600485025
    DETERMINATION OF DIPHENYLAMINE  IN  INDUSTRIAL
              AND MUNICIPAL WASTEWATERS
      J.S.  Warner, T.M. Engel and  P.J. Mondron
           Battelle Columbus Laboratories
               Columbus, Ohio 43201
              Contract No. 68-03-2956



                  Project Officer


                  Thomas Pre ssle y
        Physical and Chemical Methods  Branch
   Environmental Monitoring and Support Laboratory
              Cincinnati, Ohio 45263
ENVIRONMENTAL MONITORING AND SUPPORT  LABORATORY
      OFFICE OF RESEARCH AND DEVELOPMENT
     U.S.  ENVIRONMENTAL PROTECTION  AGENCY
           CINCINNATI, OHIO 45268

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                                 DISCLAIMER
The information in this document has been funded wholly or in part by the
United States Environmental Protection Agency under Contract 68-03-2956
to Battelle Columbus Laboratories.  It has been subject to the Agency's
peer and administrative review, and it has been approved for publication
as an EPA document.
                                      ii

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                                   FOREWORD
    Environmental measurements are required to determine the quality of
ambient waters and the character of waste effluents.  The Environmental
Monitoring and Support Laboratory - Cincinnati, conducts research to:

    o  Develop and evaluate methods to measure the presence and
       concentration of physical, chemical, and radiological pollutants in
       water, wastewater, bottom sediments, and solid wastes.

    o  Investigate methods for the concentration, recovery, and
       identification of viruses, bacteria and other microbiological
       organisms in water; and, to determine the responses of aquatic
       organisms to water quality.

    o  Develop and operate an Agency-wide quality assurance program to
       assure standardization and quality control of systems for monitoring
       water and wastewater.

    o  Develop and operate a computerized system for instrument automation
       leading to improved data collection, analysis, and quality control.

    This report is one of a series that  investigates the analytical behavior
of selected pesticides and suggests a suitable test procedure for their
measurement in wastewater.  The method was modeled after existing EPA
methods being specific yet as simplified as possible.
                                       Robert L.  Booth, Acting  Director
                                       Environmental Monitoring and  Support
                                       Laboratory - Cincinnati
                                      111

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                                   ABSTRACT

     A method was developed for the determination of  diphenylamine in
wastewaters.  The method development program  consisted  of  a literature
review; determination of extraction efficiency  for  each compound from water
using methylene chloride; development of  a deactivated  silica  gel cleanup
procedure; and determination of suitable  gas  chromatographic (GC) analysis
conditions.

     The final method was aplied to Columbus  Publicly Owned Treatment Works
(POTW) secondary effluent in order to determine  the precision  and accuracy
of the method.  The wastewater was spiked with  diphenylamine at  levels of
5 ug/L and 50 ug/1*.  Recovery for diphenylamine  at  the  5 ug/L  level was
120 ± 25 percent.  Recovery at the 50 yg/L level was  89 ±  11 percent.  The
method detection limit (MDL) for diphenylamine  in distilled water was
1.6 yg/L (7).  In wastewaters it may be higher  due  to interfering
compounds.

     This report was submitted in partial fulfillment of contract No.
68-03-2956 by Battelle Columbus Laboratories  under  the  sponsorship of the
U.S. Environmental Protection Agency.  This, report  covers  the  period from
February 1, 1982 to April 30, 1983.
                                      IV

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                                CONTENTS

Foreword .............................    ill
Abstract .............................     iv
Figures .............................     vi

     1.  Introduction ......................      1
     2.  Conclusions .......................      2
              Extraction and Concentration ............      2
              Cleanup ......................      2
              Chroma tography. .... ..............      2
              Validation Studies. ... .............      2
     3.  Experimental ......................      3
              Extraction and Concentration ............      3
              Cleanup ......................      3
              Chroma tography ...................      4
              Validation Studies .................      4
     4.  Results and Discussion .................      5
              Extraction and Concentration ............      5
              Cleanup . . ....................      5
              Chroma tography ......... . .........      5
              Validation Studies .................      8

References ........................ ....     10
Appendix
     A.  Diphenylamine Method  620

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                                 FIGURES

 Number                                                           Page

   1     GC-AFD Chromatogram of 100 ng of Diphenylamine
         (Column 1)	          6

.   2     GC-FID Chromatogram of 200 ng of Diphenylamine              _
         (Column 2)	"	          '

   3     Analytical Curve for Diphenylamine	          9
                                    vi

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                                 SECTION 1

                               INTRODUCTION
     Diphenylamine(I) is used as a stabilizer in smokeless powders  (1,2)
as well as to 'control superficial scald in some varieties of pears  and
apples (3,4).
                            oo
                                   I

The CAS registry number for diphenylamine is  122-39-4  and  its  IUPAC  name
is N-phenylbenzeneamine.  It has a melting point of 53-54°C, a boiling
point of 302°C, and an oral LD50 in rats of 300-1000 mg/kg.  Common
synonyms for diphenylamine include "Anilinobenzene", "DFA",  and "DPA".  A
literature review described extractions of diphenylamine from  water  with
methylene chloride (5) and a cleanup procedure using Bio-Beads S,-X2(6).
Several columns were used for determination of diphenylamine by GC
including 32 SP-1000 (5), 152 UC W-98 and 32  OV-17(2),  32  OV-17 with 0.022
Epikote 1001(4), and 102 OV-101, 62 OV-17, and 52 07-225(6).   Detectors
used for GC analyses included a Hall detector (6), a rubidium  bead  alkali
flame detector (AFD) (4,6), and a mass spectrometer (MS) (2,5).
Electron-impact (El) and chemical-ionization  (CI) mass  spectra were  also
reported (1,2).

     Diphenylamine is stable in water at neutral pH, can be  extracted from
water with methylene chloride, and contains nitrogen.   For these reasons,
the selected approach to the determination of diphenylamine  in water
included extraction from water using continuous extraction with methylene
chloride, cleanup using silica gel chromatography, and analysis using
packed column GC-AFD.  Standard concentration techniques using
Kuderna-Danish  (K-D) equipment were used.  The  final method  is included  in
Appendix A of this report.

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                                  SECTION 2

                                 CONCLUSIONS
EXTRACTION  AND  CONCENTRATION

     Diphenylamine can be extracted from water using methylene  chloride
with greater than 90 percent recovery by means of continuous  extraction
techniques.  Use of K-D concentration equipment  to perform extract
concentrations did not significantly affect compound recoveries.

CLEANUP

     Diphenylamine  elutes  from deactivated  silica  gel  in six percent ethyl
ether  in petroleum  ether with  greater  than  90  percent  recovery.  This use
of deactivated  silica  gel  was  an effective  cleanup  procedure for extracts
from a columbus  POTW secondary effluent,  but was not assess  for extracts
from any relevant wastewater samples.

CHROMATOGRAPHY

     Two packed  GC  columns,  3Z SP-2250  and  32  SP-1000, were  found to be
acceptable  for  the  GC-AFD  analysis  of  diphenylamine.   The 3% SP-2250 gave
better peak shape and  was  used as the  primary  column.   The  32 SP-1000
column was  designated  as the  alternate  column.

VALIDATION  STUDIES

     Recoveries  of  diphenylamine from  distilled  water  in the 10 to
1000 ug/L concentration range  were  greater  than  85  percent.   Analytical
curves constructed  from this  data were  linear.   The  MDL in  distilled water
was 1.6 yg/L.   Recoveries  of  diphenylamine  from  Columbus secondary POTW
effluent at the  5 and  50 ug/L levels were  120  ±  25  percent  and 89 ± 11
percent, respectively.

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                                  SECTION  3

                               EXPERIMENTAL
     Studies were performed  to  determine  if  extractions  with  separator/
funnel and continuous extractors,  cleanup by silica gel  adsorption
chromatography, concentration using  K-D equipment,  and analysis  using
packed column GC-AFD would be applicable  techniques for  the determination
of diphenylamine in water.   Since  recovery data  and literature references
indicated that diphenylamine is  relatively stable  in water, stability
studies were not performed.

EXTRACTION AND CONCENTRATION
 *
     Extraction of diphenylamine from water  was  studied  using both
separatory funnel and continuous extraction  techniques.   In both cases one
liter of distilled water  was used.  The sample was  adjusted to pH 7 by
addition of 6N sodium hydroxide  or 6N sulfuric acid.  For the
separatory funnel studies, the  distilled  water was  spiked with
diphenylamine at the 5,  10,  50,  100,  500, and 1000  ug/L  level and
extracted three times with 60 mL. each of  methylene  chloride.   For the
continuous extractor studies the water was spiked  at the 10 and 100 ug/L
levels.and extracted with methylene  chloride overnight.   These studies
were done in duplicate.   The extracts were dried by passing them through
10 cm of anhydrous granular  sodium sulfate,  concentrated to one mL and
analyzed by GC-AFD.

CLEANUP
                                                               •
     Activated silica gel, 20 grams,  was  stirred with 100 mL  of acetone
and 1.2 mL of reagent water  for  30 minutes.   The slurry  was transferred to
a chromatographic column, and the  solvent was allowed to elute and
discarded.  The column, was then  sequentially washed with 20 aL of
methylene chloride and 30 mL of  petroleum ether. Petroleum ether, 50 mL,
was suspended over the silica gel.  Amounts  of either 10 or 100  ug
diphenylamine dissolved  in 5 mL  of methylene chloride was added to the
above mentioned petroleum ether.   This solvent plus an additional 50 mL of
petroleum ether were eluted  from the  column  and  collected (Fl).   Nine
additional 25-mL solvent  elutions  were collected: six percent ethyl ether
in petroleum ether (F2);  15  percent  ethyl ether  in  petroleum  ether (F3);
50 percent ethyl ether in petroleum  ether (F4);  ethyl ether (F5); six
percent acetone in ethyl  ether  (F6);  15 Percent  acetone  in ethyl ether
(F7); 50 percent acetone  in  ethyl  ether (F8); acetone (F9); and six
percent methanol in acetone  (F10).  Each  fraction was concentrated to
approximately 4 mL after  addition  of  2.5  mL  of toluene.   The  fractions
were then transferred to  5-mL volumetric  flasks  and diluted to volume with
toluene.
                                      3

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CHROMATOGRAPHY

     Two columns were evaluated for the determination  of  diphenylamine,  32
SP-2250 on 100/120 mesh Supelcoport and 3Z SP-1000 on  100/120  mesh
Supelcoport.  The 3Z SP-2250 column has a maximum temperature  limit  of
300°C.  The 32 SP-1000 column has a maximum  temperature  limit  of  only 250°
C.

VALIDATION STUDIES

     The HDL for diphenylamine was determined  by analyzing seven  replicate
distilled water samples spiked at the  5  ug/L concentration level  (7).  The
sample extracts were cleaned up using  the  silica gel cleanup procedure
prior to analysis.  The amounts recovered  were determined by external
standard calibration and  the MDL was calculated from these data.


     Distilled water was  also spiked in duplicate at the  10, 50,  100, 500,
and 1000 ug/L concentration levels and recoveries of the  diphenylamine was
determined as described earlier.  An analytical  curve  was  generated  by
plotting the amount spiked into the samples  versus the amount  recovered
from the samples.

     A relevant wastewater was not available for diphenylamine and
Columbus POTW secondary effluent was used  for  wastewater  validation
studies.  Seven replicates of the wastewater were analyzed  to  determine
the background levels.  The wastewater was spiked with diphenylamine at
the 5 and 50 ug/L concentration levels, processed and  analyzed.   Seven
replicate extractions were performed at each concentration  level.

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                                 SECTION 4

                          RESULTS AND DISCUSSION
EXTRACTION AND CONCENTRATION

     Data from separatory funnel extractions of diphenylamine  from reagent
water indicated that recoveries of diphenylamine were unacceptably low
below the 100 \ig/mL conentration level.  Recovery data are given  in Table
TABLE 1.  RECOVERY OF DIPHENYLAMINE FROM WATER USING SEPARATORY
          FUNNEL TECHNIQUES


     Amount Spiked,              Amount Recovered,
          Ug/L                       ug/L

           5                       0.4 ± 0.1 (a)
          10                       2.7 ± 0.3 (b)
          50                      29   ± 2.6 (b)
         100                      94   ± 7.7 (b)
         500                     390   ± 42  (b)

(•a)  Average of seven extractions; second figure is relative
     standard deviation.
(b)  Average of two extractions; second figure is relative range.

Use of continuous extractors improved recoveries of diphenylamine at  lower
concentration levels as demonstrated by the recovery data in Table  2.

TABLE 2.  RECOVERY OF DIPHEYLAMINE FROM WATER USING CONTINUOUS
          EXTRACTION TECHNIQUES
       Amount Spiked,                Amount Recovered,
            Ug/L                          Ug/L
5
10
50
100
500
1000
3.6 ± 0.5 (a)
8.6 ± 1.2 (b)
49 ± 1.7 (b)
95 ± 1.9 (b)
500 ±11 (b)
950 ± 8.0 (b)
(a)  Average of seven extractions; second figure is relative standard
     deviation.
(b)  Average of two extractions; second figure is relative range.

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Use of continuous extractors as opposed  to  separator?  funnels  for
extraction of diphenylamine from water improved  recoveries.   For  this
reason, continuous extractors were used  for all  diphenylamine  method
validation studies.

CLEANUP

     Diphenylamine eluted from deactivated  silica gel  in  fraction  3  (six
percent ethyl ether in petroleum ether).  Recoveries of  10 and 100 yg  of
diphenylamine were 108 and 102 percent,  respectively.

CHROMATOGRAPHY

     Both the 32 SP-2250 and 3% SP-1000  columns  were satisfactory  for  the
GC determination of diphenylamine.  The  32  SP-2250  column, however,  gave
slightly better peak shape and was chosen as  the primary  column.   The
following conditions were used for the columns:
     Column:                1.8m x 2mm ID 32  SP-2250 on  100/120
                            mesh Supelaoport  or  1.8 m  x  2mm  ID 3%
                            SP-1000 on 100/120 mesh Supelcoport
     Detector:              Alkali flame
     Injector Temperature:  280°C
     Detector Temperature:  300 °C
     Oven Temperature:       80aC for 4  minutes; programmed  from
                            80 °C to 300°C at  8 C/minute;  held  at
                            300°C for 4  minutes  (SP-2250  column)
                            80°C for 4 minutes;  programmed  from
                            80°C to 250°C at  88C/minute;  held.
                            250°C for 4  minutes  (SP-1000  column).
     Carrier Gas:.          Helium-at 30 mL/minute
Chromatograms obtained under these conditions are shown  in  Figures 1 and
2.

VALIDATION STUDIES

     Recovery of diphenylamine from distilled water at the  5 ug/L level
was 3.6 ± 0.5 ug/L.  This figure is an average of seven  replicate
analyses.  The HDL in distilled water was calculated  to  be  1.6 ug/L.
Recoveries of diphenylamine from distilled  water at the  10,  50, 100,  500,
and 1000 ug/L levels were 8.6 ± 1.2, 49  ± 1.7, 95 ± 1.9,  500 ± 11 and
950 ± 8.0 Pg/L, respectively.  These data were the  averages  of duplicate
analyses.  The resultant analytical curve is  shown  in  Figure 3.

     Recoveries of diphenylamine from Columbus POTW secondary effluent at
the 5 and 50 ug/L levels were 120 ± 25 percent and  89  ±  11  percent,
respectively.  These data were the averages of seven  replicate analyses.

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           *—i "i"!—r—r~T" ••'*!" t"»—»*t—|—v- T—»—i""f--i—i—r--i—|— i •••-•!-•f
31.8
                                                                                          33.0    33.0
                              RETENTION TIME,  minutes




Figure  i.   OC-AFD Cliromatogram of 100 ng of Diphenylamine (Column  1),

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00
                                                                                    30.0      31.5
3.3. (
                                         RETENTION TIME, minutes
               PlRiire 2.  CC-PID Chroma tog ram of 200  IIR of Diphenylamlne (Column 2).

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                                                               IStolflZ
              loonjo
                   r                                                /*
              900.0-
                   f
              aon.rj"
                   i                                         '•
              700.0.
              600.fr
Amount Recovered,  j-
     Wg/L          L
              snn.o
                   r
              400.0.
                   i
                   !
                   9

              300. (T
200.0
• •'
loo Ji X
\s.\.\.\
UNHJUilll Utef O*l« .IMHi
T OflBDSTi 3>2Z2
3A .28
.1,1,1,1.1,1.1
                                      400.0 SOU) tfO.O  TOO-O  fULa 100.0

                                Amount Spiked, ug/L

                        Figure  3.   Analytical Curve  for Diphenylamine

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                                REFERENCES
1.   Ala, A. Mass Spectra of Diphenyiamine Compounds with Nirto  and
    Nitroso Substituents and of Tetraphenylhydrazine.  Explosivs to ffe,
    17(7):156-164, 1969.

2.   Mach, M.H., A. Polios, and P.F. Jones.  Feasibility of Gunshot
    Residue Detection Via Its Organic Constituents Part I. Analysis
    of Smokeless Powders by Combined Gas Chromatography-Cheaiical
    lonization Mass Spectrometry.  .J. Forensic Sei.,  23(3)-433-445,
    1978.

3.   Luke, B.C., and S.A. Cosseus.  Determination of Diphenylamine
    Residues in Apples.  Bull. Environm. Contam. Toxicol., 24:745-751,
    1980.

4.   Allen, J.G. and K.J. Hall.  Methods  for the Determination
    of Diphenylamine Residues in Apples.  J. Agric Food Chem.,
    28(2):255-258, 1980.

5.   Jungclaus, G.A. , L.M. Games, and R.A.. Kites.   Identification
    of Trace Organics Compounds in Tire  Manufacturing Plant  Wastewatars.
    Analytical Chemistry. 48(13):1894-1896, 1976.

6.   Diachenko, G.W.  Determiation of Several Industrial Aromatic
    Amines in Fish.  Environmental Science and Technology,
    13(3):329-333, 1979.
       *

7.   Glaser, J.A., et al.  "Trace Analysis for Wastewaters",  Environmental
    Science and Technology. 15, 1426 (1981).
                                   10

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              METHOD 620 DETERMINATION OF DIPHENYLAMINE  IN
                 MUNICIPAL AND INDUSTRIAL WASTEWATERS
                         BY GAS CHROMATOGRAPHY

1.  Scope and Application
    1.1  This method covers the determination of diphenylamine CAS  No.
         122-39-4.
    1.2  This is a  gas chromatographic (GC)  method applicable to the
         determination of diphenylamine in municipal  and industrial
         discharges.
    1.3  The method detection limit (MDL, defined in  Section.15)
         for diphenylamine is listed in Table 1.  The  MDL for a specific
         wastewater may differ from those listed, depending upon the
         nature of  interferences in the sample matrix.
    1.4  The sample extraction and concentration steps  in this method
         are similar to those of other 600 series methods.   Thus, a single
         sample may be extracted to measure the compounds included  in the
         scope of the methods.  When cleanup is required, the concentration
         levels must be high enough to permit selecting  aliquots, as
         necessary, in order to apply appropriate cleanup procedures.
    1.5  This method is restrictedto use by or under  the supervision of
         analysts experienced in the use of gas chromatography and  in the
         interpretation of gas chromatograms.  Each analyst must demonstrate
                                      11

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           the ability to  generate  acceptable results with this method using
           the procedure described  in  Section 8.2.
     1.6   When this method  is  used to analyze  unfamiliar samples for^any  or
           all of the compounds above, compound identifications should be  sup-
           ported by at least one additional qualitative technique.  This
           method describes  analytical conditions  for a second gas chrcmato-
           graphic column  that  can  be  used  to confirm measurements made with
           the primary column.   Section 14  provides  gas chromatograph/mass
           spectrometer (QC-MS) criteria appropriate for the qualitative con-
           firmation of compound identifications.
2.   Summary of Method
     2.1 '  A measured volume of sample, approximately 1 liter,  is  solvent
           extracted with  methylene chloride using a continuous extractor.
           The methylene chloride extract is dried and  concentrated  to  5.0 ml.
           Sas chromatographic  conditions are described which permit the sepa-
           ration and measurement of the compounds in the extract  by alkali
           flame detector  (AFD) gas chromatography.
     2.2   This method provides an  optional silica gel  column cleanup proce-
           dure to aid in  the elimination of  interferences  which may be
           encountered.
3.   Interferences
     3.1   Method interferences may be caused by contaminants in  solvents,
           reagents, glassware, and other sample processing apparatus  that
           lead to discrete  artifacts or elevated baselines in gas  chro-
           matograms.  All reagents and apparatus must  be routinely demon-
           strated to be free from  interferences under  the  conditions  of  the
                                        12

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      analysis  by running  laboratory  reagent  blanks  as described  in
      Section  8.5.
                                                    n
      3.1.1   Glassware must  be  scrupulously  cleaned.   Clean  all  glass-
              ware as soon as possible  after  use  by  thoroughly rinsing
              with the last solvent used  in  it.   Follow by washing with
              hot water and detergent and thorough rinsing with tap  and
              reagent water.   Drain dry,  and  heat in an oven or muffle
              furnace at 400*0 for  15 to  30 min.  Thermally stable
              materials such  as  PCBs  miqht not be eliminated by this treatment
              Thorough rinsing with acetone  and  pesticide quality hexane
              may be substituted for  the  heating. After  drying and  cool-
              ing, seal and store glassware  in  a clean  environment to
              prevent any  accumulation  of dust  or other contaminants.
              Store inverted  or  capped  with  aluminum foil.
      3.1.2   The use of high purity reagents and solvents helps  to  mini-
                           •
              raize interference  problems.  Purification of solvents  by
              distillation in all-glass systems  may  be  required.
3.2   Matrix interferences may be caused  by contaminants  that  are coex-
      tracted from the sample.  The extent of matrix interferences will
             i
      vary considerably from  source to  source,  depending  upon  the nature
      and diversity of the industrial complex or municipality being
      sampled.  The cleanup procedure in  Section 11  can be used to over-
      come many of these interferences, but unique samples may require
      additional cleanup approaches to  achieve the MDL  listed in Table  1.
                                   13

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4.   Safety
     4.1   The toxicity of carcinogenicity of each reagent used in this method
           has not been precisely defined; however, each chemical compound
           should be treated as a potential health hazard.  From this view-
           point, exposure to these chemicals must be reduced to the lowest
                            ->•
           possible level by whatever means available.  The laboratory is re-
           sponsible for maintaining a current awareness file of OSHA regu-
           lations regarding the safe handling of the chemicals specified in
           this method.  A reference file of material data handling sheets
           should also be made available to all personnel involved in the
           chemical analysis.  Additional references to laboratory safety are
           available and have been identified3"  for the information of the  .
           analyst.
5.   Apparatus and Materials
     5.1   Sampling equipment, for discrete or composite sampling.
           5.1.1   Grab sample bottle - Amber borosilicate or flint glass, 1-
                   liter or 1-quart volume, fitted with screw caps lined with
                   Teflon.  Aluminum foil may be substituted for Teflon if the
                   sample is not corrosive.  If amber bottles are not avail-
                   able, protect samples from light.  The container and cap
                   liner must be washed, rinsed with acetone or methylene
                   chloride, and dried before use to minimize contamination.
           5.1.2   Automatic sampler (optional) - Must incorporate glass
                   sample containers for the collection of a minimum of
                   250 ml.  Sample containers must be kept refrigerated at 4°C
                   and protected from light during compositing.  If the
                                           14

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              sampler uses a peristaltic pump,  a minimum length of
              compressible silicone rubber tubing may be used.   Before
              use, however, the compressible tubing should be thoroughly
              rinsed with methanol, followed by repeated rinsings  with
              reagent water to minimize the potential for contamination
              of the sample.  An integrating flow meter is required to
              collect flow proportional composites.
5.2   Glassware (All specifications are suggested.  Catalog numbers are
      included for illustration only.)
      5.2.1   Continuous extractor - 2000-mL, available from Paxton Woods
              Glass Shop, Cincinnati, Ohio or equivalent.
      5.2.2   Drying Column - Chromatographic column 400 mm long x 10 mm
              ID.
      5.2.3   Chromatographic column -  400 mm -long x 19 mm ID with 250 ml
              reservoir at the top and  Teflon stopcock (Kontes  K-420290
              or equivalent).
      5.2.4   Concentrator tube, Kuderna-Danish - 25-mL, graduated
              (Kontes K-570050-1025 or  equivalent).  Calibration must be
              checked at the volumes employed in the test.  A ground
              glass stopper is used to  prevent  evaporation of extracts.
      5.2.5   Evaporative flask, Kuderna-Danish - 500-mL (Kontes K-
              570001-0500 or equivalent).  Attach to concentrator tube
              with springs.
      5.2.6   Snyder column, Kuderna-Oanish - three-ball macro  (Kontes K-
              503000-0121 or equivalent).
      5.2.7   Snyder column, Kuderna-Oanish - two-ball micro (Kontes K-
              569001-0219 or equivalent).
                                        15

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      5.2.8   Vials - Amber glass, 10 to 15 ml capacity with Teflon lined
              screw-cap.
      5.2.9   Volumetric flask - 5-mL with glass stopper.
5.3   Boiling chips - approximately 10/40 mesh carborundum.  Heat to
      400°C for 4 hours or extract in a Soxhlet extractor with methylene
      chloride.
5.4   Water bath - Heated, capable of temperature control i2°C).  The
      bath should be used in a hood.
5.5   Balance - Analytical, capable of accurately weighing to the nearest
      0.0001 g.
5.6   Gas chromatograph - Analytical system complete with gas chromato-
      graph suitable for on-column injection and all required accessories
      including syringes, analytical columns, gases, detector, and strip-
      chart recorder.  A data system is recommended for measuring peak
      areas.
                                                                    •
      5.6.1   Column 1 - ISO cm long x 2 mm ID glass, packed with 3%
              SP2250 on Supelcoport (100/120 mesh) or equivalent.  This
              column was used to develop the method performance state-
              ments in Section 15.  Guidelines for the use of alternate
              columns are provided in Section 12.1.
      5.6.2   Column 2 - ISO cm long x 2 mm ID glass, packed with 3/J SP-
              1000 on Supelcoport (100/120 mesh) or equivalent.
      5.6.3   Detector - Alkali-flame detector (AFD), sometimes referred
              to as a nitrogen-phosphorous detector (NPD) or a thermionic
              specific detector (TSD).  This detector has proven effec-
              tive in the analysis of wastewaters for the compounds
              listed in the scope and was used to develop the method
                                    16

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                   performance statements in Section 15.   Alternative
                   detectors, including a mass spectrometer, may be used in
                   accordance with the provisions described in Section 12.1.
6.   Reagents
     6.1   Reagent water - Reagent water is defined as a water in which an
           interferent is not observed at the method detection limit of each
           parameter of interest.
     6.2   Methylene chloride, acetone, methanol, petroleum ether, ethyl
           ether, toluene-distilled-in-glass quality or equivalent.  Ethyl
           ether must be free of peroxides as indicated by EM Quant Test
           Strips (available from Scientific Products Co., Catalog No.
           P1126-8, and other suppliers).  Procedures recommended for removal
           of peroxides are provided with the test strips.
     6.3   Sodium sulfate (ACS) granular, anhydrous; heated in a muffle fur-
           nace at 400°C overnight.
     6.4   Silica gel - Davison Grade  923, 100-200 mesh;  activated by heating
           for 24 hours at 150°C.
     6.5  ei^Sulfuric Acid - Slowly add 16.7 mL of cone.  l^SO/v (94%)  to
          about 50 mL of reagent water.  Dilute to 100 mL with reagent water.
     6.6  6N_ Sodium hydroxide - Dissolve 24.0 grams of sodium hydroxide in
          100 mL of reagent water.
     6.7  Stock standard, solutions  (1.00 vg/tiL)  - Stock standard  solutions
          can be prepared from pure standard materials or purchased  as certi-
          fied solutions.
          6.7.1   crepare stock  standard solutions  by accurately  weighing
                  about 0.0100 grams  of  pure material.  Dissolve  the material
                  in distilled-in-glass  quality  methanol  and-dilute  to  volume
                                          17

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                   in a 10-mL volumetric flask.   Larger volumes can be used at
                   the convenience of the analyst.   If compound purity is cer-
                   tified at 96% or greater,  the weight can be used without
                   correction to calculate the concentration of the stock
                   standard.  Commercially prepared stock  standards can be
                   used at any concentration  if  they are certified by the man-
                   ufacturer or by an independent source.
           6.7.2   Transfer the stock standard solutions into Teflon-sealed
                   screw-cap bottles.  Store  at  4°C and protect from light.
                   Frequently check stock standard  solutions for signs of
                   degradation or evaporation, especially  just prior to pre-
                   paring calibration standards  from them.
           6.7.3   Stock standard solutions must be replaced after six months
                   or sooner if comparison with  check standards indicates a
                   problem.
7.   Calibration
     7.1   Establish gas chromatographic operating parameters equivalent to
           those indicated in Table 1.  The gas  chromatographic system may be
           calibrated using either the external  standard technique (Section
           7.2) or the internal standard technique (Section 7.3).
     7.2   External standard calibration procedure:
           7»2.1   For each compound of interest, prepare calibration
                   standards at a minimum of  three concentration levels by
                   adding volumes of one or more stock' standards to a
                   volumetric flask and diluting to volume with toluene.  One
                   of the external standards  should be at  a concentration
                   near, but above, the method detection limit.  The other
                                           18

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               concentrations  should  correspond to the expected range of
               concentrations  found in real  samples or should define the
               working  range of  the detector.
      7.2.2   Using injections of 2 to 5 ul of each calibration standard,
              tabulate peak height or area responses against the mass
              injected.  The results can be used to prepare a calibration
              curve for each parameter.  Alternatively,  the ratio of the
              response to the mass injected, defined as  the calibration
              factor (CF), can be calculated for each compound at each
              standard concentration.  If the relative standard deviation
              of the calibration factor is less than 10% over the working
              range, the average calibration factor can  be used in place
              of a calibration curve.
      7.2.3   The working .calibration curve or calibration factor must  be
              verified on each working shift by the measurement of one  or
              more calibration standards.  If the response for any com-
              pound varies from the predicted response by more than +10*,
              the test must be repeated using a fresh calibration
              standard.  Alternatively,  a new calibration curve or
              calibration factor must be prepared for that parameter.
7.3   Internal standard calibration procedure.  To use this approach, the
      analyst must select one or more internal standards similar in ana-
      lytical behavior to the compounds  of interest.   The analyst must
      further demonstrate that the measurement of the internal  standard
      is not affected by method or matrix interferences.  Due  to  these
      limitations,  no internal standard  applicable to all  samples can be
      suggested, although carbazole has  been used successfully  in some
      instances.
                                    19

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7.3.1   Prepare calibration standards at a minimum of three concen-
        tration levels for each parameter of interest by adding
        volumes of one or more stock standards to a volumetric
        flask.  To each calibration standard, add a known constant
        amount of one or more internal standards, and dilute to
        volume with toluene.  One of the standards should be at a
        concentration near, but above, the method detection limit.
        The other concentrations should correspond to the expected
        range of concentrations found in real samples, or should
        define the working range of the detector.
7.3.2   Using injections of 2 to 5 ul of each calibration standard,
        tabulate the peak height or area responses against the con-
        centration for each compound and internal standard.  Calcu-
        late response factors (RF) for each compound as follows:
                   RF  -  (AsCis)/(AiSCs)
        where:
             As  » Response for the compound to be measured.
             A-JS * Response for the internal standard.
             Cis * Concentration of the internal standard in vq/l.
             Cs  « Concentration of the compound to be measured
                   in vg/L.
        If the RF value over the working range is constant, less
        than 105t relative standard deviation, the RF can be assumed
        to be invariant and the average RF can be used for calcula-
        tions.  Alternatively, the results can be used to plot a
        calibration curve of response ratios, As/AjS against RF.
                                  20

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           7.3.3   The working calibration  curve  or  RF must  be  verified  on
                   each working shift by the measurement  of  one or  more  cali-
                   bration standards.  If the response for any  compound  varies
                   from the predicted response by more than  ^10%, the  test
                   must be repeated using a fresh calibration standard.
                   Alternatively,  a new calibration  curve must  be prepared  for
                   that compound.
     7.4   Before using any cleanup procedure, the analyst must process  a
           series of calibration standards  through the procedure to validate
           elution patterns and the absence of interferences from the
           reagents.
8.   Quality Control
     8.1   Each.laboratory using this method is required  to  operate a  formal
           quality control program.  The minimum requirements of this  program
           consist of an initial demonstration of laboratory capability  and
           the analysis of spiked samples as a continuing check on
           performance.  The laboratory is  required to maintain performance
           records to define the quality of data that is  generated.
           8.1,1   Before performing any analyses, the analyst  must demon-
                   strate the ability to generate acceptable accuracy  and
                   precision with this method.  This ability is established as
                   described in Section 8.2.
           8.1.2   In recognition of the rapid advances  occurring  in chroma-
                   tography, the analyst is permited certain options to  improve
                   the separations or lower the cost of  measurements.   Each
                   time such modifications  tc the method  are made,  the analyst
                   is required to repeat the procedure  in Section  8.2.
                                         21

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      8.1.3   The laboratory must spike and analyze a minimum of 10* of
              all samples to monitor continuing laboratory performance.
              This procedure is described in Section 8.4.
8.2   To establish the ability to generate acceptable accuracy and preci-
      sion, the analyst must perform the following operations.
      8.2.1   Select a representative spike concentration  for each com-
              pound to be measured.  Using stock standards, prepare a
              quality control check sample concentrate in  methanol 1000
              times more concentrated than the selected concentrations.
      8.2.2   Using a pipet, add 1.00 ml of the check sample concentrate
              to each of a minimum of four 1000-mL aliquots of reagent
              water.  A representative wastewater may be used in place of
              the reagent water, but one or more additional aliquots must
              be analyzed to determine background levels,  and the spike
              level must exceed twice the background level for the test
              to be valid.  Analyze the aliquots according to the method
              beginning in Section 10.
      8.2.3   Calculate the average percent recovery (R),  and the
              standard deviation of the percent recovery (s), for the
              results.  Wastewater background corrections  must be made
              before R and s calculations are performed.
      8.2.4   Using the appropriate data from Table 2, determine the
              recovery and single operator precision expected for the
              method, and compare these results to the values measured in
              Section 8.2.3.  If the data are not comparable, the analyst
              must review potential problem areas and repeat the test.
                                      22

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8.3   The analyst must calculate method performance criteria and define
      the performance of the laboratory for each spike concentration and
      parameter being measured.
      8.3.1   Calculate upper and lower control limits for method perfor-
              mance as follows:
                   Upper Control Limit (UCL) » R + 3 s
                   Lower Control Limit (LCL) » R « 3 s
              where R and s are calculated as in Section 8.2.3.  The UCL
              and LCL can be used to construct control charts  that are
              useful in observing trends in performance.
      8.3.2   The laboratory must develop and maintain separate accuracy
              statements of laboratory performance for wastewater
              samples.  An accuracy statement for the method is defined
              as R £ s.  The accuracy statement should be developed by
              the analysis of four aliquots of wastewater as described in
                •
              Section 8.2.2, followed by the calculation R and s.
              Alternately, the analyst must use four wastewater data
              points gathered through the requirement for continuing
              quality control in Section 8.4.  The accuracy statements
              should be updated regu-larly.
8.4   The laboratory is required to collect in duplicate a portion of
      their samples to monitor spike recoveries.  The frequency of spiked
      sample analysis must be at least 10% of all samples or one sample
      per month, whichever is greater.  One aliquot of the sample must be
      spiked and analyzed as described in Section 8.2.  If the recovery
                                    23

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           for a particular compound does not fall within the control limits
           for method performance, the results reported for that compound in
           all samples processed as part of the same set must be qualified as
           described in Section 13.3.  The laboratory should monitor the fre-
           quency of data so qualified to ensure that it remains at or below
           5*.
     8.5   Before processing any samples, the analyst should demonstrate
           through the analysis of a 1-liter aliquot of reagent water that all
           glassware and reagents interferences are under control.  Each time
           a set of samples is extracted or there is a change in reagents, a
           laboratory reagent blank should be processed as a safeguard against
           laboratory contamination.
     8.6   It is recommended that the laboratory adopt additional quality as-
           surance practices for use with this method.  The specific practices
           that are most productive depend upon the needs of the laboratory
           and the nature of the samples.  Field duplicates may be analyzed to
           monitor the precision of the sampling technique.  When doubt exists
           over the identification of a peak on the chromatogram, confirmatory
           techniques such as gas chromatography with a dissimilar column,
           specific element detector, or mass spectrometer must be used.
           Whenever possible, the laboratory should perform analysis of
           standard reference materials and participate in relevant
           performance evaluation studies.
9.   Samples Collection, Preservation, and Handling
     9.1   Grab samples must be collected in glass containers.   Conventional
           sampling practices  should be followed; however, the bottle must
                                         24

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           not be prerinsed with  sample  before collection.  Composite samples
           should be collected  in refrigerated glass  containers  in accordance
           with the requirements  of the  program.  Automatic sampling equipment
           must be as free as possible of  plastic and other potential sources
           of contamination.
     9.2   The samples must be  iced or refrigerated at 4°C from  the time of
           collection until extraction.
     9.3   Adjust the pH  of the sample to  6  to 8 with 6N. sodium  hydroxide or
           6N sulfuric acid immediately  after sampling.
10.  Sample Extraction
     10.1  Assemble continuous  extraction  apparatus  by placing 5-10
           carborundum chips into the 500-mL round-bottom flask and  attaching
           to the extraction flask.
     10.2  Add 400 ml methylene chloride to  the  extraction flask. Some
           methylene chloride should  displace into  the round-bottom  flask.
     10.3  Mark the water meniscus on the  side of the sample  bottle  for later
           determination  of sample volume.  Pour the  entire sample into the
           extraction flask and add sufficient distilled water to fill  the
           extraction flask (two  liters  total volume  aqueous  phase).
     10.4  Check the pH of the  sample with wide  range pH paper and adjust to  6
           to 8 with 6 N  sodium hydroxide  or 5 N sulfuric acid.
     10.5  Connect the stirring apparatus  to the extraction flask without the
           frit touching  the sample.   Heat methylene  chloride in round-bottom
           flask to continuous  reflux and  continue  heating for 30 minutes to
           one hour, until frit is thoroughly wetted with methylene chloride.
                                        25

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10.6  Lower frit until it just touches the sample and start the stirring
      apparatus rotating.
10.7  Continuously extract sample for 18-24 hours.
10.8  Assemble a Kuderna-Danish (K-D) concentrator by attaching a 10-ml
      concentrator tube to a 500-mL evaporative flask.  Other concentra-
      tion devices or techniques may be used in place of the K-D if the
      requirements of Section 8.2 are met.
10.9  Pour the extract from the round-bottom flask through a drying
      column containing about 10 cm of anhydrous sodium sulfate, and
      collect the extract in the K-D concentrator.  Rinse the flask and
      column with 20 to 30 ml of methylene chloride to complete the
      quantitative transfer.  Once the flask rinse has passed through the
      drying column, rinse the column with 30 to 40 ml of methylene
      chloride.
10.10 Add 1 to 2 clean boiling chips to the evaporative flask and attach
                                      •
      a three-ball Snyder column.  Prewet the Snyder column by adding
      about 1 mL methylene chloride to the top.  Place the K-D apparatus
      on a hot water bath, 60 to 65°C, so that the concentrator tube is
      partially immersed in the hot water, and the entire lower rounded
      surface of the flask is bathed with hot vapor.  Adjust the vertical
      position of the apparatus and the water temperature as required to
      complete the concentration in 15 to 20 min.  At the proper rate of
      distillation, the balls of the column will actively chatter but the
      chambers will not flood with condensed solvent.  When the apparent
      volume of liquid reaches approximately 4 ml, remove the K-0 appara-
      tus and allow it to drain and cool for at  least 10 min.
                                   26

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     10.11 Remove the Snyder column and flask and adjust the volume of the
           extract to 5.0 raL with methylene chloride.   Stopper the
           concentrator tube and store refrigerated if further processing will
           not be performed immediately.  If the extract is to be stored
           longer than two days, transfer the extract  to a screw capped vial
           with a Teflon-lined cap.  If the sample extract requires no further
           cleanup, proceed with solvent exchange to toluene and  gas chromatographic
           analysis as described in sections 11.5 and  12  respectively.   If the
           sample requires cleanup, proceed to Section  11.
     10.12 Determine the original sample volume  by refilling the sample bottle
           to the mark and transferring the water to a 1000-mL graduated
           cylinder.  Record the sample volume to the nearest 5 ml.
11.  Cleanup and Separation
     11.1  Cleanup procedures may not be necessary for a relatively clean
           sample matrix.  The cleanup procedure recommended in this method
           has been used for the analysis of various clean waters and indus-
           trial effluents.  If particular circumstances demand the use of an
           alternative cleanup procedure, the analyst must determine the elu-
           tion profile and demonstrate that the recovery of each compound of
           interest is no less than 85JJ.
     11.2  Stir 20 g of silica gel in 100 mL of acetone and 1.2 ml of reagent
           water for 30 minutes on a stirring plate.  Transfer the slurry to a
           chromatographic column (silica gel may be retained  with  a plug of
           glass wool).  Wash the column with 20 mL of methylene  chloride and then
           with 30 ml of petroleum ether.  Use a column flow rate of 2 to 2.5
           ml/min throughout the wash and elution profiles.  Add an additional
           50 ml of petroleum ether to the head of the column.
                                        27-

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     11.3  Add the extract from Section 10.11 to the head of the column.
           Allow the solvent to elute from the column until  the Florisil  is
           almost exposed to the air.  Elute the column with 50 ml of 6%  ethyl
           ether in petroleum ether.  Discard this fraction.
     11.4  Elute the column with 100 ml of 15% ethyl ether in petroleum ether
           and collect in a KD apparatus.
     11.5  Add 2.5 ml of toluene to the fraction.  Concentrate the fraction to
           approximately 4 mL with the water bath at 75-80°C as described in
           Section 10.10.  Transfer the sample to a 5-mL volumetric flask and
           dilute to 5 mL with toluene.  Proceed with gas chromatographic
           analysis.
12.  Gas Chromatoaraphy
     12.1  Table 1 summarizes the recommended operating conditions for the gas
           chromatograph.  Included in this table are estimated retention
           times and method detection limits that can be achieved by this
           method.  An example of the separations achieved by Column 1 and
           Column 2 are shown in Figures 1 and 2.  Other packed columns,
           chromatographic conditions, or detectors may be used if the
           requirements of Section 8.2 are met.   Capillary (open-tubular)
           columns may also be used if the relative standard deviations of
           responses for replicate injections are demonstrated to be less than
           6% and the requirements of Section 8.2 are met.
     12.2  Calibrate the gas chromatographic system daily as described in
           Section 7.
     12.3  If an internal standard approach is being used, the analyst must
           not add the internal  standard to the  sample extracts until
           immediately before injection into the instrument.  Mix thoroughly.
                                       28

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     12.4  Inject 2 to 5 nl of the sample extract  using  the solvent flush
           technique.®  Record the volume injected to the nearest  0.05-al,  and
           the resulting peak sizes in area or peak height units.
     12.5  The width of the retention time window  used to make identifications
           should be based upon measurements of actual retention time varia-
           tions of standards over the course of a day.   Three times the
           standard deviation of a retention time  for a  compound can be used
           to calculate a suggested window size; however, the experience of
           the analyst should weigh heavily in the interpretation  of
           chromatograms.
     12.6  If the response for the peak exceeds the working range  of the
           system, dilute the extract and reanalyze.
     12.7  If the measurement of the peak response -is prevented by the
           presence of interferences, further cleanup is required.
13.  Calculations
     13.1  Determine the concentration of individual compounds in  the sample.
           13.1.1  If the external standard calibration  procedure  is used,
                   calculate the amount of material injected from  the peak
                   response using the calibration  curve  or calibration factor
                   in Section 7.2.2.  The concentration  in the sample can  be
                   calculated as follows:
                        Concentration, ag/L »
                   where:
                                     . 29

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                       A   » Amount of material  injected  in nanograms.
                       V-f  • Volume of extract  injected in ul.
                       Vt  * Volume of total extract  in ^L.
                       Vs  * Volume of water extracted in mL.
           13.1.2  If  the internal standard calibration procedure was used,
                   calculate the  concentration  in  the sample using the
                   response factor  (RF)  determined in Section  7.3.2  as
                   follows:

                       Concentration, ,g/l *
                   where:
                        As   » Response for the compound to be measured. ,
                        A-fs a Response for the internal standard.
                        Is   * Amount of internal  standard  added to each
                                 extract in */g.
                        V0   » Volume of water extracted in liters.
     13.2  Report results  in micrograms per liter without  correction for
           recovery data.   When duplicate and spiked samples are analyzed,
           report all data  obtained with the sample results.
     13.3  For samples processed as part of a set where the laboratory spiked
           sample recovery  falls outside of the control limits in Section 8.3,
           data for the affected compounds must be labeled as suspect.
14.  SC-MS Confirmation
     14.1  It is recommended that GC-MS techniques be judiciously employed to
           support qualitative identifications made with this method.  The
           mass spectrometer should be capable of scanning the mass range from
                                       30

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      35 amu  to a mass 50 amu above the  molecular weight  of  the compound.
      The instrument must be capable of  scanning  the mass range at  a rate
      to produce at least 5 scans  per peak  but  not to exceed 7  seconds
      per scan utilizing a 70-V (nominal) electron energy in the electron
      impact  ionization mode.  A GC to MS interface constructed of  all-
                                               >
      glass or glass-lined materials is  recommended.  When using a  fused
      silica  capillary column, the column outlet  should be threaded
      through the interface to within a  few mm  of the entrance  to the
      source  ionization chamber.  A computer system should be interfaced
      to the  mass spectrometer that allows  the  continuous acquisition and
      storage on machine readable media  of  all  mass spectra obtained
      throughout the duration of the chromatographic program.
14.2  Gas chromatographic columns and conditions  should be selected for
      optimum separation and performance.   The  conditions selected  must
      be compatible with standard GC-MS  operating practices.  Chromato-
      graphic tailing factors of less than  5.0  must be achieved. The
      calculation of tailing factors is  illustrated in Method 625.
14.3  At the  beginning of each day that  confirmatory analyses are to be
      performed, the GC-MS system must be checked to see  that all DFTPP
                                        q
      performance criteria are achieved.
14.4  To confirm an identification of a  compound, the background
      corrected mass spectrum of the compound must be obtained from the
      sample.extract and compared with a mass spectrum from a stock or
      calibration standard analyzed under the same chromatographic
      conditions.  It is recommended that at least 25 nanograms of
      material be injected into the GC-MS.   The criteria  below must be
      met for qualitative confirmation.

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           14.4.1  The molecular Ion and all other ions that are present above
                   10X relative abundance in the mass spectrum of the standard
                   must be present in the mass spectrum of the sample with
                   agreement to plus or minus 103S.  For example, if the rela-
                   tive abundance of an ion is 30% in the mass spectrum of the
                   standard, the allowable limits for the relative abundance
                   of that ion in the mass spectrum for the sample would be
                   20-40*.
           14.4.2  The retention time of the compound in the sample must be
                 .  within 30 seconds of the same compound in the standard
                  .solution.
           14.4.3  Compounds that have very similar mass spectra can be
                   explicitly identified by GC-MS only on the basis of reten-
                   tion time data.
     14.5  Where available, chemical ionization mass spectra may be employed
           to aid in the qualitative identification process.
     14.6  Should these MS procedures fail to provide satisfactory results,
           additional steps may be taken before reanalysis.  These may include
           the use of alternate packed or capillary GC columns or additional
           cleanup (Section 11).
15.  Method Performance
     15.1  The method detection limit (MDL) is defined as the minimum concen-
           tration of a substance that can be measured and reported with 99%
           confidence that the value is above zero.  The MDL concentrations
           listed in Table 1 were obtained using reagent water.   Similar
           results were achieved using representative wastewaters.
                                                            •
                                         32

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15.2  This method has been tested for linearity of recovery from spiked
      reagent water and has been demonstrated to be applicable over the
      concentration range from 10 x MDL to 1000 x MDL.
15.3  In a single laboratory, Battelle Columbus Laboratories, using
      spiked wastewater samples, the average recoveries presented in
      Table 2 were obtained.  Seven replicates of each of two different
      wastewaters were spiked and analyzed.  The standard deviation of
      the percent recovery is also included in Table 2.
                                   33

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                                  REFERENCES


1.  "Development of Methods for Pesticides in Wastewaters," Report for EPA
    Contract 68-03-2956 (In preparation).

2.  ASTM Annual Book of Standards, Part 31, D3694, "Standard Practice for Prep-
    aration of Sample Containers and for Preservation," American Society for
    Testing and Materials, Philadelphia, PA, p. 679, 1980.

3.  "Carcinogens - Workinc with Carcinogens," Department of Health, Education,
    and Welfare, Public Health Service, Center for Disease Control, National
    Institute for Occupational Safety and Health, Publication No. 77-206,
    August, 1977.

4.  "OSHA Safety and Health Standards, General Industry," (29 CFR 1910), Occu-
    pational Safety and Health Administration, OSHA 2206 (Revised, January
    1976).

5.  "Safety in Academic Chemistry Laboratories," American Chemical Society
    Publications, Committee on Chemical Safety, 3rd Edition, 1979.

6.  "Handbook for Analytical Quality Control in Water and Wastewater Laborar
    tories," EPA-600/4-79-019, U.S. Environmental Protection Agency, Environ-
    mental Monitoring and Support Laboratory - Cincinnati, Ohio 45268, March
    1979.

7.  ASTM Annual Book of Standards, Part 31, D3370, "Standard Practice for Sam-
    pling Water," American Society for Testing and Materials, Philadelphia,
    PA, p. 76, 1980.

8.  Burke, 0. A., "Gas Chromatography for Pesticide Residue Analysis; Some
    Practical Aspects," Journal of the Association of Official Analytical
    Chemists, 48, 1037 (1965).

9.  Eichelberger, J. W., Harris, L. E., and Budde, W. L., "Reference Compound
    to Calibrate Ion Abundance Measurement in Gas Chromatography - Mass Spec-
    trometry," Analytical Chemistry. 47, 995 (1975).
                                       34

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       TABLE  1.   CHROMATOGRAPHIC CONDITIONS AND METHOD DETECTION LIMITS
Parameter                Retention Time (min)           Method Detection  Limit
                         Column 1Column 2                    ("9/L)


Diphenylamine             18.1         19."3                       1.6
Column 1 conditions:  Supelcoport  (100/120 mesh) coated with  2%  SP-2250  packed
in a 1.8 rn long x 2 mm ID glass column with helium carrier  gas at  a  flow rate
of 30 mL/min.  Column temperature  is held at 80°C for 4 minutes, programmed
from 80°C to 300°C at S^C/min and  held at 300°C for 4 minutes.

Column 2 conditions:   Supelcoport (100/120 mesh)  coated  with 3%  SP-1000 packed
in a 1.8 m long x 2 mm ID glass  column  with helium carrier gas at a flow rate
of 30 mL/min.  Column temperature is held at 80°C for 4  minutes,  programmed
from 80°C to 250°C at 80°C/min,  and held  at 250°C for 4  minutes.
                                          35

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             TABLE 2.  SINGLE LABORATORY ACCURACY AND PRECISION(a)
•


Parameter
Diphenylamine


Average
Percent
Recovery
120
39
Relative
Standard
Deviation,
%
25
11
••^•••^••••••••MMBMM
Spike
Level
(«g/L)
5.0
50

Number
of
Analyses
7
7


Matrix
Type(b)
1
1
(a)   Column 1 conditions were used.
(b)   1 * Columbus secondary POTW effluent,
                                      36

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               Dlphenylnralne
             r-f-j—r— r"f"i"|-•!••»—»— t—p- r- r—i—r-f"*"''—r " •"" j—i--i—t—v-|~i—r-i—r- -|--r-r--i'-r*i- •••-«•• i—i—|
                10.0       21.0       23. a       23. i       27.0       20.0       31,0       33. B     33. B

                              RETENTION TIME,  minutes

KJyure  1.   CC-AFD  Chromatogram of  100 ng of  Dlphenylamlne (Column  1).

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u>
00
r-|-

 10. B
21. •
                                   22.8
    24. •       23.9


RETENTION TIME,  minutes
                                                                           28. 9
              Figure 2.  CC-FID Cllromatogram of  200 ng of Diphenylamlne (Column 2).

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c~
TECHNICAL REPORT DATA
(Please read Instructions on the reverse bclore completing!
1. REPORT NO. 2.
4. TITLE ANO SUBTITLE
Determination of Diphenylamine in Industrial and
Municipal Wastewaters
7. AUTHORS J>s> Warner, T.M. Engel and
P.J. Mondron
9. PERFORMING ORGANIZATION NAME AND AOORESS
BatteH e Columbus Laboratories
505 King Avenue
Columbus, Ohio 43201
12. SPONSORING AGENCY NAME ANO AOORESS
U.S. Environmental Protection Agency
Environmental Monitoring and Support Laboratory
Cincinnati, Ohio 45268
3. RECIPIENT'S ACCESSION NO.
5. REPORT DATE
6. PERFORMING ORGANIZATION COOS
8. PERFORMING ORGANIZATION REPORT N(
10. PROGRAM ELEMENT NO.
CBECIC
11. CONTRACT/GRANT NO.
68-03-2956
13. TYPE OF REPORT ANO PERIOD COVEREC
14. SPONSORING AGENCY CODE
EPA 600/06
15. SUPPLEMENTARY NOTES
16. ABSTRACT
     A method  was  developed  for the determination of diphenylamine  in  wastewaters.
 The  method  development  program consisted of a literature review: determination of
 extraction  efficiency  for each compound from water using roethylene  chloride;
 development of  a  deactivated silica gel cleanup procedure; and  determination  of
 suitable gas  chromatographic (GC)  analysis conditions.

     The final method was  applied to Columbus POTW secondary effluent  in  order to
 determine the precision  and accuracy of the method.  The wastewater was  spiked with
 diphenylamine at  levels of  5 ug/L  and 50 ug/L.  Recovery for diphenylamine  at the 5
 ug/L level  was  120  * 25  percent.  Recovery at the 50 ug/L level was 89 ± 11
 percent.  The method detection limit (MDL) for diphenylamfne in distilled water was
 1.6  ug/L.   In wastewaters it may be higher due to interfering compounds.
17. KEY WORDS ANO DOCUMENT ANALYSIS
a. DESCRIPTORS

18. DISTRIBUTION STATEMENT
Release to Public
b.lOENTIFI6RS/OPEN ENDED TERMS

19. SECURITY CLASS ( Tins Report}
Nonclassified
2O. SECURITY CLASS (Tint page I
Nonclassified
T f -• <
c. CDSATI Field/Group

21. NO. OF PAGES
39
22. PRICS
EPA form 2270-1 (T?«». 4-77)   P*«viou* COITION is OB*ouETe

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