United States Robert S. Kerr EPA-600/2-89-011
Environmental Protection Environmental Research Laboratory
Agency Ada, OK 74820 March 1989
Research and Development
$EPA Treatability Potential for
EPA Listed Hazardous
Wastes in Soils
~-\ —.
• W £*5'?ff l/£p.
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PB89-166581
EPA/600/2-89/011
March 1989
TREATABILITY POTENTIAL FOR EPA LISTED
HAZARDOUS WASTES IN SOIL
by
Raymond C. Loehr
Environmental Engineering Program
The University of Texas at Austin
Austin, Texas 78712
Project CR-812819
Project Officer
Scott G. Muling
Extramural Activities and Assistance Division
Robert S. Kerr Environmental Research Laboratory
Ada, Oklahoma 74820
ROBERT S. KERR ENVIRONMENTAL RESEARCH LABORATORY
OFFICE OF RESEARCH AND DEVELOPMENT
U.S. ENVIRONMENTAL PROTECTION AGENCY
ADA, OKLAHOMA 74820
or THE
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NOTICE
The information in this document has been funded wholly or in part by the United
States Environmental Protection Agency under Cooperative Agreement CR-812819 to
The University of Texas at Austin. It has been subjected to the Agency's peer and
administrative review, and it has been approved for publication as an EPA document.
Mention of trade names or commercial products does not constitute endorsement or
recommendation for use.
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FOREWORD
EPA is charged by Congress to protect the Nation's land, air, and water systems.
Under a mandate of national environmental laws focused on air and water quality, solid
waste management and the control of toxic substances, pesticides, noise and radiation,
the Agency strives to formulate and implement actions which lead to a compatible
balance between human activities and the ability of natural systems to support and
nurture life.
The Robert S. Kerr Environmental Research Laboratory is the Agency's center
of expertise for investigation of the soil and subsurface environment. Personnel at the
Laboratory are responsible for management of research programs to: (a) determine the
fate, transport and transformation rates of pollutants in the soil, the unsaturated and the
saturated zones of the subsurface environment; (b) define the processes to be used
in characterizing the soil and subsurface environment as a receptor of pollutants; (c)
develop techniques for predicting the effect of pollutants on groundwater, soil, and
indigenous organisms; and (d) define and demonstrate the applicability and limitations
of using natural processes, indigenous to the soil and subsurface environment, for the
protection of this resource.
Soil treatment systems that are designed and managed based on a knowledge
of soil-waste interactions may represent a significant technology for simultaneous
treatment and ultimate disposal of selected hazardous wastes in an environmentally
acceptable manner. Decisions pertaining to which wastes and chemicals are amenable
in
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to this technology must take into account: (1) the long-term uncertainties associated
with the land disposal option; (2) the goal of managing hazardous wastes in an
appropriate manner; and (3) the persistence, toxicity, mobility, and propensity to
bioaccumulate hazardous wastes and their hazardous constituents. There is currently
a lack of scientifically derived fate and transport information for the wide range of
hazardous chemicals for which such decisions can be made. This report presents
information pertaining to the quantitative evaluation of the treatment potential in soil of
specific listed hazardous organic chemicals as identified by the United States
Environmental Protection Agency (EPA), and waste sludge from explosives production
and its related chemicals.
Clinton W. Hall
Director
Robert S. Kerr Environmental
Research Laboratory
IV
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ABSTRACT
This study developed comprehensive screening data on the
treatability in soil of: (a) specific listed' hazardous organic
chemicals, and (b) waste sludge from explosives production (KO44)
and related chemicals. Laboratory experiments were conducted
using two soil types, an acidic soil (Mississippi soil) with less than
one percent organic matter, and a slightly basic sandy loam soil
(Texas soil) containing 3.25% organic matter. These experiments
evaluated the: (a) relative toxicity of the chemicals and waste using
the Microtox® bioassay method, (b) degradation of the chemicals and waste in
the soils, (c) adsorption characteristics of the chemicals in the two soils, and (d)
toxicity reduction that occurred during degradation.
The major conclusions were:
1. The chemical structure of the compounds evaluated affected their relative
toxicity. With chlorophenols, the relative toxicity was related to the
position of the chlorine group on the phenol ring. The order of relative
toxicity was para>meta>ortho. The same order appeared to occur for
methylphenols and nitrophenols. The chemical substituted on the
phenol ring appeared to have an effect on toxicity. Nitro-substituted
phenols appeared to be less toxic than the methyl- or chloro-substituted
phenols. Mixing of the chemicals with the soils did not affect the relative
toxicity of the chemicals in the two soils.
2. Data characterizing the chemical loss in the soil and in the water soluble
fraction (WSF) extracted from the soil as well as the toxicity reduction in
the WSF could be represented satisfactorily by either first or zero order
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kinetics. In most cases, the data were represented by either kinetic
parameter with high correlation coefficients.
3. The rates of chemical loss were higher in the Texas soil. Chlorophenols
with chlorine substituted in the meta position had greater half-lives and
lower loss rates. Chemicals with a nitro group substituted in the phenol
ring appeared to have a lower loss rate.
4. The Freundlich equation described the adsorption of most of the
chemicals with the two soils satisfactorily. The values of the Freundlich
constant (Kf) for the chemicals in the two soils were different. For the acid
extractables, the Kf values generally were greater in the Mississippi soil.
For the amines and alcohols, the Kf values were greater in the Texas soil.
5. The loss of the applied chemical in the soil and in the WSFas well as
the reduction of the WSF toxicity were compared for nine of the
chemicals. The chemical loss in the WSF was about 1.5 times faster than
the chemical loss in the soil. The WSF toxicity decreased at about the
same rate as the WSF chemical concentration. No enhanced
mobilization of the applied chemical occurred during degradation.
VI
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CONTENTS
SECTION PAGE
NOTICE ii
FOREWARD iii
ABSTRACT v
FIGURES x
TABLES xi
ACKNOWLEDGEMENTS xiv
SECTION 1. INTRODUCTION 1
Scope of Study „ 1
Designated Chemicals and Waste 2
SECTION 2. CONCLUSIONS 5
A. Chemical Toxicity 5
B. Degradation Studies 6
C. Adsorption Studies 6
D. Toxicity Reduction 7
E. Munitions Chemicals and Wastes 8
SECTION 3. GENERAL MATERIALS AND METHODS 8
Chemicals 9
Soil 9
Instrumentation 11
Gas Chromatography 11
High Pressure Liquid Chromatography 13
QA/QC Procedures 13
Toxicity 13
SECTION 4. RELATIVE TOXICITY AND CHEMICAL LOADING 14
Objectives 14
Background 14
Microtox 15
EC5Q Evaluation 16
Acceptable Loadings 19
Conclusions 27
SECTION 5. DEGRADATION STUDIES 29
Introduction 29
Materials and Methods 31
Data Analysis 33
VII
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CONTENTS, Continued
SECTION PAGE
Results 37
Recovery Efficiency 37
Kinetic Parameters 37
Conclusions 46
SECTION 6. ADSORPTION EXPERIMENTS 49
Introduction 49
Adsorption Equilibria 50
Soil Organic Carbon 51
Soil pH 52
Materials and Methods 54
Adsorption Method 54
Stock Solutions 54
Standard Solutions 54
Soil Moisture 56
Soil:Solution Ratio ; 56
Solute Stability 58
Other Factors 58
Data Analysis 59
Results 60
Conclusions 67
SECTION 7. TOXICITY REDUCTION 69
Approach 69
Results 70
Chemical Loss in Soil 70
WSF Chemical Loss 72
WSF Toxicity Reduction 76
Comparison of Chemical Losses and Toxicity Reduction 76
Conclusions 81
SECTION 8. MUNITONS WASTES AND CHEMICALS 82
Relative Toxicity and Loading Evaluation 82
Adsorption 83
Methods 83
Results 84
Degradation Studies 87
Munitions Wastewater Treatment Sludge 87
Nitrogen and COD 87
Metals 91
GC/MS Analysis 91
Relative Toxicity 91
Conclusions 95
SECTION 9. REFERENCES 97
VIII
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CONTENTS, Continued
SECTION PAGE
APPENDIX A. THE MICROTOX© TOXICITY ASSAY USED
IN THIS STUDY 101
Introduction 101
Evaluation of Chemical Loss Using Microcosms 101
Water Extraction of Microcosms 101
Toxicity Assay 102
EC5Q Determination 102
Assay Procedure 102
Chemical Loading on Soil 103
Microtox© Analysis 104
APPENDIX B. QUALITY CONTROL/QUALITY ASSURANCE
PROCEDURES 107
Toxicity 107
Degradation Studies 107
Adsorption Studies 109
QA/QC For Analytical Instruments 111
APPENDIX C. VOLATILIZATION ESTIMATES 114
APPENDIX D. PUBLICATIONS 117
IX
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FIGURES
NUMBER PAGE
1. Schematic of the Phenol Ring and Possible Substitution Positions 18
2. Loss of Phenol in Texas Soil at 20° C 42
3. Loss of Phenol in Mississippi Soil at 20° C 42
4. Loss of 2,6-Dichlorophenol in Texas Soil at 20° C 43
5. Loss of 2,6-Dichlorophenol in Mississippi Soil at 20° C 43
6. Adsorption of 2,3-Dichlorophenol and Toluenediamine in Texas
Soil at 20° C 61
7. Adsorption of 2,3-Dichlorophenol and Toluenediamine in Mississippi
Soil at 20° C 62
8. Loss of Chemical in Two Experiments When the Higher Loading Rates
Were Used 74
9. Loss of Chemical in the WSF at Two Loading Rates 75
10. Toxicity Reduction in the WSF from the Soil Microcosms 78
11. Comparison of Chemical Loss in the Soil and the WSF for Phenol and
Eight Chlorophenols 79
12. Decrease of Soil and WSF Chemical Concentration and of WSF
Toxicity as a Function of Time-Degradation Study of
2,4-Dichlorophenol 80
13. Comparison of the WSF Chemical Loss and the WSF Toxicity Reduction
For Phenol and Eight Chlorophenols 80
14. Loss of 2,6-Dinitrotoluene in Texas and Mississippi soils at 20° C 89
15. Wastewater Treatment Process Flow Diagram for the Holston Army
Ammunition Plant 90
B-1. Representative Recovery Data for Phenol in Texas Soil 110
B-2. Representative Recovery Data for o-Cresol in Texas Soil 110
B-3. Representative Recovery Data for 2,4-Dichlorophenol in Texas Soil 111
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TABLES
NUMBER PAGE
1. Chemicals That Were Evaluated in This Study 3
2. The Hazardous Waste and Related Chemicals That Were Evaluated 4
3. Characteristics of Soils Used in This Study 10
4. Operating Conditions for the Gas Chromatographic Analysis
of Compounds in Methylene Chloride 12
5. EC5Q for Chemicals Evaluated in This Study 17
6. Relative Toxicity of Chlorinated Phenols 20
7. Relative Toxicity of Non-Chlorinated Phenols 20
8. Comparative Relative toxicity of Chloro-, Methyl,
and Nitrophenols 22
9. Method to Determine Acceptable Initial Chemical Loadings 22
10. Acceptable Non-Inhibitory Loading Rates -- Texas Soil 23
11. Acceptable Non-Inhibitory Loading Rates -- Mississippi Soil 24
12. Comparative Acceptable Loading Rates -- Chlorinated Phenols 25
13. Experimental Procedures Used in the Degradation Studies 33
14. Chemicals Whose Soil Concentration Was Evaluated
Using Shake Extraction 34
15. Shake Extraction Procedure 34
16. Recovery Efficiencies for Specific Chemicals (%) 38
17. Acceptable and Actual Loading Rates Used in the Degradation
Studies (mg/kg of Soil) 39
18. Loss Rates, Correlation Coefficients and 95% Confidence
Intervals for Specific Chemicals -- Texas Soil 40
19. Loss Rates, Correlation Coefficients and 95%
Confidence Intervals for Specific Chemicals
--Mississippi Soil 41
XI
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TABLES, Continued
NUMBER . PAGE
20. Chemical Half-Lives in Texas and Mississippi Soils (days) 45
21. Effect of Substitution Position on Degradation Rates --
Chlorinated Phenols 47
22. Effect of Substitution Position on Degradation Rates --
Non-Chlorinated Phenols 47
23. Comparative Degradation Rates of Chloro-, Methyl- and Nitrophenols....48
24. Procedure for Determining Solubility Limit of an Organic Compound
In Water 55
25. List of Maximum Solubilities (20° C) in Water Determined as Part of the
Adsorption Studies 55
26. Procedure for Measuring Soil Moisture Content. 56
27. Procedure for Determination of Optimum SoikSolution Ratio 57
28. Batch Sorption Isotherm Data - Texas Soil -- Freundlich Equation
Parameters 63
29. Batch Sorption Isotherm Data -- Mississippi Soil -- Freundlich Equation
Parameters 64
30. Chemical Concentration Range Evaluated During the Batch Adsorption
Experiments -- Texas Soil 65
31. Chemical Concentration Range Evaluated During the Batch Adsorption
Experiments -- Mississippi Soil 66
32. Comparison of Freundlich Adsorption Coefficients (Kf) for the Texas and
Mississippi Soils 68
33. Chemical Loss in Soil -- Kinetic Parameters 71
34. Loss of Water Extractable Chemical -- Kinetic Parameters 73
35. WSF Toxicity Reduction -- Kinetic Parameters 77
36. ECso Data and Acceptable Loading Rates for Munitions Manufacturing
Chemicals 83
XI I
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TABLES, Continued
NUMBER PAGE
37. Chemical Concentration Range Evaluated During the Batch
Adsorption Experiments - Munitions Chemicals 85
38. Freundlich Isotherm Data - Texas and Mississippi Soils -- 2,4- and
2,6-Dinitrotoluene 85
39. Adsorption Data for TNT, HMX and RDX in Texas Soil at 20° C 86
40. Adsorption Data for TNT, HMX and RDX in Mississippi
Soil at 20° C 86
41. Loading Rate and Recovery Efficiency Data from the Munitions
Chemical Degradation Studies 88
42. Chemical Loss Rate Data for 2,6-Dinitrotoluene and TNT in the
Degradation Studies 88
43. Nitrogen and COD Concentrations of Munitions Waste Sludge 91
44. Metals in Munitions Sludge and Sludge Filtrate 92
45. GC/MS Analysis of Munitions Sludge 93
46. Munitions Waste Toxicity Data -- Undiluted Samples 95
B-1. Accuracy Data: Recovery Efficiencies (%) for Specific Chemicals
As Determined From Day Zero Degradation Study Experiments 108
B-2. Precision Analysis Data: Texas Soil Sorption Data 112
C-1. Loss of Methanol and 1-Butanol in Volatilization Experiments 115
XIII
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ACKNOWLEDGEMENTS
This report represents the hard work and dedication of the undergraduate
students, graduate students, staff and faculty who assisted in this project. The
contribution of the following individuals deserves specific recognition and is
greatly appreciated:
Barnes Bierck Michael McFarland
Srinivasa Dasappa Joseph F. Malina, Jr.
Carol English Wan Nam-Koong
David Erickson Lynn Sanders
Lisa Gilmour-Stallsworth Karen Spaniel
Nadine Gordon John Stephenson
Frank Hulsey Eric White
R. Krishnamoorthy Chun Yoon
We also appreciate the assistance of the EPA-RSKERL project officers,
Mr. John E. Matthews and Mr. Scott G. Huling, who had responsibility for the
project throughout its duration.
XIV
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SECTION 1
INTRODUCTION
SCOPE OF STUDY
This study was designed to provide comprehensive screening data on the
treatability in soil of: (a) EPA listed hazardous organic chemicals, and (b) a specific
hazardous waste and related chemicals. The results of the study provide data that can
be used when permitting decisions are made related to: (a) management of spills, (b)
remediation of contaminated soils, and (c) the use of land as a waste management
alternative. The degradation and partitioning data can be used as input to predictive
models that estimate the movement of chemicals in the unsaturated zone of the soil.
Examples of such models include RiTZ (Regulatory and Investigative Treatment Zone
Model)(1-2), VIP (Vadose Zone Interactive Processes Model)(3» 4), and KOPT
(Kinematic Oily Pollutant Transport Model^).
These models were developed to understand the treatment potential of organic
chemicals in soil. The models integrate the processes that affect chemicals in soil
(degradation and partitioning) so that an assessment can be made of the extent to
which protection of human health and the environment occurs. The understanding
that results from the use of such models allows the identification of chemicals and
wastes that require control to reduce or eliminate their hazard potential prior to
application to soil.
Laboratory studies were conducted to determine: (a) degradation kinetics, (b)
sorption, (c) toxicity of the chemicals and waste, and (d) the reduction in toxicity that
occurs during degradation. The results of these studies are discussed in subsequent
sections.
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DESIGNATED CHEMICALS AND WASTES
The chemicals and specific waste that were part of this study are identified as
hazardous wastes under CFR Sections 261.32 and 261.33. These chemicals can be
expected to be components of many industrial compounds and wastes that enter the
soil from spills and inadequately sealed impoundments (pits, ponds and lagoons) and
as part of wastes applied to operating land treatment units.
The chemicals that were evaluated are identified in Table 1. The specific
hazardous waste, and chemicals related to that waste, that were evaluated are noted
in Table 2.
Samples of the explosives waste sludge (KO44) and the chemicals TNT, RDX,
and HMX were obtained with the help of the U.S. Army Toxic and Hazardous Materials
Agency (USATHAMA). A sample of wastewater treatment sludge resulting from the
manufacture and processing of explosives was obtained from the Holston Army
Ammunition Plant with the assistance of USATHAMA. This material was stored at 4° C
until required for analysis and use.
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TABLE 1. CHEMICALS THAT WERE EVALUATED IN THIS STUDY
EPA Hazardous
Compound _ Formula _ Waste Number
Acid Extrtctabl»»
Phenol [[[ CgHgO ........................................... U188
o-Cresol [[[ CyHgO ........................................... U052
p-Cresol [[[ 07830 ........................................... U052
m-Cresol [[[ C?H8O ........................................... U052
2-Chtorophenol ......................................... CgH5CIO .......................................... U048
3-Chkjrophenol ......................................... CgH5CIO ........................................... NOS
4-Chloroph«nol ......................................... CgHgCIO ........................................... NOS
2.3-Dichtorophenol .................................... CgH4CI2O .......................................... NOS
2,4-Dichtorophenol .................................... C6H4CI2O ......................................... U08.1
2.5-Dichloroph9nol .................................... CgH4CI2O .......................................... NOS
2,6-Dichtorophenol .................................... CgH4CI2O ......................................... U082
3.4-Dichlorophenol .................................... CgH4CI2O .......................................... NOS
2.4.5-Trtehlorophenol ................................. CgH3CI3O ......................................... U230
2,4.6-Trfchlorophenol ................................. CgH3CI3O ......................................... U231
Pentachlorophenol .................................... CgHCI5O .......................................... U242
2,4-Dimethylphenol ..................................... C8H10° .......................................... U101
2-Methyl-4-Chlorophenol ............................. C7H7CIO ........................................... NOS
3-Methyl-4-Chlorophenol ............................. C7H?CIO .......................................... U039
3-Methyl-6-Chlorophenol ............................. C7H7CIO ........................................... NOS
p-Nitroph«nol ............................................ Cgh^NO-j ......................................... U170
2.4-Dinitrophenol ...................................... CgH4N2O5 ........................................ P048
4,6-Dinrtro-o-Cresol .................................. Cr^W^S ........................................ Po48
Thtophenol ............................... , ................ -CgHgS ........................................... U014
Diphenylamina .......................................... C^H^N ......................................... X016
m-Phenyl«nediamine ................................... C6H8N2 .......................................... *017
Tolu«nBdiamin« ....................................... C ....................................... U221
Brucin« ................................................ C23 26N2°4
Alcohols
Isobutyl alcohol ......................................... C4H10° .......................................... U14°
Allyl alcohol ................................................ C3H6O ........................................... POOS
Propargyl alcohol ........................................ C3H4O ........................................... P102
1-Butanol .................................................. C4H10° .......................................... Uo31
2,3-Dichloropropanol ................................... ^HgC^ .......................................... X006
Methanol [[[ CH4O ............................................ U154
Othir
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TABLE 2. THE HAZARDOUS WASTE AND RELATED CHEMICALS
THAT WERE EVALUATED
Specific Hazardous Waste
KO44 - Wastewater treatment sludge from the manufacturing and processing of explosives
Explosive and Munitions Manufacturing Chemicals
EPA Hazardous
Compound
2,4-Dinitrotoluene
2,6-Dinitrotoluene
TNT (2,4.6-Trinitrotoluene)
RDX+
HMX++
Formula
C7H6N204
C7H6N204
C7H5N306
C3H6N606
C4H8N808
Waste Number
U105
U106
-
-
—
+ RDX - Hexahydrotrinitrotriazine
++ HMX- Cyctotetramethylenetetranitramine
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SECTION 2
CONCLUSIONS
The major results were the following:
A. CHEMICAL TOXICITY
1. The Microtox® biological assay represents an appropriate method with which
to evaluate the EC$Q toxicity of a chemical or waste.
2. Comparison of the £650 data indicated that: (a) the alcohols were less toxic
than the acid extractable compounds, and (b) within chemical categories, there
were considerable differences in relative toxicity.
3. The chemical structure of the compounds evaluated affected the relative toxicity
of a compound. With chlorophenols, the relative toxicity was related to the
substitution position of the chlorine group on the phenol ring. The order of
relative toxicity was para>meta>ortho. The £650 data suggested that the same
order occurred for methylphenols and nitrophenols.
4. The chemical that was substituted on the phenol ring appeared to have an
effect on toxicity. Nitro-substituted phenols, even when substituted in the para
position, appeared to be less toxic than the methyl- or chloro- substituted
phenols.
5. When the chemicals were mixed with two different soils, and the EC50 value of
the water soluble fraction (WSF) of the soil mixtures was measured, the values
also indicated that chemicals with the chlorine in the para position had the
greater toxicity. Mixing of the chemicals with the soils did not affect the relative
toxicity of the chemicals in the two soils.
6. In general, the acceptable non-inhibitory chemical loading rates for the
Mississippi soil were lower than those for the Texas soil. There was no
consistent pattern for the differences.
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B. DEGRADATION STUDIES
7. The chemical or waste loading procedure (Table 9, Section 4)
resulted in chemical loadings that did not inhibit the non-acclimated
organisms in the laboratory microcosms, except in one case (4,6-
Dinitro-o-Cresol). This procedure provided a good estimate of
initial, acceptable chemical loadings that can be used in laboratory
degradation studies.
8. Both zero and first order kinetics provided adequate representation
of the data. For most of the chemicals, the data could be fit to
either kinetics with high correlation coefficients.
9. The rates of chemical loss were higher in the Texas soil than in the
Mississippi soil. There did not appear to be any pattern to the
differences in rates in the two soils.
10. Chlorophenols with the chlorine substituted in the meta position had
greater half-lives and therefore lower chemical loss rates. This
was particularly evident with the mono-, di-, and trichlorophenols
in the Texas soil.
11. Chemicals that had a nitro group substituted on the phenol ring
appeared to have a lower loss rate.
C. ADSORPTION STUDIES
12. The Freundlich equation described the adsorption of the chemicals on
the two soils satisfactorily, with high correlative coefficients,
except for a few chemicals.
13. The range of chemical concentrations evaluated ranged from the low
mg/l concentrations to near or at saturation concentrations, and for
most chemicals covered two to three orders of magnitude. For these
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concentration ranges, a linear adsorption relationship, i.e., n • 1, did
not occur.
14. The values of the Freundlich constant (Kf) for the chemicals in the two soils
were different. For the acid extractables, the Kf values generally were greater in
the Texas soil which had the higher pH and the greater organic carbon content.
For the amines and alcohols, the Kf values were greater in the Mississippi soil,
which had the lower pH and the lower organic carbon content.
D. TOXICITY REDUCTION
15. Two loading rates, the Texas soil, and nine chemicals (phenol and eight
chlorinated phenols) were used in this study. Both first and zero order Kinetics
satisfactorily fit the water soluble fraction (WSF) chemical loss data and the
toxicity reduction data.
16. The higher chemical loading rates resulted in higher chemical concentrations in
the WSF and higher WSF toxicities at the beginning of the experiments.
17. The higher chemical loading rates generally resulted in slower chemical losses
(higher half lives) and slower toxicity reduction. However, at both loading rates
for each chemical, the chemicals were degraded and the toxicity was reduced.
No differences due to the loading rates were apparent in zero order kinetics.
18. The loss of the chemicals in the WSF was about 1.5 times faster than the loss of
the chemical in the soil.
19. The WSF toxicity for each chemical decreased as the soil chemical and the
WSF chemical concentrations decreased.
20. The WSF toxicity decreased at about the same rate as the WSF chemical
concentration when the data for all nine chemicals were compared.
21. No enhanced mobilization of the applied chemicals occurred as the
degradation and detoxification occurred.
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22. No water soluble toxic products appeared to be formed as the chemicals were
degraded in the soil.
E. MUNITIONS CHEMICALS AND WASTES
23. The Freundlich equation described the sorption of 2,4- and 2,6-Dinitrotoluene in
the two soils satisfactorily. It did not do so for TNT, RDX, or HMX.
24. No loss of 2,4-Dinitrotoluene occurred over a 47-day study even though the
loading rate used was determined to be acceptable using procedures
discussed in Section 4. Degradation loss rates were obtained for 2,6-
Dinitrotoluene and TNT. First order kinetics were a better representation for
TNT than were zero order kinetics.
25. The half life of TNT in the Mississippi soil was shorter, and the loss faster, than
in the Texas soil. No difference in the loss rates in the two soils for 2,6-
Dinitrotoluene was apparent.
26. The sludge resulting from the manufacture and processing of explosives
contained: (a) high concentrations of nitrogen and COD, (b) concentrations
generally less than 10 mg/l for heavy metals, and (c) no TNT, RDX or HMX.
27. The munitions sludge had a high toxicity as measured by the Microtox®
procedure. The constituents causing the relative toxicity were in the soluble
phase of the sludge.
8
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SECTION 3
GENERAL MATERIALS AND METHODS
CHEMICALS
The chemicals evaluated in this study were identified in Section 1. To the
extent possible, analytical grade chemicals were used in the experiments and as
spikes and controls in the analytical procedures. The chemicals were purchased
either from Aldrich Chemicals or Sigma Chemicals. Specific explosive and munitions
manufacturing chemicals were obtained from sources identified in Section 8.
SOILS
The intent of this study was to provide comprehensive screening data on the
treatability of specific chemicals and a hazardous waste in soil. The characteristics of
the soil will affect the degradation, sorption, and treatment potential and two soils with
different characteristics were used. One was an acid soil with a low organic content
and the other was a basic soil with a higher organic content and cation exchange
capacity (CEC).
The acid soil was obtained from an area near Wiggins, Mississippi, and was
supplied by researchers at Mississippi State University. This soil is referred to as
Mississippi soil in this report. The characteristics of this soil are presented in Table 3.
The analyses were conducted by staff at Mississippi State University using appropriate
methods(6).
The basic soil was obtained from an area near Austin, Texas, that, to the
knowledge of the personnel of this project, had not been exposed to industrial
chemicals or wastes. This soil is referred to as Texas soil. The characteristics of the
soil are presented in Table 3. The analyses were conducted by staff at the soil testing
and characterization laboratory, Texas A&M University, College Station, Texas.
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Because independent laboratories provided separate analyses of the soils, the
data in Table 3 are not always directly comparable. However, the data provide
pertinent information on the important characteristics of the two soils.
Both soils had initial microorganism counts that were typical for an agricultural
soil with an active microbial population.
TABLE 3. CHARACTERISTICS OF SOILS USED IN THIS STUDY
Characteristic
Texture
Classification
pH
CEC(meq/100g)
Organic Carbon
Sodium
Potassium
Calcium
Magnesium
Hydrogen
Carbonate (CO3)
Bicarbonate (HCO3)
Sulfate
Chloride
Particle Size Fraction (%)
Sand
Silt
Clay
Moisture Content by Weight (%)
1/3 atmosphere
15 atmosphere
Mississippi
Soil
sandy loam
Typic Paleudults
4.8
6.35
0.94%
0.02 meq/100 g
0.07 meq/1 00 g
0.29 meq/100 g
0.06 meq/100 g
5.9 meq/100 g
-
-
-
-
68.0
23.4
8.6
12.4
8.2
Texas
Soil
sandy silt loam
Mollisal, Cumulic
7.8
10.8
3.25%
0.3 meq/L+
3.6 meq/L
9.5 meq/L
1 .3 meq/L
-
0.0 meq/L
7.2 meq/L
1 .9 meq/L
2.8 meq/L
61.5
31.1
7.4
17.0
6.2
+ -- Saturated paste extract
10
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After the soils were received at the Environmental and Water Resources
Engineering Laboratory at The University of Texas, they were air dried, sieved and
stored at 4° C in the dark. Prior to each experiment, soil samples were taken for use,
the moisture content increased to near saturation, and the indigenous organisms
allowed to establish equilibrium concentrations.
INSTRUMENTATION
The principal analytical instruments used were the Microtox© analyzer, the gas
chromatograph (GC), and the high pressure liquid chromatograph (HPLC). The
Microtox© unit, which was used for chemical toxicity evaluation, is discussed in detail
in Section 4. The following describes the operational procedures employed for the QC
and HPLC analyses.
Gas Chromatoqraphv
Two types of gas chromatographs were used to quantify chemical
concentrations. The choice of GC depended on the volatility of the compound and the
solvent used with the compound.
Compounds extracted in Methylene Chloride were analyzed by capillary
column gas chromatography. The analytical procedure followed is outlined in EPA-
SW 846 (Method 8040)(7). The method consisted of injection of a 1 jiL sample into
the gas chromatograph (Hewlett Packard Model 5890A) which was equipped with an
electronic integrator (Hewlett Packard Model 3392A), a methyl silicone capillary
column, and a flame ionization detector. The chromatographic system was calibrated
using an internal standard technique each day.
The operating conditions of the capillary column gas chromatograph are
presented in Table 4. The initial temperature and temperature progress rate were
selected based on the retention time of the test compound and the internal standard.
The integrator options were set to minimize the tailing of the chromatographic peaks,
which affected the peak area calculations. The initial temperature was between 30° C
11
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and 100° C depending on the retention time of the compound, the temperature
progress rate was between 15° C and 30° C/minute, and the final temperature was
180° C. A two-minute final time was utilized at the termination of each run in order to
ensure that the column was clean.
TABLE 4. OPERATING CONDITIONS FOR THE GAS CHROMATOGRAPHIC ANALYSIS
OF COMPOUNDS IN METHYLENE CHLORIDE
Condition
Description
Capillary Column
Temperature Initial
Detector
Carrier Gas
Final
Injection Port
Detector
Hydrogen Gas Rate
Air Rate
Helium Rate
methyl silicone (dimensions: 5m length x 0.53 mm I.D. x
2.65 urn film thickness)
30° C-100° C at 0 minutes with progress rate
of 15-30° C per minute.
180° C at 2 minutes
200° C
250° C
Flame lonization
30 mL/minute
400 mL/minute
20 mL/minute
Compounds extracted in either water or Methanol (e.g., alcohols) were
quantified on a packed column gas chromatograph (Tracer 550) equipped with a six-
foot column packed with 5% Carbowax 1500, 80/100 Carbopack K-C (Supeico, Inc.).
The GC was operated isothermally at 130° C with a flame ionization detector. The
electronic output signal was converted to concentration units by interfacing the GC
with an integrator (Spectraphysics Model 4290). The GC was calibrated by the
internal standard technique outlined in Method 8040 (EPA-SW846)(7).
12
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High Pressure Liquid Chromatographv
Phenolic compounds contained in filtered water (i.e., as part of the sorption
study, Section 6) and the compounds of low volatility (Brucine, Thiourea, RDX, HMX
and TNT) were analyzed by high pressure liquid chromatography (HPLC).
The HPLC (Waters Associates Model 440) was operated at room temperature
and utilized a C-18 reversed phase column. Aqueous solutions (50 (J.L) of various
samples were eluted with 50:50:0.1 of acetonitrile:deionized distilled wateracetic acid
solution. The UV absorption detector wavelength was 254 nm. The, flow rate and
chart speed were maintained at 3.0 ml/min and 0.1 inch/min respectively. The
attenuation varied from 0.01 to 2.0 depending on the relative compound absorption at
254 nm. Duplicate samples were analyzed to ensure instrument accuracy.
Standard solutions for each compound were run prior to sample analysis for
external standard calibration. The peak height on the chart paper for each standard
solution was measured and linear regression analysis (Lotus 1-2-3, IBM/PC) used to
determine the relationship between peak height (cm) and concentration (mg/L).
Comparison of peak heights of samples to peak heights determined for standard
solutions allowed estimation of sample chemical concentrations.
QA/QC PROCEDURES
Care was taken to assure that sound, representative data were obtained. The
specific quality assurance and quality control procedures that were followed and
information on precision and accuracy are presented in Appendix B.
TOXICITY
The relative toxicity of the chemicals and the specific waste as well as of the
water soluble fraction of soil-chemical mixtures was measured by the Microtox®
system. Details of the system and how it was used are presented in Section 4.
13
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SECTION 4
RELATIVE TOXICITY AND CHEMICAL LOADING
OBJECTIVES
A major objective of this study was to obtain information on the degradation
kinetics of these chemicals and specific waste in soil. To do so, it was important that
the chemicals be in the soil in concentrations that would not be toxic or inhibitory to the
soil microorganisms. Therefore, it was necessary to determine: (a) the relative toxicity
of the chemicals and specific waste prior to adding them to the soils, and (b) the mass
loading rates of the chemicals that would not be inhibitory.
The relative toxicity tests that were conducted were not intended to provide
information on toxicity from a human hearth or safety or from an environmental
standpoint. Rather, these tests were used as a relative toxicity screening method.
Such tests also can be used to identify the relative toxicity reduction that occurs when
chemicals and waste are managed by the land treatment process.
BACKGROUND
The usual procedures to quantify toxicity of a chemical are toxicity assays which
measure the effect of the chemical on a test species under specified test conditions.
The toxicity of a chemical is proportional to the severity of the chemical on the
monitored response of the test organism(s). Toxicity assays utilize test species that
include rats, fish, invertebrates, microbes and seeds. The assays may use single or
multiple species of test organisms. The need to unequivocally measure the effect of
the toxicant on the monitored activity has favored the use of a single specie as the test
organism in a toxicity assay.
Toxicity assays using bacteria as the test organism are gaining popularity due
to their rapidity, ease in handling, cost effectiveness and the use of a statistically
significant number of test organisms^. 9). The Microtox® assay is a microbial assay
14
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used for toxicity measurement, hazard assessment, and quantitative-structure activity
relationship (QSAR) studies of environmental pollutants^"11).
Although no single bioassay procedure can provide a comprehensive toxicity
evaluation of a chemical, a valid toxicity screening test can provide information about
the relative toxicity of a compound and can help predict non-inhibitory chemical
application rates. The Microtox® system is a relatively simple, rapid and inexpensive
test and was used as the toxicity screening method in this project. The use of the
Microtox© procedure to screen and predict the treat ability potential of waste in soil has
been evaluated and found to be satisfactory(12-13).
Microtox®
The Microtox® system is a standardized toxicity test system which utilizes
marine luminescent bacteria (Photobacterium phosphoreum) as indicator organisms.
Bio luminescence of this test organism depends on a complex chain of biochemical
reactions involving the luciferin-luciferase system. Chemical inhibition any of the
involved biochemical reactions causes a reduction in bacterial luminescence. The
Microtox® toxicity assessment considers the physiological effect of a toxicant, and not
just mortality.
The system utilizes an instrumental approach in which the indicator organisms
are handled as chemical reagents. Suspensions of about one million bioluminescent
organisms are "challenged" by addition of serial dilutions of an aqueous sample. A
temperature controlled photometric device quantifies the light output in each
suspension before and after sample addition. Reduction of light output reflects
physiological inhibition which indicates presence of toxic constituents in the sample.
The small sample volume required (-10 ml_) is a positive aspect of the system.
The instrument used in this project was the Beckman Microtox® Model 2055
Toxicity Analyzer System. Except for slight modifications, procedures indicated in the
15
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Microtox© System Operating ManuaK14) were followed. A summary of the procedure
that was used is included in Appendix A.
EC«;n Evaluation
The ECso is the concentration of a chemical that causes a 50% decrease of
light produced by luminescent bacteria in the Microtox® test. The method provides for
simultaneous testing of a control and dilutions of a chemical. The percent light
decrease after 5 minutes is plotted against sample concentration. The concentration
that diminishes the light output by 50% is designated the ECso under the defined
conditions of exposure time and test temperature. When calculating ECso data,
responses for all concentrations are normalized against blank responses by
multiplying the initial light output of each concentration by the mean blank ratio for time
f. This normalization corrects for effects of light drift and offsets in light output due to
dilution. An explanation of the analytical and calculation methods used to obtain
EC5Q data is presented in Appendix A.
ECso evaluations for the specific chemicals are presented in Table 5. Results
for the munitions chemicals and sludge are presented in Section 8. These ECso
values represent values for the chemicals and sludge as a solution of the chemical
and of the as-received sludge. The values are not comparable to the £650 values that
might occur in a soil-chemical or soil-sludge mixture. These £650 values are useful
as: (a) a relative screening evaluation for the chemicals to estimate which chemicals
have a greater toxicity potential, and (b) input to decisions about concentrations that
can be applied to soil that will be non-inhibitory to the soil microorganisms.
In Table 5, the 95% confidence values (95% Cl) of the EC5Q values also are
presented. The 95% Cl was calculated in a standard statistical manner. Specific
details about the procedure are presented in Section 5. The 95% Cl identifies the
16
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TABLE 5. EC5Q DATA FOR CHEMICALS EVALUATED IN THIS STUDY
Compound
<50-
Values lma/l)
Value
95% Cl
Acid Extractables
Phenol 26.7 25.4-28.1
o-Cresol 22.4 21.8-23.0
p-Cresol 1.1 1.07-1.16
m-Cresol 5.8 5.6-6.1
2-Chtorophenol 16.1 15.1-17.1
3-Chlorophenol 3.8 3.4-4.3
4-Chlorophenol 1.0 0.9-1.0
2,3-Dichlorophenol 4.8 4.7-5.0
2,4-Dichlorophenol 2.8 2.7-2.9
2,5-Dichloropnenol 8.2 7.8-8.6
2.6-Dichlorophenol 15.9 14.8-17.1
3,4-Dichlorophenol 0.5 0.4-0.5
2,4.5-Trichtorophenol 0.9 0.8-1.0
2,4.6-Trichlorophenol 9.3 8.6-10.1
Pentachlorophenol 1.1 1.1-1.2
2.4-Dimethylphenol 3.8 3.4-4.3
2-Methyl-4-Chtorophenol 1.1 1.06-1.14
3-Methyl-4-Chtorophenol 0.3 0.2-0.3
3-Methyl-6-Chtorophenol 5.8 5.7-6.0
p-Nitrophenol 7.0 6.7-7.3
2,4-Dinitrophenol 13.5 11.0-16.6
4,6-Diritro-o-Cresol 10.5 9.1-12.1
Thiophenol 2.8 2.6-3.1
Compound
«50-
Values fma/n
Value
95% Cl
Amines
Diphenylarrine 2 1.9-2.1
m-Phenylenediamine 112 99-122
Toluenediamine 44 41-47
Brucine 213 197-230
Alcohols
Isobutyl alcohol 1740 1590-1920
AByl alcohol 1120 960-1320
Propargyl alcohol 106 99-115
1-Butanol 2400 2180-2640
2,3-Dichloropropanol 120 104-140
Methanol >84.500 -
Other
Carbon bisulfide 30...
2-NHropropane 49...
Thiourea 4530.
27-32
41-59
.4050-5080
-------
range of values within which the estimated true mean EC$Q value should occur with
the probability of error being 0.05. The relatively narrow 95% Cl values indicate the
low scatter of the data obtained.
Low ECso values indicate a higher toxicity potential and the higher £650
values indicate a low toxicity. In Table 5, the EC$Q data indicate that phenol (£650 -
26.7) has a lower toxicity potential than p-Cresol (EC$Q = 1.1).
The comparison of the EC$Q data (Tab|e 5) indicates that: (a) the alcohols are
less toxic than the acid extractables, and (b) within particular categories, there are
considerable differences in relative toxicity. It appeared that there was a relationship
between the relative toxicity and the chemical structure of the chemical. This was
explored using EC$Q values for several of the acid extractables.
In discussing these relationships, an understanding of the structure of the
phenol compound and where substitutions can occur is helpful. Figure 1 indicates the
basic structure of phenol and indicates that substitutions can occur at five locations
that can be identified by number or name. Thus, a 2-Chlorophenol indicates a
chlorine compound is substituted at the 2- or ortho position. A p-nitrophenol indicates
that a nitro- (NC>2) group is located at the 4- or para position.
6 (ORTHO)
5 (META)
•2 (ORTHO)
3 (META)
4 (PARA)
Figure 1. Schematic of the Phenol Ring and Possible Substitution Positions
18
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Data for fourteen chlorinated phenols were evaluated to identify whether a
relationship between chemical structure and relative toxicity existed in this study. The
EC5Q values for these compounds provide a reasonable data base to consider the
effect of chemical structure. When the £659 values were compared (Table 6), the
relative toxicity of these compounds appeared related to the substitution position of the
chlorine group on the phenol ring. The order of toxic potential was para>meta>ortho.
The order was particularly evident with the mono- and di-chlorophenols.
This order also appeared when the methylphenols were compared (Table 7).
There were not enough data to infer that the same order occurred with nitrophenols,
although it was suggested (Table 7).
The chemical compound that is substituted on the phenol ring also appeared to
have an effect on toxicity (Table 8). Nitro-substituted phenols, even when substituted
in the para position, appear to be less toxic than the methyl- or chloro-substituted
phenols.
While it appears that the chemical structure of a compound, such as a
substituted phenol, affects the relative toxicity of the compound, this study was not
designed to determine such effects or the reason differences occur. The effect of
substitution position and chemical may be due to the influence of physicochemical,
electronic and/or steric properties of the chemical.
ACCEPTABLE LOADINGS
Microbial degradation is the major organic removal mechanism in the soil.
Therefore, microbial inhibition by a chemical or waste can be a limiting factor when
chemicals are added to the soil. A chemical and waste loading determination protocol
for land treatment demonstrations to estimate the non-inhibitory loading has been
proposed^12'13). This protocol combines the leaching potential of the waste and the
toxicity of the leachate to arrive at a non-inhibitory loading. In this study, the
acceptable loading rates were determined by following this protocol which is outlined
19
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TABLE 6. RELATIVE TOXICITY OF CHLORINATED PHENOLS
Compound EC50 Value Substitution
(mg/l) Position
2-Chlorophenol 16.1 ortho
3-Chlorophenol 3.8 meta
4-Chlorophenol 1.0 para
2,3-Dichlorophenol 4.8 ortho, meta
2,4-Dichlorophenol 2.8 ortho, para
2,5-Dichlorophenol 8.2 ortho, meta
2,6-Dichlorophenol 15.9 ortho, ortho
3,4-Dichlorophenol 0.5 meta, para
2,4,5-Trichlorophenol 0.9 ortho, para, meta
2,4,6-Trichlorophenol 9.3 ortho, para, ortho
Pentachlorophenol 1.1 all
2-Methyl-4-Chlorophenol 1.1 ortho, para
3-Methyl-4-Chlorophenol 0.3 meta, para
3-Methyl-6-Chlorophenol 5.8 meta, ortho
TABLE 7. RELATIVE TOXICtTY OF NON-CHLORINATED PHENOLS
Compound ECso Value Substitution
(mg/l) Position
Methylphenols
o-Cresol (2-Methylphenol) 22.4 ortho
p-Cresol (4-Methylphenol) 1.1 para
m-Cresol (3-Methylphenol) 5.8 meta
2,4-Dimethylphenol 3.8 ortho, para
Nitrophenols
p-Nitrophenol 7.0 para
2,4-Dinitrophenol 13.5 ortho, para
20
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in Table 9. The logic behind this procedure is that other studies^, 13) have shown
that the detoxification of water soluble organics in soil did not occur or proceeded very
slowly when the ECso value of the initial mixture was less than 20%.
When determining an acceptable chemical loading, it is assumed that any
toxicity that results is due to the added chemical and not the soil. This assumption was
evaluated and the toxicity of the WSF of the Texas and Mississippi soils was
determined using the Microtox© procedure. No toxicity of either soil was found.
The acceptable loading rates that resulted when the specific chemicals were
mixed with two soils are noted in Tables 10 and 11. These acceptable loading data
should be viewed as initial screening data for degradation and land treatment
demonstration studies. The protocol employs only an acute toxicity testing. The
soiksolution ratio used (1:4) is low and simulates only short-term leaching effects. In
the degradation studies (Section 5), the actual loading rates used were equal to or
less than those noted in Tables 10 and 11.
The loading limit of several of the alcohols was not determined. This was
because the large quantities of such soluble chemicals that were needed saturated
the soil and caused nonrepresentative land treatment conditions.
The effect of chemical structure on the acceptable loading rate also was
evaluated. The chlorinated phenol data (Table 12) indicated that the chemicals with
chlorine in the para position generally required the lower non-inhibitory loading rates.
The mixing of the chemicals with the two different soils did not appear to affect the
relative toxicity order of the chemicals discussed earlier.
In general, the acceptable loading rates for the Mississippi soil were lower than
those for the Texas soil. There was, however, no consistent pattern for the differences.
Some of the rates were close and some differed by an order of magnitude. The
experiments were not planned to investigate the reasons for differences with different
21
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TABLE 8. COMPARATIVE RELATIVE TOXICITY OF CHLORO-,
METHYL- AND NITROPHENOLS
Compound EC50 Value
(mg/l)
3-Chlorophenol 3.8
m-Cresol (3-Methylphenol) 5.8
p-Nitrophenol 7.0
p-Cresol (4-Methylphenol) 1.1
4-Chlorophenol 1.0
2,4-Dinitrophenol 13.5
2,4-Dimethylphenol 3.8
2,4-Dichlorophenol 2.8
TABLE 9. METHOD TO DETERMINE ACCEPTABLE INITIAL
CHEMICAL LOADINGS
1. Prepare several different ratios of a waste-soil mixture or a chemical-soil mixture. This results
in different loading rates.
2. Obtain a water soluble fraction (WSF) for each mixture, i.e., loading rate.
3. Determine the EC50 for each WSF.
4. Determine relative toxicity units using the following equation: TU = 400/EC5Q.*
5. Prepare a log-log plot of TU values versus loading rate.
6. The interception point for 20 toxicity units (20TU) is the lower loading limit for the soil. Since
there can be a window of acceptable lower loading rates, a value of twice the tower limit is
identified as the upper limit for this acceptable window.*
See Appendix A for discussion of these items.
22
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TABLE 10. ACCEPTABLE NON-INHIBITORY LOADING RATES -- TEXAS SOIL
ro
oo
Compound
Loading Rate*
(mg/kg of soil)
Acid Extractabtes
Phenol
o-Cresol
p-Cresol
m-Cresol
2-Chlorophenol
3-Chlorophenol
4-Chlorophenol
2,3-Dichlorophenol
2,4-Dichlorophenol
2,5-Dichlorophenol
2,6-Dichlorophenol
3,4-Dichlorophenol
2,4,5-Trichlorophenol
2,4,6-Trichlorophenol
Pentachlorophenol
2,4-Dimethylphenol
2-Methyl-4-Chk>rophenol.
3-Methyl-4-Chk>rophenol.
3-Methyl-6-Chtorophenol.
p-Nitrophenol
2,4-Dinitrophenol
4,6-Dinitro-o-Cresol
Thiophenol
...720
...490
...100
...124
...380
...120
....88
...130
....90
...300
...630
30
....40
...300
30
88
48
18
...150
....250
...1380
....350
....160
Compound
Loading Rate*
(mg/kg of soil)
Amines
Diphenylamine 230
m-Phenylenediamine 7900
Toluenediamine 1080
Brucine 3600
Alcohols
Isobutyl alcohol ++
Alfyl alcohol ++
Propargyl alcohol 8200
1-Butanol ++
2,3-Dichloropropanol 1050
Methanol ++
Other
Carbon disulfide 400
2-Nitropropane 1620
Thiourea 6580
+ lower loading rate as determined using the procedure in Table 9
++ not determined, see text
-------
TABLE 11. ACCEPTABLE NON-INHIBITORY LOADING RATES - MISSISSIPPI SOIL
ro
Compound
Loading Rate*
(mg/kg of soil)
Acid Extractables
Phenol 320
o-Cresol 250
p-Cresol 50
m-Cresol 130
2-Chlorophenol 290
3-Chlorophenol 60
4-Chlorophenol 45
2,3-Dichlorophenol 60
2,4-Dichlorophenol 45
2,5-Dichlorophenol 30
2,6-Dichlorophenol 50
3,4-Dichlorophenol 23
2,4,5-Trichtorophenol 30
2,4,6-Trichlorophenol 215
Pentachtorophenol 55
2,4-Dimethylphenol 45
2-Methyl-4-Chtorophenol 38
3-Methyl-4-Chtorophenol 10
3-Methyl-6-Chtorophenol 140
p-Nitrophenol 45
2,4-Dinitrophenol 270
4,6-Dinitro-o-Cresol 26
Thiophenol 330
Compound
Loading Rate*
(mg/kg of soil)
Amines
Diphenylamine 110
m-Phenylenediamine *
Toluenediamine 1400
Brucine 3200
Alcohols
Isobutyl alcohol ++
Allyl alcohol ++
Propargyl alcohol 5100
1-Butanol ++
2,3-Dichtoropropanol 700
Methanol ++
Other
Carbon disulfide 1540
2-Nitropropane 1170
Thiourea 910
+ lower loading rate as determined using the procedure in Table 9
++ not determined, see text
* chemical deterioration of this chemical in this soil prevented evaluation of mass loading data
-------
TABLE 12. COMPARATIVE ACCEPTABLE LOADING RATES
CHLORINATED PHENOLS
Loading Rate (ma/ka)
Compound Texas Mississippi Substitution
Soil Soil Position
2-Chlorophenol 380 290 ortho
3-Chlorophenol 120 60 meta.
4-Chlorophenol 88 45 para
2,3-Dichlorophenol 130 60 ortho, meta
2,4-Dichlorophenol 90 45 ortho, para
2,5-Dichlorophenol 300 30 ortho, meta
2,6-Dichlorophenol 630 50 ortho, ortho
3,4-Dichlorophenol 30 23 meta, para
2,4,5-Trichlorophenol 40 30 ortho, para, meta
2,4,6-Trichlorophenol 300 215 ortho, para, meta
Pentachlorophenol 30 55 al
2-Methyl-4-Chlorophenol 48 38 ortho, para
3-Methyl-4-Chlorophenol 18 10 meta, para
3-Methyl-6-Chlorophenol 150 140 meta, ortho
soils. Therefore, it was not possible to identify the fundamental factors causing the
observed differences. However, there are several possibilities.
The concept of determining an acceptable chemical loading for a soil is to
identify an application rate or range of rates that will ensure degradation of waste
without inhibiting the microorganisms in the soil. The chemical loading determination
procedure consists of two aspects: (i) quantity of the chemical partitioning into the
water soluble fraction (WSF) under standard test conditions, and (ii) toxicity of WSF.
The first aspect is a function of sorption characteristics of the chemical under the
test conditions. The second aspect depends on the relative toxicity of the chemical,
the impact of soil characteristics on the toxicity, and the method used to measure
toxicity.
25
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The sorption of a chemical in soil depends on: (i) soil characteristics such as
pH, organic content, particle size distribution, and presence of other chemicals, (ii)
chemical characteristics such as solubility, partition coefficient, and pK value, and (iii)
test conditions such as soiksolution ratio, test temperature and agitation.
The sorption data obtained in this study are presented in Section 6 and there
are differences in the sorption characteristics of the two soils. Sorption increases the
persistence of a chemical in the soil and reduces the migration potential of the
chemical. Therefore, a chemical that has greater sorption characteristics will be more
tightly bound to the soil and less likely to be in solution and therefore in the WSF. The
basis of the procedure to estimate the acceptable chemical or waste loading rate
(Table 9) is the assumption that the WSF of the chemical or waste poses the
immediate and significant threat to soil microorganisms and to groundwater.
The results that were obtained were similar to those obtained by other
researchers. Sims(1^) reported waste mass loading data for petroleum and wood-
preserving wastes with clay loam and sandy loam soils. The same procedure (Table 9)
was used to obtain the data. In the clay loam soil, higher chemical loadings were
possible than with the sandy loam soil. This was attributed to the higher adsorption
and hence lower leaching of the wastes in clay loam soil. Pentachlorophenol (PCP)
waste showed a lower loading than the other wastes containing creosote and
polynuclear aromatic hydrocarbons (PAH's). This may have been due to the higher
toxicity of PCP using the Microtox® assay.
The pH of the Microtox® method also may have had an effect on speciation of
the chemicals that were available for sorption and the resultant relative toxicity. The
acceptable pH of the sample in the Microtox® assay is the range of 6.0-8.0. The pH of
the samples analyzed in this study was within this range. The chemicals evaluated
have pKa values that range widely. As an example, the pKa of pentachlorophenol
(PCP) is about 5.0 while that of phenol is about 9.3. When a sample is adjusted to the
26
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Microtox© pH range (6.0-8.0), the speciation will vary. For instance, at this pH range,
POP would be completely dissociated, whereas the dissociation of phenol would be
minimal.
The method (Table 9) used to identify acceptable, non-inhibitory chemical and
waste loading rates is conservative in that acclimation of the organisms to the waste or
chemicals is not considered, in the field the actual non-inhibitory waste and chemical
loading rates to the soil may be higher than those noted in this study since microbial
acclimation will occur with time.
The higher the chemical or waste loading that can be successfully treated on
the land, the more economical will be the land treatment option. Huddleston et al.(16)
reported increased soil respiration rates and waste removal rates with an increase in
oil waste application in land treatment investigations. The microbial acclimation may
significantly increase the degradation of a waste at high waste application
concentrations^ 5> 17) However, the method in Table 9 does result in an initial
chemical and waste loading that will not inhibit the soil microorganisms. Such
loadings will allow the soil treatability of chemicals and wastes to be determined.
CONCLUSIONS
1. The Microtox© biological assay represents an inexpensive and expedient
method with which to evaluate the EC$Q toxicity of a chemical or waste. This
method was used to estimate the £650 values of specific chemicals and to
estimate the non-inhibitory soil loadings of the chemicals and waste that were
evaluated.
2. The comparison of the EC$Q data indicated that: (a) the alcohols were less toxic
than the acid extractable compounds, and (b) within chemical categories, there
were considerable differences in relative toxicity.
27
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3. The chemical structure of the compounds evaluated affected the relative toxicity
of a compound. With chlorophenols, the relative toxicity was related to the
substitution position of the chlorine group on the phenol ring. The order of
relative toxicity was para>meta>ortho. The EC$Q data suggested that the same
order occurred for methylphenols and nitrophenols.
4. The chemical that was substituted on the phenol ring appeared to have an
effect on toxicity. Nitro-substituted phenols, even when substituted in the para
position, appeared to be less toxic than the methyl- or chioro-substituted
phenols.
5. When the chemicals were mixed with two different soils, and the £€50 value of
the water soluble fraction (WSF) of the soil mixtures was measured, the values
indicated that chemicals with the chlorine in the para position had the greater
toxicity. Mixing of the chemicals with the soils did not affect the relative toxicity
of the chemicals in the two soils.
6. In general, the acceptable non-inhibitory chemical loading rates for the
Mississippi soil were lower than those for the Texas soil. There was no
consistent pattern for the differences. The differences were likely due to
different sorption characteristics of the chemicals in the two soils.
28
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SECTION 5
DEGRADATION STUDIES
INTRODUCTION
These experiments were conducted to determine the removal kinetics of the
designated chemicals and wastes (Table 1) in soil. Several of the chemicals could not
be evaluated because of chemical reactions in the soils which made analytical
detection impossible. These were Diphenylamine, m-Phenylenediamine and
Thiophenol.
Biodegradation is believed to be the most important removal mechanism for
organic compounds in soil systems. Biodegradation of organics is accomplished in a
series of biochemical reactions through which a parent compound is changed or
transformed to organic and inorganic end products. Complete degradation is the term
used to describe the process whereby constituents are mineralized to inorganic end
products, including carbon dioxide, water, and inorganic nitrogen, phosphorus, and
sulfur compounds.
Aerobic soil bacteria possess the ability to biochemically catalyze the oxidation
of organic compounds. For this reason, and because the zone of incorporation at land
treatment sites generally is aerobic, the protocol used in this study allowed aerobic
conditions and aerobic biodegradation reactions to occur.
In the mixed microbial population of soil systems, one metabolic group of
microorganisms may partially metabolize a compound and may furnish a suitable
growth substrate for another group. Organic compounds also may be partially
degraded or transformed to organic intermediates that may be recalcitrant and/or toxic.
The primary goal of biodegradation testing is to obtain an overall estimate of the
rate at which a compound will biodegrade in a soil environment. While few
compounds appear in the environment in pure form, a common approach for studying
29
-------
removal rates of organic compounds has been to evaluate individual compounds.
Although this approach provides an understanding of the removal rates for specific
compounds, it is recognized that during actual land treatment chemicals normally are
applied as mixtures. Interactions between compounds in a mixture within the soil
matrix may promote or inhibit their removal from soil.
In this study, the chemicals were evaluated as individual compounds and the
data should be understood in that context.
Methods used to evaluate the biodegradation of organics in the environment
commonly use indirect measures such as oxygen consumption, CC>2 evolution, and
dissolved organic carbon (DOC) loss, to assess the persistence of compounds in test
environments, to determine chemical loss rates and to predict the relative importance
of biodegradation.
While these procedures provide a qualitative assessment of biodegradation,
they do not determine quantitative rates of degradation for specific constituents. Such
chemical specific rates are essential for the assessment of compound fate and
transport as well as for risk analyses. For quantitative assessment of the rate of
biodegradation of an individual constituent in a soil system, it is necessary to measure
changes in parent compound concentration with time, and the loss of a chemical due
to methods other than biodegradation. In addition, the immobilization potential
(partitioning into soils, liquid, and gaseous phases) provides additional information for
assessing the soil treatment potential of hazardous constituents.
In this study, no distinction was made between specific loss mechanisms. Thus
removal rates can be due to biodegradation, chemical degradation, hydrolysis,
photolysis and volatilization. The chemicals that were evaluated did not have a high
volatilization potential and volatilization was not considered an important removal
mechanism in these degradation experiments. The logic for this statement is
30
-------
presented in Appendix C. Thus, the degradation rates presented in this section were
not corrected for volatilization.
MATERIALS AND METHODS
In this study, the rate of degradation was experimentally determined by
measuring the difference between the amount of compound initially added to a soil
and that which was recovered after specified time intervals. The general protocol used
to determine these differences is presented in Table 13. The protocol notes that the
soils should be maintained at a moisture content which is about 80% of field capacity.
This will provide adequate moisture for the biodegradative reactions but avoid
saturated conditions. For the two soils that were used, the equivalent of 80% field
capacity was a moisture content of about 12% for the Mississippi soil and about 16%
for the Texas soil.
Soil used in this experimental program: (a) had not had previous exposure to
industrial chemicals or wastes, and (b) did not receive any pretreatment such as soil
amendments or specially acclimated biological cultures prior to these experiments.
Thus, the naturally occurring soil microbial consortium was responsible for the
bioremediated removal of the chemicals.
Chemical mass loadings were determined as part of the toxicity screening
evaluations (Section 4). These determinations ensured that the loadings at which the
chemicals were applied did not inhibit soil microbial activity. Soil pH was not adjusted
nor were supplementary organic substrates used. The control beaker (blank) was
carried through the experimental procedure to ensure quality control of the
instrumental analysis.
Various techniques can be used to apply test compounds to soil in laboratory-
scale studies. When the test compounds were not highly water soluble, the test
compound was dissolved in a small aliquot of solvent before application into a soil. In
31
-------
these experiments, the chemicals were added to the soil as a solution of water or of
Methylene Chloride depending on their solubility characteristics.
To minimize the possible toxic effect of the Methylene Chloride, a small volume
(100 jiL) of the solvent, which contained an appropriate amount of chemical for a
specific initial concentration, was applied to each soil sample and mixed thoroughly
immediately after application with glass stirrers. Each beaker contained a glass stirrer
to prevent possible cross-contamination. A 20 JJ.L microdispenser was utilized to
distribute the 100 JJ.L solvent/chemical at five different points on the surface of the soil
sample. The Methylene Chloride also was added in the above volume to the control
beakers. Prior to applying the aluminum foil cover, a brief time was allowed for
volatilization of the Methylene Chloride.
As noted in Table 13, at selected time intervals the concentration of the
remaining chemical in a set of beakers was determined. The time intervals were
based on estimates of the half-life of the chemical and were chosen to provide at least
five data points to establish the removal rates. At these sampling periods, a sample
set (four beakers - one blank and three with chemical) was taken from the constant
temperature room and extracted for sixteen hours with either Methylene Chloride for
phenolic compounds or Methanol for amines (including Brucine and Thiourea) in a
Soxhlet extraction apparatus (Method 3540)(7).
The extract was concentrated in a Kuderna-Danish extraction unit attached to a
three-ball Snyder column (Method 3540)(7). The concentration step was conducted in
a water bath maintained at 60° C for phenolic compounds and at 78° C for amines.
The concentrated extracts were dried by passing them through disposable sodium
sulfate columns and then refrigerated at 4° C until analysis by gas chromatography
(GC) or high pressure liquid chromatography (HPLC).
Several of the chemicals were extracted using water as a solvent and the shake
32
-------
TABLE 13. EXPERIMENTAL PROCEDURES USED IN THE DEGRADATION STUDIES
Each experiment consisted of eight sample sets. Each sample set contained four beakers
(triplicates for a sample and one blank). The experimental procedure for each chemical was as follows;
(a) Place 10 g of air-dried soil, which has been kept at 4° C, into each 150 ml beaker and adjust the
soil moisture content to 80% of field capacity with distilled deionized water. Place a glass stirrer in
each beaker. Record all weights, including beaker, soil, water and glass stirrer.
(b) Mix soil and water thoroughly, cover the beakers with aluminum foil and place the beakers in the
dark at 20° C. The cover minimizes water loss and the possible addition of contaminated dust.
(c) Wait for 10 days to allow soil microorganisms to equilibrate to experimental conditions.
(d) Prepare solutions of the test chemicals so that 100 uL of the solution gives the desired mass
loading rate. The mass loadings are based on toxicity screening results.
Add 100 uL of solution into each sample beaker using a 20 u.L micropipet. Do this five times to
distribute the solution to five points on the soil surface and mix thoroughly. Return the beakers to
the constant temperature room.
(e) Sample beakers should be arranged so that the chemical solution is added to the day 0 beaker
last. For example, if samples are scheduled at day 0, 2, 4, 8,16, 24, 32, and 64, the order for
adding a chemical to soil is sample sets for day 64,32,24,16,8,4,2 and day 0.
(f) The beakers are incubated in the dark to prevent photodegradation of the added chemicals.
(g) During the study, adjust the moisture content in each beaker weekly and maintain between 60%
and 80% of field capacity.
(h) Sacrifice sample sets at the selected time intervals for test chemical analysis.
(i) When experimental data show that the chemical remaining in the soil is below GC or HPLC
detection limits, the remaining sample beakers can be discarded.
extraction procedure. The compounds extracted in this manner are noted in Table 14
and the extraction procedure is summarized in Table 15.
DATA ANALYSIS
Biodegradation information can be used to identify the rate of loss of an organic
chemical in a soil. Such information results from treatability studies such as those
conducted in this study. Biodegradation rates were determined experimentally by
applying the chemical of interest to a soil microcosm and monitoring concentration
over time. A plot of the disappearance of a constituent originally present in the
chemical/soil mixture versus treatment time provided the following: (a) the type of
reaction (generally zero or first order), (b) the reaction rate constants for the zero or first
order reactions, and (c) the half-life (l-\/2) time of each constituent of concern.
33
-------
TABLE 14. CHEMICALS WHOSE SOIL CONCENTRATION WAS EVALUATED USING
SHAKE EXTRACTION
Compound Solvent Used
Methanol water
1 -Butanol water
Isobutyl Alcohol water
2,3-Dichloropropanol Methylene Chloride
2-Nitropropane water
Allyl Alcohol water
Propargyl Alcohol water
TABLE 15. SHAKE EXTRACTION PROCEDURE
Each experiment consisted of eight sample sets. Each sample set contained four beakers
(triplicates for a sample and one blank). The experimental procedure for each chemical was as follows.
(a) Place 40 g of air-dried soil, which has been kept at 4° C, into each of the 200 ml mason jars and adjust
the soil moisture content to 80% of field capacity with distilled deionized water. Record all weights,
including beaker, soil, water and glass stirrer.
(b) Mix soil and water thoroughly then cap reactors with screw-on top and place in 20° C incubator.
(c) Wait for 10 days to allow soil microorganisms to equilibrate to experimental conditions.
(d) Prepare solutions of the test chemicals so that 100 u.L of the solution gives the desired mass loading
rate. The mass loadings are based on toxicity screening results.
Add 100 u.L of solution into each sample beaker using a 20 ul micropipet and mix thoroughly. Return
the reactors to incubator.
(e) On day of analysis, four reactors are removed. Each is filled with 200 milliliters of the appropriate
extracting solvent. The reactors are sealed with the gas tight cap and placed on the shaker apparatus.
(f) The shaker apparatus is operated at 250 rpm for one hour. After this period, the reactors are allowed
to settle for 15 to 30 minutes.
(g) The supernatant from the reactors is decanted and centrifuged at 5000 rpm for 30 minutes.
(h) After centrifugation, the supernatant is filtered using a 0.45 ^m pore size filter and the filtrate stored at
4° C until analysis on the gas chrornatograph.
34
-------
First and zero order kinetic relationships were used to model data from the
degradation studies. The following describes both kinetic relationships.
The expression for first order kinetics is:
-dC/dt =kC (1)
where C is the chemical concentration (mg of compound/kg of soil), t is time (days),
and k is the first order kinetic constant (day '1). The integrated form of Equation 1 is:
C = C0 exp (-kt) (2)
where Co is the initial concentration of chemical.
Taking the natural logarithm of Equation 2 results in:
In C = (In C0) -kt (3)
Using linear regression, the relationship between In C and t (time) data allows
determination of the loss coefficient k (which is the slope of In C versus t curve).
Half life (t-|/2) is defined as the time required for the amount of chemical to
decrease to half of its initial value. Half life values based on first order kinetics are
obtained by rearranging Eqn. 3:
ti/2 = lrt2/k (4)
The common expression for zero order kinetics is:
-dC/dt m K (5)
where K is the zero order kinetic constant (mg/Kg/day). The integrated form of
Equation 5 is:
C= (C0) -Kt
The zero order kinetic constant K may be estimated by analyzing C versus t data by
linear regression.
Data obtained from each sampling interval were used to calculate both first order
and zero order kinetic parameters. Half-life data were calculated based on first order
kinetics. Total loss rate data (mg chemical/kg dry soil/day) were reported for zero
order kinetics.
35
-------
The zero and first order kinetic parameters and the correlation coefficient were
obtained from a least squares fit of the data. To estimate the 95% confidence interval
for the data, the standard deviation and the student t statistic were computed based on
the data and used as
t«s
95% Cl = (k or K) ±
where: k or K = least squares calculated first or zero order kinetic parameter,
t = student t statistic for the data and 95% confidence,
s = sample standard deviation, and
ssx = sum of squares for the data.
Recovery efficiency data for each chemical were acquired and used to
determine the chemical concentration that actually remained in the soil. The day zero
extraction data identified the extraction efficiency of the chemical under the conditions
of that specific experiment. It was assumed that the day zero extraction efficiency was
constant throughout the specific experiment. The recovery efficiencies were used to
calculate the actual soil concentration prior to extraction as follows. Assume that: (a)
the recovery efficiency was 50% as shown by the day zero data and (b) the extracted
soil concentration for the chemical at sampling time three was 100 mg/kg soil. The
actual recovery efficiency corrected chemical concentration at this sampling time was
calculated to be 200 mg/kg (extraction concentration divided by extraction efficiency).
The recovery efficiency corrected chemical concentrations were used in all
calculations to determine the kinetic relationships and constants.
Because of time constraints, the appropriate analytical protocol for measuring
Carbon Disulfide (CS2) was not developed. Thus, this chemical is not included as
part of these experiments.
36
-------
RESULTS
Recovery Efficiency
Recovery efficiency data for the selected chemicals are given in Table 16.
There were differences in recovery efficiencies for the two soils but there were no
consistent patterns. The differences may be due to the different sorption
characteristics of the soils with specific chemicals.
Kinetic Parameters
The loading rates for the chemicals to the soil were at or below the acceptable
loading rates determined as described in Section 4 (Table 17). In some cases, the
actual loading rates were slightly above the lower acceptable loading rate but were
within the acceptable loading rate window. In one case (4,6-Dinitro-o-Cresol in
Mississippi soil) (Table 17), the actual loading rate inadvertently was considerably
higher than the acceptable loading rate. Except in the latter case, the loading rates
should not have been such to inhibit the microorganisms in the two soils during the
degradation experiments.
The first and zero order rate constants, correlation-coefficients, and 95%
confidence intervals for the selected chemicals in Texas and Mississippi soils are
given in Tables 18 and 19 respectively. Chemical loss curves illustrative of the type
obtained in this study are shown in Figures 2-5.
In reviewing the kinetic parameters, it should be recalled that a small half-life
and a large zero order constant indicate a faster degradation rate. For example, in the
Texas soil, p-Cresol had a higher degradation rate and was removed faster in the
microcosms than was phenol.
In general, high correlation coefficients were obtained with the data. Higher
correlation coefficients were obtained with the acid extractables in both soils. Based
on the correlation coefficient data and a visual inspection of the chemical loss curves,
37
-------
TABLE 16. RECOVERY EFFICIENCIES* FOR SPECIFIC CHEMICALS (%)
Texas Mississippi
Compound Soil Soil
Acid Extractable*
Phenol 88 78
o-Cresol 57 63
p-Cresol 78 97
m-Cresol 81 88
2-Chtorophenol 23 25
3-Chlorophenol 94 85
4-Chtorophenol 97 85
2,3-Dichlorophenol 115 104
2.4-Dichtorophenol 82 92
2,5-Dichtorophenol 88 84
2,6-Dichlorophenol 80 71
3,4-Dichtorophenol 115 104
2,4,5-Trichlorophenol 82 92
2,4,6-Trichlorophenol 88 84
Pentachlorophenol 115 106
2,4-Dimethylphenol 80 81
2-Methyl-4-Chlorophenol 91 101
3-Methyl-4-Chlorophenol 99 99
3-Methyl-6-Chlorophenol 100 100
p-Nitrophenol 110 110
2,4-Oinitrophenol 95 132
4,6-Dinrtro-o-Cresol 84 100
Amln»*
Toluenediamine 100 72
Brucine 65 100
Alcoholt
Isobutyl alcohol 92 94
Allyl alcohol ,119 110
Propargyl alcohol 95 97
1-Butanol 90 89
2,3-Dichloropropanol 100 86
Methanol 100 106
Other
2-Nitropropane 60 55
Thiourea 88 93
+ Average of results from triplicate samples
38
-------
TABLE 17. ACCEPTABLE AND ACTUAL LOADING RATES USED IN THE
DEGRADATION STUDIES (mg/kg of SOIL)
Texas Soil Mississippi Soil
Compound Acceptable* Actual Acceptable* Actual
Add Extnctable*
Phenol 720 700 320 350
o-Cresol 490 500 250 250
p-Cresol 100 100 50 45
m-Cresol 124 120 130 130
2-Chtorophenol 380 400 290 300
3-Chtorophanol 120 120 60 55
4-Chtorophenol 88 90 45 50
2,3-Dichtoroph«nol 130 130 60 60
2,4-Dichbroph«K>l 90 90 45 40
2,5-Dichtoroph«nol 300 300 30 30
2,6-Dfchlorophenol 630 630 50 48
3,4-Dichk>rophenol 30 30 23 22
2.4,5-Trichlorophenol 40 40 30 30
2.4.6-Trichloroph«nol 300 300 215 220
Pentachlorophenol 30 30 55 50
2,4-Dimethylphenol 88 90 45 40
2-Methyl-4-Chlorophenol 48 50 38 40
3-Methyl-4-Chloroph«nol 18 20 10 10
3-Methyl-6-Chloroph«nol 150 150 140 140
p-Nitrophenol 250 250 45 50
2,4-Dinitroph«nol 1380 140 270 330
4.6-Dinitro-o-Cresol 350 300 26 105
Amines
Toluenediamine 1080 500 1400 680
Brucine 3600 180 3200 150
Alcohol*
Isobutyl alcohol ++ 925 ++ 940
Allyl alcohol ++ 750 ++ 700
Propargyl alcohol 8200 980 5100 930
1-Butanol ++ 870 -H- 850
2,3-Dichloropropanol 1050 1200 700 700
Methanol ++ 740 ++ 740
Other
2-Nitropropane 1620 640 1170 540
Thiourea 6580 660 910 100
+ As determined by the procedure described in Section 4, data from Table 10
' As determined by the procedure described in Section 4. data from Table 11
++ Sea text, Section 4: acceptable loading rate was considerably greater than 1000 mg/kg.
39
-------
TABLE 18. LOSS RATES, CORRELATION COEFFICIENTS AND 95% CONFIDENCE
INTERVALS FOR SPECIFIC CHEMICALS --
TEXAS SOIL
COMPOUND
KINETIC PARAMETERS
FIRST ORDER
HALF LIFE
(day)
Acid Extractables
Phenol
o-Cresol
p-Cresol
m-Cresol
2-Chlorophenol
3-Chterophenol
4-Chtorophenol
2,3-Dichlorophenol
2,4-Dichtorophenol
2,5-Dichlorophanol
2,6-Dichtorophenol
3,4-Dichbrophenol
2,4,5-Tr ichlorophenol
2,4,6-Trichlorophenol
Pentachlorophenol
2,4-Dimethylphenol
2-Methyl,4-Chlorophenol
3-Methyl-4-Chlorophenol
3-Methyl, 6-Chlorophanol....
p-Nitrophenol
2,4-Dinitrophenol
4,6-Dintro-o-Cresol
Amln»»
Toluanediamine
Brucine
Alcohol*
Isobutyl Alcohol
Allyl Alcohol
Propargyl Alcohol
1-Butanol
2,3-Dichloropropanol
Methanol
Othfr
2-Nitropropane
Thiourea
.4.1
.1.6
..(1)
.0.6
.1.7
.21.8.,..
.1.0
.8.3
.1.5
.16.6....
.2.4
.3.2
.14.6....
.5.3
.6.7
.0.7
.2.9
.1.4
.2.1
.10.2....
.4.6
. (2) ....
.11.9....
.23.1....
.2.4
,10.2....
.12.6....
.1.0
.23.1....
.1.0
.0.5
.12.8....
*
r
0.92.
0.83.
«,.
0.87.
0.98.
0.97.
0.91.
0.97.
0.94.
0.99.
0.91.
0.89.
0.95.
0.94.
0.92.
0.99.
0.90.
0.99.
0.98.
0.81.
0.92.
..
0.65.
0.86.
0.96.
0.95.
0.94.
0.75.
0.97.
0.75.
0.97.
0.86.
95% C.I,
3.1-6.1..
1.5-1.7..
„
0.4-1.4..
1.5-1.9..
19.6-24.6
0.7-1.3..
7.4-9.3..
1.1-1.9..
15.6-17.7
2.0-3.0..
2.3-4.9..
12.2-18.1
4.6-6.3..
5.5-8.5..
0.6-0.8..
2.7-3.0..
1.2-1.6..
1.9-2.4..
7.5-16.3.
2.9-11.7.
-M
7.5-28.5.
18.0-33.0
2.1-2.9..
8.8-12.2.
10.7-15.2
0.7-2.4..
20.9-26.0
0.68-2.4.
0.4-0.6..
9.9-18.0.
ZERO ORDER
mg/Kg/day r*
59.3...
62.2...
>100 (1)
28.2...
26.2...
2.3....
14.5...
3.6....
14.1...
6.1....
41.8...
2.7....
1.4....
10.4...
1.0... .
27.3...
3.7....
3.8....
11.2...
6.8....
12.9...
(2)
5.9....
3.1....
172...
30.0...
29.5...
184...
22.2...
184...
155...
2.1....
0.96.
0.95.
,.
0.96.
0.90.
0.95.
0.91.
0.98.
1.00.
0.96.
0.97.
0.95.
0.96.
0.94.
0.91.
0.90.
0.94.
0.93.
0.94.
0.92.
0.96.
__
0.47.
0.81.
0.96.
0.93.
0.97.
0.90.
0.94.
0.94.
0.77.
0.95.
95% C.I
46.9-71.4
48.5-75.9
„
20.5-36.9
20.4-32.0
2.0-2.6
9.7-19.2
3.2-3.9
13.0-15.2
5.4-6.8
36.7-46.9
2.1-3.3
1.2-1.6
8.8-12.1
0.8-1.2
18.1-36.5
3.0-4.4
2.1-5.4
9.2-13.1
5.3-8.3
7.5-18.3
,,
0.7-11.2
2.4-4.2
144-200
24.3-35.6
26.2-32.9
130-239
18.7-25.6
140-228
79-230
1.8-2.4
(1) No chemical was detected after one day.
(2) No loss during the 65-day experiment
' Correlation coefficient
40
-------
TABLE 19. LOSS RATES, CORRELATION COEFFICIENTS AND 95% CONFIDENCE
INTERVALS FOR SPECIFIC COMPOUNDS - MISSISSIPPI SOIL
COMPOUND
KINETIC
FIRST ORDER
HALF LIFE
(day)
Acid Extractables
Phenol
o-Crasol
p-Cresol
m-Cresol
2-Chlorophenol
3-Chtorophenol
4-Chlorophenol
2,3-Dichlorophanol
2,4-Dichloroph«nol
2,5-Dichk>roph«nol
2,6-Dichk>roph«nol
3,4-Dichtoroph«nol
2,4,5-Trichlorophenol
2,4,6-Trichlorophanol
Pentachlorophenol
2 4-Oirnathylphenol
2-Mathyl,4-Chlorophanol....
3-Mathyl-4-Chlorophanol....
3-Mathyl, 6-Chlorophenol...
p-Nitrophanol
2,4-Dinitrophenol
4 6-Dintro-o-Crasol
Amlnet
Toluenadiamina
Brucine
Alcohol*
Isobutyl Alcohol
Allyl Alcohol
Propargyl Alcohol
1-Butanol
2,3-Dichloropropanol
Methanol
Other
2-Nitropropane
Thiourea
..23.0
..5.1
..0.5
..11.3
..7.2
..15.1
..2.5
..18.3
..3.5
..18.5
..16.2
..18.3
..22.3
..6.3
..12.0
..1.4
..6.3
..4.2
..12.5
..2.5
..32.1
(1)
..6.5
..37.1 „
..11.3
..9.5
..13.0
..8.5
..55.3
..3.2
..0.66
..18.7
*
r
...0.95.
...0.88.
...0.93.
...0.97.
...0.95.
...0.97.
...0.96.
...0.84.
...0.95.
...0.94.
...0.94.
...0.90.
...0.95.
...0.93.
...0.94.
...0.99.
...0.94.
...0.92.
...0.93.
...0.81.
...0.96.
»
...0.85.
...0.66.
...0.66.
...0.93.
...0.89.
...0.60.
...0.92.
...0.73.
...0.95.
...0.45.
95% C.I.
19.6-27.8
4.0-6.7
0.4-0.8
9.7-13.4
6.3-8.7
13.6-17.1
2.1-3.1
13.9-26.8
2.9-4.6
16-22
13.9-19.5
15.0-23.4
19.1-26.9
5.3-7.8
10.2-14.6
1.2-1.6
5.2-7.7
3.2-5.9
10.6-15.3
1.4-5.8
25.9-42.1
„
:4.9-9.7
21.7-126.3
7.7-20.8
7.9-11.8
10.6-16.6
5.3-21.2
45.8-69.5
2.3-5.6
0.56-0.79
-8.3to+76.2
PARAMETERS
ZERO ORDER
mg/Kg/day
7.0
6.4
23.2
5.0 .-.
8.4
1.3
5.6
1.0
2.3
5.2
8.9
0.5
0.6
5.7
1.1
6.7
1.8
0.9
3.0
3.6
3.7
(1)
12.0
2.7
37.7
26.9
23.7
39.6
10.9
68
115
2.7
r*
.0.93..
.0.97..
.0.88..
.0.96..
.0.88..
.0.95..
.0.90..
.079..
.0.85..
.0.87..
.0.88..
.0.91..
.0.92..
.0.94..
.0.94..
.0.96..
.0.87..
.0.88..
.0.85..
.0.93..
.0.89..
„
.0.57..
.0.59..
.0.73..
.0.96..
.0.96..
.0.79..
.0.91..
.0.96..
.0.84..
.0.50..
95% C.I
5.6-8.4
5.7-7.1
11.8-34.5
4.1-5.9
6.2-10.6
1.1-1.5
3.6-7.5
0.6-1.3
1.3-3.3
3.9-6.5
6.5-10.6
0.4-0.6
0.5-0.7
4.7-6.7
0.9-1.3
4.9-8.5
1.2-2.3
0.5-1.2
2.1-3.9
1.9-5.3
1.5-5.5
2.7-21.2
0.4-4.9
23.1-52.3
23.0-30.8
20.6-26.7
25.9-53.3
8.5-13.4
59-76
76-154
....-0.22 to +5.6
(1) No loss during the 65-day experiment
Correlation coefficient
41
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Loss of Phenol from Texas Soil
o
OT
*»
o
D>
0»
O
4>
Q.
First Order Half Life . 4.1 days
Zero Order Total Loss Rate - 59.3 mg/Kg/day
Figure 2. Loss of Phenol In Texas Soil at 20° C.
Moisture content was maintained between 12 and 16%.
The initial chemical loading was 700 mg/kg.
Loss of Phenol from Mississippi Soil
400
(0
O
300-
at
E
- 200-
O
O
Q.
"5
u
,_ 100-
O
o
Figure 3.
First Order Half Life . 23.0 days
Zero Order Total Loss Rate » 7 mg/Kg/day
Loss of Phenol In Mississippi Soil at 20° C.
Moisture content was maintained between 12 and 16%.
The initial chemical loading was 350 mg/kg.
42
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Loss of 2,6-Dlchlorophenol from Texas Soil
= 600
o
at
o
O)
o»
§
a
o
o
df
o
u
C
o
o
500
400
Jl
300
200-
100-
O
First Order Half Life -2.4 days
Zero Order Total Loss Rate -41.8 mg/Kg/day
a
i
10
40
50
20 30
Days
Figure 4. Loss of 2,6-Dlchlorophenol In Texas Soil at 20° C.
Moisture content was maintained between 12 and 16%.
The initial chemical loading was 630 mg/kg.
Loss of 2,6-Dlchlorophenol from Mississippi Soil
o
o
Q.
O
o
v « i ' i
10 12 14 16 18 20
Days
Loss of 2,6-Dlchlorophenol In Mississippi Soil at 20° C.
Moisture content was maintained between 12 and 16%.
The initial chemical loading was 48 mg/kg.
43
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it was not possible to discern whether zero or first order kinetics were a better
representation of the data.
Generally the rates of chemical loss were higher in the Texas soil than the
Mississippi soil (Table 20). There were a few situations in which the chemical loss
rates were about the same or greater in the Mississippi soil than those in the Texas
soil. There did not appear to be any particular pattern to the differences of rates in the
two soils. Because the study was not designed to determine the reasons that such
differences occur, it was not possible to identify the factors causing the differences.
However, there are several possibilities. These include: (a) different sorption
characteristics and solubilities and therefore different availability of the chemical for
microbial degradation, and (b) the different pH in the soils and the effect of pH on
chemical dissociation and the form of the chemical available for degradation. Some
discussion of these possibilities was presented in Section 4.
Care should be taken in extrapolating these chemical loss rates to field
conditions, since such conditions can be different than those in the laboratory
microcosms. For example, these loss rates were obtained at 20° C, with no nutrient
additions, with a reasonable but narrow range of moisture, without pH control, without
acclimated microorganisms, under quiescent conditions, and each chemical was
evaluated separately. At field sites, moisture and temperature can vary considerably
over the seasons, nutrients may be added to assure microbial growth, and soils may
be limed. Typically, chemicals are added as mixtures and acclimated microorganisms
will exist at the site after repeated waste applications. In addition higher chemical
loading rates can occur. Thus, in attempting to utilize these loss rates to predict
chemical loss at specific sites, the differences between laboratory and field conditions
should be recognized and taken into account.
In Section 4, the data indicated that the substitution position of a chemical on
the phenol ring and the type of chemical group that was substituted affected the
44
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TABLE 20. CHEMICAL HALF-LIVES IN TEXAS AND MISSISSIPPI SOILS (days)
Texas Mississippi
Compound Soil Soil
Acid Extractables
Phenol 4.1 23.0
o-Cresol 1.6 5.1
p-Cresol (1) 0.5
m-Cresol 0.6 11.3
2-Chlorophenol 1.7 7.2
3-Chlorophenol 21.8 15.1
4-Chlorophenol 1.0 2.5
2,3-Dichlorophenol 8.3 18.3
2,4-Dichlorophenol 1.5 3.5
2,5-Dichlorophenol 16.6 18
2,6-Dichlorophenol 2.4 16.2
3,4-Dichlorophenol 3.2 18.3
2,4,5-Trichlorophenol 14.6 22.3
2,4,6-Trichlorophenol 5.3 6.3
Pentachlorophenol 6.7 12.0
2,4-Dimethylphenol 0.7 1.4
2-Methyl-4-Chlorophenol 2.9 6.3
3-Methyl-4-Chlorophenol 1.4 4.2
3-Methyl-6-Chlorophenol 2.1 12.5
p-Nitrophenol 10.2 2.5
2,4-Dinitrophenol 4.6 32.1
Amines
Toluenediamine 11.9 6.5
Brucine 23.1 37.1
Alcohols
Isobutyl alcohol 2.4 11.3
Allyl alcohol 10.2 9.5
Propargyl alcohol 12.6 13.0
1-Butanol 1.0 8.5
2,3-Dichloropropanol ?3.1 55.3
Methanol 1.0 3.2
Other
2-Nitropropane 0.5 0.66
Thiourea 12.8 18.7
(1) No chemical was detected after one day.
45
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relative toxicity and the acceptable loading rate data. Each chemical group and
substitution position also was evaluated to see their effect on the degradation rate
data.
When the half-life data are compared for the fourteen chlorinated phenols with
chlorine in different positions on the phenol ring (Table 21), it appeared that the
compounds with the chlorine substituted in the meta position had the greater half-lives
and therefore the lowest loss rate. This was particularly evident with the mono-, di-,
and trichlorophenols. Using the data in Table 21, it was not possible to discern
whether there were any real differences when the chlorine was in the ortho and para
positions. These results are somewhat different from those in Section 4 where relative
toxicity of these compounds appeared related to the substitution position of the
chlorine group with the order being para>meta>ortho.
When the data from non-chlorinated phenols were compared (Table 22), all of
the methylphenols were lost very rapidly in the Texas soil. In the Mississippi soil, the
chemical loss rate was considerably slower when the methyl group was substituted in
the meta position.
In Table 23, data for chemicals with different compounds substituted on the
phenol ring were compared. The data suggested that compounds that had a nitro
compound substitution on the phenol ring had a lower loss rate. No other differences
in terms of type of compound substitution were apparent.
CONCLUSIONS
1. The chemical or waste loading procedure (Table 9, Section 4) resulted in
chemical loadings that did not inhibit the non-acclimated organisms in the
laboratory microcosms, except in one case (4,6-Dinitro-o-Cresol). Thus, this
procedure provided a good estimate of initial, acceptable chemical loadings
that can be used in laboratory degradation studies.
46
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TABLE 21. EFFECT OF SUBSTITUTION POSITION ON DEGRADATION RATES
CHLORINATED PHENOLS
Compound Chemical Half-Life (davs) Substitution
Texas Soil Mississippi Soil Position
2-Chlorophenol 1.7 7.2 ortho
3-Chlorophenol 21.8 15.1 meta
4-Chlorophenol 1.0 2.5 para
2,3-Dichlorophenol 8.3 18.3 ortho, meta
2,4-Dichlorophenol 1.5 3.5 ortho, para
2,5-Dichlorophenol 16.6 18.5 ortho, meta
2,6-Dichlorophenol 2.4 16.2 ortho, ortho
3,4-Dichlorophenol 3.2 18.3 meta, para
2,4,5-Trichlorophenol 14.6 22.3 ortho, para, meta
2,4,6-Trichlorophenol 5.3 6.3 ortho, para, ortho
Pentachlorophenol 6.7 12.0 al
2-Methyl-4-Chlorophenol 2.9 6.3 ortho, para
3-Methyl-4-Chlorophenol 1.4 4.2 meta, para
3-Methyl-6-Chlorophenol 2.1 12.5 meta, ortho
TABLE 22. EFFECT OF SUBSTITUTION POSITION ON DEGRADATION RATES
NON-CHLORINATED PHENOLS
Compound Chemical Half-Life (days) Substitution
Texas Soli Mississippi Soil Position
Methylphenols
o-Cresol (2-Methylphenol) 1.6 5.1 ortho
p-Cresol (4-Methylphenol) (1) 0.5 para
m-Cresol (3-Methylphenol) 0.6 11.3 meta
2,4-Dimethylphenol 0.7 1.4 ortho, para
Nltrophenols
p-Nitrophenol 10.2 2.5 para
2,4-Dinitrophenol 4.6 32.1 ortho, para
(1) No chemical was detected after one day.
47
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TABLE 23. COMPARATIVE DEGRADATION RATES OF CHLORO-, METHYL-
AND NITROPHENOLS
Compound Chemical Half-Life (davs)
Texas Soil Mississippi Soil
3-Chlorophenol 21.8 15.1
m-Cresol (3-Methylphenol) 0.6 11.3
p-Nitrophenol 10.2 2.5
p-Cresol (4-Methylphenol) (1) 0.5
4-Chlorophenol 1.0 2.5
2,4-Dinitrophenol 4.6 32.1
2,4-Dimethylphenol 0.7 1.4
2,4-Dichlorophenol 1.5 3.5
(1) No chemical was detected after one day.
2. Based on the data, it was not possible to discern whether zero or first order
kinetics provided a better representation of data. For most of the chemicals the
data could be fit to either kinetics with high correlation coefficients.
3. In general, the rates of chemical loss were higher in the Texas soil than in the
Mississippi soil. There did not appear to be any pattern to the differences in
rates in the two soils.
4. Chlorophenols with chlorine substituted in the meta position had greater half-
lives and therefore lower chemical loss rates. This was particularly evident with
the mono-, di- and trichlorophenols in the Texas soils.
5. Chemicals that had a nitro group substituted on the phenol ring appeared to
have a lower loss rate.
48
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SECTION 6
ADSORPTION EXPERIMENTS
INTRODUCTION
The persistence of hazardous organic compounds in soils is related to reactions
that affect the transport and fate of such chemicals. One of the most important
reactions is adsorption.
Adsorption is the process by which ions or molecules present in one phase tend
to concentrate at a surface or interface. The process can occur at an interface
between any two phases, such as liquid-liquid, gas-liquid, or solid-liquid interfaces.
The adsorbed substance is the adsorbate while the adsorption phase is referred to as
the adsorbent. The tendency of organic molecules to adsorb on soil is determined by
the physical and chemical characteristics of the chemical compound and the soil to
which it is added. The two driving forces for adsorption are the lyophobic (solvent-
disliking) character of a solute relative to a particular solvent, and the affinity of the
solute for the solid, such as electrical attraction or van der Waal's attraction/19).
Adsorption is the major retention mechanism for most organic and inorganic
compounds in soils. As a result, the leaching potential of a chemical in soil is, in
general, proportional to the magnitude of the adsorption (partitioning) coefficient of that
chemical in a soil. The adsorption potential of a chemical is governed by the
properties of both the soil and the chemical. Important properties of the chemical that
affect adsorption include: (a) chemical structure, (b) acidity or basicity of the molecule
(pKa or pKfc), (c) water solubility, (d) permanent charge, (e) polarity, and (f) molecule
size.
To estimate the environmental movement of a chemical, values of the
adsorption coefficient and other sorption equation coefficients can be compared to the
49
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value of other chemicals whose behavior in soil and sediment systems is well
documented.
Adsorption Equilibria
At equilibrium, the solute remaining in solution is in dynamic equilibrium with
that of the soil surface. At this point, there is a defined distribution of solute between
the liquid and solid phases. The preferred form for depicting this distribution is to
express the quantity qe (amount of solute sorbed per unit weight of solid sorbent) as a
function of the equilibrium solution concentration (Ce) at a fixed temperature. An
expression of this type is an adsorption isotherm.
One of the oldest adsorption equations that has been used widely for solid-
liquid systems is the Freundlich equation:
qe = KfCe1/n (6)
where qe is the equilibrium distribution coefficient (mg of chemical/gm of adsorbent),
Ce is the equilibrium chemical concentration (mg/liter of solvent), and Kf and 1/n are
constants. The constant, Kf, is related to the capacity or affinity of the adsorbent and
the exponential term, 1/n, is an indicator of the intensity, or how the capacity of the
adsorbent varies with the equilibrium solute concentration. The Freundlich isotherm
has had success in describing sorption behavior of organics^) and the adsorption
data generated in this study were compared to this empirical model.
The Freundlich constants may be determined statistically when the equation is
expressed in linear form by a logarithmic transformation:
log qe = log Kf + 1/n log Ce (7)
The constants, Kf and 1/n, can be obtained, respectively, from the intercept and slope
of log-log plots of qe vs Ce.
The Freundlich equation that results from specific experiments should not be
extrapolated beyond the experimental range of data used in its construction. This is
50
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because the Freundlich equation predicts infinite adsorption at infinite concentrations,
and that any soil or clay has an unlimited capacity to retain chemicals dissolved in
water. Such an infinite capacity is not only thermodynamically inconsistent, but
experience has shown that the extent of adsorption ultimately is limited by the surface
area of the adsorbent. Thus, the Freundlich equation cannot be extrapolated with
confidence beyond the experimental range used in its construction and will not yield a
maximum capacity term. The latter term is a convenient single-valued number that
estimates the maximum amount of adsorption beyond which the soil surfaces are
saturated and no further net adsorption can be expected.
Soil Organic Carbon
Sorption of nonionic organic compounds from water onto soil has been
shown(20) t0 occur primarily by partition onto the soil organic phase. Adsorption by
soil minerals is relatively unimportant in wet soils, presumably because of strong
dipole interaction between the soil minerals and water, which excludes neutral organic
solutes from this portion of soil. Therefore, the more organic matter in soil, the more
adsorption is expected. Soil organic matter has been the single best predictor of the
adsorption isotherm parameter(21-23), and the use of the soil organic matter-water
partition coefficient, Kom, rather than the adsorption partition coefficient, Kp, has been
proposed as more appropriate:
K0m - Qom/C (8)
where qom = mg adsorbed/g soil organic matter and C = liquid phase concentration,
mg/L.
Organic matter content can be obtained by measurement or by multiplying an
experimentally determined organic carbon concentration by an appropriate
conversion factor. Because various researchers used different conversion factors, the
soil organic carbon content has been proposed(20, 23) f0 normalize adsorption
partition coefficients. This parameter (Koc) is:
51
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= (KD/%OO 100 (9)
where qoc = mg sorbed/g soil organic carbon, %OC = (mass of organic carbon/mass
of soil) 100, %OM m %OC (f), and f = conversion factor ranging from 1.7 to 2.0.
Rao et al.(24) performed an exhaustive literature search, showed that coefficient
of variation (CV) values for Koc were much lower than those of KD or K (Freundlich
coefficient), and suggested that Koc be used as a universal adsorption partition
coefficient. This relationship is valid when the organic carbon content of the soil is
more than 0.1%.
With regard to inorganics, the sorptive effect of the inorganic matrix was
indicated to be negligible at an organic content level of 1% and above(25). However,
for some sorbents, chemical interaction with the inorganic matrix may be important. In
the absence of organic carbon, the specific surface area and the nature of the mineral
surface have a greater impact on the degree of sorptionC2^).
Soil pH
The pH of a soil-water system can affect the sorption of organic solutes.
Because the extent of ionization of an acidic or basic compound affects its adsorption,
pH affects adsorption in that it governs the degree of ionization(1^). Except with ion
exchange adsorption, ions tend to be less readily adsorbed than neutral species.
Many organics form negative ions at high pH, positive ions at low pH, and neutral
species in intermediate pH ranges. Generally adsorption is increased at pH ranges
where the species are neutral in charge. pH also affects the charge on the soil
surface, altering its ability to adsorb materials^27).
In general, adsorption of organic pollutants from water increases with
decreasing pH. In many cases, this may result from neutralization of negative charges
at the soil surface with increasing hydrogen ion concentration, thereby reducing
hindrance to diffusion and making more available the active surface of the
. Usually, organic acids are more adsorbable at low pH, whereas the
52
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adsorption of organic bases is favored by high pH. The optimum pH for any
adsorption process must be determined by laboratory testing.
Sorption of the non-dissociated chemical species and also of their anionic
species can occur^S). When the pKa of a species, such as pentachlorophenol, is
relatively low compared to a natural water system, anionic species adsorption can
occur. For highly chlorinated phenols, prediction of overall distribution ratios on
simple partitioning of the nondissociated species can be in error(28).
Another soil adsorption characteristic which is influenced by soil pH is the
Cation Exchange Capacity (CEC). At very low pH values, only a small portion of the
positive ions held by the clays and organic colloids can be replaced by cation
exchange. As pH increases, the hydrogen held by organic colloids and silicate clays
becomes ionized and can be exchanged by positively charged organic molecules.
Ion exchange is hypothesized to dominate the sorption process in acidic
soil(29). The higher sorption in the acidic soil, as compared to basic soil, reflects
stronger sorption of the protonated organic cations. Competitive adsorption occurs
between compounds in an acidic soil where the protonated compound species
predominates in solution, in contrast, competition is minimal in a basic soil when the
compounds are neutral. Non-ionic organic compounds may sorb independently on
soil from a mixture^20).
When the protonated and neutral species coexist, site-specific sorption of the
cation is preferred because of the electrostatic attraction between the base and the
negatively charged soil surface^), when anionic and neutral species coexist,
neutral species sorption occurs because of the electrostatic repulsive forces of the
anionic species^). Maximum adsorption is attained near the point where pH = pKa,
and sorption capacity drops rapidly at pH values above pKa.
53
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MATERIALS AND METHODS
Adsorption Method
The adsorption of the specific chemicals was investigated in batch adsorption
experiments. A list of the chemicals investigated and a description of the soil types
used were presented earlier (Tables 1 and 3). The batch adsorption technique^31)
consists of mixing an aqueous solution containing a solute of known composition and
concentration with a given mass of sorbent (soil) for a period of time. The solution is
then separated from the sorbent and chemically analyzed to determine changes in
solute concentration. The amount of solute sorbed is assumed to be the difference
between the initial concentration (before contact with the sorbent) and the solute
concentration after mixing.
Stock Solutions
The solvent used was distilled deionized water (DDW) obtained by passage
through a Barnstead Water Purification Cartridge #DO 809. The stock solution of each
organic compound contained the maximum soluble amount of the compound in DDW
at room temperature (25°C). The procedure used to prepare the stock solutions and to
determine the solubility limit of specific chemicals is noted in Table 24.
The solubility limits of chemicals determined as part of this study are included in
Table 25. This table contains only those chemicals whose maximum aqueous
solubilities were determined as part of this research. The solubilities of chemicals not
listed in Table 25 were obtained from the literature^).
Standard Solution
To prepare a calibration curve for a chemical, an accurately prepared standard
solution is required. The following procedure was used to prepare the standard
solution of each compound. A calculated amount of the compound and 100 ml of
DDW was added to an oven-dried 100 ml volumetric flask. The solution was mixed
54
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TABLE 24. PROCEDURE FOR DETERMINING SOLUBILITY LIMIT OF AN ORGANIC
COMPOUND IN WATER
(a) Put distilled deionized water into a teflon-capped bottle (4 L).
(b) Add enough organic compound to the bottle to observe some nonsoluble (liquid) or paniculate (solid)
organic compound in the water phase.
(c) Stir the solution at room temperature (20° C) with magnetic stirrer.
(d) Add enough compound to produce a saturated solution. This will be identified when there is residual
insoluble compound remaining in the bottle after the mixing period.
(e) Before the filtration, prewash a filter with about 100 ml_ of water, then rinse the prewashed filter with
about 10 ml of the saturated solution. The prewashing procedure can remove any so-called wetting
agent from the filter.
(f) Filter solution with 0.45u.m pore size membrane filter.
(f) Determine the concentration of organic compound in the filtrate by HPLC.
(g) The HPLC result is the solubility limit in water. The filtrate is transferred into a teflon-capped amber
bottle and stored at 4° C room. This is the stock solution for adsorption test. Prior to use, this solution
is mixed at 20° C.
TABLE 25. LIST OF MAXIMUM SOLUBILITIES (20° C) IN WATER DETERMINED AS
PART OF THE ADSORPTION STUDIES
Compound
Concentration (mg/L)
Acid Extractables
2-Chlorophenol 20,300
3-Chlorophenol 17,000
4-Chlorophenol 8,650
2,3-Dichlorophenol 3,900
2,4-Dichlorophenol 2,600
2,5-Dichtorophenol 3,600
2,6-Dichlorophenol 1,500
3,4-Dichlorophenol 520
2,4,5-Trichlorophenol 930
2,4,6-Trichlorophenol 415
Pentachlorophenol 13
2,4-Dimethylphenol 7,500
2-Methyl-4-Chlorophenol 2,500
3-Methyl-4-Chlorophenol 3,900
3-Methyl-6-Chlorophenol 1,400
p-Nitrophenol 5,300
2,4-Dinitrophenol 500
Compound
Concentration (mg/L)
Amines
Diphenylamine 2.5
Toluenediamine 590
Brucine 310
Alcohols
Isobutyl alcohol 3,700
Allyl alcohol 4,300
Propargyl alcohol 4,900
1-Butanol 4,200
2,3-Dichloropropanol 7,600
Other
2-Nitropropane 2,800
Thiourea 5,000
Explosive Chemicals
2,4-Dinitrotoluene 160
2,4,6-Dinitrotoluene 610
RDX 42
HMX 3.8
55
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well until no paniculate matter was visible. This known concentration was diluted to
five concentrations ranging from 10 mg/L to 100 mg/L (usually 10, 30, 50, 70 and 100
mg/L) since five data points are needed to construct the calibration curve. The five
standard solutions for each compound were stored at 4° C and used whenever soil
extracts containing the compound were analyzed by HPLC.
Soil Moisture
The soil moisture content is needed to calculate the amount of chemical sorbed
per unit of dry soil. The standard ASTM procedure(33) for measuring soil moisture
content was followed (Table 26).
SoihSolutlon Ratio
A single soihsolution ratio may not be satisfactory for all organic compounds.
This is because a weakly adsorbed compound may not result in a measurable
concentration change after contact with soil. Conversely, a compound's affinity for soil
may be so strong that the final solution concentration is below analytical detection
limits.
TABLE 26. PROCEDURE FOR MEASURING SOIL MOISTURE CONTENT
(Taken from ASTM D2216)<33>
(a) Place aluminum pans in an 105° oven for 24 hours. Transfer them to a dessicator at room
temperature (20° C) for approximately 1 hour. Measure and record the weight of an aluminum pan
on the analytical balance (wt grams). Precision of the fourth decimal place is required.
(b) Weigh out approximately thirty grams of the sorted soil (w2 grams) into dried aluminum pans.
(c) Dry soil at 105° C for 48 hours.
(d) Measure and record the weight of the dried soil and the aluminum pan (W3 grams).
(e) Calculate soil moisture content, W (%) using the following equation:
w = weight of moisture 1QO _ W2 ' W3 x1QO
weight of oven-dried soil w3 - w1
To determine an optimum soil:solution ratio for each compound, batch sorption
tests were performed using several soil:solution ratios. To evaluate the optimum
56
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soiksolution ratio, the adsorption characteristics of the compounds at two different
initial compound concentrations were evaluated. The maximum concentration used for
the adsorption measurements was the solubility limit of the compound in water. The
minimum concentration used was about one order of magnitude higher than the
lowest detection limit determined by high pressure liquid chromatography. The
procedure for determining the optimum soil:solution ratio is outlined in Table 27.
TABLE 27. PROCEDURE FOR DETERMINATION OF OPTIMUM
SOILrSOLUTION RATIO
(a) Set soil-to-solution ratio at 1 :1 to 1 :X. Required mass of the sorted soil, Ms (gram), and volume of
solution, V (ml), are calculated by:
Ms(1 -(
V + MS.(W/100)*X
where W - moisture content of the soil.
(b) Weigh out the desired amount of sorted soil (Ms grams) into an amber bottle.
(c) Add V ml of solution to the amber bottle. This is designated as a sample. Prepare another bottle
which will not contain any soil. This is designated as the blank.
(d) Place bottles in rotating tumbler in the 20° C room and tumble continuously for 24 hours at 30 rpm.
(e) After tumbling, centrifuge the bottles for 30 minutes at 4,000 rpm.
(f) After centrifugation, filter the supernatant with a glass fiber filter and then with a 0.45 u.m pore size
membrane filter.
(g) Analyze the concentrations of the filtrate by HPLC.
(h) Calculate the percent absorption P(%):
f f8
cfb
100
where Cfb - final concentration of blank (mg/L), and
Cfs - final concentration of sample (mg/L).
(j) Use another soil-to-solution ratio, and repeat (a) through (h).
(k) The optimum soil solution ratio is one that satisfies the condition that P(%) is between 20% and
80%.
57
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Solute Stability
In conducting batch adsorption experiments, it is important to consider the
stability of the solute in solution. Processes such as photolysis, hydrolysis, and
microbial degradation can cause a decrease in solute concentration leading to errors
in adsorption results.
In this study, photolysis was minimized by using amber bottles to limit
transmission of light during solution/soil mixing. By limiting contact time to one day
and using an unacclimated soil, chemical losses due to chemical and microbial
degradation were minimal. Chemical losses due to hydrolysis were not quantified and
thus are included in adsorption calculations.
Other Factors
Other factors which affect adsorption efficiency include temperature and ionic
strength. The sorption experiments were conducted at 20° C. All experiments were
conducted at the pH of the soil:solution which was approximately that of the respective
soils. No pH adjustment of the soil solution was made.
Adsorption is not instantaneous and mixing for a specific amount of time is
necessary to assure equilibrium. The equilibration time should be the minimum time
beyond which relatively insignificant changes in the solute concentration will occur. In
this study, a separate set of experiments was conducted to determine the effect of
mixing time on the amount of chemical adsorbed. The results indicated that the
amount of adsorption, q (mg sorbed/g soil) slightly increased with time. However, the
q values did not vary greatly with time after 24 hours. The data representing the
amount of chemical adsorbed after 24 hours were analyzed statistically. The slope of
q with time was not-statistically different from zero and q was shown not to vary with
time after 24 hours. This implied that 24 hours of mixing was enough for equilibrium,
and thus mixing for one day was used in the batch adsorption studies.
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Data Analysis
At least four data points are needed to construct the Freundlich isotherms and to
determine the appropriate coefficients. In this study, a minimum of four and frequently
as many as eight data points were used to develop the isotherms. The isotherm data
were fit to the Freundlich adsorption equation (Eqn. 6). A log-log scale was used to
plot the Freundlich isotherm. The abscissa was the equilibrium (final) concentration in
liquid phase and the ordinate was the amount adsorbed per unit of soil.
The data were analyzed by least-square linear regression methods with 95%
confidence interval (Cl) determined as described in Section 4. Routine statistical
methods were used(1®). First, the linear relation between Xj (= log Cj) and Yj (= log qj)
was determined by the Lotus 1-2-3 program:
Y = a + b X (7)
where a = intercept and b - slope of the line.
Second, the variances of a and b, Sa2 and S^2 were calculated, respectively
by:
Sa2 . Sy2 {1/n + Xavg2/(IXj - Xavg)2} (8)
SD2 = Sy2/(LXj - Xavg)2 (9)
where
- YUnt j)2/(n-2)}0-5
Xj, YJ = individual data point of log Cj, log qj, respectively,
Xavg m average of Xjt
Y|jnj = estimated Yj value with respect to Xj from the linear relationship
Y = a+bX,and
n = number of samples.
The 95% Cl for the estimates of a and b were determined by a-t Sa to a+t Sa
and b-t S^ to b+t SD, respectively, where the t value can be found in a typical statistical
text for a two-sided test with a degree of freedom = n-2, and alpha = 5% level of
significance.
59
-------
RESULTS
Following the procedures described above, the adsorption isotherms and
constants were determined. Illustrative sorption patterns and isotherms are noted in
Figures 6 and 7. The isotherm data are presented in Tables 28 and 29. Based on the
correlation coefficients calculated for the data, the Freundlich equation described the
adsorption satisfactorily for all but a few chemicals in both soils.
As described earlier, the pH of the solution can affect the sorption
characteristics. In this study, no attempt was made to control the pH of the extracts to
any set value. Therefore, the pH of the extracts varied between soil types and
between samples of the same soil type (Tables 28 and 29). In the Texas soil, the pH
range varied from 7.5 to 8.0 while in the Mississippi soil it ranged from 4.5 to 7.0. In
evaluating and using the sorption data, the different pH ranges should be recognized.
As noted earlier, the adsorption coefficients represent the results obtained over
a specific chemical concentration range and the Freundlich equation that results for a
chemical should not be extrapolated beyond that range. Tables 30 and 31 indicate
the chemical concentration ranges for which the data in Tables 28 and 29 are valid.
A wide range of chemical concentrations was evaluated, i.e., from low mg/l
concentrations to near or at saturation concentrations. In many cases, the range
covered two to three orders of magnitude. Thus, the Freundlich adsorption coefficients
are appropriate for concentrations found at sites with low as well as with high
concentrations of these chemicals.
As discussed, the adsorption potential of a chemical is governed by
characteristics of both the chemical and the soil to which it is exposed. The
characteristics include: (a) chemical structure, (b) solubility, (c) pH of the solution, (d)
ionization potential of the chemical, (e) polarity, and (f) molecular size. The complex
interaction of these factors will affect the adsorption that does occur. These
60
-------
Adsorption of 2,3-Dlchlorophenol in Texas Soil at 20° C
1.0-
•8
c«
*
i
6
^»/
cr
if
0.51
0.0
-0.5-
-1.0-
-1.5-
-2.0
1
= 0.99
2 3
log Concentration (mg/1)
Adsorption of Toluenedlamlne In Texas Soil at 20° C
B
N-W
cr
j?
-0.6
•0.8-
-1.0-
-1.2-
•1.4-
-1.6-
-1.8
R-0.96
a
1 2
log Concentration (mg/1)
Figure &. Adsorption of 2,3-Dichlorophenol and Toluenedlamlne
In Texas Soil at 20° C
61
-------
Adsorption of 2,3-Dlchlorophenol In Mississippi Soil at 20° C
0.0-
j? -1.5-
-2.0
1 2 3
log Concentration (mg/1)
Adsorption of Toluenedlamlne In Mississippi Soil at 20° C
o
I
E
s
•0.4-
-0.8
-1.2
-1.6-
-2.0
R = 0.94
Q
1 2
log Concentration (mg/1)
Figure 7. Adsorption of 2,3-Dlchlorophenoi and Toluenediamlne
In Mississippi Soil at 20° C.
62
-------
TABLE 28. BATCH SORPTION ISOTHERM DATA - TEXAS SOIL
FREUNDLICH EQUATION PARAMETERS
COMPOUND Kg 1_/n T PH**
Acid Extractables
Phenol 3.44x10'3 0.53 0.97 7.9
o-Cresol 2.65x10'3 0.58 0.97 7.9
p-Cresol 9.53x10'3 0.55 0.98 ++
m-Cresol 3.19x10"6 0.59 0.97 7.9)
2-Chlorophenol 1.01x10'3 0.87 0.99 , ++
3-Chlorophenol 2.93x10'3 0.76 0.99 7.9
4-Chlorophenol 5.50x10'3 0.72 0.98 7.8
2,3-Dichlorophenol 1.71x10'3 0.91 0.99 , 7.8
2,4-Dichlorophenol 4.92x10'3 0.77 0.99 7.6
2,5-Dichlorophenol 2.71x10"3 0.82 0.99 8.0
2,6-Dichlorophenol 6.84x10'4 0.90 0.99 7.5
3.4-Dichlorophenol 7.16x10'3 0.75 0.99 8.0
2,4,5-Trichlorophenol 1.46x10"3 1.02 0,96 7.9
2,4,6-Trichlorophenol 1.76x10'3 0.73 0.99 8.1
Pentachlorophenol 5.94x10'3 0.68 0.99 8.1
2,4-Dimethylphenol 3.94x10"3 0.76 0.99 8.1
2-Methyl-4-Chlorophenol 4.97x10'3 0.75 0.99 7.4
3-Methyl-4-Chlorophenol 4.37x10"3 0.79 0.99 7.4
3-Methyl-6-Chlorophenol 1.56x10"3 0.88 0.99 7.4
p-Nitrophenol 3.20x10'3 0.99 0.96 7.4
2,4-Dinitrophenol 1.14x10'3 0.58 0.96 7.7
4,6-Dinitro-o-Cresol 2.30x10'3 0.57 0.96 8.1
Amines
Toluenediamine 1.74x10"2 0.44 0.96 7.7
Brucine 1.24 0.03 0.47 7.4
Alcohols
Isobutyl alcohol I.46x10'3 0.59 0.94 8.0
Allyl alcohol 4.50x10'3 0.45 0.91 8.0
Propargyl alcohol 4.60x10'3 0.73 0.85 8.0
1-Butanol 6.20x10'3 0.42 0.81 8.0
2,3-Dichloropropanol 1.10 0.0004 0.78 ++
Methanol 9.30 0.28 0.79 8.0
Other
2-Nitropropane 3.54x10"3 0.66 0.94 8.0
Thiourea 2.54x10*4 0.81 0.98 8.2
* -- correlation coefficient; ** - pH of adsorption extract, ++ -- not measured
63
-------
TABLE 29. BATCH SORPTION ISOTHERM DATA -- MISSISSIPPI SOIL
FREUNDLICH EQUATION PARAMETERS
COMPOUND Kjj -\_ljn r PH**
Acid Extractables
Phenol 1.30x10'3 0.71 0.98 5.8
o-Cresol 6.00x10'4 0.87 0.98 ++
p-Cresol 1.21x10"2 0.47 0.99 ++
m-Cresol 8.60x10'4 0.75 1.00 ++
2-Chlorophenol 1.60x10'3 0.77 1.00 ++
3-Chlorophenol 4.50x10"3 0.63 0.98 5.5
4-Chlorophenol 9.90x10"3 0.55 0.99 5.6
2,3-Dichlorophenol 8.5x10"3 0.65 1.00 6.9
2,4-Dichlorophenol 6.80x10'3 0.65 1.00 ++
2,5-Dichlorophenol 6.90x10'3 0.80 0.99 ++
2.6-Dichlorophenol 6.70x10"3 0.61 1.00 ++
3.4-Dichlorophenol 4.70x10'3 0.78 0.96 5.6
2,4,5-Trichlorophenol 3.60x10"3 0.91 0.97 ++
2,4,6-Trichlorophenol 9.80x10'3 0.57 1.00 ++
Pentachlorophenol 1.6xlO'3 0.51 1.00 6.2
2,4-Dimethylphenol 5.10x10"5 1.28 0.92 5.1
2-Methyl-4-Chlorophenol 6.40x10'3 0.65 1.00 ++
3-Methyl-4-Chlorophenol 7.30x10"3 0.68 0.99 7.0
3-Methyl-6-Chlorophenol 3.00x10'3 0.77 0.99 6.2
p-Nitrophenol 3.20x10'3 0.64 1.00 5.7
2,4-Dinitrophenol 1.10x10~3 0.78 0.95 ++
4,6-Dinitro-o-Cresol 8.60x10"4 0.86 0.99 4.5
Amines
Toluenediamine 5.07x10'5 1.28 0.94 5.3
Brucine 6.18x10'2 0.48 0.97 ++
Alcohols
Isobutyl alcohol 5.10x10'5 1.28 0.92 5.7
Allyl alcohol 3.30x10'4 0.72 0.96 5.7
Propargyl alcohol 6.30x10'4 0.61 0.89 5.8
1-Butanol 2.80x10"3 0.43 0.85 5.7
2,3-Dichloropropanol S.OxlO"5 0.90 0.64 ++
Methanol 1.40x10'3 0.66 0.82 5.9
Other
2-Nitropropane 3.54x10"2 0.66 0.94 5.7
Thiourea 1.50x10"4 0.92 0.98 5.1
* - correlation coefficient; " -- pH of adsorption extract, ++ - not measured
64
-------
TABLE 30. CHEMICAL CONCENTRATION RANGE EVALUATED DURING THE BATCH
ADSORPTION EXPERIMENTS •• TEXAS SOIL
Compound Concentration Range (mg/l)
Acid Extractables
Phenol 9-10,300
o-Cresol 20-1,000
p-Cresol 11-6,400
m-Cresol 8-3,200
2-Chlorophenol 26-20,300
3-Chlorophenol 27-7,800
4-Chlorophenol 18-8,600
2,3-Dichlorophenol 40-3,900
2,4-Dichlorophenol 19-2,300
2,5-Dichlorophenol 16-3,600
2,6-Dichlorophenol 26-1,500
3,4-Dichlorophenol 21-520
2,4,5-Trichlorophenol 19-910
2,4,6-Trichlorophenol 20-410
Pentachlorophenol 3-13
2,4-Dimethylphenol 90-7,400
2-Methyl-4-Chlorophenol 18-2,400
3-Methyl-4-Chlorophenol 25-3,800
3-Methyl-6-Chlorophenol 36-1,400
p-Nitrophenol 10-5,200
2,4-Dinitrophenol 10-500
4,6-Dinitro-o-Cresol 18-130
Amines
Toluenediamine 27-550
Brucine 15-300
Alcohols
Isobutyl alcohol 40-3,600
Allyl alcohol 15-4,300
Propargyl alcohol 45-4,900
1-Butanof 67-3,800
2,3-Dichloropropanol 78-7,600
Methanol 49-4,200
Other
2-Nitropropane 58-2,800
Thiourea 17-4,800
65
-------
TABLE 31. CHEMICAL CONCENTRATION RANGE EVALUATED DURING THE BATCH
ADSORPTION EXPERIMENTS -- MISSISSIPPI SOIL
Compound Concentration Range (mg/l)
Acid Extractables
Phenol 9-10,300
o-Cresol 20-1,000
p-Cresol 12-6,400
m-Cresol 100-13,000
2-Chlorophenol 34-20,300
3-Chlorophenol 25-8,800
4-Chlorophenol 34-8,600
2,3-Dichlorophenol 30-2,600
2,4-Dichlorophenol 18-2,500
2,5-Dichlorophenol 22-2,800
2,6-Dichlorophenol 26-1,500
3,4-Dichlorophenol 18-520
2,4,5-Trichlorophenol 22-930
2,4,6-Trichlorophenol 16-310
Pentachlorophenol 7-13
2,4-Dimethylphenol 18-6,300
2-Methyl-4-Chlorophenol 15-2,500
3-Methyl-4-Chlorophenol 43-3,900
3-Methyl-6-Chlorophenol 12-1,400
p-Nitrophenol 10-5,300
2,4-Dinitrophenol 4-260
4,6-Dinitro-o-Cresol 23-130
Amines
Toluenediamine 160-580
Brucine 30-300
Alcohols
Isobutyl alcohol 40-3,700
Allyl alcohol 15-4,300
Propargyl alcohol 45-4,900
1-Butanol 67-3,800
2,3-Dichloropropanol 78-7,600
Methanol 49-4,000
Other
2-Nitropropane 59-2,800
Thiourea 21-5,000
66
-------
experiments were conducted under reasonable real world conditions. Soils of
different characteristics (pH, CEC and organic carbon) different chemicals, and
different chemical concentrations were used. As a result it was not possible to
determine the relative effect of such parameters.
However, the data do allow overall effects to be identified. When the Freundlich
Kf values for the two soils, i.e., the capacity or affinity of the soils, were compared
(Table 32), in general, the Texas soil had the greater values and therefore the greater
sorption capacity for the acid extractables. However, the opposite was true for the
amines and alcohols for which the Mississippi soil had the greater Kf values, in some
cases greater by a factor of 10 or 100.
CONCLUSIONS
1. The Freundlich equation described the adsorption of the chemicals on the two
soils satisfactorily, i.e., with high correlation coefficients, except for a few
chemicals.
2. The range of chemical concentrations evaluated ranged from the low mg/l
concentrations to near or at saturation concentrations and for most chemicals
covered two to three orders of magnitude. Thus, the adsorption data are
appropriate for concentrations found at sites with low as well as high
concentrations of these chemicals.
3. For these concentration ranges, a linear adsorption relationship, i.e., n = 1, did
not occur.
4. The Freundlich Kf values for chemicals in the two soils were different. For the
acid extractables, the Kf values generally were greater in the Mississippi soil.
For the amines and alcohols, the Kf values were greater in the Texas soil.
67
-------
TABLE 32. COMPARISON OF FREUNDLICH ADSORPTION COEFFICIENTS (Kf) FOR
THE TEXAS AND MISSISSIPPI SOILS
Tsxas Mississippi
Compound
Add Extractables
Phenol
o-Cresol
p-Cresol
m-Cresol
2-Chtorophenol
3-Chlorophenol
4-Chtorophenol
2,3-Dichlorophenol
2,4-Dichlorophenol
2,5-Dichtorophenol
2,6-Dichlorophenol
3,4-Dichtorophenol
2,4,5-Trichlorophanol
2,4,6-Trichlorophenol
Pentachlorophanol
2,4-Dimethylphenol
2-Methyl-4-Chforophenol
3-Methyl-4-Chk>rophenol
3-Methyl-6-Chtorophenol
p-Nitrophenol
2,4-Dinitrophanol
4,6-Dinrtrophenol
Soil
3.4x1 0'3
2.7x10'3
9.5x1 0'3
3.2x1 0'6
1.0x10'3
2.9x1 0'3
5.5x1 0'3
1.7x10"3
4.9x1 0'3
2.7x1 0'3
j
6.8x1 0'4
7.2x1 0'3
1.5x10"3
1.8x10'3
5.9x1 0'3
3.9x1 0'3
5.0x1 0'3
4.4x1 0'3
1.6x10'3
3.2x1 0"3
1.1x10'3
2.3x1 0'3
Soil
1.3x10"3
6.0x1 0"4
1.2x10"2
8.6x1 0'4
1.6x10'3
4.5x1 0'3
9.9x1 0'3
8.5x1 0"3
6.8x1 0'3
6.9x1 0"3
1%
6.7x1 0'3
4.7x1 0'3
3.6x1 0'3
9.8x1 0"3
1.6x10-3
5.1x1 0'5
6.4x1 0'3
7.3x1 0"3
3.0x1 0'3
3.2x1 0'3
1.1xiO'3
8.6x1 0'3
Compound
Amlnet
Toluenediamine
Brucina
Alcoholi
Isobutyl alcohol
Allyl alcohol
Propargyl alcohol
1 -Butanol
2,3-Oichloropropanol
Methanol
Othir
2-Nitropropane
Thiourea
Texas Mississippi
Soil
1.7x10'2
1.24
1.5x10"3
4.5x1 0"3
4.6x1 O*3
6.2x1 0'3
1.10
9.3
3.5x1 0"3
2.5x1 0'4
Soil
5.1x1 0"5
o
6.2x1 0'2
5.1x10'5
3.3x1 0'4
6.3x1 0'4
2.8x1 0"3
5.0x1 0'5
1.4x10"3
3.5x1 0'2
1.5x10'4
68
-------
SECTION 7
TOXICITY REDUCTION
A major objective of this study was to provide comprehensive screening data on
the treatability of specific organic chemicals in soil. Hazardous constituents that enter
the soil are to be detoxified or immobilized.
When a chemical is added to the soil, it is transformed into other products through
chemical and biological reactions with or without complete detoxification and
immobilization. Measuring the loss of the parent compound, such as was presented in
Section 4, does not assure that complete detoxification and immobilization occurs.
Intermediate degradation products, which may be more mobile and/or toxic than the
parent compound, may be generated as the parent compound degrades.
Additional information on the transformation and/or detoxification of a chemical is
necessary to establish that the loss of the parent compound leads to the complete
detoxification of the chemical or waste. Such information can be obtained using either
chemical or bioassay analyses.
Chemical analysis of detoxification products may yield information about
biochemical degradation pathways, but it is time consuming and expensive.
Bioassays have been used successfully to demonstrate detoxification of the applied
waste in the soiK12> 13> 15) and are less expensive and time consuming. Such
bioassays also have been used as a screening tool to evaluate the soil treatment
potential of a chemical or wasted ^« 34)
APPROACH
In this study, the reduction of toxicity that occurred in selected degradation studies
was evaluated by determining the toxicity of the water soluble fraction (WSF) of the
chemical/soil mixture at the same sampling intervals used to obtain the degradation
data. The chemical compounds that can be extracted with water represent the
69
-------
potentially leachable fraction of the chemical or any intermediate chemical
detoxification products. The WSF of the chemical poses the greatest threat to
groundwater contamination. Hence, evaluating the loss of the potentially leachable
fraction of a chemical is important.
In addition, the concentration of the parent chemical in the WSF also was
determined. This concentration was expressed in terms of quantity of chemical that
was water extractable per kg of the soil. The procedures for: (a) obtaining the WSF, (b)
determining the toxicity of the WSF using the Microtox© method, and (c) determining
the chemical concentration were the same as those described in the earlier sections.
To put the toxicity reduction data in perspective, the WSF toxicity reductions, the
WSF chemical concentration reductions and the soil chemical concentration
reductions were compared.
RESULTS
This toxicity and chemical reduction comparison was done for phenol and eight
different chloro- substituted phenols: phenol; 2-, 3- and 4-Chlorophenol; 2,3-, 2,4-, and
2,6-Dichlorophenol; 2,4,6-Trichlorophenol; and Pentachlorophenol. Texas soil was
used in each degradation study.
Two loading rates were used. One was the acceptable loading rate identified in
Section 4 (Table 11) and the other was twice those loading rates. Based upon
previous work(12- 13), loading rates twice the acceptable rates were not expected to
completely inhibit degradation. However, the higher rates could slow degradation
rates and may result in detoxification by-products if incomplete detoxification occurred.
Chemical Loss In Soil
The kinetic parameters for the loss of chemical in the soil microcosms are
presented in Table 33. Data that represented concentrations that were zero or below
the detectable limit were not included in the kinetic analyses because it was not clear
70
-------
TABLE 33. CHEMICAL LOSS IN SOIL - KINETIC PARAMETERS
Initial Cone First Order Kinetics
Chemical mg/kg+
Phenol
2-Chlorophenol
3-Chlorophenol
4-Chlorophenol
2,3-Dichlorophenol
2,4-Dichlorophenol
2,6-Dichlorophenol
2,4,6-Trichlorophenol
Pentachlorophenol
1400
700
760
380
235
120
176
88
260
130
180
90
1260
630
600
300
60
30
Half Life
(days)
19.5
4.1
4.1
1.7
16.2
21.8
1.5
1.0
27.2
8.3
3.7
1.5
12.8
2.4
11.0
5.3
3.6a
6.7
r*
0.96
0.92
0.97
0.98
0.93
0.97
0.92
0.91
0.91
0.97
0.89
0.94
0.99
0.91
0.92
0.94
1.00
0.92
95%CI"
16.5-23.9
3.2-5.7
3.5-4.9
1.5-1.9
13.1-21.5
19.6-24.6
0.9-3.6
0.7-1.3
21.2-37.9
8.1-8.5
2.7-5.8
1.1-2.4
12.0-13.7
2.0-3.0
8.7-15.0
4.60-6.3
5.5-8.5
•*
?ero Order Kinetics
mg/kg-d
32.7
59.3
37.0
26.1
6.0
2.3
21.5
14.5
4.7
3.6
8.4
6.1
34.3
41.8
18.0
10.4
4.4a
0.99
r
0.95
0.96
.0.90
0.90
0.92
0.9,5
0.92
0.91
0.88
0.98
0.98
0.96
0.96
0.97
0.88
0.94
0.88
0.90
95% Cl
25.7-39.6
48.5-70.1
24.2-49.8
20.4-32.0
4.4-7.6
2.0-2.6
8.6-34.3
9.7-19.2
3.0-6.3
3.5-3.7
7.2-9.6
5.4-6.8
27.9-40.7
36.7-46.9
11.9-24.2
8.8-12.1
0.80-1.2
+ Initial concentration of chemical in soil at time zero, mg/chemical per kg of dry soil, lower concentration is the
acceptable loading rate as discussed in Section 4 and as noted in Table 11.
* Correlation coefficient
' * 95% confidence interval for the kinetic parameter
a Data from initial lag phase (through day 21) were excluded in calculating these values; data points were too few
to calculate 95% confidence intervals.
when the chemical actually disappeared. The data fit both first and zero order kinetics
satisfactorily as indicated by the high correlation coefficients.
The data for the lower loading rates were the same as that obtained in the
degradation studies (Section 5, Table 18) since they were done at the same time as
71
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the previous studies. In most cases, the kinetic parameters at the higher loading rates
were larger than those at the lower loading rates.
Example chemical loss patterns for 2,4-Dichlorophenol and Pentachlorophenol
(PCP) at the higher loading rates are noted in Figure 8. The initial PCP concentration
of 60 mg/kg resulted in a lag phase up to 21 days after the chemical loading.
However, the microcosm extracted on day 29 indicated a reduced concentration of
PCP and that acclimation had occurred. Because there was a rapid chemical loss
after the lag period (Figure 8), kinetic parameters were obtained using data from days
21 and 29. No lag periods occurred for the other chemicals at any of the loading rates.
WSF Chemical Loss
The change of chemical concentration in the WSF also was analyzed and both
first and zero order kinetic parameters were determined. The kinetic data are
summarized in Table 34. First and zero order kinetics satisfactorily represented the
data. Due to rapid loss of 4-Chlorophenol, data on the WSF reduction of this chemical
were limited. As with the soil data, PCP concentrations in the WSF exhibited a lag
phase at the higher loading and only zero order kinetics could be calculated from the
data obtained after the end of the lag phase.
With respect to the higher loading rates, Table 34 indicates that: (a) higher
chemical concentrations were in the WSF initially at the higher loading rates, and (b)
for the first order kinetics, the WSF chemical concentrations decreased at a slower rate
(greater half life) when the chemicals were applied at the higher loading rate. No
difference was apparent in zero order kinetics due to the difference in loading rates.
Examples of WSF chemical loss patterns for two chemicals are presented in
Figure 9. The losses for both initial concentrations are indicated.
72
-------
TABLE 34. LOSS OF WATER EXTRACTABLE CHEMICAL -- KINETIC PARAMETERS
Initial Cone
Chemical mg/kg+
Phenol
2-Chlorophenol
3-Chlorophenol
4-Chlorophenol
2,3-Dichlorophenol
2,4-Dichlorophenol
2,6-Dichlorophenol
2,4,6-Trichlorophenol
Pentachlorophenol
+ Milligram of chemical
replicates
' Correlation coefficient
1150
575
610
305
155
75
93
32
130
68
98
55
1080
580
570
310
25
11
First Order Kinetics
Half Life
(days)
10.9
3.7
4.0
2.2
14.6
9.5
1.6
1.6
17.0
7.9
3.3
0.93
9.5
8.3
11.5
6.5
— initial
3.9
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
r*
.80
.60
.98
.95
.98
.93
.99
.97
.89
.97
.91
.85
.92
.99
.92
.98
lag phase —
0.84
that was water soluble at
time zero
95%CI"
7.8-18.1
2.0-36.8
3.7-4.4
1.8-2.9
13.2-16.4
7.9-11.9
£
£
13.5-22.9
7.0-9.1
2.6-4.6
0.6-1.9
7.9-12.0
7.7-8.9
9.5-14.7
5.9-7.4
2.7-7.3
per kilogram
Zen
mg/kg-d
35.5
46.8
21.7
30.6
4.
2.
11
5.
3.
3.
4.
8.
32
27
17
16
2.
1.
of dry
4
9
.9
3
2
6
8
8
.4
.9
.0
.4
2
2
soil,
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
a Order Kinetics
r
.85
.87
.86
.95
.95
.91
.95
.74
.90
.95
.94
.97
.98
.97
.93
.98
.99
.88
average
95% Cl
23.5-47.5
27.9-65.7
15.3-28.2
23.6-37.6
3.6-5.1
2.3-3.6
:
-#~
_a_
ff
2.4-4.0
3.0-4.1
3.8-5.8
7.0-10.6
30.
24.
13.
14.
0-35.8
2-31.6
6-20.4
6-18.1
0.76-1.7
of two
or three
95% confidence interval for the kinetic parameter
* Insufficient data to calculate 95%
confidence intervals due to rapid loss of chemicals
73
-------
o
(A
••»
O
O»
J£
O>
—
"o
c
0
.c
Q.
O
£
£
O
O
<*
en
"5
«
"5
CT
01
§,
a
0
200-
i
150-
•
100-
50-
0.
Loss of 2,4-Dichlorophenol
a
a
Initial chemical concentration in soil = 1 80 mg/kg
a
D
D
a
m ^
i • i i • i • i
0 5 10 15 20 25
Time in Days
80-
60 <
•
40-
20-
-i
Loss of Pentachlorophenol
B
B Initial PCP concentration in soil = 60 mg/kg
a a a Q
a
B
0- 5 101520253035
Time in Days
Figure 8. Loss of Chemical in Two Experiments When the Higher
Loading Rates Were Used
74
-------
Loss of 2-Chlorophenol in the WSF of the Soil Microcosms
^^
~0
in
"5
CD
£
Q.
O
CM
"5
U.
(fi
800-
600 J
400-
i
200-
o-
C
I a = initial concentration = 760 mg/kg of soil
» = initial concentration = 380 mg/kg of soil
Q
a
D
B
9 • a m
*D « q g
) 5 10 15 20 25 30
Tim* in Days
Loss of 2,4,6-Trichlorophenol in the WSF of the Soil Microcosms
'o
o
0
O)
Q.
O
40
CM"
"5
u.
w
600-
500-
400-
300-
200-
100-
o-
0
g H = initial concentration - 600 mg/kg of soil
H « = /n/'f/a/ concentration = 300 mg//cg of so//
t Q
* • * B 5 i
• ° B
* a
10 20 30
Tim* In Day*
Figure 9. Loss of Chemical in The WSF at Two Loading Rates
75
-------
WSF Toxicltv Reduction
The evaluation of loss of applied chemical in the soil microcosms and in WSF did
not address the existence of possible toxic transformation products. A study of the
toxicity of the constituents in the WSF can indicate the presence and/or accumulation
of any potentially teachable toxic intermediate products.
The toxicity of the WSF was measured by the Microtox® procedure and was
expressed as soil toxicity units (Section 4). The resulting data was analyzed by both
first and zero order kinetics to provide results that were consistent with the soil
chemical loss and the WSF chemical loss. Data on these kinetic parameters, in terms
of toxicity reduction, are presented in Table 35. Both first and zero order kinetics
appeared to represent the data satisfactorily.
Examples of the toxicity reduction patterns that occurred are presented in Figure
10. The WSF toxicity reductions that were noted were the result of the detoxification
that occurred in the soil microcosms as they were incubated at 20° C for the indicated
time periods.
With Pentachlorophenol, the WSF toxicity reduction indicated a slow reduction for
the first days of incubation, followed by a rapid decrease (Figure 10). No other lag
periods in toxicity reduction were observed for the other chemicals.
As had been noticed with the WSF chemical loss and the soil chemical loss, the
soil chemical loading rates did cause different results. With respect to the toxicity
reduction data (Table 35), the higher loading rates: (a) resulted in a higher initial WSF
toxicity, and (b) larger toxicity reduction half-lives. No difference in the zero order
kinetics as a function of loading rate was apparent.
Comparison of Chemical Losses and Toxicltv Reduction
The loss of the chemical in the soil and in the WSF and the WSF toxicity reduction
data were compared using the first order kinetic data (half-lives). Figure 11 compares
the half-life of the soil and the WSF chemical loss for all of the nine chemicals tested at
76
-------
both low and high loadings. The correlation shows that, in general, the soil chemical
half-life was about 1.5 times greater than the WSF chemical half-life. For these
chemicals, this indicates that the loss of the chemical in the WSF was about 1.5 times
faster than the loss of the chemical in the soil. This correlation also indicated that no
enhanced mobilization of applied chemical occurred as the degradation and
detoxification took place.
TABLE 35. WSF TOXICITY REDUCTION -- KINETIC PARAMETERS
First Order Kinetics
Chemical Chemical
Initial Cone
rna/ko+
Phenol
2-Chlorophenol
3-Chlorophenof
4-Chlorophenol
2,3-Oichlorophenol
2,4-Dichlorophenol
2,6-Dichlorophenol
1400
700
760
380
235
120
176
88
260
130
180
90
1260
630
2,4,6-Trichlorophenol600
Pentachlorophenol
300
60
30
Initial
Toxicity
40
13
20
10
28
15
46
6
37
19
52
22
31
14
29
12
31
13
Half Life
(davsl r*
14.9
10.2
5.8
5.8
14.5
8.9
1.8
0.6
18.7
7.7
5.3
2.4
11.2
13.3
12.1
5.8
24.3
7.3
0.87
0.5
0.96
0.95
0.88
0.91
0.87
0.97
0.92
0.90
0.95
0.87
0.95
0.79
0.92
0.94
0.78
0.88
95%cr
11.6-20.6
Zero Order
Sol
JJJffl r
1.17
— 1.17
5.0-7.0
4.6-7.8
11.5-19.5
7.3-11.3
1.1-4.0
-@-
15.5-23.6
6.2-10.2
4.5-6.5
1.7-4.1
9.6-13.4
9.3-23.7
9.9-15.3
5.0-7.1
16.4-46.6
5.3-11.9
0.90
0.76
0.76
0.
6
4
0.
1
2
2
0.
0.
0.
0.
0.
0.
62
.0
.5
75
.1
.2
.9
95
49
79
64
65
90
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
.90
.81
.91
.95
.92
.95
.90
.88
.89
.89
.98
.93
.95
.81
.89
.94
.76
.88
Kinetics
95% Cl
0.89-1
0.65-1
0.69-1
0.57-0.
0.60-0.
0.52-0.
3.0-9.
2.3-6.
0.56-0.
0.8-1.
2.0-2.
2.1-3.
0.80-1
0.29-0.
0.59-0.
0.53-0.
0.31-1
0.55-1
.5
.7
.7
95
91
95
0
6
94
3
5
8
.1
70
99
76
.0
.3
+ Initial concentration of chemical in soil at time zero, mg/chemical per kg of dry soil, lower concentration is the
acceptable loading rate as discussed in Section 4 and as noted in Table 11
++ Toxicity of the WSF at time zero in toxicity units (Section 4)
Correlation coefficient
'' 95% confidence interval for the kinetic parameter
a Data from initial lag phase (through day 21) were excluded in calculating these values; data points were too few
to calculate 95% confidence intervals.
77
-------
50
40
S
&J 30
n 20
1 10
n.
2,3-Dichlorophenol
• * initial concentration
g • * initial concentration
t ™ 0 B
•t; • " g :
- 260 mg/kg of soil
m 130 mg/kg of soil
10
20
30
40
Time In Days
Pentaohlorophenol
40 n
§ 30-
I 20-
>-
S 10-
o-
c
i
• _ • - initial concentration m 60 mg/kg of soil
m • - //?/f/a/ concentration m 30 mg/kg of soil
• S B
8
1 10 20 30
Time In Days
Figure 10. Toxlclty Reduction In the WSF from the Soil Microcosms
78
-------
to
w
o
"m <" 20
ra Q *"
o°
E —
62
= 2 «1
-1.39+ 1.47x
R-0.87
10 21
Half Life In Days
WSF Chemical Loss
30
Figure 11. Comparison of Chemical Loss In the Soil and the WSF
for Phenol and Eight Chlorophenols
The toxicity reduction study was conducted to evaluate the possibility of toxic
intermediate chemicals and their effect, if any, on the treatment of the chemical applied
to the soil. The WSF of the land applied hazardous constituents pose the immediate
threat to soil microbes and groundwater, and the identification of any potentially
leachable toxic constituents is important to evaluate the performance of a HWLT.
The toxicity of the WSF could be a result of: (a) the chemical added to the soil,
(b) intermediate transformation products that are potentially teachable, and (c)
background toxicity from the soil. The analyses of blank samples (soil only) did not
show any background toxicity from the Texas soil used in this research. The toxicity of
the WSF extracted on day zero was contributed only by the applied target chemical,
since degradation had not yet occurred. The subsequent reduction in toxicity of the
WSF was a result of detoxification and immobilization reactions in the soil.
The chemical loss in the soil and the WSF and the WSF toxicity reduction were
compared. In each case, the WSF toxicity decreased as the soil chemical and the
79
-------
WSF chemical concentrations decreased. Figure 12 provides an example of the
decreases for 2,4-Dichlorophenol.
«^
o
o
w
0
0
o
E
250-
200
150
100
SO
0 .
•
I »
1
" '
<_« '
It
i
O
in
o
- WSF toxicity reduction r> » ui
- chemical toss in soil
-WSF chemical toss
-
n
• • *
L t
0
*
45
H
30 5
c
3
15 >,
*;
_u
• n K
10 20
Time In Days
so
Figure 12. Decrease of Soil and WSF Chemical Concentration and of WSF Toxicity as a
Function of Time-Degradation Study of 2,4-Dlchlorophenol
Figure 13 compares the WSF chemical loss and the WSF toxicity reduction for
all nine tested chemicals at both low and high loadings. The correlation of 0.90
indicates that the WSF toxicity can be attributed to the target chemical concentration in
the WSF and that no water extractable toxic intermediate products were formed. Thus,
these chemicals were detoxified in the soil.
3 >»
10
(0
y-2.0 + 0.95x R-0.90
0 10 20
Half Life In Days
WSF Chemical Loss
Figure 13. Comparison of WSF Chemical Loss and the WSF Toxicity Reduction for
Phenol and Eight Chlorophenols
80
-------
CONCLUSIONS
The loss of chemical in the soil and in the WSF extracted from the soil and the
reduction of toxicity of the WSF were evaluated for nine chemicals that were part of this
study: phenol and eight chlorophenols. Two loading rates and one soil (Texas soil)
were used. The results were:
1. Both first and zero order kinetics satisfactorily represented the chemical loss
and toxicity reduction data.
2. The microcosms with Pentachlorophenol resulted in a lag phase in chemical
loss and WSF toxicity reduction, especially at the higher loading rate. No lag
periods were observed with any of the other chemicals at either loading rate.
3. The higher chemical loading rates resulted in higher chemical concentrations in
the WSF and higher WSF toxicities at time zero.
4. The higher chemical loading rates generally resulted in slower chemical losses
(higher half-lives) and toxicity reduction. However, at both loading rates for
each chemical, the chemicals were degraded and the toxicity was reduced.
5. No differences were apparent in zero order kinetics due to the loading rates.
6. The loss of the chemicals in the WSF was about 1.5 times faster than the loss of
the chemical in the soil.
7. The WSF toxicity for each chemical decreased as the soil chemical and the
WSF chemical concentrations decreased.
8. The WSF toxicity decreased at about the same rate as the WSF chemical
concentration when the data for all the nine chemicals were compared.
9. No enhanced mobilization of the applied chemical occurred as the degradation
and detoxification occurred.
81
-------
SECTION 8
MUNITIONS WASTES AND CHEMICALS
As part of this cooperative agreement, the soil treatablility potential of a hazardous
waste generated in the explosives industry and several chemicals used in that
industry were evaluated (Table 2). The toxicity and adsorption behavior of these
chemicals and the waste were determined using procedures outlined in Sections 3
and 4. Because of limited quantities of ROX and HMX available, these compounds
were not evaluated as part of the degradation studies. RDX and HMX were obtained
from the United States Army Toxic and Hazardous Materials Agency (USATHAMA).
TNT was obtained commercially from Chem Services, Inc. in Pennsylvania. 2,4- and
2,6-Dinitrotoluene were purchased from Aldrich Chemicals, Milwaukee, Wisconsin.
In addition to these chemicals, wastewater treatment sludge from the
manufacturing of explosives at the Holston Army Ammunition Plant was evaluated for
its land treatability potential. This sludge was obtained with the help of USATHAMA.
The following sections summarize the toxicity, adsorption and degradation
characteristics of the pure compounds and the land treatability potential of munitions
waste treatment sludge.
RELATIVE TOXICITY AND LOADING EVALUATION
Results of the toxicity evaluation are summarized in Table 36. As part of the
adsorption experiments, the maximum solubility of HMX was established as 3.8 mg/l.
This low solubility made the toxicity evaluation difficult. The toxicity of hydrophobic
compounds cannot be properly evaluated using the Microtox© system. Because of the
limited quantities of HMX and RDX that were obtained, neither chemical was able to
be evaluated for soil mass loading ranges.
82
-------
TABLE 36. EC5Q DATA AND ACCEPTABLE LOADING RATES FOR MUNITIONS
MANUFACTURING CHEMICALS
ECgQ Value (mg/h Loading Rates (mg/kg of soil)*
Compound Value 95% Cl Texas Soil Mississippi Soil
2,4-Dinitrotoluene 31.2 29.7-32.8 500 165
2,6-Dinitrotoluene 4.4 4.3-4.5 86 74
TNT (2,4,6-Trinitrotoluene) 1.0 0.7-1.3 14 12
RDX 76.1 60.0-97.0 * *
HMX " " * *
* Loading rate data for RDX and HMX could not be evaluatod since sufficient quantities of
material could not be obtained.
'' Due to its insolubility, EC5Q data for HMX could not be obtained.
+ Lower loading rate as determined using the procedure in Table 9
ADSORPTION
Methods
The procedure followed for evaluation of the adsorption behavior of RDX, HMX,
and TNT was a modification of that outlined in Section 3. The following section
describes the protocol used to evaluate the sorption behavior of these chemicals.
RDX and HMX were received as aqueous solutions consisting of approximately
100 mg of chemical in five milliliters of water. Prior to use, the compounds were dried
at 105° C. After drying, each chemical was used to prepare standard stock solutions
using a 50:50 mixture of waterMethanol as the solvent. This protocol improved the
accuracy of chemical detection. The final concentrations of the RDX and HMX stock
solutions were 41.8 and 3.8 mg/l, respectively. The resulting concentration of HMX
was very close to the detection limit of this compound by high pressure liquid
chromatography (~1 mg/l).
83
-------
TNT was received as a solid consisting of 20% moisture. The moisture content
was taken into account in preparing a stock solution of 700 mg/l TNT using a 50:50
mixture of watenmethanol as the solvent.
External standard solutions were prepared from the stock solutions by a serial
dilution technique utilizing the 50:50 watermethanol solvent mixture. These external
standards were used to evaluate the concentration of aqueous chemical stock
solutions used in adsorption tests. Chemical concentrations were determined using
the HPLC and a 50:50 watermethanol solution as the eluent for the RDX and HMX.
Methanol was used as the eluent for TNT.
Results
The range of chemical concentrations evaluated is noted in Table 37. The
adsorption results for 2,4- and 2,6-Dinitrotoluene are presented in Table 38. The
Freundlich equation described the sorption of these chemicals on the two soils
satisfactorily, i.e., with high correlation coefficients. However, such was not the case
for the other munitions chemicals.
The sorption data for TNT, RDX and HMX are presented in Tables 39 and 40.
With TNT, for both soils, the value of qe reached a maximum of about 0.045 mg
TNT/gm dry soil at a solution concentration of about 15 mg/l TNT and then decreased
at higher solution concentrations. This is not in agreement with the Freundlich
equation in which qe should increase as the compound concentration approaches its
solubility limit(31). Data for the other chemicals (RDX and HMX) also produced poor
correlation with the Freundlich equation. The correlation coefficients ranged from 0.0
to 0.44.
As discussed in Section 6, the adsorption of a chemical is affected by a number
of characteristics, none of which were evaluated in these experiments. For the two
chemicals for which the Freundlich equation did seem to fit the adsorption data (Table
38), the Texas soil appeared to have a greater affinity (higher Kf value) for 2,4-
84
-------
TABLE 37. CHEMICAL CONCENTRATION RANGE EVALUATED DURING THE BATCH
ADSORPTION EXPERIMENTS •• MUNITIONS CHEMICALS
Concentration Range fma/ko)
Compound
2,4-Dinitrotoluene
2,6-Dinitrotoluene
(2,4,6-Trinitrotoluene) TNT
RDX
HMX
Texas Soil
32-160
37-600
0.1-60
8-30
0.9-3.6
Mississippi Soil
15-150
14-200
0.3-90
11-32
1.5-3.8
TABLE 38. FREUNDLICH ISOTHERM DATA - TEXAS AND MISSISSIPPI SOILS
2,4- AND 2,6-DINITROTOLUENE
Compound
'correlation coefficient
1/n
r*
PH
Texas Soil
2,4-Dinitrotoluene
2,6-Dinitrotoluene
Mississippi Soil
2,4-Dinitrotoluene
2,6-Dinitrotoluene
9.2x1 0'3
5.0x1 0'3
6.3x1 0"3
1.4x10"2
0.59
0.61
1.08
0.68
0.88
0.90
0.99
0.95
7.6
8.1
5.2
5.6
85
-------
TABLE 39. ADSORPTION DATA FOR TNT, HMX AND RDX
IN TEXAS SOIL AT 20° C
2,4,6-Trlnltrotoluene (TNT) (pH=7.8)
Concentration fmg/l) q£ (mg sorbed/ g dry soin
0.1 0.0031
6.5 0.0238
13.2 0.0475
60.8 0.0311
RDX (pH=8.0)
Concentration (mg/l) q^ (mg sorbed/ g dry soil)
8.6 0.0077
10.7 0.0056
11.7 0.0151
13.7 0.0106
26.6 0.0154
HMX (pH=7.9)
Concentration (mg/l) q^ (mg sorbed/ g dry soil)
0.9 0.00322
1.6 0.00086
2.3 0.00322
3.6 0.00036
TABLE 40. ADSORPTION DATA FOR TNT, HMX AND RDX
IN MISSISSIPPI SOIL AT 20° C
2,4,6 Trinitrotoluene (pH=5.8)
Concentration (mg/h q^ (mg sorbed/ a dry soin
0.4 0.00275
10.6 0.0197
17.9 0.0428
90.7 0.0012
RDX (pH=5.8)
Concentration (mg/h q^ (mg sorbed/ g dry sol!)
11.3 0.00498
11.9 0.00439
12.7 0.0142
19.2 0.00506
32.3 0.00973
HMX (pH=5.9)
Concentration (mg/n q^ (mg sorbed/ g dry soil)
1.5 ' 0.00092
1.7 0.00069
3.8 0.00022
86
-------
Dinitrotoluene while the Mississippi soil appeared to have a greater affinity for the 2,6-
Dinitrotoluene.
DEGRADATION STUDIES
The experimental procedures for these degradation studies were identical to
those described in Section 5. The degradation of RDX and HMX could not be
evaluated due to the limited quantities that were available.
The recovery efficiencies and the loadings that were used in these degradation
studies are presented in Table 41. Even though the loading rate for 2,4-Dinitrotoluene
was in the acceptable range, no loss of this chemical occurred over a 47-day study.
However, losses of 2,6-Dinitrotoluene and 2,4,6-Trinitrotoluene (TNT) did occur. The
loss pattern for 2,6-Dinitrotoluene in both soils is presented in Figure 14. The loss
rates for the two chemicals are indicated in Table 42.
The data indicate that first order kinetics were a better representation for TNT
than were zero order kinetics. The data also indicate that the first order half-life of TNT
in the Mississippi soil was less, and the loss faster, than in the Texas soil. No
differences in the loss rates for 2,6-Dinitrotoluene for the two soils were apparent.
MUNITIONS WASTEWATER TREATMENT SLUDGE
In addition to the compounds noted above, the land treatability potential of
wastewater sludge resulting from the manufacture and processing explosives at the
Holston Army Ammunition Plant (HAAP) was evaluated. Figure 15 indicates the
wastewater treatment process flow diagram at HAAP. The sludge evaluated was
obtained from the Area B raw wastewater settling basin. Prior to use, the
characteristics of this sludge were determined.
Nitrogen and COD -- Nitrogen and COD concentrations of the sludge (Table
43) were determined according to procedures described in Standard Methods(36).
87
-------
TABLE 41. LOADING RATE AND RECOVERY EFFICIENCY DATA FROM THE
MUNITIONS CHEMICAL DEGRADATION STUDIES
Compound
Loading Rate (trig/kg of soil)
Acceptable* Actual
Average Recovery
Efficiency /%1++
Texas Soil
2,4-Dinitrotoluene 500 500 100
2,6-Dinitrotoluene 86 85 91
2,4,6-Trinitrotoluene 14 14 :...100
Mississippi Soil
2,4-Dinitrotoluene 165 150 92
2,6-Dinitrotoluene 74 70 85
2,4,6-Trinitrotoluene 12 12 101
+ data from Table 36
** average of three replicates __
TABLE 42. CHEMICAL LOSS RATE DATA FOR 2,6-DINITROTOLUENE AND TNT
IN THE DEGRADATION STUDIES
KINETIC PARAMETERS
COMPOUND
Texas So//
2,6-Dinitrotoluene
2,4,6-Trinitrotoluene
HALF LIFE
(day)
72
7.7
FIRST
*
r
0.91
0.94
ORDER
95% C.I.
59-90
6.5-9.4
7ERO ORDER
mg/Kg/day
0.7
0.5
r*
0.91
0.81
95% C.I
0.6-0.9
0.3-0.6
Mississippi Soil
2,6-Dinitrotoluene
2,4,6-Trinitrotoluene
92
5.7
0.78
0.93
66-153
4.6-7.2
0.5
0.4
0.80
0.80
0.3-0.6
0.2-0.6
correlation coefficient
88
-------
Concentration of 2 6-DNT
(mg/Kg of soil)
oo
to
c
o
c*
o
«•»
M
0>
6
o
o
c
2
s
en
fit
a
(0
(0
(0
(0
•o
•o
(A
O
O
o
O
Concentration of 2,6-DNT
(mg/Kg of soil)
en -
1
a
O
10
o
to
en
u
O
CO
en
NJi
5" 8s
0)
o
I
-------
AREA B
RAW WASTE
WATER
Nutrient Additions
and oH Control
Stilling Bttln
Denltriflcatlon by
Submerged Anaerobic
Filters
AREA A
RAW WASTE
WATER
PH
/
\
Equtllzttlan
Buln
Control
Nutrient
Additions
& pH
Control
'Aerobic^
/Fixed Fllm\
Reactors
V (Trickling /
\Fllters)/
Aerobic Suspended
Growth Reactors
(Activated Sludge)
Figure 15. Wastewater Treatment Process Flow Diagram for the Holston Army
Ammunition
90
-------
TABLE 43. NITROGEN AND COD CONCENTRATIONS OF MUNITIONS WASTE SLUDGE
Total Kjeldahl Nitrogen 4820 mg/L
Ammonia Nitrogen 2300 mg/L
C.O.D 1.27 x 105 mg/L
Metals -- The metals in the munitions sludge were analyzed at the USEPA Robert
S. Kerr Environmental Research Laboratory in Ada, Oklahoma. Results were obtained
using procedures outlined in EPA SW 846 for both the total sludge and the liquid
fraction. The results are contained in Table 44.
GC/MS Analysis -- One goal of this study was to determine if quantities of
hazardous organics were present in the munitions waste sludge. If significant
quantities were present, degradation experiments would be conducted to estimate
loss rates of these constituents. The organics present in the sludge were determined
as follows. The organics were extracted from the sludge using a shake extraction
procedure and Methylene Chloride as the extracting solvent (EPA method 3450)^).
The extract was analyzed on a Finnigan-MAT4000 gas chromatograph/mass
spectrometer (GC/MS) located in the Department of Chemistry at The University of
Texas at Austin. Results are summarized in Table 45. The listed compounds
represent the fourteen most significant peaks of the chromatogram. Many small peaks
were observed but they had a peak intensity of the same magnitude as instrument
noise. None of the small peaks corresponded to TNT, HMX or RDX. Because of the
absence of these compounds in the munitions waste sludge, degradation experiments
with the sludge were not conducted.
Relative Toxlcltv -- The toxicity of the munitions sludge was evaluated using
the Microtox© procedure. It is possible that the chemicals contained as part of the
sludge solids could cause toxicity as the solids decomposed. To determine if the
91
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TABLE 44. METALS IN MUNITIONS SLUDGE AND SLUDGE FILTRATE
Element
Na
K
Ca
Mg
Fe
Mn
Co
Mo
Al
As
Se
Cd
Be
Cu
Cr
Ni
Zn
Ag
Tl
Pb
Li
Sn
V
Ba
B
Ti
mg/l
20.7
18.8
45.8
12.4
400
1.10
0.02
<.01
<.1
<.03
<.1
<.003
<.003
0.03
0.01
0.01
0.10
<.03
<.04
<.02
<.01
0.17
<.03
0.01
0.23
<.1
Filtrate
Std. Dev.
2.0
1.9
4.5
1.2
40
0.09
0.01
-
-
-
-
-
-
0.01
0.01
0.01
0.01
-
-
-
-
0.01
-
0.01
0.03
_
Total
mg/kg
wet wt.
92.8
202
711
196
3940
14.7
0.81
0.69
1740
5.8
<1
0.19
0.21
13.9
7.85
2.85
35.9
<.3
<.7
10.4
1.15
3.59
4.76
11.0
0.88
6.7
Sample
Std. Dev.
9.2
20
71
1.9
390
1.3
0.15
0.26
170
2.50
-
0.04
0.04
1.3
0.77
0.28
3.6
-
-
1.5
0.14
0.35
0.71
1.1
0.31
1.4
92
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TABLE 45. GC/MS ANALYSIS OF MUNITIONS SLUDGE
COMPOUND MOLECULAR WT.
6-Methyl. 1-H Indole
CgHgN
Benzenemethanlmlne
C/H7N
1.3 Benzoxazlne
CfgHi7ON
8-Methyl. Decanolc Acid
C12H24O2
2.2-Dlmethylcvclohexanol
C8H16OCI
10-Undecenoyl Chlorlda
CiiHigOCI
8-Methvl Paeanolc Acid
C12H2402
1-Methyl. 4-Nltrosoplperazlne
C5HnON3
Sulfur
S8
Undecenal
7- Methyl. Nonanolc acid
Hexanedlolc Acid
C14H26O4
3-Nltro. 1.2-Benzana-
DIcarboxyHc Acid
CsHsOgN
Thlocyanlc Acid
C-| iH-jsONS
131
105
239
200
128
202
202
129
256
168
186
258
21 1
207
% FIT* PEAK INTENSITY**
93.8
89.1
73.1
79.0
83.8
84.3
87.3
70.2
84.8
82.7
85.4
84.1
85.7
81.4
268
81
.402
338
172
170
290
242
641
148
240
223
443
254
* The % fit refers to how well the library fit matched the ion chromatograph generated from the sample.
* * Peak intensity refers to the relative concentrations of the compounds in the extract. For example, sulfur (Sg)
had the largest concentration of the fourteen compounds listed.
93
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solids may have included chemicals that exhibited a toxic effect, one set of samples
was homogenized and the other was not before the toxicity was determined. With the
Microtox© procedure, the soluble constituents exert the greatest effect.
Preparation of both sets of samples consisted of taking aliquots of
homogenized and aliquots of nonhomogenized sludge (100 ml each), diluting them
with distilled deionized water to obtain different concentrations of the sludge
constituents and centrifuging them at 2000 rpm for 15 minutes. Fifty milliliters of the
supernatant liquid from each sample were discarded and replaced by fifty milliliters of
a 2% Sodium Chloride/distilled deionized water solution. The addition of the salt
solution maintained the proper environment for the marine bioluminescent bacteria
used in the toxicity test. This procedure was in accordance with standard Microtox©
operating methods (Section 4). The samples were then centrifuged at 2000 rpm for an
additional 15 minutes. Twenty five milliliters of the supernatant were withdrawn from
each sample and filtered (0.45 |im pore size filter). Within thirty minutes after filtering,
the samples went from clear to a brownish red color suggesting the oxidation of some
metal species. Because of the color interference, a color absorbance correction was
necessary to obtain toxicity results. The results of the toxicity evaluation are
summarized in Table 46.
As noted above, the homogenized and nonhomogenized sludge samples
were diluted to provide varying concentrations of sludge constituents for
the toxicity evaluation. This resulted in the toxicity results being known
in terms of percent of the original sludge. If the sludge were nontoxic,
the ECso value would be around 100%. These evaluations (Table 46)
indicated that high dilutions were needed to obtain £659 values and that,
therefore, the sludge was toxic to the Microtox® organisms.
94
-------
Because these samples demonstrated significant toxicity, a second set of
samples was processed in the same way and the relative toxicity determined to verify
the previous results. The results from this second evaluation were comparable to
those presented in Table 46.
TABLE 46. MUNITIONS WASTE TOXICITY DATA - UNDILUTED SAMPLES
Sample H&C(D*
EC50(5min., 15°C) 0.92%
95% confidence interval 0.52 -1 .65%
pH of the sample 5.6
H&C(2)*
1 .30%
1.25 -1.35%
5.7
cm
1.22%
1.16-1.28%
5.7
C(2)
1 .47%
1.37-1.57%
5.7
* H - homogenized, C - centrifuged; (1) and (2) are replicates.
The relative toxicity results indicated that: (a) the munitions sludge exhibited
considerable relative toxicity, and (b) there was no difference in relative toxicity
between the homogenized and the nonhomogenized samples. The latter statement
indicates that the constituents causing the toxicity effect were in the soluble and not the
solid phase. The low pH of the sludge (Table 46) may have contributed to the relative
toxicity.
CONCLUSIONS
1. The Freundlich equation described the sorption of 2,4- and 2,6-Dinitrotoluene in
the two soils satisfactorily. However, it did not do so for TNT, RDX, or HMX.
2. No loss of 2,4-Dinitrotoluene occurred over a 47-day study, even though the
loading rate used was determined to be acceptable using procedures
discussed in Section 4.
3. Because of the small amounts of RDX and HMX that were received, no
degradation studies could be conducted.
95
-------
4. Degradation loss rates could be obtained for 2,6-Dinitrotoluene and TNT. First
order kinetics were a better representation for TNT than were zero order
kinetics.
5. The half-life of TNT in the Mississippi soil was shorter, and the loss faster, than
in the Texas soil. No differences in the loss rates in the two soils for 2,6-
Dinitrotoluene were apparent.
6. The sludge resulting from the manufacture and processing of explosives
contained: (a) high concentrations of nitrogen and COD, (b) concentrations
generally less than 10 mg/l for the heavy metals, and (c) no amounts of TNT,
RDX, or HMX.
7. The munitions sludge had a high toxicity as measured by the Microtox©
procedure. The constituents causing the relative toxicity were in the soluble
and not the solid phase of the sludge.
96
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SECTION 9
REFERENCES
1. Short, Thomas E. "Modeling of Processes in the Unsaturated Zone". In: B.C.
Loehr and J.F. Malina, Jr., Editors, Land Treatment: A Hazardous Waste
Management Alternative. Water Resources Symposium 13, Center for
Research in Water Resources, The University of Texas at Austin, Austin, Texas
78712, 1986, pp. 211-240.
2. Nofziger, D.L, Williams, J.R. and T.E. Short. "Interactive Simulation of the Fate of
Hazardous Chemicals During Land Treatment of Oily Wastes: RITZ Users'
Guide". Robert S. Kerr Environmental Research Laboratory, U.S. Environmental
Protection Agency, Ada, Oklahoma, 1988, 59 pages.
3. Caupp, C.L., Grenney, W.J., and P.J. Ludvigsen. "VIP -- A Model for the
Evaluation of Hazardous Substances in Soil -- Version 2". Civil and
Environmental Engineering Department, Utah State University, Logan, Utah,
1988, 25 pages.
4. Grenney, W.J., Caupp, C.L, Sims, R.C. and T.E. Short. "A Mathematical Model
for the Fate of Hazardous Substances in Soil: Model Description and
Experimental Results". Haz. Waste and Haz. Materials 4:223-240,1987.
5. Charbeneau, R.J., Weaver, J.W. and V.J. Smith. "Kinematic Modelling of
Multiphase Solute Transport in the Vadose Zone". Final Report, Cooperative
Agreement CR-813080, Robert S. Kerr Environmental Research Laboratory, U.S.
Environmental Protection Agency, Ada, Oklahoma, 1989,108 pages.
6. McGinnis, G.D., Borazjani, H., McFarland, L.K., Pope, D.F. and D.A. Strobel.
"Characterization and Laboratory Soil Treatability Studies for Creosote and
Pentachlorophenol Sludges and Contaminated Soil". Final Project Report,
Robert S. Kerr Environmental Research Laboratory, U.S. Environmental
Research Laboratory, U.S. Environmental Protection Agency, Ada, Oklahoma,
1988, 138 pages.
7. U.S. Environmental Protection Agency. Test Methods for Evaluating Solid
Waste SW-846, Volumes 1-4 (Third Edition). National Technical Information
Service, Springfield, Virginia, 1986.
8. Dutka, B.J. and G. Bitton. Toxicity Testing Using Microorganisms. Boca
Raton, Florida, CRC Press, Inc., 1986, 395 pages.
9. Liu, D. and B.J. Dutka (Editors). Toxicity Screening Procedures Using
Bacterial Systems. Marcel Dekker, Inc., New York, 1984, 470 pages.
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10. Hermans, J., Busser, F., Leeuwangh, P. and A. Musch. "QSAR's and Mixture
Toxicity of Organic Chemicals in Photobacterium Phosphoreum: The Microtox©
Test". Ecotox. Environ. Safety 9:17-25,1985.
11. Kamlet, M.J., Doherty, R.M., Veith, G.D., Taft, R.W., and M.H. Abraham. "Solubility
Properties in Polymers and Biological Media 7. An Analysis of Toxicant
Properties That Influence Inhibition of Bioluminescence in Photobacterium
Phosphorium (The Microtox Test)". Environ. Sci. Tech.: 690-695, 1986.
12. Matthews, J.E. and A.A. Bulich. "A Toxicity Reduction Test System to Assist in
Predicting Land Treatability of Hazardous Organic Wastes". In: J.K. Petros, Jr. et
al., Editors, Hazardous and Industrial Solid Waste Testing: Fourth
Symposium. Philadelphia, ASTM/STP 886, 1984.
13. Matthews, J.E. and L Hastings. "Evaluation of Toxicity Test Procedure for
Screening Treatability Potential of Waste in Soil". Toxicity Assessment 2. 265-
281,1987.
14. Beckman Instruments, Inc.. Microtox® System Operating Manual. Beckman
Instruments, Inc., Carlsbad, California, 1982, 52 pages.
15. Sims, R.C. "Loading Rates and Frequencies for Land Treatment Systems in Land
Treatment". In: R.C. Loehr and J.F. Malina, Jr., Editors, Land Treatment: A
Hazardous Waste Management Alternative. Water Resources Symposium
13, Center for Research in Water Resources, The University of Texas at Austin,
Austin, Texas 78712, 1986, 370 pages.
16. Huddleston, R.L, Bleckmann, C.A. and J.R. Wolfe. "Land Treatment Biological
Degradation Processes". In: R.C. Loehr and J.F. Malina, Jr., Editors, Land
Treatment: A Hazardous Waste Management Alternative. Water
Resources Symposium 13, Center for Research in Water Resources, The
University of Texas at Austin, Austin, Texas 78712,1986, 370 pages.
17. Edgehill, R.V. and R.K. Finn. "Microbial Treatment of Soil to Remove
Pentachlorophenol". Appl. Environ. Microbiol. 45:1122-1125, 1983.
18. Kennedy, J.B. and A.M. Neville. Basic Statistical Methods for Engineers
and Scientists (Third Edition). Harper and Row, Inc., New York, 1986, 495
pages.
19. Weber, W.J., Jr. Physicochemical Process for Water Quality Control.
John Wiley and Sons, Inc., New York, 1972, 640 pages.
20. Chiou, C.T., Porter, P.E. and D.W. Schmedding. "Partition Equilibrium of
Nonionic Organic Compounds Between Soil Organic Matter and Water". Environ.
Sci. Technol 17:227-231, 1983.
21. Sheets, T.J., Crafts, A.S. and H.R. Drever. "Influence of Soil Properties on the
Phytotoxicity of s-triazines". J. Agric. Food Chem. 70:458-462,1962.
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22. Savage, K.E. and R.D. Wauchope. "Fluometuron Adsorption-Desorption
Equlibria in Soils". Weed Sci. 21-A 06-110, 1974.
23. Hamaker, J.W. and J.M. Thompson. "Adsorption". In: Goring and Hamaker,
Editors, Organic Chemicals in the Environment. Marcel Dekker, Inc., New
York, 1972.
24. Rao, P.S.C. and J.M. Davidson. "Estimation of Pesticide Retention and
Transportation Parameters Required in Nonpoint Source Pollution Models".
Environmental Impact of Nonpoint Source Pollution, Ann Arbor Science
Publications, Ann Arbor, Michigan, 1980, pages 23-67.
25. McCarty, P.L, Reinhard, M. and B.E. Rittmann. "Trace Organics in Groundwater".
Environ. Sci. Technol. 75:40-51, 1981.
26. Schwarzenbach, R.P. and J. Westell. "Transport of Nonpolar Organic
Compounds from Surface Water to Groundwater". Environ. Sci. Technol.
75:1360-1367, 1981.
27. Sawyer, C.N. and and P.L. McCarty. Chemistry for Environmental
Engineers (Third Edition). McGraw-Hill, Inc., New York, New York, 1978, 370
pages.
28. Schellenberg, K., Leuenberger, C. and R.P. Schwarzenbach. "Sorption of
Chlorinated Phenols by Natural Sediments and Aquifer Materials". Environ. Sci.
Technol. 18:652-657, 1984.
29. Zachara, J.M., Alnsworth, C.C., Cowan, C.E. and B.L Thomas. "Sorption of
Binary Mixtures of Aromatic Nitrogen Heterocyclic Compounds on Subsurface
Materials". Environ. Sci. Technol. 21:397-402, 1987.
30. Murin, C.J. and V.L Snoeyink. "Competitive Adsorption of 2,4-Dichlorophenol
and 2,4,6-Trichlorophenol in the Nanomolar to Micromolar Concentration Range".
Environ. Sci. Technol. 73:305-311, 1979.
31. U.S. Environmental Protection Agency. "Batch Type Adsorption Procedures for
Estimating Soil Attenuation of Chemicals". Technical Resource Document
(EPA/530-SW-87-006), Office of Solid Waste and Emergency Response,
Washington, D.C. , 1986, 183 pages.
32. Verschueren, K. Handbook of Environmental Data on Organic
Chemicals, Second Edition. Van Nostrand Reinhold Company, Inc., New
York , 1983, 1310 pages.
33. American Society for Testing Materials. Annual Book of ASTM Standards,
Part 19. Soil and Rock, Building Stone. Philadelphia, Pennsylvania,
ASTM D2216, 1982, pp. 338-339.
99
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34. U.S. Environmental Protection Agency, Office of Solid Waste. Permit
Guidance Manual on Hazardous Waste Land Treatment
Demonstrations - Final Version. National Technical Information Service,
Springfield, Virginia , 1986, 98 pages.
35. Burrows, R.D. Letter to R.C. Loehr, December 1986.
36. American Public Health Association. Standard Methods for the
Examination of Water and Wastewater, 16th Edition. American Public
Health Association, Washington, DC, 1985, 1268 pages.
37. Qureshi, A.A., Coleman, R.N., and J.H. Paran. "Evaluation and Refinement of the
Microtox® Test". In: D. Liu and B.J. Dutka, Editors, Toxlcity Screening
Procedures Using Bacterial Systems. New York, Marcel Dekker, Inc.,
1984, pages 1-22.
100
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APPENDIX A
THE MICROTOX® TOXICITY ASSAY
USED IN THIS STUDY
INTRODUCTION
The Microtox® toxicity assay was used for toxicity screening due to its simplicity,
rapidity, cost effectiveness, and because the assay procedure had undergone
evaluation and standardization for this purpose^ 2- 13). The test organism is a
marine bioluminescent bacteria (Photobacterium phosphoreum Strain NRRL B-
11177). The Microtox® Model 2055 Toxicity Analyzer System and Microtox®
accessories used in this research were obtained from Beckman Instruments, Inc., and
Microbics, Inc., Carlsbad, California.
EVALUATION OF CHEMICAL LOSS USING MICROCOSMS
The batch type microcosms used to simulate aerobic soil conditions were 150
ml_ glass beakers. The chemicals were extracted from the soil microcosms with
Methylene Chloride using a Soxhlet extraction apparatus. The extract was
concentrated using a Kuderna-Danish apparatus. The final extract was analyzed with
a Model 5890 (Hewlett Packard) Gas Chromatographic System.
WATER EXTRACTION OF MICROCOSMS
For the Microtox® analyses, a water soluble fraction (WSF) is needed. The
contents of a microcosm were transferred into an amber glass bottle having a Teflon-
lined cap and 40 mL (1:4 by soil weight:water volume) of distilled deionized water was
added. The soil-water mixture was extracted in a rotary extractor for about 24 hours at
30 rpm at room temperature. Immediately after the extraction, the sample was
centrifuged at about 2000 rpm for 15 minutes to separate the WSF. The WSF was
vacuum filtered through a 0.45 ^im membrane filter. Ten milliliters of the filtered
sample were adjusted to 2% NaCI using reagent grade dry NaCI and tested for
toxicity. In addition, the pH of the composite sample of triplicates of a chemical
101
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loading at a sampling time was measured. The remaining portion of the WSF was
stored in a sample vial with minimum head space. The vial was frozen to < -20° C to
maintain the integrity of the chemical composition of the sample until further analysis
by HPLC.
TOXICITY ASSAY
The toxicity assay was used to: (a) determine the Effective Concentration
(EC50) of a chemical solution or of a waste, (b) determine the EC50 of the WSF to
determine the safe initial chemical loading on soil, and (c) study the toxicity reduction
of the WSF.
EC50 DETERMINATION
The EC5Q of a sample is defined as the concentration of the sample required to
reduce the initial luminescence of the bacterial suspension by 50% under standard
test conditions. To determine the EC50 of the chemicals that were evaluated, a
solution of a known concentration of a chemical was prepared in distilled deionized
water (DDW). The DDW used in the sample preparation did not exhibit any toxicity as
determined by the Microtox® assay. The solution was mixed using a magnetic stirrer
for 24 hours or until the chemical was completely dissolved. To the extent possible,
the solubility of the chemical was estimated from the literature, before attempting
sample preparation. The chemical solution was adjusted to 2% Sodium Chloride
(NaCI) using either reagent grade dry NaCI or Microtox© Osmotic Adjusting Solution
(MOAS -- 22% NaCI solution).
The sample preparation for EC$Q values that were part of the acceptable
loading determination and toxicity reduction studies followed the same procedure.
ASSAY PROCEDURE
This section.briefly describes the standard Microtox® toxicity assay procedure.
A detailed description of the assay can be found elsewhere (14> 37).
102
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The lyophilized Microtox© bacteria are reconstituted with 1.0 ml of
reconstitution solution (ultra-pure water) maintained at 2-4° C in a glass cuvette.
Aliquots of 10jiL of the reconstituted bacterial suspension are equilibrated for about 15
minutes at 15° C in 0.5 mL of Microtox© diluent (2% NaCI solution) in glass cuvettes.
At the end of the equilibration period, the initial luminescence intensity is measured as
light units using the Microtox© System. 0.5 ml of the sample and three serial dilutions
of the sample are added to the respective cuvettes. The final luminescence readings
are taken at the end of the desired exposure time intervals: 5 minutes, 15 minutes, 30
minutes or longer.
The test sample and its dilutions were tested in duplicate. Blank cuvettes
containing just the diluent also are read to correct for any shift in the luminescence that
may be caused by the bacteria or the machine. From the observed results a dose-
response curve is obtained and the EC50 value calculated.
CHEMICAL LOADING ON SOIL
The chemical concentration applied to the soil should be within the assimilative
capacity of the soil to avoid toxic effects that may limit the microbial degradation of the
chemical. The safe chemical loading determination is based on the water soluble
fraction (WSF) of the chemical and the toxicity of WSF using the Microtox© assay.
One hundred grams of air-dried soil were weighed into glass jars and mixed
with 400 mL ( 1:4 by soil weight : water volume) of DDW. Immediately, the organic
chemical (solution prepared in ethanol) was added to the soil-water mixture, at
concentrations of 2x, 5x, 10x and 20x where x is the £650 of the chemical. All
chemical loadings were prepared in duplicate. The maximum concentration of the
ethanol in the water was limited by the required chemical loading and the ease with
which the chemical dissolved in ethanol. The maximum concentration of ethanol in
water was 2500 to 5000 ppm. The £€59 value of ethanol using the Microtox© assay
is 30,000-35,000 ppm. Hence, the concentrations of ethanol used in these
103
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experiments did not have a significant influence on the toxicity of WSF. The chemical
was extracted with DDW for about 24 hours in a tumbler shaker rotating at 30 rpm at
room temperature (usually 21-25° C).
After the extraction, the soil-water mixture was allowed to settle for about 20
minutes. About 50 ml_ of the supernatant was transferred into small glass bottles
which were centrifuged at about 2000 rpm for 15 minutes to separate the WSF from
the soil particles. The WSF was filtered through a 0.45 p.m filter using a vacuum. The
filtered sample was osmotically adjusted to 2% NaCI with dry NaCI and analyzed
using the Microtox© system. These results were used to indicate the safe lower
chemical loading on the soil. The upper limit was set as two times the lower chemical
loading.
MICROTOX© ANALYSIS
The sample analysis using the Microtox© system was done as described
above. The luminescence readings are converted into toxicity measurements as
indicated in the system operating manuaK14):
where
Ij = Initial luminescence reading
If = Final luminescence reading and,
X = Blank Ratio = If + Ij from the blank cuvette luminescence reading
G = Relative toxicity measurement
The concentration of the sample, as ppm or % of the sample solution, is plotted
against the toxicity measurements, G, on a log-log scale. The concentration of the
sample corresponding to a toxicity measurement of 1.0 is termed the EC§Q of the
sample, as shown below.
In the chemical loading determination and toxicity reduction study, the EC$Q of
the water extract was converted to soil toxicity units (soil TU) in the following
104
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manner^13- 37); TU = 400 + £€50- The chemical loading and the corresponding soil
TU were plotted on a log-log graph.
An Example Plot of EC50 Determination
Chemical: Pentachlorophenol
Chemical concentration in ppm
As has been described(13), the point at which the loading rate intercepted the
value of 20 toxicity units (TU), i.e., £650 = 20, was used as the lower limit, or toxic
floor, of the initial loading rate window. The available logic and data(13) indicated that
chemicals or waste that exhibited an £650 less than 20 were likely to cause inhibition
of the microorganisms in the soil and result in no detoxification. An example plot is
shown below.
105
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10
An Example Plot of Chemical Loading Determination
Chemical: 3-chlorophenol
3 _
:f
I10
20 soil TU
Low loading ~ 12mg/1 OOg soil
(calculated value-11.9mg/1 OOg soil)
10° 101
Chemical Loading in mg/100g of soil
10
106
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APPENDIX B
QUALITY CONTROL/QUALITY ASSURANCE PROCEDURES
The objective of this study was to evaluate the relative toxicity, degradation
kinetics and sorption of specific EPA listed hazardous chemicals and a specific
hazardous waste and related chemicals. The data generated consisted of physical
and chemical analyses performed by the project personnel. All data and observations
were recorded in permanent laboratory notebooks. Replicates and standards were
part of the analytical protocol. The following sections describe measures used to
determine the accuracy and precision of the measurements.
TOXICITY
Toxicity screening and relative toxicity evaluations were performed using the
Microtox© system to establish ECso values for each chemical. Procedures for the
Microtox© system outlined in the operation manual were followed^4). ECso values
were determined graphically by evaluating duplicate samples at each chemical
concentration. To evaluate randomness in toxicity data, ECso data and 95%
confidence intervals were reported.
Water soluble fraction (WSF) samples were obtained following procedures
outlined in EPA SW-846(7). Duplicate samples of WSF at each chemical
concentration were used to graphically determine the chemical loading rate as
outlined in Section 3.
DEGRADATION STUDIES
Accuracy and precision of degradation data were monitored routinely. The
procedures and analytical techniques included replicate analyses and determination
of recovery efficiencies as part of the overall quality control effort. Replicate analysis of
three samples at each sampling time avoided basing degradation kinetic results on
one data point which could be an outlier.
107
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Illustrative recovery efficiency data are presented in Table B-1. The analytical
data were reviewed frequently to discover possible anomalies or omissions. If suspect
data were discovered, the raw data used to calculate the results were reviewed and
the experiment in question was repeated if necessary. To determine the conformity of
the degradation data to zero and first order kinetic models, 95% confidence intervals
were determined for all chemicals.
TABLE B-1. ACCURACY DATA: RECOVERY EFFICIENCIES (%) FOR SPECIFIC
CHEMICALS AS DETERMINED FROM DAY ZERO DEGRADATION STUDY
EXPERIMENTS
Compound
#2
#3
Average Std. Dev. C.v.
Texas Soil
Phenol
m-Cresol
2-Chlorophenol
2,4,5-Trichlorophenol
Pentachlorophenol
2,4-Dimethylphenol
3-Methyl-4-Chlorophenol
p-Nitrophenol
Mississippi Soil
Phenol
m-Cresol
2-Chlorophenol
2,4.5-Trichlorophenol
Pentachlorophenol
2,4-Dimethylphenol
3-Methyl-4-Chlorophenol
p-Nitrophenol
97
78
73
99
118
84
89
122
80
93
27
110
108
83
99
100
84
85
71
95
117
80
105
101
75
86
23
107
105
80
101
116
83
81
76
98
110
76
104
108
80
84
24
107
106
80
98
114
88
81.3
73.3
97.3
115
80
99.3
110.3
78.3
87.7
24.7
108
106.3
81
99.3
110
7.8
3.5
2.5
2.1
4.4
4.0
9.0
10.7
2.9
4.7
2.1
1.7
1.5
1.7
1.5
8.7
8.9
4.3
3.4
2.1
3.8
5.0
9.0
9.7
3.7
5.4
8.4
1.6
1.4
2.1
1.5.
7.9
+ standard deviation
++ coefficient deviation (%)
108
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Degradation study procedures, extraction and analytical methods, and data
analysis procedures were described in Section 5. The day zero percent recovery
value for each chemical was assumed to remain unchanged for extractions at later
sampling periods.
To understand the typical variation that could occur with the day zero recovery
data (chemical recovery efficiencies), multiple extractions of three chemicals
representative of those used in this study were conducted. The chemicals were:
Phenol, o-Cresol, and 2,4-Dichlorophenol. In each case, each chemical was added to
the Texas soil and immediately extracted and analyzed using the procedures
presented in Sections 3 and 5. From eighteen to thirty-one extractions and analyses
were conducted with these chemicals.
The results that were obtained are shown in Figures B-1, B-2 and B-3. Data in
the figures include the average percent recovery, two standard deviations of the data
as upper and lower warning limits, and three standard deviations as the upper and
lower control limits. Ideally, the data should fall within two standard deviations of the
mean. Based on the data in these figures, the recovery efficiencies used in the
degradation study calculations (Table 16) fall within satisfactory limits. Since the
recoveries for these three chemicals were within satisfactory limits, it was assumed
that the recoveries for the other chemicals (Table 16) also were within satisfactory
limits.
ADSORPTION STUDIES
Procedures used to obtain the adsorption data were described in Section 6.
Isotherms were determined using soil-chemical mixtures with different initial
concentrations of chemical. Replicate samples of the low and high concentrations
were run. Precision was monitored using replicate data points. These replicate
results were averaged and standard deviations and corresponding coefficients of
109
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110
too J
90-
80 i
70-
60-
Uppw Control Limit
Upper Warning Umit
. LOMT Warning Umtt
Lo««r Control Umtt
80 H 1 1 P
-1 1 1 1 1 1 1—
8 12 16 20
—I 1 « «—
24 28 32
Figure B-1. Representative Recovery Data for Phenol In Texas Soil
110
i
100-
90-
60-
70-
60-
SO
40
30
Upp*r Control Umlt
Warning Umtt
WG.
LMMT Warning LMk
Control Unit
•T 1 1 1 1 1 1 « "!"
3 S 7 9 11
13 13 17
Figure B.2. Representative Recovery Data for o-Cresol In Texas Soil
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10
7 »
Sompto !
Figure B-3. Representative Recovery Data for 2,4-Dlchlorophenol In Texas Soil
variation (CV) for each sample pair were calculated. Illustrative data are presented in
Table B-2 for Texas soil. The results indicated close agreement among duplicates.
QA/QC FOR ANALYTICAL INSTRUMENTS
Daily Logs of Instrument Usage
Individual logs were maintained for each major analytical instrument (gas
chromatograph, high pressure liquid chromatography, Microtox®. The log contained
information pertaining to instrument conditions and analyses performed. Unusual
events or circumstances were noted and reported if instrument performance was
affected.
Calibration Procedures
Prior to each day's analysis the instruments were calibrated using known
standards. Both internal and external standards were used depending upon
application. Instrument responses of greater than 10% of that expected for standard
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TABLE B-2
PRECISION ANALYSIS DATA+: TEXAS SOIL SORPTION DATA
CHEMICAL SAMPLE A
(mg/l)
3-Chtorophenol
4-Chtorophenol
2,4-Dichtarophenol
2,5-Dichtorophenol
2,6-Dichlorophenol
3,4-Dichlorophenol
2,4,5-Trichlorophenol
2,4,6-Trichlorophenol
Pentachlorophenol
2,4-Dimethylphenol
2-Methyl,4-Chlorophenol
3--Methyl,4-Chlorophenol
3-Methyl,6-Chlorophenol
p-Nitrophenol
2,4-Din'rtrophenol
4,6-DinJtro-o-Cresol
Thiourea
9.7
5540
2.7
5810
3.9
1180
4.4
1980
16.3
1380
4.3
659
24.7
508
21.8
298
0.8
4.5
33.4
4770
7.2
1730
5.8
1930
72.6
2570
6.3
3520
5.6
445
49.4
112
42.4
4380
SAMPLE B
(mg/l)
10.0
5550
2.7
6250
3.9
1260
5.2
2040
16.8
1480
5.3
659
24.9
508
22.1
298
0.8
4.6
34.0
4950
7.8
1780
5.8
1950
67.8
2680
6.5
3720
6.0
463
52.0
120
43.5
4700
MEAN
(rng/l)
9.9
5545
2.7
6030
3.9
1220
4.8
2010
16.6
1430
4.8
659
24.8
508
22.0
298
0.8
4.6
33.7
4860
7.5
1760
5.8
1940
70.2
2630
6.4
3620
5.8
454
50.7
116
43.0
4540
STANDARD
DEVIATION
(ma/I)
0.21
6.2
0.00
313
0.00
56
0.57
42
0.35
65
0.71
0.00
0.14
0.00
0.21
0.00
0.00
0.07
0.42
133
0.42
37
0.00
11
3.4
75
0.14
144
0.28
12.3
1.84
5.8
0.78
225
COEFF. OF
VARIATION
(%)
2.15
0.11
0.00
5.19
0.00
4.59
11.8
2.14
2.15
4.58
14.7
0.00
0.57
0.00
0.97
0.00
0.00
1.55
1.26
2.75
5.66
2.11
0.00
0.59
4.83
2.85
2.21
3.99
4.88
2.71
3.63
4.97
1.81
4.96
The two values for each chemical represent tow and high concentrations used in the sorption studies.
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solutions required setting new chemical standards and/or making appropriate
adjustments to the instrument. This action was the responsibility of the project analyst.
All reagents for chemical analysis were prepared using analytical reagent
grade (AR) chemicals. For all analyses requiring reagent water, organic free DDW
was used.
Data calculation, manipulaitons and analysis were performed using calculators
and computers. Computer programs used for data reduciton and analysis were
validated before use.
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APPENDIX C
VOLATILIZATION ESTIMATES
The results of the degradation studies (Section 5) represent the overall
chemical loss that occurred in the soil microcosms. While the main loss mechanism
was expected to be microbial degradation, it was possible that chemical degradation,
hydrolysis and volatilization could have contributed to the removals that were
observed. Most of the chemicals evaluated had low vapor pressures and volatilization
losses were assumed to be negligible.
However, several chemicals such as Methanol and 1-Butanol had higher vapor
pressures, and it was possible that appreciable volatilization losses could have
occurred during the degradation studies. To evaluate this possibility, a series of
simple experiments was conducted to estimate the amount of volatilization that might
occur.
These experiments consisted of adding the chemicals to two soil microcosms,
one of which had a constant air sweep to remove any volatilized chemical, and the
other of which did not have any air sweep. The experiments were conducted for two
and twenty-four hours. These times were used because the greatest volatilization
potential occurred when the chemical concentration in the soil was the highest, i.e., at
the beginning of a study. Twenty-four hours was the longest period that could be used
since degradation also would be occurring during the experiments, thus decreasing
the chemical concentration.
Texas soil was used for these experiments. The chemicals were added to the
soil to achieve a concentration of about 1000 mg/kg which was close to but higher
than the concentrations of Methanol and 1-Butanol used in the degradation studies.
The air sweep was held constant at 100 ml/minute during the experiments. The results
are presented in Table C-1.
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TABLE C-1. LOSS OF METHANOL AND 1-BUTANOL
IN VOLATILIZATION EXPERIMENTS
Condition Time Period Chemical Chemical
(hours) Added (mg)+ + Remaining (mg)
Methanol
no air sweep 2 40 25.0
continuous air sweep"1" 2 40 23.5
1-Butanol
no air sweep 2 40 34.2
continuous air sweep"1" 2 40 34.8
no air sweep 24 40 35.2
continuous air sweep"1" 24 40 14.2
* A constant air flow of 100 ml/min was maintained over the surface of the microcosms
throughout the noted time period.
** This amount of chemical was added to 40 grams of soil resulting in an initial soil concentration of 1000 mg/kg.
The data indicate that there was little change due to the air sweep and little
enhanced volatile loss during a two-hour time period. However, over a twenty-four
hour period the continuous air sweep did increase the loss of 1-Butanol.
The driving force for volatilization is related to the concentration gradient at the
boundary layer (soil surface) established by the concentration in the soil and the
concentration in the air layer immediately above the soil. With an air sweep, any
volatile constituents in the air layer above the soil are continuously removed, the
concentration gradient will be maximum, and the volatilization potential will be the
greatest. In the degradation experiments, quiescent conditions were maintained and
no air sweep was used. Under quiescent conditions, if any volatilization occurs, the
concentration above the soil builds up, the concentration gradient is less, the
volatilization potential is the least, and the volatile flux from the soil is suppressed.
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For this reason, even for chemicals in this study with a high vapor pressure, little
volatilization was expected under the conditions used for the degradation
experiments.
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APPENDIX D
PUBLICATIONS
In addition to this report, a technical report and several technical articles, two
M.S. theses, one Ph.D. dissertation and several presentations have resulted from this
research. These are:
TECHNICAL REPORT
One of the objectives of this research effort was an assessment of the terrestrial
and aquatic bioaccumulation that could result by the chemicals evaluated in this study.
The following draft report was submitted to the EPA project officer, Mr. John E.
Matthews:
Loehr, R.C. and R. Krishnamoorthy, "Bioaccumulation Potential of Designated
Hazardous Organic Chemicals", July 1987.
The report was reviewed and accepted by the project officer and therefore is not
included in this final report.
TECHNICAL ARTICLES
Loehr, R.C. and R. Krishnamoorthy, "Terrestrial Bioaccumulation Potential of Phenolic
Compounds", Hazardous Waste and Hazardous Materials 5:109-120 (1988).
Nam-Koong, W., Loehr, R.C. and J.F. Malina, Jr., "Kinetics of Phenolic Compounds in
Soil", Hazardous Waste and Hazardous Materials 4:321-328 (1988).
M.S. THESES
•
Yoon, C.G., "Multisolute Adsorption of Toxic Organic Compounds Onto Soil", The
University of Texas at Austin, May 1988.
Dasappa, S.M., "Detoxification and Immobilization of Chlorophenols in Soil", The
University of Texas at Austin, September 1988.
PH.D. DISSERTATION
Nam-Koong, W., "Removal of Phenolic Compounds in Soil", The University of Texas at
Austin, May 1988.
PRESENTATIONS
Nam-Koong, W., Loehr, R.C. and J.F. Malina, Jr., "Removal of Phenolic Compounds in
Soils", Texas Water Pollution Control Association Conference, Corpus Christi,
Texas, June 1987.
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Dasappa, S.M. and R.C. Loehr, "Detoxification and Immobilization of Chlorophenols in
Soil", Texas Water Pollution Control Association Conference, Houston, Texas,
June 1988.
Nam-Koong, W., Loehr, R.C., and J.F. Malina, Jr., "Removal of Phenolic Compounds in
Soil", Joint CSCE-ASCE National Conference on Environmental Engineering,
Vancouver, BC, Canada, July 1988.
Dasappa, S.M. and R.C. Loehr, "Detoxification and Immobilization of Chlorophenols in
Soil", 61st Annual Conference, Water Pollution Control Federation, Dallas,
Texas, October 1988.
Nam-Koong, W., Loehr, R.C. and J.F. Malina, Jr., "Effects of Mixture and Acclimatio' n
Removal Rate of Phenolic Compounds in Soil", 61st Annual Conference, v sr
Pollution Control Federation, Dallas, Texas, October 1988.
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