United States
Environmental Protection
Agency
Environmental Monitoring and
Support Laboratory
Cincinnati OH 45268
Research and Development
EPA-600/S4-84-001 Mar. 1984
&EPA 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
7,7,7 -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 10 mL priortoanalysiswith 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 bydeterminingthe
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
/ug/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-
Table III. Accuracy and Precision for Purgeable Organics
Three POTW Sludges
Spike Recovery Spike Recovery
Two Industrial Sludges
Spike Level
US/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
160
100
170
150
130
140
160
130
120
120
Standard
Deviation
55
58
53
33
38
51
69
47
36
26
Spike
"9
Min
1
2
5
3
5
5
2
16
2
5
Level
/L
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 -Dichloroet hene
Trichloroethene^
1,1,1 - Trichloroethane
Chlorobenzene
Ethyl Benzene"
Spike
Level
(ngj
250
250
250
250
250
250
250
250
250
250
10-mL Aliquot
Unspiked Sludge
fng>
49.2
NO"
43.6
123
NO
ND
19.0
118
Spike
2.5 mL
133
77
101
112
102
102
135
Recovery (%)
5 mL
142
68
123
165
114
117
108
10 mL
148
55
122
194
152
133
61
" ND * not detected.
b Ion plots for all unspiked and spiked samples exhibited saturated peaks.
c 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 solids.
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(GPC) 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: 50 mL 10% 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, VA22161
Telephone: 703-487-4650
For information, contact Robert W. Slater at:
Environmental Monitoring and Support Laboratory
U.S. Environmental Protect/on Agency
Cincinnati, OH 45268
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