
isomer specificity for 2,3,7,8-TCDO --  60
5  m x  0.32 0.02  mm  i.d.;  0.25  urn  5X
phenyl,  94X  methyl,  1X  vinyl   silicone
bonded phase fused silica capillary column
(J & W OB-5,  or equivalent).

GC  Column   for  isomer  specificity  for
2,3,7,8-TCDF  --  30 5  m x  0.32  tO.02  mm
i.d.;  0.25  urn  bonded phase  fused silica
capillary  column   (J  &   W  DB-225,   or
equivalent).

Mass spectrometer  --  28  -  40 eV  electron
impact   ionization,   shall   repetitively
selectively monitor 11 exact m/z's minimum
at  high  resolution  (>10,000)  during  a
period of approximately 1 second.

The  groups  of  m/z's  to  be  monitored are
shown   in  Table  3.     Each   group  or
descriptor    shall    be    monitored    in
succession as  a function of GC retention
time to ensure  that  all PCODs  and PCOFs
are  detected.    The  theoretical abundance
ratios  for  the  m/z's are  given  in Table
3A, along with  the control  limits of each
ratio.

The mass spectrometer shall be operated  in
a   mass  drift  correction  mode,  using
perfluorokerosene  (PFK)  to  provide  lock
masses.   The lock mass  for each  group of
m/z's is shown  in Table 3.  Each lock mass
shall be  monitored and shall  not  vary by
more  than   10  percent  throughout  its
respective    retention    time    window.
Variations of  the  lock mass  by more than
10   percent   indicate   the  presence  of
coeluting     interferences     that    may
significantly  reduce  the  sensitivity  of
the  mass spectrometer.    Re-injection  of
another aliquot of the sample extract will
not  resolve   the   problem.    Additional
cleanup of the  extract  may be required  to
remove the interferences.

GC/MS  interface --  The  mass spectrometer
shall  be interfaced  to  the  GC  such that
the end of the  capillary  column terminates
within 1 cm of  the ion source but  does not
5 ml with                intercept the electron or  ion beams.   All
                         portions of  the  column which  connect the
                         GC to  the  ion source  shall   remain at  or
                         above   the   column   temperature   during
                         analysis to preclude condensation  of  less
                         volatile compounds.

                  5.12    Data system  --  Shall  collect  and record
                         and store MS data.

               5.12.1    Data acquisition  --  The  signal   at  each
                         exact m/z shall be  collected repetitively
                         throughout    the   monitoring   period   and
                         stored  on a  mass  storage device.

               5.12.2    Response     factors     and     multipoint
                         calibrations -- The  data  system  shall  be
                         used  to  record   and  maintain  lists  of
                         response  factors  (response  ratios   for
                         isotope    dilution)     and    multi-point
                         calibration   curves.       Computations  of
                         relative standard  deviation (coefficient
                         of variation) are used to test calibration
                         linearity.   Statistics on initial  (Section
                         8.2)    and    ongoing    (Section     U.5)
                         performance  '  shall   be    computed   and
                         maintained.

                     6    REAGENTS AND STANDARDS

                   6.1    pH adjustment and back extraction

                 6.1.1    Potassium  hydroxide  --   Dissolve  20  g
                         reagent grade KOH  in 100 ml reagent water.

                 6.1.2    SuIfuric acid  --  Reagent  grade (specific
                         gravity 1.84).

                 6.1.3    Sodium chloride -- Reagent grade,  prepare
                         a five  percent  (w/v) solution  in reagent
                         water.

                   6.2    Solution drying and evaporation
                 6.2.1    Solution drying -- Sodium sulfate, reagent
                         grade, granular anhydrous  (Baker 3375, or
                         equivalent),    rinsed   with    methylene
                         chloride  (20  ml/g),   baked  at 400  C for
                         one hour minimum,  cooled in a dessicator,
                         and stored  in a  pre-cleaned  glass bottle
                         with screw cap that prevents moisture  from
                         entering.

                 6.2.2    Prepurified nitrogen

                   6.3    Solvents -- Acetone,   toluene, cyclohexane,
                         hexane,   nonane,    methanol,    methylene
                         chloride, and  nonane: distiI led-in-glass,
                         pesticide  quality,  lot  certified to be
                         free of interferences.

-------
    6.4   GPC   calibration  solution   --  Solution
          containing  300 mg/mL  corn oil,  15 mg/mL
          bis(2-ethylhexyl)  phthalate,   1.4  mg/mL
          pentachlorophenol, 0.1 mg/mL perylene, and
          0.5 mg/mL sulfur

    6.5   Adsorbents  for sample cleanup

  6.5.1   Silica gel

6.5.1.1   Activated silica  gel --  Bio-Sil  A, 100  -
                                                          6.5.3.1    Activated   carbon   --
                                                                    Development   Company,
                                                                    equivalent).   Prewash
                                                                    dry in vacuo at 110 C.
                                  AX-21    (Anderson
                                 Adrian,   MI,   or
                                with  met Hanoi  and
                                      131-1340,    or
                                    with   methylene
                                    C  for  one hour
                                    dessicator,  and
          200    mesh     (Bio-Rad
          equivalent),    rinsed
          chloride,  baked  at  250
          minimum,   cooled   in  a
          stored  in  a  pre-cleaned glass bottle with
          screw  cap  that  prevents  moisture  from
          entering.

6.5.1.2   Acid  silica   gel  (30  percent  w/w)  --
          Thoroughly  mix  4.4  g  of  concentrated
          sulfuric acid with 10.0 g activated silica
          gel.  Break  up aggregates with a stirring
          rod  until  a uniform mixture  is obtained.
          Store   in   a   screw-capped   bottle  with
          Teflon-lined cap.

6.5.1.3   Basic silica gel  --  Thoroughly mix 30 g of
          1N   sodium  hydroxide   with   100   g  of
          activated silica  gel.  Break up aggregates
          with  a  stirring  rod  until  a  uniform
          mixture  is obtained.   Store  in  a screw-
          capped bottle with Teflon-lined cap.

  6.5.2   Alumina

6.5.2.1   Neutral  alumina  --   Bio-Rad  Laboratories
          132-1140   Neutral   Alumina   Ag    7   (or
          equivalent).   Heat  to 600 C for 24 hours
          minimum.   Store  at   130  C in  a covered
          flask.   Use  within five days of baking at
          600  "C.

6.5.2.2   Acid  alumina  -- Bio-Rad Laboratories 132-
          1340  Acid  Alumina  AG 4  (or equivalent).
          Activate by heating  to 130 C for 12 hours
          minimum.

6.5.2.3   Basic alumina  --  Bio-Rad  Laboratories 132-
          1240  Basic Alumina AG 10 (or equivalent).
          Activate by heating  to 600 C for 24 hours
          minimum.     Alternatively,   activate  by
          heating  alumina in a tube furnace at 650  -
          700  C  under an  air  flow of approximately
          400  cc/min.  To avoid melting the alumina,
          do not  heat  over  700 C.   Store at 130 C
          in a covered flask.   Use within five days
          of baking.

  6.5.3   AX-21/Celite
                                                          6.5.3.2
                                                          6.5.3.3
        Celite   545
        equivalent).
(Supelco   2-0199,   or
        Thoroughly  mix  5.35  g  AX-21  and  62.0  g
        Celite 545  to produce a 7.9%  w/w mixture.
        Activate  the  mixture at  130  C  for  six
        hours minimum.   Store in a dessicator.
6.5.4   White  quartz  sand,  60/70   mesh   --   For
        Soxhlet/Dean-Stark   extraction,   (Aldrich
        Chemical  Co,  Milwaukee    UI     Cat   No.
        27,437-9, or  equivalent).   Bake at  450 "C
        for four hours minimum.

  6.6   Reference matrices

6.6.1   Reagent water  --  Water in which the PCDDs
        and  PCDFs  and interfering  compounds  are
        not detected by this method.

6.6.2   High solids reference matrix  --  Playground
        sand  or  similar  material  in  which   the
        PCDDs and  PCDFs  and interfering compounds
        are not  detected by this  method.    May be
        prepared  by   extraction  with  methylene
        chloride and/or  baking at 450 C  for  four
        hours minimum.

6.6.3   Filter   paper  --   Gel man   type   A   (or
        equivalent)  glass  fiber   filter  paper  in
        which the PCDDs  and PCDFs and  interfering
        compounds are  not detected by this method.
        Cut the paper  to simulate  the surface  area
        of the paper sample being  tested.

6.6.4   Other  matrices  --  This  method   may  be
        verified  on any matrix by performing  the
        tests given in Section 8.2.   Ideally,  the
        matrix  should be  free  of the  PCDDs  and
        PCDFs, but  in  no case  shall the background
        level  of   the PCDDs   and PCDFs   in   the
        reference  matrix  exceed   three  times  the
        minimum  levels given  in  Table 2.    If  low
        background  levels  of  the  PCDDs and PCDFs
        are present in the  reference matrix,  the
        spike   level   of   the  analytes   used   in
        Section 8.2 should be  increased to provide
        a  spike-to-background  ratio  in the range
        of 1/1 to 5/1  (Reference  15).

  6.7   Standard    solutions    --   Purchased    as
        solutions  or  mixtures  with  certification
        to   their   purity,   concentration,   and
        authenticity,  or  prepared from materials
        of  known   purity  and  composition.    If
8

-------
        compound purity  is 98 percent or greater,
        the weight  may be used without  correction
        to   compute   the   concentration  of   the
        standard.   When  not being used, standards
        are stored  in  the  dark at  room  temperature
        in  screw-capped  vials  with Teflon-lined
        caps.  A mark  is placed on the  vial  at  the
        level  of  the  solution  so   that  solvent
        evaporation  loss  can  be detected.    If
        solvent  loss  has  occurred,   the solution
        should be replaced.

  6.8   Stock solutions

6.8.1   Preparation  -- Prepare in nonane  per  the
        steps   below   or   purchase   as    dilute
        solutions (Cambridge  Isotope Laboratories,
        Cambridge,  MA,  or equivalent).   Observe
        the  safety  precautions in Section  4,  and
        the recommendation in Section 4.1.2.
6.8.2
6.8.3
  6.9
 6.10
 6.11
Dissolve an appropriate  amount  of assayed
reference  material   in  solvent.     For
example, weigh 1 - 2 mg of 2,3,7,8-TCDD to
three  significant  figures  in  a  10  ml
ground  glass   stoppered  volumetric  flask
and fill  to  the mark with  nonane.   After
the TCDO is completely dissolved, transfer
the solution  to a  clean 15 ml  vial  with
Teflon-lined cap.

Stock standard solutions should be checked
for  signs  of  degradation  prior  to  the
preparation of  calibration  or performance
test standards.   Reference  standards  that
can be  used  to determine the  accuracy of
calibration standards  are  available  from
Cambridge Isotope Laboratories.

Secondary   standard    --    Using   stock
solutions (Section 6.8), prepare secondary
standard    solutions    containing    the
compounds  and   concentrations   shown  in
Table 4 in nonane.

Labeled compound  spiking  standard --  From
stock   standard  solutions   prepared  as
above, or from purchased mixtures, prepare
this  standard   to   contain  the  labeled
compounds at  the concentrations  shown in
Table  4  in  nonane.     This  solution  is
diluted with acetone prior to use (Section
10.3.2).

Cleanup standard  -  Prepare    Cl,-2,3,7,8-
TCDD at the concentration shown in Table 4
in nonane.
 6.12    Internal   standard  --   Prepare   at   the
         concentration shown in Table 4 in nonane.
  6.13   Calibration standards (CS1 through CSS)  --
         Combine  the  solutions  in  Sections  6.9,
         6.10, 6.11,  and 6.12 to  produce the  five
         calibration solutions shown  in Table  4  in
         nonane.     These   solutions  permit   the
         relative  response  (labeled  to  unlabeled)
         and  response  factor  to be  measured as  a
         function   of   concentration.     The   CS3
         standard    is   used    for   calibration
         verification  (VER).

  6.14   Precision  and recovery standard (PAR)  --
         Used for determination  of  initial  (Section
         8.2) and  ongoing (Section 14.5)  precision
         and recovery.   This solution contains  the
         analytes  and   labeled  compounds  at   the
         concentrations   listed   in   Table  4   in
         nonane.    This   solution   is  diluted  with
         acetone prior to use (Section 10.3.4).

  6.15   GC   retention    time   window    defining
         solutions  --  Used to define the  beginning
         and ending  retention times for  the  dioxin
         and furan  isomers.

6.15.1   DB-5  column  window defining  standard  --
         Cambridge  Isotope Laboratories  ED-1732-A,
         or  equivalent,   containing  the   compounds
         listed in Table 5.

  6.16   Isomer specificity  test standards --  Used
         to demonstrate  isomer specificity for  the
         2,3,7,8-tetra-   isomers   of   dioxin   and
         furan.

6.16.1   Standards for the DB-5  column  --  Cambridge
         Isotope Laboratories ED-908, ED-908-C,  or
         ED-935,   or   equivalent,   containing   the
         compounds  listed in Table  5.

6.16.2   Standards   for    the  DB-225   column
         Cambridge  Isotope  Laboratories  EF-937  or
         EF-938,   or   equivalent,   containing   the
         compounds  listed in Table  5.

  6.17   Stability   of   solutions   --    Standard
         solutions  used   for  quantitative  purposes
         (Sections  6.9  -  6.14)  shall  be  analyzed
         within 48 hours  of preparation  and on  a
         monthly  basis   thereafter  for   signs  of
         degradation.      Standards   will   remain
         acceptable   if   the  peak  area  at   the
         quantitation   m/z   remains    within   15
         percent  of   the  area   obtained  in   the
         initial  analysis  of the  standard.     Any
         standards  failing to meet this  criterion
         should   be   assayed   against    reference
         standards,  as  in  Section  6.8.3.,  before
         further use.

-------
     7    CALIBRATION

   7.1    Assemble  the   GCMS   and   establish   the
          operating conditions  necessary to meet the
          relative retention time  specifications  in
          Table 2.

  7.1.1    The following GC  operating  conditions may
          be  used  for  guidance  and  adjusted  as
          needed to meet the relative retention time
          specifications in Table 2:
          Injector temp:  270 C
          Interface temp:   290  "C
          Initial temp and time: 200  C,  2 min
          Temp Program:  200-220 "C at 5  "C/min
                         220 C for 16 min
                         220-235 C at 5  "C/min
                         235 "C for 7 min
                         235-330 "C at 5  "C/min

  7.1.2    Obtain a  selected ion current  profile  of
          each  analyte  in  Table  4  at  the  exact
          masses specified in Table 3 and at >10,000
          resolving power by injecting  an authentic
          standard  of   the  PCDDs  and PCDFs  either
          singly or  as part of a mixture  in which
          there  is  no  interference  between closely
          eluted components, using the  procedure  in
          Section 13.

   7.2    The ion abundance  ratios,  minimum levels,
          and absolute retention times -- Inject the
          CS1 calibration solution (Table 4) per the
          procedure in Section  13 and the conditions
          in Table 2.

  7.2.1    Measure the  selected  ion  current profile
          (SICP) areas  for  each analyte  and compute
          the  ion  abundance  ratios  specified  in
          Table  3.   Compare the computed  ratio  to
          the theoretical  ratio given in Table 3.

  7.2.2    All PCDDs and PCDFs  shall  be within their
          respective  ratios;  otherwise,  the  mass
          spectrometer  shall  be  adjusted  and  this
          test  repeated until   the m/z  ratios  fall
          within  the  limits  specified.    If  the
          adjustment alters  the  resolution  of  the
          mass  spectrometer,  resolution  shall  be
          verified (Section 7.1) prior  to repeat  of
          the test.

  7.2.3    Verify that the HRGC/HRMS instrument meets
          the minimum  levels in Table 2; otherwise,
          the  mass  spectrometer  shall   be  adjusted
          and this  test repeated until  the minimum
          levels in Table 2 are met.

  7.2.4    The retention times  of   C12-1,2,3,4-TCDD
          and    C12-1,2,3,7,8,9-HxCDF  (the internal
        standards,  Section 6.12)  shall  exceed  27
        and 38  minutes,  respectively,  on the  DB-5
        column,  and the retention  time of    C._-
        1,2,3,4-TCDD  shall exceed  17  minutes  on
        the  DB-225   column;   otherwise,  the   GC
        temperature program  shall  be adjusted  and
        this  test   repeated   until  the  minimum
        retention time criteria are met.

  7.3   Retention   time  windows  --  Analyze   the
        window  defining  mixtures   (Section  6.15)
        using the procedure in Section  13 (Figures
        2A - 20).

  7.4   Isomer specificity

7.4.1   Analyze   the   isomer   specificity    test
        standards   (Section   6.16)    using    the
       . procedure in Section  13.

7.4.2   Compute  the percent  valley between the  GC
        peaks  that  elute  most  closely  to   the
        2,3,7,8-  TCDD  and TCDF  isomers, on their
        respective columns, per Figure 3.

7.4.3   Verify   that  the  height   of   the  valley
        between  the most  closely  eluted isomers
        and the  2,3,7,8-  isomers  is  less than  25
        percent  (computed as  100 x/y in  Figure 3).
        If  the  valley exceeds  25  percent,  adjust
        the analytical  conditions  and   repeat  the
        test   or   replace   the  GC   column   and
        recalibrate (Section  7.2 through 7.4).

  7.5   Calibration   with   isotope  dilution
        Isotope  dilution is  used  when   1) labeled
        compounds are  available,  2) interferences
        do  not  preclude its  use,  and  3) the  SICP
        area  for  the analyte  at  the  exact   m/z
        (Table 3)  is  in the  calibration range  for
        the analyte.   The  reference compound  for
        each native and  labeled compound is shown
        in  Table 6.   Alternate labeled compounds
        and quant i tat ion m/z's may be   used based
        on  availability.    If  any  of   the  above
        conditions  preclude   isotope dilution,  the
        internal  standard  method (Section 7.6)  is
        used.

7.5.1   A   calibration  curve   encompassing   the
        concentration  range   is  prepared for  each
        compound to be  determined.   The relative
        response   (native    to    labeled)     vs
        concentration  in  standard solutions  is
        plotted   or   computed   using   a   linear
        regression.    Relative  response  (RR)  is
        determined  according  to   the   procedures
        described below.   A  minimum of five  data
        points are  employed for calibration.
10

-------
               6-MAY-88    Sir: Voltage 705    Sys: DB5US
    Sample 1 Injection 1     Group 2    Mass 303.9016
100
 80
 60
 40
 20
            1,3,6,8-TCDF
                                                              Norm:    3044
                                             1,2,8,9-TCDF
   25:20  26:40  28:00  29:20  30:40  32:00  33:20  34:40  36:00  37:20   38:40
              6-MAY-88    Sir: Voltage 705    Sys: DB5US
   Sample 1 Injection 1    Group 2    Mass 319.8965
100
 80
 60
 40
 20
          1,3,6,8-TCDD
                                                               Norm:    481
                                             1,2,8.9-TCDD
   25:20 26:40  28:00  29:20   30:40- 32:00/33:20  34:40  36:00  37:20   38:40
         FIGURE 2A  First and Last Eluted Tetra- Dioxin and Furan Isomers
                                                                                 11

-------
                     6-MAY-88    Sir: Voltage 705    Sys: DB5US
          Sample 1 Injection 1     Group 2    Mass 339.8597
         100
         80
         60-
         40
         20-
                                                  Norm:    652
                  1,3,4,6,8-PeCDF
1,2,3,8,9-PeCDF
           29:20  30:40  32:00  33:20  34:40  36:00   37:20  38:40
                      6-MAY-88    Sir: Voltage 705    Sys: DB5US
           Sample 1 Injection 1    Group 2    Mass 355.8546
        100
         80
         60
         40
         20
                        1,2,4,7,9-PeCDD
                                                  Norm:    503
                                                     1,2,3,8,9-PeCDD
 \
           29:20  30:40   32:00  33:20  34:40   36:00  37:20  38:40
       FIGURE 2B  First and Last Eluted Penta- Dioxin and Furan Isomers
12

-------
                 6-MAY-88    Sir: Voltage 705    Sys: DB5US
      Sample 1 Injection 1     Group 3    Mass 373.8208
 100
 80-
 60'
 40-
 20-
                                                               Norm:    560
           1,2,3,4,6,8-HxCDF
                                                      1,2,3,4,8,9-HxCDF
     39:30 40:00 40:30  41:00 41:30  42:00 42:30  43:00  43:30  44:00  44:30
               6-MAY-88    Sir: Voltage 705    Sys: DB5US
    Sample 1 Injection 1    Group 3    Mass 389.8156
100
 00
 60
 40
 20
                   1,2,4,6,7,9/1,2,4,6,8,9-HxCDD
                                                               Norm:    384
                                                   1,2,3,4,6,7-HxCDD
    39:30  40:00 40:30  41:00  41:30  42:00  42:30  43:00  43:30  44:00  44:30
          FIGURE 2C  First and Last Eluted Hexa- Dioxin and Furan Isomers
                                                                                 13

-------
                   6-MAY-88    Sir: Voltage 705   Sys: DB5US
        Sample 1  Injection 1    Group 4    Mass 407.7818
       100
        80
        60'
        40'
        20
                   1,2,3,4,6,7,8-HpCDF
        \s
                                                                      Norm:    336
                         1,2,3,4,7,8,9-HpCDF
           45:20   46:40  48:00   49:20   50:40  52:00   53:20   54:40   56:00   57:20

                   6-MAY-88    Sir: Voltage 705    Sys: DB5US
        Sample 1 Injection 1    Group 4    Mass 423.7766
                    1,2,3,4,6,7,9-HpCDD
lOOn
 80
 60
 40
 20
  0
                                                                      Norm:    282
                            1.2,3.4.6.7.8-HpCDD
           45:20   46:40  48:00   49:20   50:40   52:00   53:20   54:40   56:00   57:20

                   6-MAY-88   Sir: Voltage 705    Sys: DB5US
        Sample 1 Injection 1     Group 4   Mass 441.7428
100

 80

 60

 40

 20-I

  0
                                                         OCDF
                                                                      Norm:    13
          45:20   46:40   48:00  49:20   50:40   52:00   53:20   54:40   56:00   57:20
 Sample 1

100

 80

 60

 40

 20

  0
                   6-MAY-88    Sir: Voltage 705    Sys: DB5US
                 Injection 1    Group 4    Mass 457.7377
                                                        OCDD
                                                                      Norm:
          45:20   46:40   48:00   49:20   50:40   52:00   53.20   54:40   56:00   57:20


                FIGURE 2D  First and Last Eluted Hepta- Dioxin and Furan Isomers
14

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            3A   DB225 Column
                       21-APR-88    Sir: Voltage 705    Sys: DB225
            Sample 1 Injection 1    Group 1    Mass 305.8987
            Text: COLUMN PERFORMANCE
           100
           80
           60
           40
           20
                                             2.3,7,8-TCDF
                                 2,3,4,7-TCDF
Norm:    3466
                                                          1,2,3,9-TCDF
              16:1016:20^6:30 16:40 16:50 17:00 17:10 17:20 17:30 17:40 17:50 18:00
          3B    DBS Column
FIGURE 3  Valley between 2,3,7,8- Tetra Dioxin and Furan Isomers and Other Closely Eluted Isomers
                                                                                   15

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  7.5.2    The relative response  of  a  PCDO  or  PCDF  to
          its  labeled  analog  is  determined   from
          isotope   ratio '  values    computed   from
          acquired data.   Three isotope  ratios are
          used in this process:

          Rx = the  isotope  ratio  measured  for the
               pure pollutant.

          Ry = the  isotope  ratio  measured  for the
               labeled compound.

          Rm = the  isotope  ratio,  of  an  analytical
               mixture  of  pollutant  and   labeled
               compounds .

          The m/z's are selected such that Rx  > Ry.
          If Rm  is not between  2Ry  and  O.SRx, the
          method does  not apply and  the sample  is
          analyzed by the internal  standard method.
                                                         7.5.4
7.5.3
          When there  is  no  overlap  between the  GC
          peaks or the quant i tat ion m/z's,  as  occurs
          with nearly all  of the PCODs  and  PCDFs  and
          their respective  labeled analogs, the  RR
          is calculated per the  following:
          Rx  =   [area m1/z]
                       i

                  at  the  retention  time  of
                  native compound.
                                               the
          Ry  =   	1
                  [area m2/z]

                  at  the  retention  time  of   the
                  labeled compound (RT2).

          Rm  =   [area at m1/z (at RT2)]
                  [area at m2/z (at RT1)]
                  as measured in the mixture of  the
                  native   and   labeled   compounds
                  (Figure 4) (RT1).
                                AREA AT
                                  M,(Z
    FIGURE 4   Selected Ion Current Profiles for
    Chromatographically Resolved Labeled (m.2/z)
    and Unlabeled (m-j/z)  Pairs.
                                                         7.5.5
                                                             7.6
                                                           7.6.1
                                                         7.6.1.1
                                                       7.6.1.2
To  calibrate  the  analytical  system  by
isotope dilution, inject  a  1.0 uL aliquot
of calibration  standards CS1  through CSS
(Section  6.13  and  Table  4)  using  the
procedure in Section 13 and the conditions
in  Table  2.     Compute   the   RR  at  each
concentration.

Linearity  --   If the  ratio   of  relative
response to concentration for any compound
is   constant    (less   than    20   percent
coefficient of variation) over the 5-point
calibration  range,  an  averaged  relative
response/concentration ratio  may be  used
for that compound; otherwise,  the complete
calibration curve for  that  compound shall
be  used   over   the  5-point   calibration
range.

Calibration by   internal  standard  --  The
internal  standard  method  is  applied  to
determination   of   compounds  having  no
labeled  analog,  and  to  measurement  of
labeled  compounds   for   intra- laboratory
statistics (Sections 8.4  and 14.5.4).

Response factors -- Calibration  requires
the determination of response factors (RF)
defined by the following  equation:
                                                                 RF
        (A
                                                                                Cis>
                                                                           (Ais x V
                                                                 where,
                                                                 A   is the area  of the  exact  m/z for  the
                                                                 compound  in the calibration standard.
                                                                 A.   is  the area of  the  exact  m/z for  the
                                                                 internal  standard.
                                                                 Cis
     is  the  concentration  of  the  GCHS
internal standard  (Section  6.12  and Table
4) in pg/uL.
C  is the concentration of the compound in
the calibration standard in pg/uL.

The  response  factor  is determined  for at
least  five  concentrations  appropriate to
the  response  of   each  compound  (Section
6.13); nominally,   0.5,  2,  10,  40,  and 200
ng/mL.   The  amount  of  internal  standard
added  to each calibration  solution  and
extract  is  the same  (100 ng/mL)  so that
Cis  remains  constant.   The RF  is  plotted
vs concentration for  each  compound in the
standard  (Cs)  to  produce  a  calibration
curve.

Linearity --  If  the  response  factor (RF)
for any compound is constant (less  than 35
16

-------
        percent coefficient of variation) over the
        5-point  calibration  range,  an  averaged
        response  factor  may  be  used  for  that
        compound;    otherwise,    the    complete
        calibration curve  for  that compound shall
        be used over the 5-point range.

  7.7   Combined    calibration    --    By    using
        calibration  solutions  (Section 6.13  and
        Table  4)  containing  the  unlabeled  and
        labeled   compounds,   and  the  internal
        standards, a single set of analyses can be
        used to produce calibration curves for the
        isotope  dilution  and  internal  standard
        methods.   These curves are  verified each
        shift  (Section  U.3)  by  analyzing  the
        calibration  verification  standard  (VER,
        Table  4).    Recalibration is  required  if
        calibration verification criteria (Section
        14.3.4) cannot be met.

    8   QUALITY ASSURANCE/QUALITY CONTROL

  8.1   Each  laboratory  that uses  this method  is
        required  to   operate   a   formal  quality
        assurance  program  (Reference  16).    The
        minimum   requirements   of   this  program
        consist  of  an  initial   demonstration  of
        laboratory capability, analysis of samples
        spiked with  labeled compounds  to evaluate
        and document data quality, and  analysis of
        standards and  blanks as tests  of continued
        performance.    Laboratory  performance  is
        compared    to    established    performance
        criteria  to determine  if the  results  of
        analyses   meet  the   performance   charac
       1 teristics of the method.   If the method is
        to   be  applied   routinely   to  samples
        containing  high solids  with  very   little
        moisture   (e.g.,   soils,  filter   cake,
        compost)  or to  an alternate  matrix,  the
        high   solids   reference  matrix  (Section
        6.6.2)  or  the alternate  matrix (Section
        6.6.4)   is  substituted   for   the  reagent
        water   matrix   (Section   6.6.1)   in  all
        performance tests.

8.1.1   The   analyst   shall   make   an  initial
        demonstration  of  the  ability  to generate
        acceptable  accuracy   and precision  with
        this  method.   This ability is  established
        as described in Section 8.2.

8.1.2   The  analyst is  permitted  to  modify this
        method to  improve  separations  or lower the
        costs   of   measurements,   provided  all
        performance  specifications are met.   Each
        time  a modification is made to the method,
        the  analyst   is  required  to  repeat  the
        procedures  in  Sections 7.2 through 7.4 and
        Section    8.2
        performance.
to   demonstrate   method
8.1.3   Analyses   of   blanks   are   required   to
        demonstrate   freedom  from   contamination
        (Section   3.2).     The   procedures   and
        criteria   for   analysis   of   a   blank  are
        described  in Section 8.5.

8.1.4   The  laboratory  shall  spike  all   samples
        with  labeled compounds  to monitor  method
        performance.    This  test  is  described  in
        Section 8.3.  When results of these spikes
        indicate  atypical method performance  for
        samples,  the  samples are  diluted  to bring
        method   performance   within   acceptable
        limits.    Procedures^ for dilutions  are
        given in Section  16.4.

8.1.5   The  laboratory  shall,  on  an ongoing basis,
        demonstrate       through       calibration
        verification  and  the  analysis   of   the
        precision  and  recovery  standard  that  the
        analytical  system is  in  control.   These
        procedures are  described  in  Sections  14.1
        through 14.5.

8.1.6   The  laboratory shall  maintain  records  to
        define   the   quality  of  data   that   is
        generated.      Development   of   accuracy
        statements is described  in Section 8.4.

  8.2   Initial   precision  and   accuracy  --   To
        establish    the   ability   to   generate
        acceptable   precision  and  accuracy,   the
        analyst   shall   perform   the   following
        operations.

8.2.1   For  low solids  (aqueous  samples),  extract,
        concentrate,   and  analyze   four   1-liter
        aliquots-of  reagent  water spiked  with  the
        diluted  precision  and  recovery  standard
        (PAR)  (Sections 6.14 and 10.3.4) according
        to the procedures  in  Sections  10 through
        13.   For an alternate sample matrix,  four
        aliquots  of  the alternate matrix are used.
        All   sample   processing   steps,   including
        preparation    (Section   10),   extraction
        (Section   11),  and  cleanup  (Section  12)
        that are  to  be  used  for  processing samples
        shall be  included in this test.

8.2.2   Using results of  the set of  four  analyses,
        compute  the  average recovery (X)  in ng/mL
        and the  standard  deviation of the recovery
        (s)  in ng/mL for  each  compound, by  isotope
        dilution   for  PCDDs  and PCDFs   with  a
         labeled  analog,  and  by  internal  standard
        for labeled  compounds and PCDDs and PCDFs
        with no  labeled analog.
                                                                                                          17

-------
 8.2.3    For  each compound,  compare s  and X with
          the   corresponding   limits  for   initial
          precision  and accuracy in  Table  7.   If s
          and   X   for   all   compounds   meet   the
          acceptance  criteria,  system performance is
          acceptable   and  analysis   of   blanks   and
          samples  may  begin.     If, however,   any
          individual  s  exceeds  the   precision  limit
          or  any  individual  X  falls  outside  the
          range  for  accuracy, system  performance  is
          unacceptable for that  compound.    Correct
          the  problem and  repeat  the test  (Section
          8.2).

   8.3    The  laboratory shall  spike  all  samples  and
          QC   aliquots  with  the  diluted   labeled
          compound spiking standard   (Sections 6.10
          and  10.3.2)  to assess method  performance
          on the sample  matrix.

 8.3.1    Analyze  each  sample  according  to   the
          procedures  in  Sections 10 through  13.

 8.3.2    Compute  the  percent  recovery  (P) of  the
          labeled  compounds in the labeled  compound
          spiking  standard and the cleanup  standard
          using    the   internal   standard    method
          (Section 7.6).

 8.3.3    Compare  the labeled compound  recovery  for
          each   compound   with  the  corresponding
          limits in  Table 7.   If  the  recovery of  any
          compound falls  outside  its limit,  method
          performance   is   unacceptable  for  that
          compound in that sample.  To overcome such
          difficulties,   water  samples  are  diluted
          and  smaller  amounts  of  soils,   sludges,
          sediments    and    other    matrices    are
          reanalyzed per Section 17.

   8.4    Method  accuracy  for  samples  shall   be
          assessed and records  shall  be  maintained.

 8.4.1    After  the  analysis of five samples of a
          given  matrix  type  (water, soil,  sludge,
          pulp,  etc)  for which the  labeled  compound
          spiking   standards   pass   the  tests   in
          Section  8.3,  compute the  average percent
          recovery (P) and the standard  deviation of
          the  percent recovery (sp)  for  the labeled
          compounds   only.    Express  the  accuracy
          assessment as  a  percent recovery  interval
          from P  - 2sp to P + 2sp for  each matrix.
          For  example,  if  P  = 90% and sp = 10X  for
          five  analyses   of  pulp,  the   accuracy
          interval is expressed as 70 -  110%.

 8.4.2    Update  the accuracy  assessment   for  each
          compound in each matrix on a regular basis
          (e.g.,  after   each  5-10 new  accuracy
        measurements).

  8.5   Blanks   --  Reference   matrix  blanks  are
        analyzed   to  demonstrate    freedom   from
        contamination (Section  3.2).

8.5.1   Extract  and  concentrate a  1-liter  reagent
        water  blank  (Section  6.6.1), high  solids
        reference  matrix  blank  (Section  6.6.2),
        paper  matrix  blank   (Section  6.6.3)   or
        alternate  reference matrix blank  (Section
        6.6.4)   with   each  sample   set   (samples
        started  through  the extraction process  on
        the same 12-hour shift,  to  a  maximum of  20
        samples).   Analyze  the blank  immediately
        after   analysis   of   the  precision   and
        recovery   standard   (Section   14.5)   to
        demonstrate freedom from contamination.

8.5.2   If any of  the  PCODs or  PCOFs  (Table  1)  or
        any  potentially  interfering  compound  is
        found in blank at greater than the  minimum
        level  (Table   2),   assuming  a   response
        factor of  1 relative to the   c.2-1,2,3,4-
        TCOD  internal  standard for  compounds  not
        listed in  Table  1,  analysis  of  samples  is
        halted  until  the  source of  contamination
        is   eliminated   and   a  blank  shows   no
        evidence of contamination at  this  level.

  8.6   The   specifications   contained   in   this
        method can be met  if the apparatus  used is
        calibrated properly and then  maintained in
        a  calibrated state.    The  standards  used
        for  calibration (Section 7),  calibration
        verification   (Section   14.3),   and   for
        initial  (Section 8.2)  and ongoing  (Section
        14.5)  precision  and  recovery  should  be
        identical,   so  that   the    most   precise
        results   will   be   obtained.     A   GCHS
        instrument   will    provide    the    most
        reproducible  results  if dedicated  to  the
        settings  and conditions required  for  the
        analyses   of   PCODs  and   PCOFs  by  this
        method.

  8.7   Depending      on     specific      program
        requirements,   field   replicates   may   be
        collected  to  determine  the  precision  of
        the  sampling technique, and spiked samples
        may  be required to determine the  accuracy
        of the analysis  when the internal  standard
        method  is  used.

    9   SAMPLE    COLLECTION,    PRESERVATION,   ANO
        HANDLING

  9.1   Collect    samples   in   glass   containers
        following  conventional  sampling  practices
        (Reference  17).    Aqueous   samples  which
18

-------
         flow freely are collected  in  refrigerated
         bottles     using    automatic     sampling
         equipment.   Solid  samples are  collected  as
         grab samples using wide  mouth  jars.

  9.2    Maintain samples at 0  -  4 C from the time
         of   collection  until   extraction.      If
         residual chlorine  is  present  in aqueous
         samples, add 80 mg  sodium  thiosulfate per
         liter of  water.    EPA  Methods  330.4 and
         330.5 may   be  used  to  measure  residual
         chlorine (Reference 18).

  9.3    Begin sample extraction  within one year  of
         collection,   and   analyze  all   extracts
         within 40 days  of  extraction.

    10    SAMPLE PREPARATION
         The  sample  preparation  process  involves
         modifying the physical form of  the sample
         so  that  the  PCDDs   and  PCOFs   can  be
         extracted efficiently.    In  general,  the
         samples must be in a liquid form or in the
         form of finely divided solids  in order for
         efficient extraction to take  place.  Table
         8   lists   the  phase(s)   and   quantity
         extracted  for  various  sample  matrices.
         Samples  containing  a   solid   phase  and
         samples  containing  particle  sizes  larger
         than  1  mm  require  preparation prior  to
         extraction.      Because  PCDDs/PCOFs   are
         strongly associated with particulates, the
         preparation   of    aqueous   samples   is
         dependent on   the  solids  content  of  the
         sample.   Aqueous samples  containing less
         than one percent solids are extracted in a
         separatory  funnel.    A  smaller  sample
         aliquot   is   used  for   aqueous   samples
         containing  one  percent  solids  or  more.
         For  samples  expected or known  to contain
         high levels of the PCDDs and/or PCOFs, the
         smallest sample size representative of the
         entire  sample  should be  used,  and  the
         sample  extract  should   be   diluted,  if
         necessary, per Section 16.4.
  10.1    Determine percent solids

10.1.1    Weigh  5  -  10  g  of  sample  (to  three
         significant figures)  into  a tared beaker.
         NOTE:    This  aliquot  is   used  only  for
         determining  the  solids  content  of  the
         sample, not for analysis of PCDDs/PCDFs.

10.1.2    Dry overnight (12 hours minimum) at 110 5
         C, and cool in a dessicator.

10.1.3    Calculate percent solids as follows:
  10.2
% solids =
weight of sample after drying
weight of sample before drying

Determine particle size
                                         x 100
10.2.1    Spread  the  dried   sample  from  Section
         10.1.2  on  a  piece  of   filter  paper  or
         aluminum foil in a fume hood or glove box.

10.2.2    Estimate the size of the particles  in  the
         sample.    If  the  size  of   the  largest
         particles  is  greater   than  1   mm,   the
         particle size  must  be reduced to  1  mm or
         less prior to extraction.

  10.3    Preparation of  aqueous  samples containing
         less   than  one   percent  solids  --   The
         extraction  procedure for  aqueous samples
         containing  less than one  percent  solids
         involves filtering  the  sample, extracting
         the  paniculate  phase   and  the   filtrate
         separately, and combining the  extracts  for
         analysis.      The   aqueous   portion   is
         extracted   by   shaking   with  methylene
         chloride  in  a  separatory  funnel.     The
         particulate  material  is  extracted   using
         the SOS procedure.

10.3.1    Mark  the  original  level  of  the  sample on
         the  sample bottle  for   reference.    Weigh
         the sample  in  the  bottle on a top loading
         balance to 1 g.

10.3.2   Dilute a  sufficient volume of the labeled
         compound  spiking  standard by  a, factor of
         50  with  acetone.   1.0  ml  of the diluted
         solution  is  required for each sample,  but
         no  more  solution should  be prepared  than
         can  be used in one  day.   Spike 1.0  ml  of
         the   diluted   solution   into  the   sample
         bottle.  Cap the bottle  and  mix  the  sample
         by  careful  shaking.  Allow the sample  to
         equilibrate   for   1-2    hours,    with
         occasional shaking.

10.3.3   For   each  sample   or  sample set  (to  a
         maximum  of  20) to be extracted during  the
         same   12-hour  shift,  place  two  1.0  liter
         aliquots  of  reagent water in clean 2 liter
         separatory flasks.

10.3.4   Spike  1.0   ml  of   the   diluted   labeled
         compound  spiking  standard  (Section  6.10)
          into  one  reagent  water  aliquot.    This
         aliquot  will  serve  as  the blank.   Dilute
         20   uL  of  the  precision   and   recovery
         standard  (Section  6.14)  to  1.0  ml  with
         acetone.    Spike  1.0  mL of  the  diluted
         precision and  recovery standard  into  the
                                                                                                           19

-------
          remaining  reagent   water  aliquot.    This
          aliquot  will  serve  as  the  PAR  (Section
          14.5).

 10.3.5    Assemble  a Buchner  funnel   on top  of  a
          clean  1  L  filtration flask.    Apply  a
          vacuum  to  the  flask, and pour  the entire
          contents  of  the sample  bottle through  a
          glass fiber filter  (Section  5.5.4)  in the
          Buchner   funnel,    swirling   the   sample
          remaining  in  the  bottle to suspend  any
          particulates.

 10.3.6    Rinse the sample bottle twice with 5 ml of
          reagent  water  to  transfer   any  remaining
          particulates onto the filter.

 10.3.7    Rinse the  any  particulates   off the  sides
          of   the   Buchner   funnel    with   small
          quantities of reagent water.

 10.3.8    Weigh the  empty sample  bottle on a  top-
          loading  balance  to  1  g.    Determine the
          weight  of  the  sample  by difference.   Do
          not discard the bottle at this point.

 10.3.9    Extract the filtrates using  the procedures
          in Section 11.

10.3.10    Extract   the   particulates   using   the
          procedures in Section 11.

   10.4    Preparation of  samples containing greater
          than one percent solids

 10.4.1    Weigh a well-mixed  aliquot  of each sample
          (of  the same matrix type)   sufficient  to
          provide  10  g  of dry solids  (based on the
          solids  determination  in 10.1.3)  into  a
          clean beaker or glass jar.

 10.4.2    Spike   1.0  ml   of   the  diluted   labeled
          compound spiking solution (Section 10.3.2)
          into the sample aliquot(s).

 10.4.3    For  each  sample  or  sample  set  (to  a
          maximum of  20)  to  be extracted during the
          same   12-hour   shift,   weigh   two   10  g
          aliquots   of   the   appropriate  reference
          matrix  (Section 6.6) into clean beakers or
          glass jars.

 10.4.4    Spike   1.0  ml   of   the  diluted  labeled
          compound   spiking   solution   into   one
          reference  matrix aliquot.    This aliquot
          will  serve  as  the  blank.  Spike 1.0 ml of
          the   diluted   precision    and   recovery
          standard   (Section    10.3.4)   into   the
          remaining  reference matrix  aliquot.   This
          aliquot will  serve as  the  PAR  (Section
          14.5).
10.4.5   Stir   or  tumble   and   equilibrate   the
         aliquots for 1-2 hours.

10.4.6   Extract  the  aliquots  using the procedures
         in Section 11.

  10.5   Multi-phase samples

10.5.1   Pressure filter the sample, blank, and  PAR
         aliquots through Whatman GF/D glass  fiber
         filter  paper.    If necessary,  centrifuge
         these  aliquots  for 30  minutes  at greater
         than 5000 rpm prior to filtration.

10.5.2   Discard  any  aqueous  phase  (if   present).
         Remove any non-aqueous  liquid (if present)
         and  reserve  for  recombination   with  the
         extract  of   the  solid  phase  (Section
         11.1.2.5).    Prepare the  filter  papers  of
         the  sample  and  QC aliquots  for particle
         size   reduction  and   blending   (Section
         10.6).

  10.6   Sample   grinding,    homogenization,    or
         blending --  Samples  with  particle   sizes
         greater  than   1   mm   (as  determined   by
         Section  10.2.2) are subjected to  grinding,
         homogenization,  or blending.   The  method
         of reducing  particle  size  to  less   than  1
         mm is  matrix dependent.  In general, hard
         particles can be reduced by grinding with
         a mortar and pestle.  Softer particles  can
         be reduced by grinding  in a Wiley mill  or
         meat  grinder,  by  homogenization,   or   by
         blending.

10.6.1   Each  size  reducing  preparation  procedure
         on  each matrix  shall  be  verified   by
         running  the  tests  in  Section  8.2  before
         the procedure is employed routinely.

10.6.2   The  grinding,  homogenization,  or blending
         procedures shall be carried out  in a glove
         box or fume  hood to prevent particles from
         contaminating the work  environment.

10.6.3   Grinding --  Tissue samples, certain  papers
         and  pulps,  slurries,  and amorphous  solids
         can  be  ground  in a  Wiley mill  or  heavy
         duty   meat   grinder.      In  some   cases,
         reducing the temperature of the  sample to
         freezing or  to dry ice  or  liquid nitrogen
         temperatures can   aid  in the  grinding
         process.   Grind the  sample aliquots  from
         Section  10.4.5  or  10.5.2  in  a   clean
         grinder.     Do  not   allow   the   sample
         temperature  to  exceed  50  C.    Grind  the
         blank  and  reference matrix aliquots using
         a clean  grinder.
20

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  10.6.4   Homogem'zation  or  blending  --   Particles
           that   are  not   ground  effectively,   or
           particles  greater than 1 mm  in size  after
           grinding,  can often be reduced in size  by
           high   speed   homogenization  or   blending.
           Homogenize and/or blend the sample, blank,
           and   PAR   aliquots  from  Section  10.4.5,
           10.5.2, or 10.6.3.

  10.6.5   Extract  the  aliquots using  the procedures
           in Section 11.

      11   EXTRACTION AND CONCENTRATION

    11.1   Extraction

  11.1.1   Extraction of  filtrates  --  extract  the
           aqueous  samples,  blanks, and PAR  aliquots
           according  to  the  following  procedures.

11.1.1.1  j Pour  filtered aqueous  sample  into  a  2-L
           separatory funnel.   Add  60 mL  methylene
           chloride   to  the   sample  bottle,   seal,and
           shake 60   seconds  to  rinse  the   inner
           surface.

11.1.1.2   Transfer   the  solvent  to  the separatory
           funnel  and extract  the sample by  shaking
           the   funnel  for   2 minutes  with   periodic
           venting.    Allow  the   organic   layer   to
           separate   from  the  water  phase  for   a
           minimum  of 10 minutes.   If the  emulsion
           interface  between layers  is more  than one-
           third the  volume  of   the  solvent  layer,
           employ mechanical  techniques to  complete
           the   phase   separation   (e.g.   a   glass
           stirring   rod).      Drain   the   methylene
           chloride   extract   into   a   500-mL   KD
           concentrator.

11.1.1.3   Extract  the  water sample  two  more  times
           using 60  mL  of   fresh  methylene  chloride
           each  time.  Drain each  extract  into the  KD
           concentrator.  After  the third  extraction,
           rinse the  separatory funnel with  at  least
           30 mL of fresh methylene chloride.

  11.1.2   Soxhlet/Dean-Stark extraction of  solids  --
           Extract  the  solid samples,  participates,
           blanks,   and  PAR  aliquots   using    the
           following  procedure.

11.1.2.1   Charge a clean extraction  thimble with  5.0
           g of  100/200  mesh silica (Section 6.5.1.1)
           and  100  g of quartz sand  (Section  6.5.4).
           NOTE:    Do not  disturb the silica  layer
           throughout the extraction process.
11.1.2.2   Place  the  thimble  in  a  clean  extractor.
           Place  30   -  40  ml   of   toluene   in  the
           receiver and 200  - 250  mL  in the flask.

11.1.2.3   Pre-extract  the  glassware  by  heating  the
           flask  until  the toluene is boiling.   When
           properly  adjusted,  1-2 drops  of  toluene
           per  second will  fall  from the condenser
           tip   into   the  receiver.      Extract  the
           apparatus  for 3 hours minimum.

11.1.2.4   After  pre-extraction,  cool  and disassemble
           the  apparatus.   Rinse the  thimble  with
           toluene and allow to air dry.

11.1.2.5   Load  the  wet sample  from  Section  10.4.6,
           10.5.2,  10.6.3,  or  10.6.4, and any  non-
           aqueous  liquid  from Section  10.5.2  into
           the thimble and manually mix into  the sand
           layer  with  a clean metal spatula carefully
           breaking up any large  lumps of  sample.

11.1.2.6   Reassemble the  pre-extracted SOS apparatus
           and  add  a fresh  charge of toluene  to  the
           receiver and reflux flask.

11.1.2.7   Apply  power to  the heating  mantle  to begin
           refluxing.    Adjust  the  reflux  rate  to
           match  the  rate of percolation  through  the
           sand  and  silica  beds  until water  removal
           lessens  the restriction  to toluene  flow.
           Check  the  apparatus  for foaming frequently
           during the first 2 hours  of  extraction.
           If foaming occurs,  reduce  the  reflux rate
           until  foaming subsides.

11.1.2.8   Drain  the  water from the receiver  at 1  -  2
           hours  and 8-9 hours, or sooner  if  the
           receiver  fills  with  water.    Reflux  the
           sample for  a total of  16 -  24  hours.  Cool
           and disassemble the  apparatus.   Record the
           total  volume of water collected.

11.1.2.9   Remove the distilling  flask,  estimate  and
           record the  volume  of extract   (to  the
           nearest 100 mL),  and pour  the  extract from
           the  receiver  and  flask   into  a   500  mL
           separatory funnel.  Rinse  the  receiver and
           flask   with  toluene   and  add  to   the
           separatory funnel.     Proceed  with  back
           extraction per  Section  11.1.3.

  11.1.3   Back extraction with base and  acid

11.1.3.1   Spike  1.0  mL   of   the cleanup  standard
           (Section 6.11)  into  the separatory funnels
           containing  the  sample and   QC  extracts
           (Section 11.1.1.3 or 11.1.2.9).
                                                                                                             21

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11.1.3.2   Partition  the extract  against  50 mL  of
           potassium   hydroxide   solution    (Section
           6.1.1).  Shake for 2 minutes with  periodic
           venting  into  a hood.   Remove  and discard
           the  aqueous   layer.     Repeat  the  base
           washing  until  no color  is  visible in the
           aqueous  layer,   to   a   maximum  of  four
           washings.   Minimize  contact  time between
           the  extract   and  the   base  to  prevent
           degradation of the PCDDs and PCDFs.

11.1.3.3   Partition  the extract  against  50 ml  of
           sodium  chloride   solution  (Section 6.1.3)
           in the same way as with base.  Discard the
           aqueous  layer.

11.1.3.4   Partition  the extract  against  50 ml  of
           sulfuric acid  (Section  6.1.2)  in the same
           way as with base.  Repeat the acid washing
           until no color is visible  in  the aqueous
           layer, to a maximum of four washings.

11.1.3.5   Repeat   the  partitioning  against  sodium
           chloride solution and discard  the aqueous
           layer.

11.1.3.6   Pour each  extract through  a drying column
           containing 7  to  10 cm of anhydrous sodium
           sulfate.  Rinse the separatory funnel with
           30 - 50 ml of toluene and pour through the
           drying column.   Collect each extract  in a
           500  ml   round  bottom flask.   Concentrate
           and  clean  up the  samples  and  QC  aliquots
           per Sections 11.2 and 12.

    11.2   Concentration

  11.2.1   Macro-concentration   --   Concentrate   the
           extracts  in  separate 500  ml  round bottom
           flasks on a rotary evaporator.

11.2.1.1   Assemble  the rotary  evaporator  according
           to  manufacturer's instructions,  and  warm
           the  water bath   to  45  C.    On  a  daily
           basis,  preclean   the  rotary  evaporator by
           concentrating  100 ml of  clean extraction
           solvent  through  the  system.   Archive both
           the  concentrated solvent  and  the solvent
           in the catch flask for  contamination check
           if necessary.  Between  samples,  three 2 -
           3 ml  aliquots  of toluene should be rinsed
           down the feed tube into a waste beaker.

11.2.1.2   Attach  the round  bottom flask containing
           the   sample   extract   to   the   rotary
           evaporator.   Slowly  apply vacuum  to the
           system,  and  begin   rotating  the  sample
           flask.
11.2.1.3   Lower  the  flask  into  the  water  bath  and
           adjust  the  speed  of  rotation  and   the
           temperature  as  required  to  complete  the
           concentration  in  15  -  20  minutes.  At  the
           proper rate  of concentration, the flow  of
           solvent  into the receiving  flask will  be
           steady, but  no bumping or visible boiling
           of the extract will  occur.   NOTE:  If  the
           rate of concentration  is too  fast, analyte
           loss may occur.

11.2.1.4   When the liquid  in the concentration  flask
           has  reached  an  apparent  volume  of  2  ml,
           remove the  flask from the  water  bath  and
           stop the rotation.   Slowly and carefully,
           admit air into the system.  Be sure not  to
           open the valve so quickly that the sample
           is blown out of  the flask.  Rinse the  feed
           tube with approximately 2 ml of hexane.

11.2.1.5   Transfer the extract to a vial using  three
           2  -  3  mL rinses  of  hexane.   Proceed  with
           micro-concentrat ion and solvent exchange.

11.2.1.6   The  extracts   of  the  filtered  aqueous
           sample   and    its  particulates   must   be
           combined prior  to  cleanup  and   analysis.
           Transfer the concentrated  extract  of  the
           aqueous sample to the  flask containing  the
           concentrated  particulate  extract.   Rinse
           the flask twice with 5 ml toluene, and  add
           these  rinses   to  the   flask   with   the
           combined extracts.   Reattach the flask  to
           the  rotary  evaporator  and  continue   to
           concentrate  the  combined extract  until  the
           volume  is  approximately  2  ml.    Proceed
           with   micro-concentration    and   solvent
           exchange.

  11.2.2   Micro-concentration and solvent exchange

11.2.2.1   Toluene  extracts to  be  subjected  to  GPC
           cleanup  are  exchanged   into   methylene
           chloride.  Extracts that are  to be cleaned
           up using  silica  gel,  alumina,  and/or  AX-
           21/Celite   are   exchanged   into  hexane.
           Extracts  to   be  subjected   to   HPLC   are
           exchanged into nonane.

11.2.2.2   Transfer  the  vial  containing  the  sample
           extract  to a nitrogen evaporation device.
           Adjust the   flow  of  nitrogen so  that  the
           surface  of   the  solvent  is  just visibly
           disturbed.   NOTE:  A  large vortex  in  the
           solvent may  cause analyte loss.

11.2.2.3   Lower the vial  into a  45 C water bath  and
           continue concentrating.
 22

-------
11.2.2.4   When   the   volume   of   the   liquid   is
           approximately 100 uL,  add  2  -  3 mL of the
           desired  solvent  (methylene  chloride  or
           hexane)  and  continue  concentration  to
           approximately 100 uL.  Repeat the addition
           of solvent and concentrate once more.

11.2.2.5   If the extract is to be cleaned up by GPC.
           adjust the volume of the extract to 5.0 ml
           with methylene chloride.  Proceed with GPC
           cleanup (Section 12.2).

11.2.2.6   If  the extract  is  to be  cleaned  up  by
           column  chromatography  (alumina,   silica
           gel, AX-21/Celite), bring the final volume
           to  1.0  ml  with  hexane.    Proceed  with
           column cleanups (Sections 12.3 - 12.5).

11.2.2.7   For   extracts   to  be  concentrated  for
           injection  into the  HPLC  or GCMS -- add 10
          , uL of  nonane to the  vial.   Evaporate the
           solvent  to  the  level   of  the  nonane.
           Evaporate  the  hexane  in  the vial  to the
           level of the nonane.

11.2.2.8   Seal  the  vial and  label  with  the sample
           number.    Store  in  the  dark  at  room
           temperature  until ready for HPLC or GCMS.

      12   EXTRACT CLEANUP

    12.1 |  Cleanup   may   not    be   necessary   for
           relatively  clean  samples  (e.g.,  treated
           effluents,  groundwater,   drinking  water).
           If  particular  circumstances  require  the
           use  of  a  cleanup procedure,  the analyst
           may use any  or all of  the procedures below
           or   any   other    appropriate   procedure.
           Before  using  a  cleanup  procedure,  the
           analyst    must    demonstrate   that   the
           requirements  of  Section  8.2  can  be  met
           using the cleanup procedure.
  12.1.1
  12.1.2
  12.1.31
Gel  permeation   chromatography  (Section
12.2)  removes  many high  molecular  weight
interferences   that   cause   GC   column
performance  to  degrade.   It may be used
for all soil and sediment extracts and may
be  used  for  water   extracts  that  are
expected to  contain high  molecular  weight
organic    compounds     (e.g.,    polymeric
materials,  humic acids).

Acid,  neutral,  and basic silica  gel,  and
alumina (Sections  12.3  and  12.4)  are used
to     remove    nonpolar     and     polar
interferences.
AX-21/Celite  (Section  12.5)   is
remove nonpolar interferences.
                                              used   to
  12.1.4   HPLC  (Section  12.6)   is  used  to  provide
           specificity   for   the   2,3,7,8-substituted
           and other PCDD and PCOF isomers.

    12.2   Gel permeation chromatography  (GPC)

  12.2.1   Column packing

12.2.1.1   Place 70  - 75 g of SX-3 Bio-beads in a 400
           - 500 mL beaker.

12.2.1.2   Cover  the beads   with  methylene  chloride
           and  allow  to swell  overnight  (12  hours
           minimum).

12.2.1.3   Transfer  the swelled  beads  to the  column
           and pump  solvent  through the  column,  from
           bottom  to top,  at 4.5 -  5.5  mL/min prior
           to connecting the  column  to  the detector.

12.2.1.4   After purging the column with solvent for
           1-2   hours,   adjust   the  column  head
           pressure  to  7 -  10 psig  and purge  for 4  -
           5  hours to  remove air.   Maintain  a  head
           pressure  of  7  -  10  psig.    Connect  the
           column to the detector.

  12.2.2   Column calibration

12.2.2.1   Load  5  mL  of  the  calibration  solution
           (Section 6.4) into the sample  loop.

12.2.2.2   Inject the calibration solution  and record
           the signal from the detector.  The elution
           pattern   will be  corn   oil,  bis(2-ethyl
           hexyl)    phthalate,     pentachlorophenol,
           perylene, and sulfur.

12.2.2.3   Set  the "dump time"  to allow >85  percent
           removal  of   the  corn  oil  and >85  percent
           collection of the  phthalate.

12.2.2.4   Set the "collect  time" to the  peak minimum
           between perylene and sulfur.

12.2.2.5   Verify    the   calibration    with    the
           calibration   solution   after   every   20
           extracts.   Calibration is verified  if the
           recovery   of   the  pentachlorophenol   is
           greater  than 85  percent.   If calibration
           is  not   verified,   the   system   shall  be
           recalibrated   using    the   calibration
           solution,  and  the   previous  20  samples
           shall be  re-extracted  and cleaned up using
           the calibrated GPC system.

  12.2.3   Extract  cleanup -- GPC  requires  that the
           column  not   be  overloaded.    The  column
           specified  in this method  is  designed  to
           handle   a  maximum  of   0.5  g   of  high
           molecular  weight   material    in   a   5  mL
                                                                                                             23

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           extract.    If  the  extract   is  known  or
           expected to  contain more than 0.5  g,  the
           extract is split into aliquots for GPC and
           the  aliquots are  combined  after  elation
           from  the  column.   The solids content  of
           the     extract      may    be    obtained
           gravimetrically by evaporating the solvent
           from a 50 uL aliquot.

12.2.3.1    Filter  the  extract  or  load through  the
           filter  holder   to  remove   particulates.
           Load the 5.0 ml extract onto the  column.

12.2.3.2    Elute  the  extract  using  the calibration
           data   determined    in   Section   12.2.2.
           Collect the eluate in a clean 400 - 500 ml
           beaker.

12.2.3.3    Rinse  the  sample  loading  tube  thoroughly
           with  methylene  chloride  between extracts
           to prepare for the next sample.

12.2.3.4    If   a  particularly  dirty  extract   is
           encountered,  a  5.0  mL  methylene chloride
           blank  shall  be  run  through  the  system to
           check for carry-over.

12.2.3.5    Concentrate the eluate per Section 11.2.1,
           11.2.2, and  11.3.1  or  11.3.2 for further
           cleanup or for injection into the GCMS.

    12.3    Silica gel  cleanup

  12.3.1    Place  a  glass wool  plug  in  a 15  mm i.d.
           chromatography column.  Pack  the column in
           the following order  (bottom  to top):   1  g
           silica gel  (Section 6.5.1.1), four g basic
           silica gel  (Section 6.5.1.3), 1  g silica
           gel,   8   g   acid    silica   gel   (Section
           6.5.1.2),  2  g  silica  gel,  1  g  sodium
           sulfate (Section 6.2.1).   Tap  the column
           to settle the adsorbents.

  12.3.2    Pre-rinse  the column with 50 -  100  ml  of
           hexane.    Close   the  stopcock  when  the
           hexane  is  within   1  mm  of  the  sodium
           sulfate.   Discard  the eluate.   Check  the
           column for  channeling.    If  channeling  is
           present, discard  the  column and  prepare
           another.

  12.3.3    Apply  the  concentrated  extract  to  the
           column.    Open  the  stopcock  until  the
           extract  is  within  1   mm of the  sodium
           sulfate.

  12.3.4    Rinse  the   receiver  twice  with   1   mL
           portions of hexane and apply  separately to
           the  column.   Elute the  PCDDs/PCDFs  with
           100 mL hexane and collect the eluate.
12.3.5



  12.4

12.4.1


12.4.2
Concentrate the  eluate per Section  11.2.1
or  11.2.2  for  further  cleanup  or  for
injection into the HPLC or GCMS.
12.4.3
12.4.4
12.4.5
12.4.6
12.4.7


12.4.8



  12.5

12.5.1
Alumina cleanup

Place a  glass wool plug  in a  15  mm
chromatography column.
i.d.
12.5.2
Pack  the column  in  the  following order
(bottom  to   top):    1  g  neutral   alumina
(Section  6.5.2.1),   3   g  basic   alumina
(Section 6.5.2.2),  1  g  neutral alumina, 6
g  acid  alumina  (Section 6.5.2.3),   2  g
neutral  alumina,   1   g   sodium   sulfate
(Section 6.2.1).  Tap the column to settle
the adsorbents.

Pre-rinse the  column with 50  -  100 mL of
hexane.    Close  the  stopcock  when  the
hexane  is  within  1  mm  of  the  sodium
sulfate.

Discard the  eluate.   Check the column for
channeling.     If  channeling   is  present,
discard the column and prepare another.

Apply  the  concentrated  extract   to  the
column.    Open  the  stopcock  until  the
extract  is   within  1  mm of  the  sodium
sulfate.

Rinse   the   receiver  twice   with   1   mL
portions of  hexane and apply separately to
the   column.      Elute   the   interfering
compounds with  100  mL  hexane  and  discard
the eluate.

Elute  the PCDDs and  PCDFs with 20 mL of
methylene chloride:hexane (1:1 v/v).

Concentrate the eluate  per Section 11.2.1
or  11.2.2  for  further  cleanup   or  for
injection into the HPLC or GCMS.

AX-21/Celite

Cut  both ends from  a  10 mL disposable
serological   pi pet  to  produce  a  10  cm
column.   Fire  polish both ends  and flare
both ends if desired.  Insert  a glass  wool
plug at. one end, then pack the column  with
1 g of  the  activated AX-21/Celite  to  form
a 2 cm  long adsorbent bed.  Insert  a glass
wool  plug on top of  the bed  to  hold the
adsorbent in place.

Pre-rinse  the  column  with   five  mL  of
toluene   followed  by   2  mL  methylene
chloride:methanol:toluene  (15:4:1  v/v),  1
mL  methylene   chloride:cyclohexane   (1:1
 24

-------
           v/v),  and  five  ml hexane.    If  the  flow
           rate  of  eluate  exceeds  0.5  mL  per  min,
           discard  the column.

  12.5.3   When  the  solvent  is  within 1 mm  of  the
           column  packing,  apply the  sample  extract
           to  the column.   Rinse  the sample  container
           twice  with  1  ml  portions  of  hexane  and
           apply  separately to the  column.   Apply  2
           ml  of hexane to  complete  the transfer.

  12.5.4   Elute  the interfering  compounds with  2 ml
           of    hexane,    2    ml    of     methylene
           chloride:cyclohexane  (1:1 v/v),  and 2 ml
           of   methylene    chloride:methanol:toluene
           (15:4:1  v/v).  Discard the eluate.

  12.5.5   Invert  the column and elute the  PCDDs and
           PCDFs  with 20 ml  of  toluene.    If  carbon
           particles  are   present   in   the   eluate,
           filter through glass fiber filter paper.

  12.5.6   Concentrate the  eluate per  Section  11.2.1
           or  11.2.2  for   further   cleanup  or  for
           injection into the HPLC or GCMS.

    12.6   HPLC (Reference  6)

  12.6.1   Column calibration

12.6.1.1   Prepare  a calibration standard containing
           the 2,3,7,8-  isomers  and/or other  isomers
           of   interest   at   a   concentration  of
           approximately 500 pg/uL in chloroform.

12.6.1.2   Inject  30  uL of the  calibration solution
           into  the HPLC and  record the signal  from
           the detector.   Collect the  eluant  for re-
           use.  The elution order will be the tetra-
           through  octa-isomers.

12.6.1.3   Establish the  collect  time  for the  tetra-
           isomers   and  for  the other   isomers of
           interest.    Following  calibration,  flush
           the   injection    system    with    copious
           quantities  of   chloroform,    including   a
           minimum  of five  50-uL  injections  while the
           detector  is  monitored,   to  ensure  that
           residual PCDDs and PCDFs  are  removed from
           the system.

12.6.1.4   Verify     the    calibration    with    the
           calibration  solution  after  every  20
           extracts.   Calibration is verified  if  the
           recovery of the PCDDs and PCDFs  from the
           calibration standard (Section  12.6.1.1)  is
           75   -   125  percent   compared   to   the
           calibration  (Section  12.6.1.2).       If
           calibration is  not  verified,  the  system
           shall     be    recalibrated    using    the
           calibration solution,  and  the previous 20
           samples shall  be  re-extracted and cleaned
           up using the calibrated system.

  12.6.2   Extract cleanup --  HPLC  requires that the
           column  not  be overloaded.    The  column
           specified  in  this  method  is  designed to
           handle a maximum of  30 uL  of extract.  If
           the extract cannot be  concentrated to  less
           than 30 uL, -it is split into  fractions and
           the  fractions  are  combined  after elution
           from the column.

12.6.2.1   Rinse the  sides of  the vial  twice with 30
           uL of  chloroform  and  reduce  to  the  level
           of the nonane with the blowdown  apparatus.
           Rinse the  sides of  the vial  with 20 uL of
           chloroform  to  bring  the  extract  volume to
           30 uL.

12.6.2.2   Inject the 30 uL extract into the HPLC.

12.6.2.3   Elute  the   extract  using  the calibration
           data  determined  in  12.6.1.    Collect the
           fraction(s) in a  clean 20  mL concentrator
           tube  containing   5   mL of  hexane:acetone
           (1:1 v/v).

12.6.2.4   If an  extract  containing  greater than 100
           ng/mL   of   total   PCDD   or   PCDF   is
           encountered,  a  30  uL  chloroform  blank
           shall be  run through  the  system to check
           for carry-over.

12.6.2.5   Concentrate the eluate per Section 11.2.2
           for injection into the GCMS.

      13   HRGC/HRMS ANALYSIS

    13.1   Establish  the  operating conditions given
           in Section 7.1.

    13.2   Add   10   uL  of  the   internal   standard
           solution   (Section  6.12)  to  the  sample
           extract immediately  prior  to injection to
           minimize   the   possibility   of   loss  by
           evaporation, adsorption, or  reaction.   If
           an extract  is to be reanalyzed,  do not add
           more    instrument     internal    standard
           solution.    Rather,  bring  the extract  back
           to its previous volume (e.g., 19 uL)  with
           pure nonane only.

    13.3   Inject 1.0  uL  of  the  concentrated extract
           containing the internal standatd solution,
           using  on-column   or split less   injection.
           Start  the   GC  column  initial   isothermal
           hold  upon   injection.    Start   MS   data
           collection  after  the solvent peak elutes.
           Stop data collection after the octachloro-
                                                                                                             25

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          dioxin and furan  have  eluted.   Return the
          column  to  the   initial  temperature  for
          analysis of the next extract or standard.

     14   SYSTEM AND LABORATORY PERFORMANCE

   14.1   At  the beginning of  each   12-hour  shift
          during which  analyses  are performed,  GCMS
          system  performance  and  calibration  are
          verified   for  all  native   and  labeled
          compounds.   For  these  tests,  analysis of
          the  CS3  calibration  verification  (VER)
          standard  (Section 6.13  and  Table 4)  and
          the  isomer   specificity  test  standards
          (Sections 6.16  and  Table 5)  shall  be used
          to   verify   all    performance   criteria.
          Adjustment   and/or   recall* brat ion   (per
          Section 7}  shall  be performed  until  all
          performance criteria are met.   Only after
          all  performance   criteria   are  met   may
          samples,   blanks,   and   precision   and
          recovery standards be analyzed.

   14.2   Mass spectrometer resolution -- A static
          resolving  power  of  at  least   10,000  (10
          percent   valley   definition)   must   be
          demonstrated  at  appropriate  masses before
          any  analysis  is   performed.      Static
          resolving  power checks  must  be performed
          at  the  beginning and  at the end  of  each
          12-hour shift.  Corrective actions must be
          implemented whenever the resolving  power
          does not meet the requirement.

 14.2.1   The analysis  time for  PCDDs  and PCDFs may
          exceed the long-term mass stability of the
          mass spectrometer.  Because the instrument
          is  operated  in the  high-resolution  mode,
          mass drifts of  a few ppm (e.g., 5 ppm in
          mass) can  have  serious  adverse effects on
          instrument  performance.     Therefore,   a
          mass-drift  correction  is mandatory.    A
          lock-mass  ion from the reference compound
          (PFK)   is   used   for    tuning   the   mass
          spectrometer.      The   lock-mass   ion  is
          dependent  on   the   masses   of   the  ions
          monitored within  each descriptor, as shown
          in  Table  3.   The level  of  the reference
          compound   (PFK)   metered  into   the   ion
          chamber during  HRGC/HRMS analyses should
          be  adjusted so  that the  amplitude of the
          most intense selected lock-mass ion signal
          (regardless of the descriptor number) does
          not  exceed 10  percent  of  the  full-scale
          deflection  for  a given  set of  detector
          parameters.     Under   those  conditions,
          sensitivity   changes  that   might  occur
          during   the    analysis   can    be   more
          effectively monitored.   NOTE:   Excessive
          PFK (or any other reference substance) may
         cause noise  problems and contamination  of
         the ion source resulting  in an  increase  in
         time lost in cleaning the source.

14.2.2   By  using  a  PFK  molecular  leak,  tune the
         instrument  to meet  the  minimum  required
         resolving  power  of  10,000  (10   percent
         valley) at m/z 304.9824 (PFK) or any other
         reference  signal  close  to  m/z   303.9016
         (from TCDF).   By using  the peak  matching
         unit and  the  PFK reference  peak,  verify
         that the  exact mass  of m/z 380.9760  (PFK)
         is within 5 ppm of the required value.

  14.3   Calibration verification

14.3.1   Inject   the   VER   standard    using  the
         procedure in Section 13.

14.3.2   Them/z abundance ratios for all PCODs and
         PCDFs shall  be within the limits  in  Table
         3A; otherwise, the mass spectrometer  shall
         be adjusted until the m/z abundance ratios
         fall within  the  limits  specified,   and the
         verification    test     (Section    14.3.1)
         repeated.    If  the  adjustment  alters the
         resolution   of   the   mass  spectrometer,
         resolution  shall  be  verified   (Section
         14.2) prior  to  repeat of the verification
         test.

14.3.3   Compute the  concentration of  each  native
         compound by isotope dilution (Section 7.5)
         for  those  compounds  that  have   labeled
         analogs   (Table   1).       Compute   the
         concentration of  the labeled  compounds  by
         the  internal  standard  method.      These
         concentrations are  computed based on the
         averaged  relative  response  and   averaged
         response  factor  from the calibration data
         in Section 7.

14.3.4   For    each    compound,    compare   the
         concentration    with    the     calibration
         verification  limit   in  Table 7.    If all
         compounds  meet  the   acceptance criteria,
         calibration   has   been   verified.     If,
         however,    any   compound    fails,   the
         measurement   system   is   not  performing
         properly  for  that  compound.     In this
         event,    prepare   a   fresh    calibration
         standard  or  correct  the  problem   causing
         the  failure  and  repeat  the  resolution
         (Section  14.2)  and  verification  (Section
         14.3.1) tests, or recalibrate (Section 7).

  14.4   Retention times and GC resolution

14.4.1   Retention times
26

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14.4.1.1    Absolute  "The  absolute retention  times
           of   the   13C12-1,2.3,4-TCDD   and     C^-
           1,2,3,7,8,9-HxCDF  GCMS internal  standards
           shall   be  within   15   seconds  of   the
           retention    times     obtained     during
           calibration (Section 7.2.4).

14.4.1.2   Relative  --  The  relative retention  times
           of  native  and  labeled  PCDDs  and  PCDFs
           shall  be  within the limits given  in Table
           2.

  14.4.2   GC  resolution

14.4.2.1    Inject  the  isomer  specificity  standards
           (Section   6.16)    on    their   respective
           columns.

14.4.2.2   The valley height  between 2,3,7,8-TCDO and
           the other  tetra-   dioxin isomers  at  m/z
           319.8965,  and  between  2,3,7,8-TCDF  and the
           other  tetra- furan isomers at m/z 303.9016
           shall   not  exceed  25  percent  on  their
           respective columns (Figure 3).

  14.4.3   If  the absolute or relative  retention time
           of  any compound is  not  within  the  limits
           specified or  the 2,3,7,8- isomers  are not
           resolved,   the   GC   is   not   performing
           properly.   In  this event,  adjust  the  GC
           and repeat the verification  test  (Section
           14.3.1) or recalibrate (Section 7).

    14.5   Ongoing precision  and  accuracy

  14.5.1   Analyze the  extract of  the  precision and
           recovery  standard  (PAR)  (Section 10.3.4 or
           10.4.4) prior  to analysis of  samples from
           the same  set.

  14.5.2   Compute the  concentration of  each  PCOO  or
           PCDF by isotope dilution (Section 7.5) for
           those  compounds that have labeled analogs.
           Compute the concentration of  the  labeled
           compounds by the  internal  standard method.

  14.5.3   For    each     compound,     compare    the
           concentration  with the limits  for  ongoing
           accuracy  in Table  7.    If  all  compounds
           meet    the  acceptance  criteria,   system
           performance  is acceptable and  analysis  of
           blanks  and  samples  may proceed.     If,
           however,   any   individual   concentration
           falls   outside  of  the  range  given,  the
           extraction/concentration  processes  are not
           being    performed   properly    for    that
           compound.    In  this  event,  correct  the
           problem,   re-extract    the    sample   set
         (Section   10)   and   repeat   the  ongoing
         precision   and  recovery   test   (Section
         14.5).

14.5.4   Add results  which  pass the specifications
         in Section  14.5.3  to initial  and  previous
         ongoing  data  for  each  compound  in each
         matrix.    Update  QC  charts   to  form  a
         graphic    representation    of   continued
         laboratory   performance.       Develop   a
         statement of  laboratory  accuracy  for each
         PCDD  and  PCDF  in  each   matrix  type  by
         calculating  the average  percent  recovery
         (R) and  the standard deviation  of percent
         recovery (sr).   Express  the accuracy as a
         recovery interval from R - 2sr  to  R  + 2sr.
         For example,  if R  = 95X and  sr = 5X, the
         accuracy is 85  - 105%.

    15   QUALITATIVE DETERMINATION

         Identification    is    accomplished   by
         comparison  of  data  from  analysis   of  a
         sample  or  blank with data  for authentic
         standards.   For compounds for which the
         relative   retention   times   are    known,
         identification  is  confirmed  per  Sections
         15.1 and 15.2.

  15.1   Labeled  compounds   and native  PCDDs and
         PCDFs having no labeled analog

15.1.1   The  signals  for  the exact   m/z's   being
         monitored  (Table 3A)  shall be  present and
         shall   maximize  within   the   same  two
         consecutive scans.

15.1.2   Either  (1)  the ratio  of  the  background
         corrected  exact SICP areas,  or  (2) the
         corrected   relative   intensities  of  the
         exact m/z's at the GC peak maximum shall
         be within the  limits  in Table 3A.

15.1.3   For  the individual  labeled compounds and
         individual  PCDDs  and  PCDFs,  the  relative
         retention  time  shall  be  within  the  limits
         specified in Table 2.

  15.2   PCDDs and PCDFs having a labeled analog

15.2.1   The  signals  for  the exact   m/z's   being
         monitored  (Table 3)  shall  be present and
         shall   maximize  within   the   same  two
         consecutive scans.

15.2.2   The  ratio  of  the   ion abundances  of the
         exact m/z's at the GC peak maximum shall
         agree within the limits in Table 3.
                                                                                                             27

-------
15.2.3    The relative  retention  time between  the
          native compound  and  its  labeled  analog
          shall  be within  the windows  specified  in
          Table  2.

   15.3    If   identification   is   ambiguous,    an
          experienced spectrometrist   (Section  1.5)
          is to  determine the presence or absence of
          the compound.

     16    QUANTITATIVE DETERMINATION

   16.1    Isotope  dilution  --  By  adding  a  known
          amount of   a   labeled  compound  to  every
          sample prior to extraction,  correction for
          recovery  of  the  native  compound  can  be
          made because the native compound  and its
          labeled  analog  exhibit  the  same  effects
          upon  extraction,   concentration,  and  gas
          chromatography.     Relative   response  (RR)
          values for  sample  mixtures  are  used  in
          conjunction    with    calibration    data
          described  in   Section  7.5  to  determine
          concentrations    directly,    so   long   as
          labeled  compound   spiking   levels   are
          constant.

16.1.1    Because  of  a  potential  interference,  the
          labeled analog  of OCOF is not added to the
          sample.  Therefore, this native analyte is
          quant itated against the labeled OCDD.

16.1.2    Because the labeled analog of 1,2,3,7,8,9-
          HxCDD   is  used  as an  internal  standard
          (i.e., not added before  extraction of the
          sample),  it cannot be used  to quantitate
          the  native  compound.     Therefore,   the
          native  1,2,3,7,8,9-HxCDD  is  quantitated
          using  the avtrage  of  the responses of the
          labeled analogs of  the  other two 2,3,7,8-
          substituted HxCOD's, 1,2,3,4,7,8-HxCDD and
          1,2,3,6,7,8-HxCDD.

16.1.3    Any   peaks    representing    non-2,3,7,8-
          substituted   dioxins    or    furans   are
          quantitated  using  an  average   of   the
          response  factors  from  all  the  labeled
          2,3,7,8-   isomers  in   the same  level  of
          chlorination.

   16.2    Internal     standard   --    Compute   the
          concentrations  of  the  labeled analogs and
          the cleanup standard  in  the extract using
          the   response   factors   determined  from
          calibration data  (Section   7.6)  and  the
          following equation:
                                 where  C    i s
                                 compound  in
          Cex (ng/ml)  =
(A
Cis>
                          16.3
                          16.4
                        16.4.1
                        16.4.2
                        16.4.3
                          16.5
                        16.5.1
                                                       the  concentration  of   the
                                                     the  extract  and  the other
                                       terms are as defined in Section 7.6.1.
                                       The concentration  of  the  native compound
                                       in  the  solid  phase  of  the  sample   is
                                       computed  using the  concentration  of  the
                                       compound  in  the extract  and the weight  of
                                       the solids (Section 10), as follows:
                                 Concentration
                                 in solid (ng/kg)
                                                             (Cex x Vex>
                                 where,

                                   * is
                                 U  is the sample weight in Kg.
                                       V   is the extract volume in ml.
                                        cx
 If  the SICP  area  at the  quant i tat ion m/z
 for  any  compound  exceeds  the  calibration
 range  of  the  system,  a  smaller  sample
 aliquot  is extracted.

 For  aqueous  samples containing  one percent
 solids or  less,  dilute  100 ml,  10  ml,
 etc.,  of sample  to  1  liter with  reagent
 water  and extract  per Section 11.

 For  samples  containing  greater than  one
 percent   solids,   extract  an   amount  of
 sample equal  to  1/10,  1/100,  etc of  the
 amount determined in 10.1.3.   Extract per
 Section  10.4.

 If   a  smaller  sample  size  will  not  be
 representative  of   the   entire   sample,
 dilute the sample  extract by a  factor of
 10,   adjust   the   concentration   of   the
 instrument  internal standard to  100  pg/uL
 in  the extract, and  analyze  an aliquot of
 this  diluted  extract   by  the  internal
 standard method.

 Results  are  reported  to  three  significant
 figures  for  the native and labeled isomers
 found   in   all  standards,  blanks,   and
 samples.   For  aqueous samples,  the  units
 are   ng/L;    for   samples   containing   one
 percent    or   greater    solids   (soils,
 sediments,   filter,  cake,  compost),   the
-units  are ng/kg,  based on the  dry weight
 of  the sample.

 Results   for   samples  which   have   been
 diluted  are  reported at  the  least dilute
 level    at    which   the   area   at   the
 quantitat ion m/z is within the calibration
 range  (Section 16.4).
                             (A-s x RF)
28

-------
16.5.2   For  native  compounds  having  a  labeled
         analog, results are  reported at the least
         dilute  level  at  which  the  area at  the
         quant itat ion m/z is within the calibration
         range  (Section  16.4)   and  the  labeled
         compound  recovery  is  within  the  normal
         range for the method (Section 17.4).

16.5.3   Additionally,  the  total concentrations of
         all  isomers  in an   individual   level  of
         chlorination   (i.e.   total   TCDD,   total
         PeCOD,   etc.)   are   reported   to   three
         significant  figures  in  units of ng/L, for
         both  dioxins  and  furans.   The  total  or
         ng/kg  concentration   in  each  level  of
         chlorination    is    the    sum   of    the
         concentrations  of  all  isomers  identified
         in that  level, including any non-2,3,7,8-
         substituted  isomers.

    17  'ANALYSIS OF COMPLEX SAMPLES

  17.1   Some samples may  contain high levels (>10
         ng/L;  >1000 ng/kg)  of  the  compounds  of
         interest,  interfering  compounds,  and/or
         polymeric materials.    Some extracts will
         not  concentrate to  10  uL  (Section 11);
         others may  overload  the  GC col urn and/or
         mass spectrometer.

  17.2  i Analyze  a smaller aliquot  of  the sample
        , (Section  16.4) when  the  extract will not
         concentrate  to 20  uL  after  all  cleanup
        ' procedures have been exhausted.

  17.3   Interferences  at the primary m/z -- If an
         interference   occurs   at    the   primary
         quantitat ion m/z  (Table 3) for any native
         or labeled  compound,  the alternate m/z is
         used for quantitat ion.

  17.4   Recovery   of   labeled   compound  spiking
         standards  --  In most  samples,  recoveries
         of the  labeled compound spiking  standards
         will  be  similar  to  those  from reagent
         water   or   from   the   alternate  matrix
         (Section  6.6).   If recovery  is outside of
        1 the  limits  given  in  Table  7,  a diluted
         sample (Section 16.4) is analyzed.   If the
         recoveries of  the  labeled  compound spiking
         standards   in   the   diluted  sample  are
         outside  of   the  limits  (per the criteria
         above),   then   the   verification  standard
         (Section  14.3)   shall   be   analyzed  and
         calibration  verified  (Section 14.3.4).  If
         the calibration cannot  be verified,  a new
        . calibration  must  be   performed  and  the
         original  sample extract  reanalyzed.   If
         the   calibration   is   verified  and  the
         diluted  sample does  not  meet  the  limits
            for  labeled  compound recovery,  then  the
            method  does  not apply to the  sample  being
            analyzed   and  the   result   may  not   be
            reported    for    regulatory    compliance
            purposes.

      18    METHOD  PERFORMANCE

            EPA   is  in  the  process   of  developing
            performance  data  for this  draft  method.
            When   these   tests   are   complete,    the
            specifications   in   this  method  will   be
            modified  based  on  these  data,  and  the
            supporting documents will be referenced in
            this  section.
REFERENCES
       1    Tondeur,  Yves,  "Method  8290:  Analytical
            Procedures   and   Quality  Assurance   for
            Multimedia   Analysis  of   Pol/chlorinated
            Oibenzo-p-dioxins   and  Dibenzofurans   by
            High-Resolution  Gas  Chromatography/High-
            Resolution   Mass   Spectrometry,"   USEPA,
            EMSL-Las  Vegas,  Nevada,  June  1987.

       2    "Measurement   of  2,3,7,8-Tetrachlorinated
            Dibenzo-p-dioxin   (TCDD)   and   2,3,7,8-
            Tetrachlorinated  Dibenzofuran  (TCDF)   in
            Pulp,   Sludges,   Process   Samples   and
            Uastewaters  from  Pulp  and Paper  Mills",
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            45435,  June  1988.

       3    "NCASI  Procedures  for the  Preparation  and
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            10016,  Technical  Bulletin No.  551,  Pre-
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       4    "Analytical    Procedures    and    Quality
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            55804,  April  1988.

       5    Yves Tondeur, "Proposed  GC/MS Methodology
            for the  Analysis   of PCDDs  and PCDFs  in
            Special  Analytical   Services   Samples",
            Triangle    Laboratories,   Inc.,    801-10
            Capitola  Dr,  Research Triangle  Park   NC
            27713,  January  1988; updated  by  personal
            communication September  1988.
                                                                                                           29

-------
 6   Lamparski,   L.L.,   and   Nestrick,   T.J.,
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 7   Lamparski,   L.L.,   and   Nestrick,   T.J.,
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 8   Patterson,  D.G.,  et.   al.  "Control   of
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 9   Stanley,  John  S.,  and  Sack,  Thomas  M.,
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     Resolution     Gas     Chromatography/High-
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     Environmental      Monitoring      Systems
     Laboratory,  Las Vegas    NV   89114,  EPA
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10   "Working  with  Carcinogens,"  DHEW,  PHS,
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11   "OSHA Safety and Health Standards, General
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     1976).

12   "Safety      in     Academic     Chemistry
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     Safety (1979).

13   "Standard  Methods  for  the  Examination of
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     Association,    1015     15th    St,    N.W.,
     Washington     DC     20005,  Section  108
     "Safety", 46 (1985).

14   "Method 613  --  2,3,7,8-Tetrachlorodibenzo-
     p-dioxin",  40  CFR  136  (49  FR  43234),
     October 26, 1984, Section 4.1.

15   Provost,    L.P.,    and    Elder,    R.S.,
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     "American Laboratory",  15,  56-83 (1983).

16   "Handbook of Analytical Quality  Control in
     Water and Wastewater Laboratories," USEPA,
     EMSL,  Cincinnati,  OH 45268, EPA-600/4-79-
     019 (March  1979).
                                                              17    "Standard  Practice  for  Sampling  Water,"
                                                                    ASTM   Annual   Book  of   Standards,   ASTM,
                                                                    Philadelphia.  PA,  76 (1980).

                                                              18    "Methods   330.4   and  330.5   for   Total
                                                                    Residual     Chlorine,"    USEPA,     EMSL,
                                                                    Cincinnati,  OH   45268,   EPA  600/4-70-020
                                                                    (March 1979).
30

-------
                                                    Table  1
        POLYCHLORIMATED DIBENZODIOXINS AND FURANS DETERMINED BY ISOTOPE DILUTION AND INTERNAL STANDARD
               HIGH RESOLUTION GAS CHROMATOGRAPHY (HRGO/HIGH RESOLUTION MASS SPECTROMETRY (HRMS)
PCDDs/PCOFs (1)
I somer/Congener
2,3,7.8-TCDD

Total -TCDD
2,3.7,8-TCDF
Total -TCOF
1,2,3,7,8-PeCDD
Total-PeCDD
1,2,3,7,8-PeCDF
2,3,4.7.8-PeCDF
Total-PeCDF
1,2,3,4,7,8-HxCDD
1.2,3,6,7,8-HxCDD
1,2,3, 7,8, 9-HXCDD
Total-HxCDD
1,2, 3,4. 7,8- HxCDF
1,2.3,6,7,8-HxCDF
1,2,3,7,8,9-HxCDF
2,3,4,6,7,8-HxCDF

Total -HxCDF
1,2,3,4,6,7,8-HpCDD
Total -HpCOD
1, 2,3,4,6, 7,8-HpCOF
1,2,3,4,7,8,9-HpCDF
Total-HpCDF
OCDD
OCDF
(1) Polychlorinated dioxins

CAS Registry
1746-01-6

41903-57-5
51207-31-9
55722-27-5
40321-76-4
36088-22-9
57117-41-6
57117-31-4
30402-15-4
39227-28-6
57653-85-7
19408-74-3
34465-4608
70648-26-9
57117-44-9
72918-21-9
60851-34-5

35822-46-9
37871-00-4
67562-39-4
55673-89-7
38998-75-3
3268-87-9
39001-02-0
and furans
TCDD = Tetrachlorodibenzo-p-dioxin
PeCDD - Pentachlorodibenzo-p-dioxin
HxCDD - Hexachlorodibenzo-p-dioxin
HpCDD = Heptachlorodibenzo-p-dioxin
OCDD = Octachlorodibenzo-p-dioxin


Labeled Analog CAS Registry
"c12-2,3,7,8-TCDD
37Cl4-2,3',7,8-TCDD

13C12-2,3,7,8-TCDF

13C12-1,2,3,7,8-PeCDD

13C12-1,2,3,7,8-PeCDF
13C12-2,3,4,7.8-PeCDF

13C12-1,2,3,4,7,8-HxCDD
13C12-1,2,3,6,7,8-HxCDD
13C12-1,2,3,7,8,9-HxCDD(2)

13C12-1,2,3,4,7,8-HxCDF
13C12-1,2,3,6,7,8-HxCDF
13C12-1,2,3,7,8,9-HxCDF
13C,,-2,3,4,6,7,8-HxCDF


13C12-1,2,3,4,6,7,8-HpCDD

13C12-1,2,3.4,6,7,8-HpCDF
13C12-1,2,3,4,7.8,9-HpCDF

13C12-OCDD


TCDF  Tetrachlorodibenzofuran
PeCDF = Pentachlorodibenzofuran
HxCDF = Hexachlorodibenzofuran
MpCDF = Heptachlorodibenzofuran
OCDF - Octachlorodibenzofuran
76523-40-5
85508-50-5

89059-46-1

109719-79-1

109719-77-9
116843-02-8

109719-80-4
109719-81-5
109719-82-6

114423-98-2
116843-03-9
116843-04-0
116843-05-1

109719-83-7

109719-84-8
109719-94-0

114423-97-1







(2)  Labeled analog is used as  an internal standard and therefore cannot be used for quant itat ion by isotope
     dilution.
                                                                                                            31

-------
                                                   Table 2
                           RETENTION TIMES AND MINIMUM LEVELS FOR PCDDs AND PCDFs
Compound
Compounds using C.2-1,2,3
Native Compounds
2,3,7,8-TCDF
2,3,7,8-TCDD
1,2,3,7,8-PeCDF
2,3,4,7,8-PeCDF
1,2,3,7,8-PeCDD
Labeled Compounds
13C12-2,3,7,8-TCDF
13C12-1,2,3,4-TCDD
13C12-2,3,7,8-TCDD
37Cl4-2,3,7,8-TCDD
13C12-1,2,3,7,8-PeCDF
13C12-2.3,4,7,8-PeCDF
13C12-1,2,3,7,8-PeCDD
Compounds using C.p-1,2,3
Native Compounds
1,2,3,4,7,8-HxCDF
1,2,3,6,7,8-HxCDF
2,3,4,6,7,8-HxCDF
1,2,3,4,7,8-HxCDD
1,2,3,6,7,8-HxCDD
1.2,3,7,8,9-HxCDD
1,2,3,7,8,9-HxCDF
1,2,3,4,6,7,8-HpCDF
1,2,3,4,6,7,8-HpCDD
1,2,3,4,7,8,9-HpCDF
OCDD
OCDF
Labeled Compounds
13C,, -1,2, 3. 4. 7,8- HxCDF
17
C12-1,2,3,6,7,8-HxCDF
13C,,-1,2,3,4,7,8-HxCDD
13
IJC12-1,2,3,6,7,8-HxCDD
13C,.,-1,2,3f7,8t9-HxCDD
17
"c,,-!, 2, 3, 7,8,9- HxCDF
13
C,,-1,2,3,4,6.7,8-HpCDF
12 ,,,,., HV
IJC12-1,2,3,4,6.7.8-HpCDD
13C.,-1,2f3,4,7,8,9-HpCDF
C -OCDD
13cJ2-OCDF
Absolute
Retention
Time
(Minutes)
Minimum Level (2)
Retention
Time
Reference
Relative
Retention
Time (1)
Water
P9/U
ppq
Solid
ng/kg
PPt
Extract
pg/uL
ppb
,4-TCDD as internal standard

26.35
27.24
31.16
32.16
32.45

26.35
27.03
27.22
27.23
31.16
32.15
32.44
,7,8,9-HxCDD

36.19
36.29
37.19
37.30
37.36
38.07
38.23
40.55
42.27
43.01
46.56
47.05

36.18
36.27
37.29
37.38
38.06
38.23
40.54
42.27
43.01
46.55
47.04

3C12-2,3,7,8-TCDF
,C12-2,3,7,8-TCDD
"c12-1,2,3,7.8-PeCDF
"c 12-2,3,4,7,8-PeCDF
'3C12-1,2,3,7,8-PeCDD

13C12-1,2,3,4-TCDD
13C12-1,2,3,4-TCDD
13C12-1,2,3,4-TCDD
13C12-1.2,3,4-TCDD
13C12-1,2,3,4-TCDF
13C12-1,2,3.4-TCDD
13C12-1,2,3,4-TCDD
as internal standard

]3C -1,2,3,4,7,8-HxCDF
^C12-1,2.3,6,7.8-HxCDF
C12-2, 3,4, 6,7,8- HxCDF
]3C12- ,2,3,4,7,8-HxCDD
3C.2- ,2,3,6,7,8-HxCDD
"C12- ,2,3,6,7.8-HxCDD
"C.2- ,2,3,7,8,9-HxCDF
C12- ,2,3,4,6,7,8-HpCDF
3C.2- ,2,3,4,6,7,8-HpCDD
3C -1,2,3,4,7,8,9-HpCDF
]3C12-OCDD
C12-OCDD

13C,,-1,2.3,7,8,9-HxCDD
17
aC12-1,2,3,7,8.9-HxCDD
13C..,-1,2,3,7,8.9-HxCOD
13
C12-1,2,3,7,8,9-HxCDD
"c,--!, 2,3, 7,8,9- HxCOD
17
JC...-1,2,3.7,8,9-HxCDD
13
0C,,-1,2,3,7,8,9-HxCDD
13
C12-1,2,3,7,8,9-HxCDD
13C,,-1,2,3,7,8,9-HxCDD
IT F  * f 1
13C,,-1,2,3,7,8,9-HxCDD
., 12 ' '
C12-1,2,3,7.8,9-HxCDD

0.999 -
0.999 -
0.999 -
0.999 -
0.999 -

0.970 -
1.000 -
1.002 -
1.002 -
1.147 -
1.183 -
1.194 -


0.999 -
0.999 -
0.999 -
0.999 -
0.999 -
0.999 -
0.999 -
0.999 -
0.999 -
0.999 -
0.999 -
1.007 -

0.946 -
0.948 -
0.975 -
0.977 -
1.000 -
0.999 -
1.060 -
1.105 -
1.124 -
1.217 -
1.229 -

1.001
1.001
1.001
1.001
1.001

0.980
1.000
1.012
1.013
1.159
1.196
1.206


1.001
1.001
1.001
1.001
1.001
1.001
1.001
1.001
1.001
1.001
1.001
1.013

0.956
0.958
0.985
0.987
1.000
1.010
1.071
1.116
1.136
1.230
1.242

10
10
50
50
50










50
50
50
50
50
50
50
50
50
50
100
100













1
1
5
5
5










5
5
5
5
5
5
5
5
5
5
10
10













0.5
0.5
2.5
2.5
2.5










2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
5.0
5.0












(1)  Initial  specifications  are estimated  based on  isotope dilution and internal standard data from Method
     These specifications  may be revised when  further data have been collected by EPA using Method 1613.

(2)  Level at which  the analytical  system  will give acceptable SICP and calibration.
                                                                                                         1625.
32

-------
                                     Table 3
DESCRIPTORS,  MASSES. M/Z TYPES, AND ELEMENTAL COMPOSITIONS OF THE CDDs AND CDFs (1)
Descriptor
Number
1











1



2










3










Accurate
m/z (2)
292.9825
303.9016
305.8987

315.9419
317.9389
319.8965

321.8936

327.8847
330.9792
331.9368
333.9339
375.8364

339.8597

341.8567
351.9000
353.8970
354.9792
355.8546
357.8516
367.8949
369.8919
409.7974
373.8208
375.8178
383.8639
385.8610
389.8157
391.8127
392.9760
401.8559
403.8529
430.9729
445.7555
m/z
Type
Lock
M
M+2

M
M+2
M

M+2

M
QC
M
M+2
M+2

M+2

M+4
M+2
M+4
Lock
M+2
M+4
M+2
M+4
M+2
M+2
M+4
M
M+2
M+2
M+4
Lock
M+2
H+4
QC
M+4
Elemental Composition
C7F11
C12 H4 ^U 
C,, H. 37Cl. 0
12 4 4
13C,, H. 35Cl, 0
12 4 4
13 35 . 37 .
C12 H4 Cl, Cl 0
C,_ H. 35Cl. 0,
12 4 42
C,- H. 35Cl, 37Cl 0-
12 4 3 2
C12 H4 ^U 2
C7F13
13C12 H4 35C14 02
13. , 35 37
C12 H4 Cl, Cl 02
C,, H. 35CU 37Cl 0
12 4 5
C., H, 35Cl, 37Cl 0
12 3 4
C,, H, 35C13 37C12 0
13 35.. 37
C12 H3 C14 Cl 0
13C H 35cl 37cl Q
C9F13
C12 H3 35cl4 37cl 2
C12 H3 35cl3 37cl2 2
13 35 . 37..
C12 H3 C14 Cl 2
13. H 35 37
C12 H3 C13 C12 2
C12 H3 35cl6 3/Cl 
C12 H2 35C15 37Cl 0
C12 H2 ^U 37cl2 
13C12 H2 35C16 0
13C12 H, 35C15 37Cl 0
C12 H2 35cl5 3?Cl 2
C H 35Cl 37Cl 0
C12 H2 C14 C12 2
C9F15
13C H 35cl 37cl 0
13C H 35cl 37cl
C9F13

Compound
(3)
PFK
TCDF
TCDF

TCDF(4)
TCDFC4)
TCDD

TCDD

TCDOC4)
PFK
TCDD<4)
TCDD(4)
HxCDPE

PeCDF

PeCDF
PeCDF(4)
PeCDF(4)
PFK
PeCDD
PeCDD
PeCDD(4)
PeCDD(4)
HpCDPE
HxCDF
HxCDF
HxCDF(4)
HxCDF(4)
HxCDD
HxCDD
PFK
HxCDD(4)
HxCDD(4)
PFK
OCDPE
Primary
m/z?

Yes


Yes

Yes





Yes



Yes


Yes


Yes

Yes


Yes

Yes

Yes


Yes



                                                                                             33

-------
DESCRIPTORS, MASSES,  M/Z
       Table 3 (continued)
TYPES, AND ELEMENTAL COMPOSITIONS OF
                                                                           THE CDDs AND CDFs  (1)
Descriptor Accurate m/z
Number m/z (2) Type
4 407.7818 M+2

409.7789 M+4
417.8253 M
419.8220 M+2
423.7766 M+2
425.7737 M+4
430.9729 Lock

435.8169 M+2
437.8140 M+4
479.7165 M+4
5 441.7428 M+2
442.9728 Lock
443.7399 M+4
457.7377 M+2
459.7348 M+4

469.7779 M+2
471.7750 M+4
513.6775 M+4
(1) From Reference 5
(2) Nuclidic masses used:
H = 1.007825 C = 12.00000
0 = 15.994915 35Cl = 34.968853
(3) Compound abbreviations:
Chlorinated dibenzo-p-dioxins
TCDD = Tetrachlorodibenzo-p-dioxin
PeCDD = Pentachlorodibenzo-p-dioxin
HxCDD = Hexachlorodibenzo-p-dioxin
HpCDD = Heptachlorodibenzo-p-dioxin
OCDD = Octachlorodibenzo-p-dioxin

Elemental Composition
C,, H 35Cl., 37Cl 0
l O
35 37
C12 H 03C15 3 C12 0
13C H 35Cl 0
C12 H C17 
13C,, H 35Cl, 37Cl 0
l O
C12 H 35C16 37Cl O,
C12 H 35C15 37C12 02
C9F17
13 35 37
C12 H C16 Ct 2
13 35 . 37
C12 H C15 C12 2
C12 H 35C17 37C12 0
c12 35ci7 37ci o
C10 F17
C12 35cl6 3/Cl2 
c,2 35ci7 37ci o.
c,, 35ci., 37ci, o,
12 6 22
13C12 35C17 37Cl O,

C12 35cl8 37cl2 


13C = 13.003355
37Cl = 36.965903

Compound
(3)
HpCDF

HpCDF
HpCDF(4)
HpCDF(4)
HpCDD
HpCDD
PFK

HpCDD (4)
HpCDD(4)
NCDPE
OCDF
PFK
OCDF
OCDD
OCDD

OCDD (4)
OCDD (4)
DCDPE


F = 18.9984


Primary
m/z?
Yes


Yes

Yes



Yes


Yes


Yes


Yes







Chlorinated diphenyl ethers
HxCDPE =
HpCDPE =
OCDPE
NCDPE =
DCDPE =
Hexachlorodiphenyl ether
Heptachlorodiphenyl ether
Octachlorodiphenyl ether
Nonachlorodiphenyl ether
Decachlorodiphenyl ether





     Chlorinated dibenzofurans
        TCDF     =    Tetrachlorodibenzofuran
        PeCDF    =    Pentachlorodibenzofuran
        HxCDF    =    Hexachlorodibenzofuran
        HpCDF    =    Heptachlorodibenzofuran

(4)  Labeled compound
                   Lock mass and QC compound
                      PFK     =    Perfluorokerosene
 34

-------
                        Table 3A
       THEORETICAL M/Z RATIOS AND CONTROL  LIMITS
No. of
Chlorine
Atoms
4
5
6
6 (2)
7
7 (3)
8
m/z's
Forming
Ratio
M/M+2
M+2/M+4
H+2/M+4
M/M+2
M+2/M+4
M/H+2
M+2/M+4
Theoretical
Ratio
0.77
1.55
1.24
0.51
1.05
0.44
0.89
Control
Lower
0.65
1.32
1.05
0.43
0.88
0.37
0.76
Limitsd)
Upper
0.89
1.78
1.43
0.59
1.20
0.51
1.02
(1)  Represent   15X windows around the theoretical ion
     abundance  ratios.
(2)  Used for 13C-HxCDF only.
(3)  Used for 13C-HpCDF only.
                                                                                                            35

-------
                                                    Table 4
                  CONCENTRATIONS OF SOLUTIONS CONTAINING LABELED AND UNLABELED CDDS AND CDFS
Stock
Solution
(1)
Compound ng/mL
Native CDDs and COFs
2,3,7,8-TCDD
2,3,7.8-TCDF
1,2,3,7,8-PeCDD
1,2,3,7,8-PeCDF
2,3,4,7,8-PeCOF
1,2,3,4,7,8-HxCDD
1,2,3,6,7,8-HxCDD
1,2,3.7.8,9-HxCDD
1,2,3,4,7,8-HxCDF
1,2,3,6,7,8-HxCDF
1,2,3,7.8,9-HxCDF
2,3,4,6,7,8-HxCDF
1.2,3.4.6,7,8-HpCOD
1,2,3,4,6,7,8-HpCDF
1,2,3,4,7.8,9-HpCDF
OCDD
OCOF
Labeled Compound Spiking Standards
13C12-2,3,7,8-TCDD
13C12-2.3.7.8-TCOF
13C12-1.2.3,7,8-PeCDD
13C12-1,2,3.7,8-PeCOF
13C12-2,3,4.7.8-PeCDF
13C,,-1,2.3.4.7,8-HxCOD
 12 ' ' ' '
13C12-1,2.3,6,7,8-HxCDD
13C,,-1,2,3,4.7,8-HxCDF
13 12
'>C,,-1,2,3,6.7,8-HxCOF

"c12-1,2,3,7.8.9-HxCOF
13C,--2,3.4.6,7,8-HxCDF

13C12-1, 2.3,4,6, 7,8-HpCOO
13C,,-1.2,3,4,6.7.8-HpCDF

l3C12-1,2.3,4,7,8,9-HpCOF
13
C12-OCDD

-
-
-
-
-
.
-
-
.
-
-
.
-
.
-
-
-

100
100
100
100
100
100

100
100
100
100
100
100
100
100
200
Spike
Solutions
(2)
ng/mL

-
-
-
-
-
.
-
-
.
-
-
.
-
.
-
-
-

2
2
2
2
2
2

2
2
2
2
2
2
2
2
4
Calibration and Verification Solutions
CS1

0.5
0.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
5.0
5.0

100
100
100
100
100
100

100
100
100
100
100
100
100
100
200
CS2

2
2
10
10
10
10
10
10
10
10
10
10
10
10
10
20
20

100
100
100
100
100
100

100
100
100
100
100
100
100
100
200
ng/mL
VER(3)
CS3 CS4

10
10
50
50
50
50
50
50
50
50
50
50
50
50
50
100
100

100
100
100
100
100
100

100
100
100
100
100
100
100
100
200

40
40
200
200
200
200
200
200
200
200
200
200
200
200
200
400
400

100
100
100
100
100
100

100
100
100
100
100
100
100
100
200
CSS

200
200
1000
1000
1000
1000
1000
1000
1000
1000
1000
1000
1000
1000
1000
2000
2000

100
100
100
100
100
100

100
100
100
100
100
100
100
100
200
PAR(4)
ng/mL

40
40
200
200
200
200
200
200
200
200
200
200
200
200
200
400
400

-
-
-
-
-
.

-
-
-
-
-
-
-
-
-
Cleanup Standard
       37,
         Cl4-2,3,7,8-TCOD
Internal Standards
       13,
  13,
         C12-1.2.3.4-TCDD
   JC12-1.2.3.7.8,9-HxCDD
0.8


200
200
0.5

100
100
100
100
 10

100
100
 40

100
100
200

100
100
(1)  Stock solution (Section 6.10}  -  Prepared  in  nonane,  and diluted daily with acetone to prepare the spiking
     solution (Section 10.3.2).
(2)  Spiking solutions (Sections 6.11, 6.12,  8.3,  10.3.2,  and 10.4.2).
     Calibration verification solution (Section 14.3).
     Precision and  recovery standard  (Section  6.14)  - Prepared in  nonane,  and diluted daily with acetone  to
     prepare the spiking solution (Section 10.3.4).
(3)
(4)
  36

-------
                        Table 5
  GC  RETENTION TIME WINDOW DEFINING MIXTURES AND ISOMER
                SPECIFICITY TEST MIXTURES

DB-5 Column GC Retention Time Window Defining  Standard
(Section 6.15)

Congener	First Eluted	Last Eluted
TCDF
TCOD
PeCDF
PeCDD
HxXCDF
HxCDD
HpCOF
HpCDD
1,3,6,8-
1,3.6,8-
1.3.4,6.8-
1,2,4,7,9-
1,2,3,4,6,8-
1,2,4,6,7,9-
1.2,3.4,6,7,8-
1.2,3,4,6,7,9-
1,2,8,9-
1,2,8,9-
1,2,3,8,9-
1,2,3,8.9-
1,2,3,4,8,9-
1,2,3,4,6,7-
1.2,3,4,7.8,9-
1,2,3,4.6.7.8-
DB-5 TCDD Isomer Specificity Test Standard
(Section 6.16.1)
                  1.2,3.4-TCDD          1,2.3.7-TCOD
                  1,2,7,8-TCDD          1,2,3,8-TCDD
                  1,4,7,8-TCDD          2,3,7,8-TCDD

DB-225 Column TCDF Isomer Specificity Test Standard
(Section 6.16.2)
                  2,3,4,7-TCDF
                  2,3,7.8-TCDF
                  1,2,3,9-TCDF
                                                                                                             37

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                                                    Table 6
                          REFERENCE COMPOUNDS FOR NATIVE AND LABELED PCDDS AND PCDFS
                                                           Labeled PCDDs and PCDFs
Native PCDDs and PCDFs
2,3.7,8-TCDD
2,3.7,8-TCDF
1,2,3,7,8-PeCDD
1,2.3,7,8-PeCDF
2,3,4,7,8-PeCDF
1,2,3,4,7,8-HxCDD
1,2,3,6,7.8-HxCDD
1,2,3,7.8.9-HxCDD
1,2,3,4,7,8-HxCDF
1,2,3,6,7,8-HxCDF
1,2,3,7,8,9-HxCDF
2.3,4,6,7,8-HxCDF
1. 2,3,4, 6,7,8-HpCDD
1,2,3,4,6,7,8-HpCDF
1,2,3.4.7,8,9-HpCDF
OCDD
OCDF
Reference Compound
13C12-2,3.7,8-TCDD
13C12-2,3,7,8-TCDF
13C12-1,2,3,7,8-PeCDD
13C12-1,2,3,7,8-PeCDF
13C12-2,3,4,7.8-PeCDF
13C12-1,2,3,4,7,8-HxCDD
13C12-1,2,3,6,7,8-HxCDD
13C12-1.2,3,7,8,9-HxCDD
13C12-1, 2,3,4, 7,8-HxCDF
13C12-1,2,3,6,7,8-HxCDF
13C12-1,2,3,7,8,9-HxCDF
13C12-2,3,4,6,7,8-HxCDF
13C12-1,2,3,4,6,7,8-HpCDD
13C12-1,2,3,4.6,7,8-HpCDF
13C..,-1,2,3,4,7,8,9-HpCDF
12 ,,,,,, H*.
"C , -OCDD
13
13C12-OCDD
Reference Compound
C,,-2, 3, 7,8-TCDD
,, 12
C12-2.3,7,8-TCDF
13C1,-1,2,3,7,8-PeCDD
13 '
1:SC12-1,2,3,7,8-PeCDF
13
'C12-2,3,4,7,8-PeCDF
13C..,-1,2,3,4,7,8-HxCDD
13 12
C-.-I^.S^^.S-HxCDD
15 12 ' ' '
C12-1,2,3.7,8,9-HxCDD
13C,,-1,2,3,4,7,8-HxCDF
13 ''
C,--1,2,3,6,7,8-HxCDF
 12
'C12-1,2,3,7,8,9-HxCDF
13C12"2,3,4,6,7,8-HxCDF
13C12-1,2,3,4,6,7,8-HPCDD
13 '*
C.,-1,2,3,4,6,7,8-HpCDF
13 12 ^
C,--1f2,3,4,7,8,9-HpCDF

13C,,-OCDO
37
Cl^-2, 3. 7,8-TCDD
13C,,-1,2,3,4-TCDD
13
C12-1,2,3,4-TCDD
13C 2-1,2,3,4-TCDD
17
IJC12-1,2,3,4-TCDD
13C12-1,2,3.4-TCDD
13C., -1,2, 3, 7, 8,9-HxCDD
IT * * * * *
13C., -1,2, 3, 7, 8,9-HxCDD

5C12-1,2,3,7,8,9-HxCDD
13C1,-1,2,3,7f8,9-HxCDD
13 l<:
C12-1,2,3,7,8,9-HxCDD
13C12-1,2,3,7,8,9-HxCDD
13C12-1,2,3,7,8,9-HxCDD
13C,,-1, 2,3, 7,8,9- HxCDD
13
C12-1,2,3,7,8,9-HxCDD
"c-.-I^.S^.B^-HxCDD
13
C,,-1, 2,3,7,8, 9-HxCDD
13
'C12-1.2,3,4-TCDD
                                                    Table 7
                                 ACCEPTANCE CRITERIA FOR  PERFORMANCE TESTS (1)




PCDDs/PCDFs



Compound
by internal standard
13C-tetra-hepta CDD and CDF


PCDDs/PCDFs

pent a

37Cl-tetra CDD
13C-octa CDD
by isotope dilution
tetra CDD and CDF
- hepta CDD and CDF
octa CDD and CDF
Test
Concen-
t rat ion
(ng/mL)

100
40
200

40
200
400
Initial
Precision
and Accuracy
Sec
s

32
13
64

9
45
90
8.2.3
X

60 -
24 -
120 -

30 -
150 -
300 -



145
58
290

52
260
520
Labeled
Compound
Recovery
Sec 8.3
and 1
P (5

25 -
25 -
25 -

25 -
25 -
25 -

Calibration
Verification
Sec
14.5
'.) (ug/mL)

150
150
150

150
150
150

65
26
130

30
150
300

- 140
- 56
- 280

- 52
- 260
- 520
Ongoing
Accuracy
Sec
R

55
22
110

28
140
280
14.6
(%)

- 150
- 60
- 300

- 56
- 280
- 560
(1)  Based on data  from Method  1625.
 38

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                                                    Table 8
                            SAMPLE PHASE  AND  QUANTITY  EXTRACTED  FOR VARIOUS MATRICES
Sample Matrix (1)
SINGLE PHASE
Aqueous
Solid
Organic
Example

Drinking water
Groundwater
Treated wastewater
Dry soil
Compost
Ash
Waste solvent
Waste oil
Organic polymer
Percent Quantity
Solids Phase Extracted

<1 (2) 1000 mL
>20 Solid 10 g
<1 Organic 10 g
   MULTIPHASE

       Liquid/Solid

        Aqueous/solid
         Organic/solid


       Liquid/Liquid

         Aqueous/organic
        Aqueous/organic/
        sol id
Wet soil
Untreated effluent
Digested municipal sludge
Filter cake
Paper pulp
Tissue

Industrial sludge
Oily waste
In-process effluent
Untreated effluent
Drum waste

Untreated effluent
Drum waste
 1-30
1-100
Solid
Both
                  Organic
                  Organic
                  & solid
10 g
10 g
                    10 g
                                                                                                      10 g
(1)  The exact matrix may be vague for some samples.   In  general,  when the CDDs and CDFs are in contact with  a
     multiphase system in which one of the phases is water,  they will  be preferentially dispersed  in or  adsorbed
     on the alternate phase,  because of their  low solubility in  water.

(2)  Aqueous samples are filtered after spiking with labeled analogs.   The  filtrate and the material  trapped on
     the filter are extracted separately,  and  then the  extracts  are combined for analysis.
                                                                                                             39

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