United States
 Environmental Protection
 Agency
 Environmental Monitoring and
 Support Laboratory
 Cincinnati OH 45268
 Research and Development
 EPA-600/S4-84-001  Mar. 1984
 Project Summary
 Development  of Analytical
Test  Procedures for  the
 Measurement of Organic
 Priority  Pollutants  in  Sludge

Clarence L Haile and Viorica Lopez-Avila
  The tremendous diversity of sludge
characteristics presents a considerable
challenge to precise and accurate
determinations of trace levels of the
organic priority pollutants in sludges.
The primary objective of this research
program was  the development of test
methods for  the analysis of organic
priority pollutants in sludges as would
be generated from the treatment of
municipal and industrial wastewaters.
The research program was divided into
two stages. In the first stage, a prelimi-
nary sludge  analysis protocol was
developed based on existing wastewater
procedures. This was done to satisfy an
immediate need of the U.S. Environ-
mental Protection Agency  (EPA) for
test methods to be used in support of a
project  initiated  to determine the
occurrence and fate of the  organic
priority  pollutants in  representative
publicly owned treatment works (POTW).
Based on the experience in this program
and as a result of a systematic study of
sludge analysis techniques, a refined
and revised analytical protocol was
developed.
  This report details the second phase
of the project, which was a systematic
study of sludge analysis techniques. For
purposes of analysis, a separate proce-
dure was provided for the  purgeable
and extractable organic priority pollu-
tants in sludges.  In each  case, the
components were identified and quanti-
tated using GC/MS techniques. The
development of  a procedure  for the
analysis  of the volatile components
 involved an investigation of various
 purging procedures intended to improve
 recoveries, reproducibilities, and detec-
 tion limits.
  Since sludges contain a wide variety
 of extractable organics  which can
 interfere with the GC/MS analysis, the
 development of suitable cleanup pro-
 cedures was required. Selected tech-
 niques of extract cleanup were applied
 in various combinations, the  most
 successful of which are included in the
 recommended protocol.
  The project summary was developed
 by EPA's Environmental Monitoring
 and Support Laboratory, Cincinnati,
 Ohio, to announce key findings of the
 research project that is fully documented
 in a separate report of the same title (see
 Project Report ordering information at
 back).

Analysis of Purgeables from
Sludge -- Method
Development
  This aspect  of the  research was
directed toward applying and modifying
the purge and trap GC/MS procedures
developed  by the EPA  for  wastewater
analysis.
  Preliminary experiments with sludges
for various POTW indicated that most but
not all sludges could successfully be
purged if the sample aliquot was diluted
to approximately 0.5% (w/v) total solids.
Recoveries for analytes spiked into POTW
sludges and analyzed by this procedure
showed poor precision during the recovery

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determinations and a  decrease in  the
method detection limit as a consequence
of the sample dilution. Efforts to overcome
these deficiencies form the basis of this
study. Table 1  lists the priority pollutants
used to develop the methods.
Table 1.    Representative Purgeable Organ-
          ic Priority Pollutants Used for
          Method Development and Evalua-
          tion Experiments


Purgeables:
  Benzene
  Carbon Tetrachloride
  Chlorobenzene
  Chloroform
  1' ,2-Dichloroethane
  1,1 -Dichloroethene
  Ethyl Benzene
  Tetrachloroethene
  1.1.1-Trichloroethane
  Trichloroethene
  Vinyl Chloride

Extractables:
  1,4-Dichlorobenzene
  Hexachloroethane
  bis-(2-Chloroisopropyl)Ether
  bis-(2-Chloroethyl)Ether
  A cenaphthylene
  2.6-Dinitrotoluene
  Fluoranthene
  Benzidine
  3,3'-Dichlorobenzidine
  Benzyl n-Butyl Phthalate
  bis-(2-Ethylhexyl)Phthalate
  Benzo(a)pyrene
  Phenol
  2.4-Dimethylphenol
  2,4-Dichlorophenol
  Pentachlorophenol	
   The relative influences of dissolved
 solids  and suspended solids on the
 purging efficiencies of analytes from
 sludges were investigated by determining
 analyte  recoveries from  spiked  sludge
 and spiked sludge supernatant (SS). Aliquots
 of primary  POTW sludge (4.8% solids)
 were centrifuged, and the superna'tants
 were  decanted.  The supernatants and
 replicate aliquots of unfractionated
 sludge were spiked and analyzed by the
 preliminary POTW sludge method.
   The recoveries for the spiked compounds
 from the SS were both higher and less
 variable than from the unfractionated
 sludge for all compounds. Furthermore,
 the range of recoveries observed for the
 supernatant aliquots was similar to those
 typically observed for spiked clean water.
 Hence, it is likely that the presence of
 dissolved solids  in the primary sludge
 does not significantly affect the purging
 efficiencies of the spiked compounds.
  The addition  of salt or a high-boiling
hydrophilic organic compound (with  a
very low tendency to purge from water),
to sludge  aliquots prior to purging was
evaluated  as a  means of enhancing the
purging efficiencies of volatile compounds,
or at least  improving the reproducibility of
purging. The recoveries of compounds
spiked into sludge and SS indicated that
interactions of the purgeable compounds
with  the  SS  may  have a  significant
influence  on  their purging efficiencies.
The addition of salt  or high-boiling
organic was an attempt to alter favorably
those interactions to provide a consistent
matrix for purging.
  The influences of salt or an organic on
purging from  aqueous  media  were
investigated by analyzing spiked  clean
water by  purge and  trap  GC/flame
ionization detection (FID). Aliquots of
volatile-free water were spiked and
diluted to 10mL prior toanalysis with one
of three diluents: (a) volatile-free water;
(b) saturated sodium chloride in volatile-
free water; or (c) 10% ethylene glycol in
volatile-free water. The detector responses
were compared to determine the influence
of  each diluent. Ethylene  glycol was
selected as the organic additive because
of  its low volatility,  good aqueous
solubility,  and general availability.
   Recovery experiments were also con-
ducted with primary POTW sludge (2%
solids). Spiked and unspiked sludge
aliquots were  diluted  with  the salt or
ethylene  glycol solutions and analyzed
by the preliminary purge and trap GC/MS
methods.  No specific  advantages from
adding  salt  or ethylene glycol were
apparent.  Recoveries for all compounds
were good. Recoveries for carbon tetra-
chloride were  significantly higher than
typically observed using the preliminary
POTW sludge  protocol. However, both
additives appeared to increase significant-
ly the tendency of the sludge to foam dur-
ing purging.  Hence, the use of salt and
ethylene glycol in sludge dilutions did not
provide significant overall improvements
for the sludge method.


Alternative  Purging Hardware
Systems
   Alternative purging hardware systems
which provide  mechanical mixing during
purging were evaluated for tneir potential
to enhance desorption of analytes from
the sludge solids to the supernatant and
to improve transfer to the purge gas
stream.  All systems  evaluated  were
 designed to be used as direct replacements
 for the conventional purge tube in the
 purge and trap system employed in the
preliminary POTW sludge method. Four
hardware systems were evaluated. Three
configurations were fabricated using a
small round-bottom flask: stirred purging,
stirred impinging, and sonicated imping-
ing. The fourth system evaluated was a
stirred-purge  configuration using a
specially designed  bottom frit tube.
  The efficiencies  of transfer of volatile
compounds from aqueous solution to the
gas phase afforded by the stirred purging,
stirred impinger, and sonicated impinger
systems were  evaluated by  comparing
the  chromatographic  responses  for
compounds spiked into water and analyzed
with these systems. The chromatographic
responses for  identical solutions  were
analyzed via the conventional purge tube.
  The performance of the bottom frit tube
with and without mechanical stirring was
compared with that  of the conventional
purge tube for spiked water  analyzed by
purge and trap GC/FID. The responses
observed for a few compounds analyzed
in spiked water with the bottom frit tube
without  stirring were lower than those
from the conventional purge tube. The
bottom  frit tube with stirring produced
responses generally as high or higher
than the conventional tube. These results
likely  reflect  poorer contact  of the
sparging bubbles with the water  in  the
unstirred bottom frit tube relative to  the
conventional purge tube.

Recoveries
  The performance of the bottom frit tube
was further evaluated by determining  the
recoveries for compunds spiked into
aliquots of primary POTW  sludge (2%
solids) immediately prior to analysis. The
sludge aliquots were diluted and analyzed
by the  purge  and trap  GC/MS method
using the stirred bottom frit tube in place
of the  conventional  purge  tube.  Good
recovery data were  obtained for all  the
compounds except carbon tetrachloride.
The carbon tetrachloride  response was
too low to determine under the specific
GC/MS operation conditions utilized.
Recovery  determinations were very
reproducible. No foaming was observed
with the bottom frit  tube, whereas
aliquots of the same sample previously
analyzed with the conventional purge
tube foamed profusely.
   Since good data  on accuracy and
precision were obtained with the stirred
bottom frit  tube and  there was  no
appreciable foaming, the system was
evaluated for its capability to purge larger
aliquots of sludge without troublesome
foaming. Aliquots of 2.5, 5.0, and 10.0 mL
of primary sludge (2% solids) were spiked
with 250 ng of each of the purgeable

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spiking compounds just prior to analysis,
diluted as required to 10 mL with volatile-
free water, and analyzed by the prelimi-
nary POTW sludge GC/MS method. The
recoveries  of  the spiked purgeable
compounds in different volumes of sludge
are shown in Table II. Recoveries for all
compounds were good and did not appear
to be strongly dependent on the volume of
sludge purged.  No foaming  problems
were encountered, even  with  10-mL
aliquots. Evidently, the force imparted by
the magnetic stirring bar  swirling the
sludge aliquot  was  an efficient foam
dispersion mechanism.
  Recoveries did not  appear to  be
dependent on the size of  the  aliquot
purged even though the spike concentra-
tions ranged from 125 /ug/L in the 2.0-mL
aliquots. Recoveries of compounds in
10.0-mL aliquots of spiked sludge (at 25
fjg/L) analyzed with the stirred bottom frit
tube were very similar to those in 2.0-mL
aliquots of spiked sludge (at 125 /ug/L)
using the conventional purge tube. In
addition,  no foaming was  observed
stirred purging, even while purging 10.0-
mL aliquots of undiluted sludge.

Precision  and Accuracy—
Purgeables
  The precision and accuracy of the
purgeables  method were evaluated by
determining recoveries for spiked com-
pounds from five primary sludges.  The
subject sludges,  three from  POTW
receiving various fractions of wastewater
from  residential and industrial sources
and two from plants treating  industrial
wastewaters, were selected to provide a
wide  variety of sludge characteristics
with which to test the performance of the
method. The results of these determina-
tions are shown in Table III.
  Although the recoveries observed were
generally good,  many recovery determi-
Tablelll.   Accuracy and Precision for Purgeable Organics

                          Three POTW Sludges
             Two Industrial Sludges
Spike Recovery Spike Recovery
Spike Level
U9/L
Compound
Benzene
Chloroform
1, 1 -Dichloroethene
Tetrachloroethene
Vinyl chloride
1 ,2-Dichloroethane
Trichloroethene
1,1,1- Trichloroethane
Chlorobenzene
Ethyl benzene
Min
1
2
5
3
50
5
20
16
2
5
Max
100
200
170
300
500
500
200
1,600
200
500
Mean
1%)
160
100
170
150
130
140
160
130
120
120
Standard
Deviation
55
58
53
33
38
51
69
47
36
26
Spike Level
H9/L
Min
1
2
5
3
5
5
2
16
2
5
Max
100
200
500
300
500
500
200
1,600
200
500
Mean
<%)
98
76
110
150
110
100
140
110
160
150
Standard
Deviation
25
22
51
70
47
28
44
40
62
55
nations were likely influenced by relative-
ly high concentrations of the spiking
compounds in  the unspiked sludges.
Several compounds were  present in
some of the  sludges  at levels greater
than the spike level, and in some cases
concentrations in the  unspiked sludges
were more than 10 times the spike level.
Recoveries for the latter cases were not
representative of the  method precision
and accuracy and were excluded from the
table. Zero or very low recoveries were
observed for carbon tetrachloride in all
sludges except Industrial No. 2. Carbon
tetrachloride and, to some extent, chloro-
form are  frequently troublesome com-
pounds because of poor sensitivity to
mass spectrometric detection. Although
the detection limits for these compounds
were chosen based on previous work,
their detection limits in these particular
sludges are  evidently higher than is
typical. Determinations of vinyl chloride,
chlorobenzene, and 1,1,1-trichloroethane
were frequently obscured by coeluting
interferences in the POTW sludges.
  Nonetheless,  in most cases,  the
recoveries observed were both good and
reproducible.  Only 11% of all  recovery
Table II.    Recovery of Spiked Purgeable Compounds from Spiked Sludge by Stirred Purging with
          a Bottom Frit Tube; Effect of Different Volumes of Sludge.
Compound
Benzene
Carbon Tetrachloride
Chloroform
1, 1 -Dichloroethene
Tetrachloroethene*'
1 ,2-Dichloroethene
Trichloroethene"
1,1,1- Trichloroethane
Chlorobenzene
Ethyl Benzenec
Spike
Level
(ng)
250
250
250
250
250
250
250
250
250
250
10-mL Aliquot
Unspiked Sludge Spike
(ng)
49.2
ND"
43.6
123

ND

ND
19.0
118
2.5 mL
133
77
101
112

102

102
135

Recovery f%J
5mL
142
68
123
165

114

117
108

10 mL
148
55
122
194

152

133
61

' ND = not detected.
" Ion plots for all unspiked and spiked samples exhibited saturated peaks.
0 Ion plots for all spiked sludge samples exhibited saturated peaks.
determinations were less than 50%.
More than 63% fell within the range of
50-150% recovery. An additional 25% of
the recovery determinations  exceeded
150%. The relative standard deviations
(RSD) for triplicate recovery determina-
tions were generally low. More than 90%
of the RSD were 30% or less. Of those,
27%  were  less than  10%  RSD.  The
method reproducibility was surprisingly
good considering the difficulty in remov-
ing  representative aliquots from a
heterogeneous sample matrix.
Analysis of Extractables from
Sludge—Method Development
  The method development and evalua-
tion experiments described in this section
were conducted using a subset  of the
organic priority pollutants to  simplify
sample spiking and recovery determina-
tions. These compounds were selected to
represent the  physical and chemical
characteristics of most  of  the organic
priority pollutants.  The list of spiking
compounds is shown in Table I.
  The primary objective in developing the
extraction method was to adapt industrial
wastewater  screening methods  for
sludge extraction and extract cleanup, so
as to provide extracts of sufficient quality
for GC/MS analysis. Many of the extrac-
table compounds  were expected  to
associate strongly with the sludge sol ids.
It was anticipated that the wastewater
extraction method  (simple  liquid-liquid
partitioning with dichloromethane) would
not  provide  sufficient  contact of the
extracting solvent with the solids to allow
efficient extraction  of  those compounds
from sludges, and would be hindered by
formation of emulsions. A procedure
using a high speed homogenizer probe to
provide vigorous mixing  and blending of
the sludge aliquot with the extracting
solvent was evaluated.

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  Unfortunately, this vigorous homoge-
nization/centrifugation procedure also
extracted large quantities of lipids, fatty
acids, and other high molecular weight
compounds present in  POTW sludges.
These compounds can cause significant
interferences during GC/MS analysis
and necessitate extract cleanup.
  Three alternative extraction procedures
were evaluated for sludges in an attempt
to develop a more efficient, selective, and
less time consuming extraction method.
The procedures evaluated include: con-
tinous liquid-liquid extraction (CLLE),
steam distillation, and microextraction. As
expected, each of these procedures
achieved promising recoveries for con-
centrating spiked compounds from water.
  None of the tested methods resulted in
any significant improvements over the
preliminary protocol. Hence, the homo-
genization/centrifugation method was
selected  for the  sludge  protocol. This
selection necessitated additional evalua-
tion  of extract  cleanup procedures as
alternatives or supplements  to the  gel
permeation chromatography (G PC) meth-
od used in the preliminary POTW protocol.

Extract Cleanup Studies
  Successful employment of a rigorous,
nonselective extraction  procedure  for
sludges, necessitated the use of a very
selective and efficient  extract cleanup
procedure to produce extracts of sufficient
quality for reliable GC/MS determination.
Two extract  cleanup mechanisms  were
evaluated to  meet these requirements,
molecular size discrimination and polarity
selection. Various GPC procedures were
evaluated to determine the optimum
molecular size fractionation  procedure.
The polarity-based cleanup method
evaluated was adsorption chromatogra-
phy on  silica  gel and Florisil.  The
performance of adsorption procedures
was evaluated for sludge extracts both
with and without GPC precleaning.
  Five GPC  packings (Bio-Beads SX-2,
SX-3, SX-4, SX-8, and Sephadex LH-20)
were  evaluated with  three solvent
systems. The  solvents tested  were
dichloromethane, 15% cyclohexane in
dichloromethane,  and 50% cyclohexane
in dichloromethane.
   Adsorption chromatography experiments
were  performed  using  1% deactivated
Florisil or 3% deactivated silica gel. The
elution scheme was:

   • fraction I: 20 mL hexane
   • fraction II: 50mL10%dichlorometh-
     ane in hexane
   • fraction III: 50 mL dichloromethane
     in hexane
  • fraction IV: 150 mL 5% acetone in
    dichloromethane
  The elution volumes and peak profiles
data obtained  for the various gels and
solvent  combinations indicate  that the
best overall separation is obtained with
SX-3 gel when dichloromethane is the
eluting solvent. Furthermore, the selection
of dichloromethane as the elution solvent
is directly compatible with the homogeni-
zation/centrifugation of sludge extracts.
  The Base/Neutral (B/N) and acid
extracts of sludges were chromatographed
in one or more 5.0-mL injections. The
combined cleaned extract fractions were
concentrated to 1 -5 mL for GC/MS analy-
sis.  Extracts  that  were still highly
colored were cleaned by a second pass
through  the GPC column.
  The B/N  and acidic extracts were
analyzed by GC/MS according to the
procedures described in  the industrial
wastewater protocol.
  The recoveries observed for most of the
test compounds were fairly good, although
several compounds were not recovered
from the primary sludge. These compounds
were hexachloroethane, benzidene, 3,3'-
dichlorobenzidene, and pentachlorophe-
nol. Part of the loss of hexachloroethane
can be attributed to  volatilization during
extract concentrations. Dilution of the
primary  sludge extracts to reduce the
concentration of interfering coextractants
also reduced the spike levels to at or near
the detection limit.
GC/MS  Procedures
  Capillary  GC/MS procedures were
evaluated as an alternative to the packed
column  GC/MS methods described
above. Four wall-coated-open-tubular
(WCOT) capillary columns were evaluated
using standard performance test mixtures
and solutions of the B/N and acidic test
compounds. Of the four columns tested.
the SE-54 fused silica column exhibited
the best overall performance.
  The suitability of the SE-54 fused silica
column for analyses of sludge extracts
was evaluated by chromatographing the
spiked sludge  extracts prepared for the
evaluation of adsorption chromatographic
cleanup procedures. The better resolution
and inertness of  the SE-54 capillary
column (relative to the packed column)
provided GC/MS data for B/N extracts
that were easier to interpret.

Precision and Accuracy—
Extractables
  The recovery data was  shown to vary
somewhat from various sludge sample
types. Some of this effect is likely due to
the high concentration of some of the
analytes in the unspiked sludge samples.
In general, recoveries were lower for the
more polar compounds, such as benzidine.
Of all recovery determinations, 49% fell
within the range of 50-150%. An addition-
al 19% were lower than  50% recovery,
and only 8% of the recoveries were
greater than 150%.
  The accuracy of the  method was
somewhat dependent on the particular
sludge samples. However, the precisions
of the  recovery determinations were
good. The relative standard deviations for
triplicate determinations  were  30%  or
less for 95% of the measurements and
10% or less for 62% of the measurements.
In view of the complexity and diversity of
municipal and  industrial wastewater
treatment sludges, the  precision and
accuracy results presented here demon-
strate  that the protocol developed can
reliably be applied to the  analysis of the
organic priority pollutants in sludge. The
success of this protocol for the variety of
extractable compounds for which is was
developed and evaluated  indicates that
the methods included  may also be useful
for many nonpriority pollutant analytes.
   Clarence L Haile and Viorica Lopez-Avila are with Midwest Research Institute,
     Kansas City, MO 64110.
   Stephen Billets was the EPA Project Officer (see EPA contact below).
   The complete report, entitled "Development of Analytical Test Procedures for
     the Measurement of Organic Priority Pollutants in Sludge,"  (Order No. PB
     84-129 048; Cost: $26.50, subject to change) will be available only from:
           National Technical Information Service
           5285 Port Royal Road
           Springfield. VA 22161
           Telephone: 703-487-4650
   For information, contact Robert W. Slater at:
           Environmental Monitoring and Support Laboratory
           U.S. Environmental Protection Agency
           Cincinnati, OH 45268

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